r LT) icr I r-R i O JHemoirs from the fitolociical Caboratorj) OF THE JOHNS HOPKINS UNIVERSITY II THE GENUS SALPA A MONOGRAPH WITH FIFTY-SEVEN PLATES BY WILLIAM K. BROOKS, PH. D., LL. D. Professor in the Johns ffopJcins University and Director of its Marine Laboratory WITH A SUPPLEMENTARY PAPER BY MAYNARD M. METCALF Fellow of the Johns Hopkins University BALTIMORE THE JOHNS HOPKINS PRESS 1893 THE KRIEDENWALD CO., PRINTERS, BALTIMORE. \ CONTENTS. PART I. A GENERAL ACCOUNT OP THE LIFE HISTORY OP SALPA. CHAPTER I. INTRODUCTORY, PACK 1 CHAPTER II. THE DEVELOPMENT OF THE SOLI- TARY SALPA FROM THE EGG, .... SECTION 1. An Outline of the History of the Egg, . SECTION 2. The Foetal Membranes, .... SECTION 3. The Migration of the Follicle, SECTION 4. The Organs of the Embryo outlined in Follicle Cells, . SECTION 5. TheBlastodermicTissuesoftheEmbryo, SECTION 6. The Degeneration of the Follicle, . SECTION 7. The Placenta, SECTION 8. The Nutrition of the Embryo, CHAPTER III. THE MORPHOLOGICAL SIGNIFI- CANCE OF THE SALPA EMBRYO, The Embryology of Primitive Tunicates, . Has the Egg of Salpa passed through a Stage with a large Food Yolk '! The Primitive Salpa Embryo, The Origin and Significance of the Follicle of Salpa, CHAPTER IV. THE ORIGIN OF THE PROLIFEROUS STOLON, SECTION 1. Outline Sketch, SECTIONS. The Orientation of the Stolon, SECTION 3. The Ectoderm of the Stolon, 17 17 21 24. 28 32 42 46 48 54 55 56 58 59 00 6C 68 68 SECTION SECTION SECTION SECTION SECTION SECTION The Nerve Tube, 70 The Endodermal Tube, .... 71 The Blood Tubes, 73 The Perlthoracic Tubes, . . . 75 The Mesoderm of the Stolon, . . 7ti The Genital Rod, 76 SECTION 10. The Derivatives from the Parts of the Stolon, 76 CHAPTER V. THE TRANSFORMATION OF THE STOLON INTO THE SERIES OF AGGREGATED SALP.E, 78 SECTION 1. Outline Sketch, 78 The Proliferous Stolon, 78 The Segmentation of the Stolon, .... 78 The Rudimentary Chain Salpa, .... 73 The Secondary Changes, 79 The Development of the Chain Salpa, ... 80 SECTION 2. The General Characteristics of Aggre- gated Salpa. 1 , S4 SECTION 3. The Segmentation of the Stolon, . . s APPENDIX I, 363 APPENDIX 11, 364 APPENDIX III, 367 LIST OF PAPERS KEFEKKED TO IN PART IV, . . 371 PART V. EXPLANATION OP THE PLATES. PART ONE. A GENERAL ACCOUNT OF THE LIFE HISTORY OF SALPA. CHAPTERS I, II, III, IV, AND V. CHAPTER I. INTRODUCTORY. Most of the material for the researches on which this memoir is based was gathered and preserved at sea, at various points upon the eastern and western coast of the United States, by the vessels of the United States Fish Commission, under the direction of Marshall M'Donald, the Com- missioner of Fisheries. As the schooner Grampus was engaged during the summer of 1888 in work which gave, incidentally, an excellent opportunity for pelagic exploration, the commissioner invited me to avail myself of the advant- age, and I owe to him the pleasure of a cruise in the Gulf Stream under conditions which were most favorable for studying its floating fauna. I have also received from the commissioner, from time to time, col- lections of salpoe which have been gathered, under his direction, at various localities near our coast. These specimens have been preserved for microscopic research, according to approved methods, by Professor Wm. Libbey, Professor L. A. Lee and Mr. K. P. Bigelow, and I am indebted to these gentlemen for many valuable additions to my collection. My thanks are also due to the National Academy of Sciences for a grant of money from the "Bache" fund, to aid me in the prosecution of this research ; and to the Trustees of the Johns Hopkins University for the opportunity to study living salpae at many points upon our coast, and for the provision which they have made, from the funds of the Uni- versity, for the publication of this memoir. As I enter upon the preparation of this account of a research which has been, up to this point, an unfailing source of pleasant interest, I am confronted by the disagreeable prospect of inevitable controversy ; and all my pleasure is destroyed by the thought that nearly every one of my statements will contradict the published statements of some one among the numerous writers on the subject. 2 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. Scientific controversy is so unprofitable that I shall try to make it as subordinate as possible, that the reader may devote all his attention to the life-history of salpa without interruption at every point where my own observations confirm or contradict the statements of others. The story of the life-history of salpa is most interesting, but it is complicated and difficult to tell in simple words, even after it has been stripped- of all which is not essential, and I am sure that I shall promote the interest of my readers by strict adherence to the unity of my descrip- tion. I shall therefore give, in the first place, a continuous, uninterrupted account of the subject, and shall reserve the discussion of disputed points for later chapters. I know that I have built upon a foundation which has been laid by others, and that many of the facts have been the common property of naturalists for many years, but truth and error are so closely bound together in the literature of the subject that it is not possible to give to each author the credit which is his due without entering into adverse criticism at the same time, and as the history of our knowledge of Salpa has been reviewed over and over again, it does not seem necessary to enter into it here. The home of the salpas is so remote from common observation that few persons except naturalists have a true conception of their scientific interest, or of their importance in the economy of the sea, although they occupy a prominent place in the mind of every one who has enjoyed the pleasure of studying the floating fauna of the open ocean, and is thus enabled to call up a mental image of pelagic life as a whole. The older naturalists who explored the ocean in sailing vessels, when calms gave leisure for studying its wonders, found in them an unfailing subject for fascinating and delightful research. They are seldom found near the shore, however, and as they are so transparent that they are scarcely visible from the deck of a modern steamship, they are little more than a name to most of the naturalists of our own day; but to the student, even the name, salpa, itself calls up a long list of famous naturalists and explorers. Among them is the friend of Linnaeus, Peter Forskal, who lost his life while exploring Arabia in 1774. His description of the animals which he had observed in his journey to the East was edited and pub- lished by his fellow-explorer, Niebuhr ; and in it eleven forms of salpae, which he had observed and studied in his voyage through the Mediter- W. K. BROOKS ON THE GENUS SALPA. 3 ranean, are so faithfully described that most of them can still be iden- tified, and continue to bear the names which he gave them. He also proposes for them the generic name salpa, and gives the first satisfactory definition of them. We think also of Chamisso, who, in 1814, left his romances and his poetry to go round the world with Kotzebue, and who found in the life- history of salpa a story more strange and wonderful than that of Peter Schlemihl ; of Quoy and Gaimard, who, after many years of wandering among the islands of the Pacific with Freycinet and with Dumont d'Ur- ville, enriched our literature with the zoological treasures of the "Voyage autour du monde" and the "Voyage de la corvette 1' Astrolabe." These and many other famous names are associated with the zoology of salpa, but the history of the literature of the subject has been reviewed so many times that we cannot dwell upon it, and the reader may refer to the bibliography which is given by Traustedt in his Spolia Atlantica, (Vidensk. Selsk. Skr., 6th Raekke 2, viii, Kjobenhavn, 1885, pp. 341-346,) or to the historical sketch and bibliography which are given by Herdman in the introductory chapters to his various reports on the Tunicata of the Challenger Expedition, (Report upon the Tunicata collected during the Voyage of H. M. S. Challenger during the years 1873-76). I shall not duplicate these lists, but since there are a few papers to which I shall have to make frequent references, I give them here in a numbered series, and I shall refer to them hereafter by number. 1. LEUCKAKT, Dr. RUDOLF. Zur Anatoraie und Entwicklungsgeschichte der Tuni- caten. Zoologische Untersuchungen 2. Giessen, 1854. 2. SALENSKY, Dr. W. Ueber die embryonale Entwicklungsgeschichte der Salpen. Zeit. f. wiss. Zool. XXVII, pp. 179-237. Plates XIV-XVI. 3. SALENSKY, Prof. W. Ueber die Entwicklung der Hoden und iiber den Genera- tionswechsel der Salpen. Zeit. f. wiss. Zool. XXX, Supplement, pp. 375-393. Taf. XIII. 1878. 4. BARROIS, Dr. J. Memoire sur les membranes embryonnaires des Salpes. Journ. de 1'Anat. et de la Phys., pp. 455-498. Plates XVII and XVIII. 1881. 5. SALEKSKY, Prof. W. Neue Untersuchungen iiber die embryonale Entwicklung der Salpen. Mittheilungen aus der Zoologischen Station zu Neapel, I, pp. 90-171 and 327-402. Plates VI-XVII and XXII-XXVII. 1882. 6. KOWALEVSKY, A., and BARROIS, J. Materiaux pour servir a 1'histoire de 1'An- chinie. Journ. Anat. Phys. XIX, 1883, pp. 1-23. Plates I-III. 7. ULJANIN, Dr. BASILIUS. DieArten der Gattung Doliolum im Golf e von Neapel. Fauna und Flora des Golfes von Neapel, X, 1-140, with twelve plates. 1884. 8. TRAUSTEDT, M. P. A. Spolia Atlantica. Bidrag til Kundskab von Salperne. Mem. Acad. Royal Copenhagen, 6, II, 8. 1885. Ten plates. 4 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. 9. SEELIGER, Dr. OSWALD. Die Entwicklungsgeschichte der socialen Ascidien. Jenaischen Zeitschr. XVIII, N. Z. XI, 1-145. Taf. I-VIII. 1885. 10. BARROIS, Dr. JULES. Recherches sur le cycle genetique et le bourgeonnement de 1'Anchinie. Journ. Anat. Phys. XXI, 1885, pp. 193-207. Plates VIII-XII. 11. SEELIGER, Dr. OSWALD. Die Knospung der Salpen. Jenaische Zeitschr. XIX, 573-677. Plates X-XIX. 1886. 12. BROOKS, W. K. The Anatomy and Development of the Salpa-Chain. Studies Biol. Lab. Johns Hopkins Univ. 1886, pp. 451-487. Plates XXVIII, XXIX. 13. SEELIGER, Dr. OSWALD. Die Entstehung des Generationswechsels der Salpen. Jenaische Zeitschr. XXII, 398-414. 1888. - 14. HERDMAN, Dr. W. A. Report upon the Tunicata collected during the Voyage of H. M. S. Challenger during the Years 1873-7G. Part III. Report on the Scientific Results of the Voyage of H. M. S. Challenger. Vol. XXVII, Part LXXVI. 1888. 15. SEELIGER, OSWALD. Zur Entwicklungsgeschichte der Pyrosoma. Jenaische Zeitschr., pp. 595-658. Plates XXX-XXXVII. 1889. 16. DAVIDOFF, Dr. M. v. Unters. zur Ent. der Distaplia magnilarva Delia Valle. Mittheilungen aus der Zool. Station zu Neapel, IX, 113-178 and 533-562. Plates V, VI and XVIII-XXIV. 1 889-91 . 17. SALENSKY, Prof. W. Beitriige zur Embryonalentwickelung der Pyrosoma. Zool. Jahrbiicher, Band IV, 424-478. Taf. XXVI-XXVIII ; Band V, 1-98. Taf. I-VIII. 1891. 18. D'KOROTNEF, Dr. ALEXIS. La Dolchinia mirabilis. Mittheilungen aus der Zool. Station zu Neapel, X, 2, 1891, pp. 187-205. Salpa is a transparent, swimming Tunicate, and its body, Plates III and IV, is, in effect, an enormous pharynx which swims through the water, gulping in great mouthfuls at each contraction of its muscles. In shape it is subcylindrical, and its body may be compared to a barrel open at both ends, so that water flows through it without obstruc- tion. The mouth, r, occupies the anterior end of the barrel, and the lips are infolded in such a way that they act as valves which permit the water to enter between them while they prevent it from escaping, while, at the opposite end of the body, the atrial aperture, g v , affords an exit for it. Practically the chamber of the barrel is uninterrupted from one opening to the other, for while it is divided, morphologically, into the pharynx, c, and the atrium, g'", these two chambers are separated from each other only by the rod-like "gill," o, which traverses the cavity on the middle line. The " gill " is so narrow that it offers little obstruction to the water, and there is a free passage on each side of it. The body is encircled, more or less completely, by muscles which are placed like barrel-hoops, so that their contraction empties the barrel and drives the water out through the atrial aperture, and thus propels the W. K. BROOKS ON THE GENUS SALPA. 5 floating animal through the water in the opposite direction. Around the mouth the muscles are so placed that they close the lips and prevent the water from escaping in this direction when the barrel contracts. The body is inclosed in a thick outer mantle, Plates I and II, which by its elasticity antagonizes the muscles, and expands the barrel and draws in a fresh mouthful of water. In most species the atrial aperture, Plate I, Fig. 3, is encircled by sphincter muscles which constrict it and prevent the water from entering during expansion, so that the animal moves forwards, by jerks, along a column of water which passes through its body. The food of salpa consists of radiolarians, diatomes, and other micro- organisms which float in the water, and as these lodge on the inner sur- face of the barrel they are gathered up and swept through the oesophagus into the stomach in a way which will be described soon. The supply of this food is unlimited, and salpae are often found swarming at the sur- face of the ocean in number beyond description. In size they range from 6 mm. or about one-fourth of an inch, the average length of the aggregated form of Salpa democratica, Plate II, Fig. 1, to about 20 cm. or eight inches, the length of a large specimen of the solitary form of Salpa costata, Plate IV, Fig. 4. Although they are met with in the greatest variety and abundance in the warmer parts of the ocean, they are by no means confined to the tropics, and they have been found in great numbers north of Norway and Scotland and south of Cape Horn and the most southern points of Australia and New Zealand. They are abundant only after the water has been for some time undisturbed by winds; and as prolonged calms are most frequent in warm seas, these waters are most favorable for the development of these animals, which multiply with astonishing rapidity. The smaller species are often so abundant that for hundreds of miles any bucketful of water dipped up at random will be found to contain hundreds of them. In such places collecting with the surface-net becomes impracticable, for almost as soon as the net is dropped into the water it becomes choked with a mass of salpse so that nothing more can enter it. A drop from an organic infusion swarming with infusoria, seen under a low power of the microscope, bears some resemblance to the surface of the ocean when salpae are abundant, except that the water is not turbid like the infusion, but beautifully clear and transparent. 6 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. No one who has not seen these animals under favorable conditions can form any conception of the amount of animal life which pure sea- water is able to support ; and the salpae are able to multiply with great rapidity, both sexually and asexually, in order to avail themselves quickly of favorable conditions. Each species of salpa has two generations in its life-cycle, known as the solitary generation and the aggregated generation. The solitary salpa is born from an egg which is carried within the body of the aggregated salpa, whose blood nourishes the embryo during its development by means of a nutritive placenta, Plate III, Fig. 4, and Plate XXXV. The aggregated salpa3 are produced asexually from the body of the solitary salpa. Plate II shows the solitary form of Salpa democratica, and Plate XLIII, Fig. 1, shows the aggregated form of the same species. In Plate I, the solitary form of Salpa pinnata is sho"wn in dorsal view in Fig. 5, and in ventral view in Fig. 6. The aggregated community of the same species is shown in Fig. 2, and one of the members of the community in Fig. 3. Figs. 4 and 7 show the dorsal and ventral aspects of the solitary form of Salpa chamissonis, and the aggregated cormus is shown in Plate XLI, Fig. 9, and one of its members in Plate VIII, Fig. 6. The solitary form of Salpa cordiformis is shown in Plate IV, Figs. 3 and 4, and a part of a "chain" of the same species in Plate IV, Fig. 6. Plate IV, Fig. 7, is the embryo of the solitary form of Salpa scutigera, and Plate IV, Fig. 1, part of its "chain," and so on. In most species each aggregated salpa carries only one egg, so that the solitary generation consists of only one individual; but in all the species the aggregated generation consists of many hundred individuals, and there is reason to believe that it has no fixed limit, but that the solitary salpa may continue to produce aggregated salpae for an indefinite time and in unlimited numbers. The aggregated salpa3 are born in sets or cormi. In the chain-like cormi the number of individuals is usually more than a hundred, but in circular cormi it is very much smaller, and each cormus of Salpa pinnata, Plate I, Fig. 2, contains only eight or nine. In a few species, each aggregated salpa contains more than one egg, but the number is small, and seems to be fixed and constant for the species. In Salpa cordiformis, for example, there are five eggs, and each chain-salpa gives birth to five embryos, Plate III, Figs. 2 and 3, em. Salpa hexagona, Plate X, Fig. 10, also produces five embryos. W. K. BROOKS ON THE GENUS SALPA. 7 The solitary salpa of each species differs from the aggregated form of the same species in many of the details of its structure, and in many cases the difference is very considerable, but the fundamental plan is the same for all. The specific characteristics of each solitary salpa are quite different from those of its corresponding aggregated form, so that there is no way of deciding what specimens of the two forms belong together, except by actually rearing them, or by the discovery in the body of a given chain-salpa of an embryo sufficiently advanced in development to exhibit its adult characteristics ; or else by finding them associated in great numbers free from admixture with other species. The two forms, when mature, are nearly equal in size, and this is sometimes an aid in identi- fying them. The divergent modification which has produced the various species has affected the two forms of each species in different ways, but to about the same amount, so that we cannot say that the solitary salpae of different species are less or more differentiated than the aggregated salpae. This fact indicates clearly that the separation of the life-cycle into the two generations took place before the species diverged from their common ancestor. The general features of the structure of a solitary salpa are well shown in the longitudinal vertical section of an embryo of Salpa pinnata in Plate XXXV, and in the horizontal sections in Plate XIX, and in the embryos of this species in Plate XLI, Figs. 1, 2, 3 and 4, and in the embryo of Salpa hexagona in Plate III, Fig. 4. The structure of the aggregated salpa is shown in Plate VIII, where Fig. 1 shows two young aggregated salpa? of Salpa pinnata, and Fig. 2 two of Salpa cylindrica. As the figure of the solitary Salpa africana in Plate IV, Fig. 2, shows, the body is subcylindrical in shape, and the two orifices, the mouth, r, and the atrial aperture, gr v , which are usually close together in sessile tunicates, are widely separated and are nearly or quite at opposite ends of the body. In the solitary salpa? of all species, and in nearly all the aggregated salpae, the mouth is terminal, or at one end of the cylinder. In a few aggregated forms, Salpa cordiformis, for example, Plate IV, Fig. 6, it is not at the extreme end, but on the outer surface. Figure 8 of Plate XLV is a vertical section through the oral region of the embryo of Salpa pinnata just before the mouth is formed. The figure 8 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. shows that the upper lip is rounded and protuberant and thick, while the lower lip is very thin. In mature specimens this thin lower lip folds inwards so that the thickened upper lip shuts closely upon it when the mouth is closed. The flexible edge of the lower lip acts as a valve, for it is depressed by the inflowing current of water, while back pressure folds it up against the upper lip and closes the mouth. In many species there are two short muscles on the upper surface of the upper lip to open it. They lie near the middle line, and they are parallel or nearly parallel to each other. They are shown in Plate II. The muscles for closing the mouth are much more complicated, as many of the figures show : Plate IV, Fig. 2, for example. Their arrange- ment in the solitary salpa is usually different from that in the aggre- gated form, and they also vary according to the species, but in most cases some approximation to the arrangement shown in Plate XLI, Figs. 3 and 5, can be made out, although the oral musculature is seldom as well developed as it is in these figures, which represent solitary embryos of Salpa pinnata. The mouth is not yet open in these embryos, but r marks its position. A bridle-like muscle arises from about the middle of the second body-muscle, the body-muscle which crosses the upper surface of the body just behind the ganglion, s, and runs across the oral end on what is to become the angle of the lower lip, and, crossing the middle line, unites to the middle of the second body-muscle on the opposite side. At its lower end this body-muscle joins a muscle which runs upwards and forwards to the angle of the mouth, where it divides into three branches, two superior ones which cross the middle line on the convex surface of the upper lip, and one inferior one which crosses the middle line just below the mouth. These muscles are shown in section in Plate XLV, Fig. 8. In adult salpa3 the oral muscles are usually quite distinct from the body muscles in their anatomical relations, and very much smaller; but these embryos show that they belong to the system of body-muscles, or perhaps it is more near the truth to say that the body-muscles of salpa are modified oral and atrial sphincters. The body-muscles exhibit the greatest diversity of arrangement, as may be seen by comparing the more divergent forms, such as Salpa hexagona, Plate III, Fig. 4 ; Salpa scutigera, Plate IV, Fig. 7, and Salpa costata. Fig. 4. It is hard to say what number of muscle-bands is most characteristic ; nine seems to be more common than any other number, but in some species, Salpa scutigera for example, they are reduced to four, while in W. K. BROOKS ON THE GENUS SALPA. 9 Salpa costata there are twenty. They may be continuous, as in Salpa pinnata, Plate XLI, Fig. 5, or interrupted as in Salpa costata, Plate IV, Fig. 4. They may be independent as in the last species, or united in sets as in Salpa democratica, Plate I. They may be restricted to the dorsal surface of the body, as in Salpa africana, Plate IV, Fig. 2, or they may be completely closed rings passing entirely around the body like the hoops around a barrel, as in the solitary form of Salpa democratica, Plate II. Leuckart says (I, p. 15) that while the gap may be so short that it is not visible without a microscope, they are always interrupted on the middle line ; but I am at a loss to find the basis for this statement, for the fibers may be traced across the middle line, without any break, in many species. He also says, p. 16, that as they never cross the ventral middle line they are never complete rings, and this statement has been repeated over and over again until it has found its way into all the text- books and scientific memoirs, notwithstanding the fact that most of them are completely closed, dorsally and ventrally, in the most familiar and abundant species, the solitary Salpa democratica, Plate II. This error has been most persistent, and it has been made the basis of the fundamental definition of the whole Salpa family, for which Glaus has proposed the name Desmomyaria, and Herdman the name Hemi- myaria, to distinguish them from the Doliolums, for which Gegenbaur has proposed the family name Cyclomyaria. Even if this difference between Salpa and Doliolum were absolute, the selection of a characteristic so very variable as the form of the loco- motor muscles as a basis for fundamental classification would be most unwise ; and, as a matter of fact, some of the muscle-bands of Doliolum are incomplete, and some of them, in at least one species of Salpa, com- plete. In the first generation or "Amme" of Doliolum the seventh body- muscle is incomplete dorsally, and in the median "Pflegethiere" it is incomplete ventrally, while the lateral buds or "Ernahrungthiere" depart very widely from the cyclomyarian type. So far as I am aware, Traustedt is the only modern writer on Salpa who has described the muscle-bands of Salpa democratica as complete circles. In his description of this species, p. 365, and also in his descrip- tion of the variety flagellif era, p. 369, he states the facts correctly, but while his draughtsman, Cordts, has figured Salpa democratica correctly, Plate II, Figs. 25 and 26, he has followed tradition in his figure of Salpa flagelli- fera, Plate I, Fig. 12, rather than nature and Traustedt, and has drawn 10 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. all the muscle-bands as interrupted ventrally; so hard is it to correct established error. The atrial aperture is sometimes valvular, as, for example, in the aggregated form of Salpa cordiformis, Plate III, Fig. 2, g v , but it is usually a simple opening, circular or nearly so, without valves. Its mode of origin in the embryo of Salpa pinnata is shown in Plate XVII, Figs. G and 7. Quite frequently it is produced into a projecting funnel, as in Salpa costata, Plate IV, Fig. 4, g v . It may be terminal at the posterior end of the body, as in Salpa pinnata, Plate I, Figs. 1, 3, 5 and 6, or it may be on the upper surface of the body at some distance from the posterior end, as in Salpa democratica, Plate II and Plate XLIII, Fig. 1. There is little uniformity in its position. In Salpa democratica it is on the upper sur- face in both the solitary form and the aggregated form. In Salpa pin- nata it is terminal in both forms, as it is also in Salpa costata. In the solitary form of Salpa cylindrica, Plate III, Fig. 5, it is terminal, while it is on the upper surface of the aggregated form, Fig. 6, while the reverse of this is the case in Salpa hexagona, Figs. 4 and 1. The atrial aperture is usually encircled by sphincter muscles, as in Salpa cylindrica, Plate III, Fig. 5. These muscles are usually a continua- tion of the series of body-muscles, which gradually become narrow and crowded together as they approach the cloacal aperture. In these cases the atrial sphincters are often complete rings. In a few species they have a more complicated arrangement, as in Salpa costata, Plate IV, Fig. 4 and Plate VIII, Fig. 4, where they branch out in a dendritic man- ner from two lateral longitudinal trunks. The chain-form of Salpa cor- diformis, Plate IV, Fig. 6, has the same arrangement in a rudimentary or slightly developed condition. Often there is a lack of bilateral sym- metry in the posterior body-muscles of aggregated salpae. In the aggre- gated form of Salpa costata, Plate VIII, Fig. 4, the branches of one lateral trunk are very slightly developed, while those from the other almost encircle the atrial funnel. The last body-muscle of the aggregated form of Salpa cordiformis, Plate III, Fig. 2, is forked on the right side but not on the left. As Leuckart pointed out long ago (I, p. 6), the mouth and the atrial aperture are very much nearer each other in the young salpa than they are in the adult, as examination of my figures will show. Figure 2 of Plate XLI shows a young embryo of Salpa pinnata with the point where the mouth is to be formed marked r, and the position of the atrial aperture marked g v . It will be seen that the interval between W. K. BROOKS ON THE GENUS SALPA. 11 these two points is considerably less than a fourth of the circumference of the body. In Fig. 3, and in Plate XXXV, they are a little further apart ; still further apart in Fig. 5, while in the adult, Plate I, Fig. 4, they are at opposite ends of the long axis of the body. The history is the same in the aggregated form, as is shown by the series of figures of the aggre- gated Salpa pinnata in Plate VII, Figs. 4 and 5; Plate VIII, Fig. 1, and Plate I, Fig. 3. The test of salpa has never received the attention it merits. Most of the figures and descriptions of the species are from preserved specimens, and as the muscles are made opaque and distinct by alcohol, while the test remains transparent, and usually becomes more or less swollen and indefinite by the action of preserving fluids, the published descriptions contain very few references to the fact that in many species the surface of the body is delicately sculptured and is marked by ridges and serrations. In this particular most of the figures are untrue to nature, and as I have attempted to show in my figures the structure of the test, many of my illustrations of familiar species are so different from those which usually pass as correct drawings that I feel called upon to explain the dis- crepancy. In many species the test is divided by longitudinal ridges or keels into thick and thin portions, and the ridges are sometimes simple and sometimes serrated. Their physiological function is undoubtedly to give strength and stiffness to the test, that it may antagonize the muscles more effectively, and restore the shape of the body after contraction. While the different species which possess these ridges exhibit con- siderable variation, there is a general plan which can always be recog- nized. On the middle line of the dorsal surface the test is thin, and when the muscles are contracted, as they usually are in preserved specimens, this thin area, Plate III, Figs. 1, 2, 3 and 5, forms a deep longitudinal furrow, bounded on each side by the prominent, keel-like edge of the thickened portion of the test. Along each side of the body there is usually a dorso-lateral keel, and the thickened portion of the test often forms a prominent "wing" on each side of the body between this keel and the one nearer the middle line, as is shown in Fig. 5 and on the left side of Fig. 1. On the sides of the body the test is thin, and sometimes strengthened by a lateral ridge on each side. On the ventral surface the test is thick, and is bounded at the sides by two ventro-lateral keels, Fig. 7, while there is often a median keel on the middle line of the ventral surface. I have found these ridges in the solitary form of Salpa demo- 12 JOHNS HOPKINS UNIVEKSITY MORPHOLOGICAL MONOGRAPHS. cratica, Plate II ; in both the aggregated form of Salpa hexagona, Plate III, Fig. 1, and the solitary form ; in both the aggregated form of Salpa cor- diformis, Plate III, Figs. 2 and 3, and the solitary form, Plate IV, Fig. 5 ; in the solitary form of Salpa cylindrica, Plate III, Figs. 5 and 7 ; in the aggregated form of Salpa runcinata, Plate XLIII, Fig. 2, and in the solitary form, Plate XLIII, Fig. 3; and in the solitary form of Salpa africana, Plate IV, Fig. 2, and the careful study of living specimens will undoubtedly show that they exist in most of the species. The aggregated form of Salpa runcinata is shown in Plate XLIII, Fig. 3. It has the two serrated ridges on the upper surface, running from the posterior end of the body to the region of the mouth, and it has three more ridges on the lower surface. The lower surface of the solitary form of this species, Plate XLIII, Fig. 2, is so highly ornamented that I should be almost disposed to regard it as a new species, if it did not agree in all other respects with the published descriptions, and had I not found markings of the same sort in so many other species. The arrangement of the serrated ridges in this species is so much like those of Herdman's Salpa echinata that I am almost disposed to believe that this species is a Salpa runcinata. Except for a slight difference in the muscles its internal structure is like that of Salpa runcinata, and I have found great variation in the arrangement of the muscles in all the species of the run- cinata group. I have not found the ridges in my specimens of the aggregated form of Salpa cylindrica, Plate III, Fig. 6, but they are well developed in the solitary form and, except that they are not serrated, they are almost exactly as they are in Salpa runcinata. They are shown on the upper surface in Fig. 5, and on the lower surface in Fig. 7. The digestive organs of Salpa consist of the pharynx, c, which opens externally through the mouth, r, and communicates through the oeso- phagus, Plate VIII, Fig. 2, q, with the stomach, p, from which the intes- tine, p', runs to the anal orifice, p", by which the intestine opens into the atrium, g'". In most species of Salpa the digestive organs, with their accessory glands, and in the aggregated form the testis also, are bound together into a compact "nucleus," Plate IV, Fig. 2, which is so solid and opaque in the adult that its structure can be studied only by sections. The arrangement of the digestive organs is essentially like Fig. 2 of Plate VIII, however, and those of the solitary salpa are usually like those of the aggregated salpa, W. K. BROOKS ON THE GENUS SALPA. 13 The oesophagus, beginning a little to the right of the base of the "gill," o, runs towards the upper surface of the body to open into the stomach which communicates with one or two blind diverticula. The intestine arises on the lower side of the stomach, and, describing a curve, passes to the left of the oesophagus, and opens into the median atrium at or a little to the left of the middle line above the base of the "gill." In the species of the Pinnata group there is no compact "nucleus," and the anatomy of the digestive organs is quite different from that of the other species, and there is great difference between those of the soli- tary and those of the aggregated form. In the solitary Salpa pinnata, Plate I, Pig. 5, the solitary Sal pa cha- missonis, Plate I, Fig. 4, the solitary Salpa affinis, and probably the solitary Salpa dolichosema, the intestine, Plate XXXV, p, runs through the gill, and the anus is at the extreme anterior end of the median atrium. In the aggregated Salpa affinis and Salpa chamissonis, Plate VIII, Fig. 6, the digestive tract is coiled upon itself much as it is in ordinary salpae, except that the coils are not bound together into a com- pact nucleus ; but in Salpa pinnata the intestine of the aggregated form is nearly ventral in position, Plate VIII, Fig. 1, p, and the anus is far forward and close to, but on the left of the middle line. The history of its development in this species shows that when it first makes its appear- ance the digestive tract of the aggregated form is like that of ordinary salpae, and that it has the position which is shown in Salpa cylindrica, in Fig. 2, with the intestine, p, crossing to the left of the oesophagus, q, and running towards the dorsal surface to open into the cloaca above the base of the gill. As the aggregated Salpa pinnata grows, the intestine and anus move downwards along the left side of the body, and at the stage shown in Plate VII, Fig. 5, the oesophagus and intestine lie at the same level. The left-hand salpa in this figure has its right side towards the observer, and the oesophagus, q, is seen to run from the pharynx to the stomach, p', while on the left side of the right-hand salpa the intes- tine, p, and anus, p", are shown in almost exactly the same position. At an older stage, Plate VIII, Fig. 1, the intestine, p, seems in surface view to be on the middle line, but sections, Plate XXXVIII, Figs. 52 and 80, p, show that it is actually to the left of the middle line, although very close to it. In view of its history in Salpa pinnata, I think there can be no doubt that the primitive position of the digestive tract in all species of salpa is like that which is shown in Plate VIII, Fig. 2, and that, in this particular, the pinnata-like species have undergone secondary modification. 14 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. The microscopic organisms which form the food of salpa are gathered up and conveyed into the stomach by means of an apparatus which has been described so frequently that only a very brief account of it need be given, although its prominence during the development of salpa demands some acquaintance with it in order to render the account of the life-history intelligible. On the middle line of the ventral surface of the pharynx there is a longitudinal furrow, Plate XLV, Fig. 5, D, bounded by two thickened borders. This structure is the endostyle. It is shown at end in Plate IV, Fig. 2, and in many of the other figures. In the adult some of the cells of its walls are glandular, secreting an adhesive substance, while other cells carry cilia, which are so placed as to slide the adhesive excretion along the endostylic furrow to the anterior end of the body, where the endostyle ends between two ciliated bands, c&, one of which lies on each side of the inner surface of the pharynx just inside the mouth. Each ciliated band consists of two parallel rows of cilia close together, and by their activity the adhesive matter from the endostyle is drawn upwards along the sides of the pharynx, in fine threads, which stick to and entangle all the organisms which touch them as they are swept in by the current of water which passes through the pharynx. The pharynx, Plate IV, Fig. 2, c, and the atrium, g'", are in free com- munication with each other, as already noted, except 011 the middle line, where they are separated from each other by the "gill," o. In a median longitudinal section, like the one shown in Plate XXXV, the outline of each chamber is well marked. In a young embryo the "gill" is nearly horizontal, and the atrium is above the pharynx, but as the animal grows up the "gill" becomes more and more inclined, until in the adult, Plate IV, Fig. 2, its posterior end approaches the ventral side of the body, and the pharynx and atrium lie end to end ; the former extending farthest backwards on the ventral middle line, while the latter extends farthest forwards on the dorsal middle line. On each side of the "gill" the two chambers communicate with each other, and I have not been able to find in the adult any indication whatever, in their side walls, of the line where the one chamber ends and the other begins. In front of the ganglion, s, the pharynx occupies the whole cavity, as does the atrium behind the nucleus, nu, and the imaginary line where they meet is probably inclined, like the "gill," so that the cavity of the pharynx diminishes in size as we pass backwards, while the atrium increases at the same rate. In Salpa costata, where the nucleus is a considerable distance in front of the atrial aperture, the tubular portion of the atrium is very long. W. K. BEOOKS ON THE GENUS SALPA. 15 The "gill" of salpa is a respiratory organ, and a true gill in the physiological sense, but it is not homologous with the structures which in ordinary tunicates are called gills. This name is usually applied to the clefts or slits in the sides of the pharynx by which this communicates with the lateral atria or peribranchial spaces. In salpa, as the life-history shows, there is one enormous gill-slit on each side of the body, and the "gill" is simply the portion of the body cavity which lies on the middle line between the pharynx in front and below, and the atrium above and behind, while its sides are the inner edges of the two gill-slits. It therefore corresponds, as Herdman (p. 56) has pointed out, to the structure which in ordinary ascidians he has called the dorsal lamina. I shall give further on my reasons for believing that the gill-slit on each side of the pharynx of salpa has actually arisen by the coalescence of all the gill-slits of an ancestor which had a pharynx like that of ordinary ascidians. At present, however, it has lost all traces of this history, even in the embryo. Herdman says that in Salpa bicaudata there are traces of stigmata along the sides of the gill, but I have made sections through the gill of the embryo of Salpa scutigera, at the stage shown in Plate IV, Fig. 7, and find no trace of stigmata, and according to Traustedt, bicaudata is a synonym for scutigera. The central nervous system of salpa is a compact subspherical gang- lion, placed midway between" the mouth and the atrial aperture, on the dorsal surface, in the position which it occupies in the sessile tunicates, in which these two apertures are close together ; and it is so different from the elongated tubular nervous system of the primitive chordate type, as shown, for example, by the larvas of the ascidians, that we are forced to believe that it has been affected by the same influences as those which have led to its centralization in the sessile tunicata. In all respects the general plan of the structure of salpa is funda- mentally identical with that of the ordinary tunicates, and the differences are differences of detail. The atrial aperture, instead of being near the mouth, as it is in ordinary tunicates, is widely removed from it, as it is also in Doliolum and Pyrosoma ; and the atrium, instead of being wrapped around the pharynx as it is in the ordinary tunicates and, to a less degree, in Pyrosoma and Doliolum also, is placed end to end with it, but there is no reason whatever for questioning the strict homology of the atrium of Salpa with that of the other tunicates. This homology has been questioned by several recent writers on the development of Salpa and Doliolum, but I shall show that the history of 16 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. the development of both the solitary and the aggregated salpa proves that there is no basis for this opinion. On the contrary, the facts of embryology are in perfect accord with the teachings of comparative anatomy, and can be explained in only one way, that is, by unqualified acceptance of the view of the older writers, that the atrium of Salpa is the same chamber as the atrium of the ordi- nary tunicates. This short sketch is not intended to be an account of the structure of salpa, and it is given only to enable the reader to follow the account of the life-history which follows. CHAPTER II. THE DEVELOPMENT OF THE SOLITARY SALPA FROM THE EGG. SECTION 1. An Outline of the History of the Egg. The history of the eggs of Salpa before they are fertilized is so inti- mately bound up with the history of the process of asexual multiplication, that it is difficult to describe the one without continual reference to the other, and I shall therefore leave the detailed discussion of the origin and homologies of the germ cells for a later chapter, after the process of asexual multiplication has been described. It will, however, be best to speak briefly of the early history of the eggs before we enter upon the description of the process by which the ripe fertilized egg becomes con- verted into a salpa embryo. The germ cells are definitely set apart for reproduction at such an early stage, that our account of the embryology of salpa must begin with the embryo of the preceding generation ; for very early in its life, while it is still an embryo, we find in its body cavity a sharply defined mass of cells which the study of older specimens shows to be the germ of the reproductive organs. It is shown at n in Plate XXXV. At the earliest stage in which it can be identified, it lies in the body cavity of the embryo on the middle line of the ventral surface, and it marks the spot where the proliferous stolon is afterwards to be developed. As this latter is gradually formed the germinal mass is folded into it, in a way which is made clear by the successive stages shown in Plate XX, Fig. 6, Fig. 5, Fig. 7; Plate XXXV, n, and Plate XVI, Fig. 5. A series of transverse sections of the proliferous stolon which is shown in longitudinal section in the last figure, is given in Plate XX, Figs. 1, 2 and 3. As the stolon lengthens the germinal mass also elongates, as is shown at n, in Figs. 4 and 6 of Plate XLI, so that any transverse section of the former, like those given in Plate XXI, cuts some portion, m, n, of the latter. At first all the cells which enter into the composition of the germ- inal mass are alike, and its structure is homogeneous, as shown in Plate XX, Fig. 6, and Plate XLI, Fig. 7 ; but its peripheral cells soon become arranged in an epithelium, Plate XX, Fig. 2, m; Plate XXI, m, and Plate 18 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. XVI, Fig. 5, m ; which thus forms a follicle around a central core of cells, n. These central cells are the ovarian eggs. In young stolons, and at the root or proximal end of old ones, these eggs are small and crowded together, Plate XXXI, Fig. 4, but as we pass outward towards the tip of the stolon, Figs. 5, 6, 7, 8 and 9, they gradually grow larger, and the number cut by each ^transverse section grows less and less, until (Plate XXXIV, Fig. 4, m, and Plate XV, Fig. 2, m), they are pulled out into a series of single eggs. The figures in Plates XXI and XXXI are from young stolons which are carried by embryos like those which are shown in Plate XLI, Figs. 3 and 5 ; but even in mature stolons, which have set free many generations of buds, there is an undifferentiated portion of the germinal mass at the root, as is shown in Plate XLI, Fig. 8, which is a little nearer the root of the stolon than the figures on Plate XXXIV. On Plate XLI, Fig. 7, is a longitudinal section through the middle of the germinal mass of a very young embryo, before it has become differ- entiated into a central core of eggs and a peripheral follicle of epithelium. Figure 8 is a transverse section through the germinal mass of a fully grown stolon at its root, where the undifferentiated or embryonic germ cells are multiplying by karyokinesis, while Fig. 9 is from the same stolon a short distance from the root, where there is a continuous follicle, filled with ova, which latter have entirely lost the power of vegetative multiplication. In both young stolons and old ones the undifferentiated germ cells multiply by indirect division, and one or two cells with nuclear figures may be found in each section, but as we pass towards the tip of the stolon and the central cells assume the characteristics of ova, Plate XXXI, Fig. 5, from a young stolon, and Plate XLI, Fig. 9, from a mature stolon, the egg cells cease to multiply, although they increase in size, both yolk and nucleus growing rapidly. The material for this growth is furnished by follicle cells, Plate XXXI, Figs. 5 and 6, which migrate from the peripheral layer, inwards among the egg cells, where they degenerate and break down. In a stolon which is mature and ready to produce buds, the repro- ductive organ consists of a single row of fully developed ova, Plate XV, Fig. 2, n, surrounded by a follicular sheath which consists of an egg capsule of flattened cells, and an epithelium of thicker cells on the ventral or ha3mal side of the eggs. See also Plate XXXIV, Figs. 2 and 4. The flattened cells give rise to the follicular capsule of the egg, Plate X, Fig. W. K. BROOKS ON THE GENUS SALPA. 19 1, m, and to the fertilizing duct, x, by which the egg is attached to the wall of the cloaca of the chain-salpa, as shown in Fig. 10. The thickened layer of epithelium gives rise .to the testes. As the constrictions which mark out the bodies of the chain-salpae make their appearance in the walls of the stolon, the germinal mass also becomes divided up, Plate XV, Figs. 1 and 2, and Plate XXIII, Figs. 1, 2 and 5, into a series of segments, one for each chain-salpa. In Salpa pin- nata, and in most species, each of these segments contains a single egg ; but in those species which produce several embryos, such as Salpa cordi- formis and Salpa hexagona. the number of eggs is the same as the num- ber of embryos which is characteristic of the species. Thus the chain form of Salpa hexagona, Plate III, Fig. 1, normally gives birth to five embryos, and as shown in Plate XLV, Figs. 6 and 7, each segment of the genital string contains five eggs. Salpa democratica appears to be in an intermediate condition, for while it normally produces only one embryo, the segments of its genital string often contain, in addition to the single normal egg, one or two others which appear to be abortive, and which often exhibit indications of degeneration, although Salensky thinks there is reason to believe that a second embryo may be produced in this species after the first one has completed its development and has been set free. As the body cavities of the chain-salp* become separated from each other, the thickened epithelium under the egg becomes folded and pushed out to form two lateral pouches, Plate XV, Fig. 1, m, and Plate XXIII, Figs. 1 and 2, m, which are to give rise to the testes ; and at a somewhat later stage a third fold or outgrowth appears on the middle line and ultimately becomes the fertilizing duct. This outgrowth, which is indi- cated in Plate XXV, Fig. 7, H, by the letter m, is derived from the epithelium of the follicle on the dorsal or neural side of the egg. In Plate XXV, Fig. 7, (7, all three folds, the two paired testicular folds, and the unpaired median fold which represents the fertilizing duct, are shown, colored blue; and they are also shown in Plate XXVI, Fig. 1, .ff and G, and Figs. 2, E and F. As the bodies of the chain-salpse become developed and undergo changes of position, the reproductive organs also move away from their primitive positions, in a manner which will be readily understood from the figures. At the stage shown in Plate XXXIV, Fig. 4, the ovary is a continuous structure in a continuous sheath of follicle cells. At the stage shown in Plate XV, Figs. 1 and 2, and also in the reconstruction in Plate V, Fig. 1, the testicular folds, m, have appeared, and the follicle is con- 20 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. stricted into a series of segments with, in S. pinnata, and in most other species, one egg in each. This has gone a little further in the chain-salpee shown in Plate XXVI, A, B, C and Z>, and in the reconstructions which are shown in Plate V, Figs. 2, 3 and 4, although the eggs still lie in a straight line in a plane which corresponds with the middle of the stolon. At a stage a little older, Plate XXV, Figs. 5, 6 and 7, F, O and H, and in the reconstruction in Plate VI, Figs. 1, 2, 3 and 4, the single row is broken up into two, and the eggs are carried alternately to the right and to the left, with the growth of the bodies of the salpa3, until they finally assume the positions shown in Plate XXXVIII, Figs. 95 and 99, n, and Plate VII, Fig. 4, n. The path which is taken by the egg in its migration, and its relations, and that of the testes, to the other organs of the body, will be fully described in the chapter on the process of budding. All that concerns us here is the attachment of the egg to the wall of the cloaca by means of the fertilizing duct, Plate XXXIX, Fig. 4, x, which ultimately becomes a tube, Plate X, Fig. 10, x, and Fig. 1, x, through which the spermatozoa reach the eggs. Most writers state that the egg is fastened to the wall of the pharynx, and it is difficult to decide, from the exam- ination of adults alone, whether the point of attachment lies in the pharynx or in the cloaca, for there is nothing to mark the boundary between these structures, which are, however, more sharply separated in the young chain-salpa, where the duct is clearly seen to be attached to the wall of the cloaca. The chain-salpae of S. pinnata are set free in wheel-shaped or cylin- drical clusters of eight or nine individuals each, Plate I, Fig. 2. At the time of birth each of them contains an unfertilized egg, essentially like the one from Salpa hexagona which is shown in Plate X, Fig. 1. The large nucleus with its network of chromatin threads and large nucleoms is surrounded by a granular yolk, which is enclosed by a capsule of follicle cells, which are now elongated, although they were so flat as to be scarcely visible at the stage shown in Plate XXXVI, Fig. 2. At one point the follicle is continuous with the fertilizing duct, x, which has, by most writers, been termed the oviduct, although there is no good ground for the use of this name, for no ova ever pass through it ; and while it may possibly be homologous with the true oviduct of other tunicates, there is no evidence that this is the case, and I therefore prefer to use a name which at least has the merit of expressing its function, at the present day, as a channel for the spermatozoon. W. K. BROOKS ON THE GENUS SALPA. 21 SECTION 2. The Foetal Membranes. Each egg gives rise to an embryo which becomes a solitary salpa, while the chain-salpae are produced by budding from the solitary salpa. The embryo is developed within the body of a chain-salpa, and its growth begins very soon after the chain-salpa is set free and while it is very small, and it keeps pace with the growth of the chain-salpa, so that a fully grown einbryo is gigantic in comparison with the animal which carries it. Plate I, Fig. 3, is an individual of the chain-form of Salpa pinnata with its embryo, but in other species the embryo is relatively very much larger. Each chain-salpa usually contains only one embryo, as is shown in this figure, but in a few species there are several embryos in each. Plate III, Fig. 2, is a side-view, and Fig. 3 a dorsal view of the chain- form of Salpa cordif ormis, showing the embryos, e m, on the right side of the body, in the space between the fifth muscle and the sixth. Plate IV, Fig. 6, is a portion of a chain of the same species, showing, on the right side of the figure, the right sides of three salpas, with five embryos in each, arranged in a row in the space between the fifth muscle and the sixth. When there are several embryos they are in successive stages of development, as shown in Plate X, Fig. 10, which is from the chain-form of Salpa hexagona, shown in Plate III, Fig. 1, where the five embryos appear as a row of dots on the right side of the body, in the space between the last muscle and the next to the last. The egg before it is fertilized, and the embryo during the early stages of its development, lies in one of the blood-channels of the chain-salpa : the space which is marked y in Plate X, Fig. 10, and in the other figures. The egg is suspended by the fertilizing duct, x, of Plate X, Fig. 10, which is fastened to the wall of the cloaca, c, into which it opens. The sperma- tozoa which are drawn into the pharynx of the chain-salpa with the sea water, are swept past this opening by the contractions of the muscles in swimming, and some of them enter it and, penetrating to the egg, ferti- lize it. As the embryo grows it pushes in to the cavity of the cloaca, carrying its wall before it, as is shown in Plate XI, Fig. 3, where the letter y marks the blood-channel, while the c above the figure is in the cavity of the cloaca. The layer of epithelium which is marked 6' is that part of the wall of the cloaca which is pushed in before the embryo, and becoming 22 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. closely wrapped around it, forms the covering which I shall call the epithelial capsule of the embryo. The way in which the growing embryo comes to project into the cloaca is also shown in Plate IX, Figs. 1 to 9, where the blood-space, y, is colored yellow, and the cavity, c, of the cloaca red. The epithelial capsule covers the embryo during the early stages of development, and it is shown at b' in Plates XI, XII, XIII, XIV and XXII, and in cuts B, C and D. As it does not grow, it becomes distended by the growth of the embryo, and its cells grow more and more flat and farther and farther apart; and as the ectoderm is formed under it, it breaks up into separate cells which are thrown off as shown at 6' in Plate XVI, Figs. 2 and 3, and in Plate XVII, Fig. 4, and at B' in Plate XLV, Fig. 3. Plate XVI, Fig. 6, is a part of Fig. 2 very highly magnified to show the formation of the ectoderm and the molting of epithelial cap- sule. In this figure, 15 is the body cavity of the embryo, a its ectoderm, and b' the detached cells of the epithelial capsule; 21 and 22 are the outer and inner folds of the embryo sac, which is to be described soon. It will be seen that, at the stage shown in Plate XI, Fig. 3, that part of the embryo which is at the bottom is not covered by the epithelial capsule, but is directly exposed to the blood of the chain-salpa which circulates in the space y. While this uncovered area subsequently becomes smaller, as compared with the growing embryo, it never becomes covered in completely, and within it the placenta is formed. This organ, which serves to nourish the growing embryo with food derived from the blood, is shown in Plate III, Fig. 4, pi, and in Plate XLI, Figs. 1, 2, 3 and 5, pi. It is also shown in longitudinal section in Plate XXXV at y". It preserves its communi- cation with the blood-channels of the chain-salpa until the embryo is born, and as this is nourished from the blood which passes into and out of the placenta, its function and its anatomical relations are strikingly like those of the mammalian placenta, although there is a very important difference which will be described soon. The fold in the wall of the cloaca which covers the embryo and forms the epithelial capsule, soon extends down for some distance below the level of the embryo, as is shown in Plate XLV, Fig. 1, and forms the boundary of a spacious chamber, the cavity of the placenta, which opens through a constricted neck into the blood-channel of the chain-salpa. The cells which compose this wall soon become elongated and thickened, as is shown in the figure at 23, while those which cover the W. K. BROOKS ON THE GENUS SALPA. 23 embryo and form the epithelial capsule are flat, as shown at B'. This difference becomes more and more marked and the transition more and more abrupt, until the lower thickened portion, cut B, 23, becomes sharply separated from the epithelial capsule B', and forms what I shall call the supporting ring of the placenta. This is shown at various stages of development at 23, in Plate XVIII, Plate XXXV and Plate XLV, and in cuts B, C and D. One of its functions, and apparently the only one in most species, is to act as a framework for the placenta, and a support to hold the embryo in its position above the placenta ; but it also has a nutritive function in at least one species, Salpa pinnata, and its cells ultimately degenerate and become converted into food for the embryo. The rupture which sets the fully grown embryo free usually occurs around the neck of the placenta, so that the supporting ring is carried away with it and is gradually absorbed. While the embryo projects into the cloaca of the chain-salpa as I have shown, it is not at first in direct contact with the water, for it is covered, in the first place, by the epithelial capsule, and in the second place by the embryo sac, which is now to be described. This structure is often called the amnion, as it bears a certain resemblance, in its anatomical relations, to the amnion of the higher vertebrates, although it is not formed, as it is in the vertebrates, from the tissues of the embryo, but from those of the chain-salpa. It first makes its appearance as a cir- cular ridge or fold, Plate XLV, Fig. 2, 21 and 22, in the wall of the cloaca, around the area where the embryo is attached by the neck of the placenta. In some species it seems to be absent ; in others, as in Salpa hexagona, it is never any more developed than it is in the figure just referred to. More usually, however, it grows up around the embryo until this is com- pletely shut in except for a small pore or unclosed space. It is shown at 21 and 22 in transverse sections of Salpa pinnata in Plate XVIII, Figs. 1-6, in longitudinal section in Fig. 8, and in surface view in Plate XLI, Fig. 1. The space between the embryo sac and the embryo is the brood chamber. In its origin this is part of the cavity of the cloaca, but it becomes completely shut off except for the pore, which is shown in Plate XVIII, Fig. 4. The wall of the embryo sac is double, and the space between its two folds is continuous with the blood-spaces of the chain- salpa. It is plain from this description that a horizontal section, like the one in Plate XIII, Fig. 3, in the plane of the line marked xm, 3 in cut, will 24 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. pass, first, through the outer fold, 21, of the embryo sac ; second, through the space between the outer and inner fold, which is part of the body cavity of the chairi-salpa, and is, like this, colored yellow ; third, through the inner fold, 22, of the embryo sac ; fourth, through the brood pouch which is part of the cloaca of the chain-salpa and is colored red ; fifth, through the epithelial capsule, &', which is also colored red, and sixth, through the embryo. The embryo sac is formed during the early stages of development, and it becomes complete while the embryo is very small ; and as it does not increase in size, the rapid growth of the embryo soon causes it to dis- tend, and the embryo soon pushes through the small opening, stretching this and forcing its way out into the cavity of the cloaca, as is shown in Plate XLI, Fig. 2, and in Plate XXXV. As the epithelial capsule has in the meantime been cast off, the sur- face of the body of the embryo is now directly exposed to the water in the cloaca, and is fastened to its wall only around the neck of the placenta. At the stage which is shown in the last two figures the placenta is still inclosed in the embryo sac, although the embryo itself is free ; but at the stages shown in Plate XLI, Figs. 3 and 5, the placenta also is uncovered, and the embryo sac is no longer recognizable, as its folds have been flattened out and obliterated by the growth of the embryo. This brief outline of the history of the foetal appendages is enough to make these structures intelligible in the figures of the various stages of development, and we are now in a position to trace the embryology of salpa from the egg, as our description need not be interrupted by refer- ences to these structures. SECTION 3. The Migration of the Follicle. The salpa embryo consists of elements of two sorts : those derived from the fertilized egg, colored orange in the plates, and those derived from the follicle, which are, with a few exceptions, colored blue. The egg, before fertilization, Plate X, Fig. 1, is inclosed in a capsule of follicle cells, m, which are, in ultimate origin, modified germ cells, as I shall show farther on. Each follicle cell is a cell which might have become an egg, although they are not now eggs. They are not fertilized, and while the part they play in the formation of the embryo is very remarkable and interesting, their function is purely nutritive, and they do not become converted into any of the tissues of the embryo. At first W. K. BEOOKS ON THE GENUS SALPA. 25 the egg entirely fills the cavity of the follicle, as shown in Plate X, Fig. 1 ; but as soon as segmentation begins, after fertilization, Fig. 3, the cavity becomes divided into two portions, an empty one, 5, and another which is occupied by the egg. Over the half occupied by the egg the follicle cells, 10, retain their original character and their sharp boundaries, while those which form the wall of the empty half, 7, lose their distinctness, and multiply rapidly by karyokinesis, so that this half soon becomes much thicker than the other. For a short time the blastomeres, Fig. 5, 9, which are formed by the segmentation of the egg, are bounded on one side by the empty f ollicular cavity, o ; but the follicle cells soon begin to migrate inwards in the zone which is marked 8 in Fig. 3, and soon completely cover up the blastomeres, as shown in Fig. 5, 7 and 8, so that the folli- cular cavity is now bounded on all sides by follicle cells, and the blasto- meres are imbedded in a mass of follicle cells. The follicle cells in the area which is marked 10 in Fig. 3, now begin to move inwards in radial lines and to push their way in among the blastomeres, and to force these apart, as is shown in Fig. 8, and this process of migration goes on until the folli- cular cavity is obliterated, as shown in Fig. 9. At the stage shown in Fig. 8, the embryo consists of the following structures : first, an outer or somatic layer of follicle cells, 7; second, a visceral layer of follicle cells, 8; third, an area, 10, where these two layers are continuous with each other ; fourth, the blastomeres, 9 ; fifth, follicle cells between the scattered blas- tomeres, and sixth, the follicular cavity, o. In a stage a little older, Fig. 9, and Plate XI, Figs. 1 and 2, the cavity of the follicle becomes obliterated as already noted, and the follicle cells of the visceral layer begin to multiply very rapidly by direct division of the nuclei, although the somatic cells still multiply by karyokinesis, but much less actively. Plate XLII, Fig. 1, is a part of the embryo, shown in Plate XI, Fig. 2, very highly magnified. Three blastomeres, 9, and part of a fourth are shown imbedded in the mass of follicle cells of the visceral layer, 8, which is not separated by any empty space from the somatic layer, 7, of follicle cells. The blastomeres are so much larger than the follicle cells that they can be recognized without difficulty. Each of them has a very large spherical nucleus, with a complicated network of very fine and delicate threads of chromatin. The nucleus is near the center of the blastomere, which is itself nearly spherical, very much more transparent than the surrounding follicle cells, with a well marked boundary, and filled with a number of bodies which stain much more deeply than the substance of the blastomere. The boundaries 26 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. between the visceral follicle cells are almost invisible, and their nuclei are elongated in radial lines, and are irregularly pear-shaped, with an aggregation of a substance, which stains very deeply, at the central end. These elongated nuclei are often arranged in pairs, the two members of the pair lying in the same radius, and many of them are in the act of dividing into an inner and an outer portion. In Fig. 2, which is from the somewhat older embryo which is shown in Plate XI, Fig. 3, a is a folli- cular nucleus which is about to divide, b is one in the act of division, and c is one which is separated into two daughter nuclei, of which the inner one has migrated inwards to a considerable distance towards the center of the embryo. In their migration some of these nuclei push their way in to the substance of the blastomeres, and in Fig. 1 one is shown in the act of penetrating its outer wall. Some of the bodies inside the blastomeres are sharply defined, and these agree with the follicle nuclei in size, in their color in stained specimens, and in the arrange- ment of their chromatin ; and the study of sections at this and subsequent stages proves that the less sharply defined bodies are follicle nuclei in process of degeneration, and that the blastomeres are nourished by migratory nuclei from the visceral layer of follicle cells. The space between the blastomeres is also filled with these nuclei in all stages of degeneration and with the granules which have come from their disinte- gration. The multiplication of the blastomeres goes on slowly, and while they gradually become smaller and more numerous, as shown in Figs. 3, 4, 5, 6 and 8, they are seldom found in the act of division. The material which is assimilated by the blastomeres from the migratory follicular nuclei, seems for some time to be converted into chromatin ; for while the protoplasm of the blastomeres continues transparent, as shown in Figs. 2, 3 and 4, the chromatin of these nuclei increases in amount and forms a sharply defined reticulum with a large central nucleolus, and a number of smaller nucleoli around the periphery and in the meshes of the net- work. The nuclei of the follicular cells, on the other hand, become vesi- cular and transparent through repeated division, and their chromatin becomes more and more scanty, while they continue to divide so rapidly that they show a well-marked arrangement in pairs, as is shown in Fig. 3 and in the following figures. As the blastomeres continue to multiply they gradually become very granular, as is shown in Figs. 5, 6 and 8, and even after they have become nearly as small as the follicle cells, they are easily distinguishable by their large nucleoli and conspicuous network, W. K. BROOKS ON THE GENUS SALPA. 27 from the double, transparent, vesicular nuclei of the follicle cells, as is well shown in Fig. 8, where the blastomeres are marked bl and the follicle cells 7 and S. Finally, the blastomeres begin to multiply actively by karyokinesis, as shown at b in Fig. 9, and to give rise to the germ layers; but as it is difficult to understand the peculiar relation between the follicle cells and the blastomeres, without some knowledge of the history of both structures, it will now be necessary to take up the history of the follicle before we study the history of the germ layers. Stated in a word, the most remarkable peculiarity of the salpa embryo is this. It is blocked out in follicle cells which form layers and undergo foldings and other changes which result in an outline or model of all the general features in the organization of the embryo. While this process is going on the development of the blastomeres is retarded, so that they are carried into their final positions in the embryo while still in a very rudimentary condition. Finally, when they have reached the places which they are to occupy, they undergo rapid multiplication and growth, and build up the tissues of the body directly, while the scaffolding of follicle cells is torn down and used up as food for the true embryonic cells. No other animal presents us with an embryonic history quite like that of Salpa, although other Tunicates show something similar, but very much less pronounced. In another chapter I shall try to show how the life-history of Salpa has come about, but we must now confine our- selves to the facts. An imaginary illustration may help to make the subject clear. Suppose that while carpenters are building a house out of wood, that brickmakers pile clay on the boards as they are carried past, and shape the lumps of clay into bricks as they find them scattered through the building where they have been carried with the boards. Now, as the house of wood approaches completion, imagine that bricklayers build a brick house over the wooden framework, not from the bottom upwards, but here and there wherever the bricks are to be found, and that, as fast as parts of the brick house are finished, the wooden one is torn down. To make the analogy complete, however, we must imagine that all the structure which is removed is assimilated by the bricks, and is thus turned into the substance of new bricks to carry on the construction. 28 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. SECTION 4. The Organs of the Embryo outlined in Follicle Cells. The structure of the young embryo and the shares which the two sorts of cells take will be understood from the plates, and especially from the horizontal sections in Plates XII, XIII, XIV, XVI and XVII, and from the diagrammatic reconstructions in cuts A, B, C and D. The shape of the young embryo makes it difficult to control the position of sections in any plane except the horizontal ; that is, the plane which is parallel to the bottom of the page in Plate XXXV. I have therefore paid especial attention to serial sections in this plane, and have figured a series from young embryos at successive stages of development, in Plates XI, XII, XIII, XIV, XVI and XVII. Longitudinal sections like Plate XXXV, and vertical transverse sections like those in Plates XVIII, XXII and XLV, are much more intelligible if they are perfectly symmetrical and exactly at right angles to the horizontal plane, but as the slightest deviation makes them hard to interpret, I have not been able to obtain, from young embryos, any which are exact enough to be useful for illustration. They have been valuable to me in the interpre- tation of the horizontal sections, but an attempt to describe them would complicate the description so much that I have not drawn them, but have, instead, reconstructed from them, and from the horizontal sections which are figured, the series of diagrams of vertical transverse sections which is shown in cuts A, B, C and D. We left the follicle at the stage, Plate XI, Fig. 1, at which its cavity is entirely filled up by the visceral layer, 8, which is in direct contact with the somatic layer, 7, although the inner ends of the somatic cells are sharply defined. These two layers soon become separated again by a space, Fig. 3, 15, which persists from this time as the permanent body cavity of the embryo. It is colored purple in all the figures, and is marked 15. It is probably the original cavity of the follicle, opened a second time by the growth of surrounding parts, and at the stage of Fig. 3, and for a long time in the later history of the embryo, it is bounded on all sides by follicle cells. A diagram, constructed from a series of trans- verse sections of an embryo, like Fig. 3, is shown in cut A on page 29. In this, as in the plates, y is the body cavity of the chain-salpa, B is the epithelial capsule, 15 is the body cavity of the embryo, 9 and H' are the blastomeres, and 7 and 8 the somatic and visceral layers of follicle cells. On each side of the middle line the somatic layer is invaginated to form a pit which opens into the space between the embryo and the epithe- W. K. BROOKS ON THE GENUS SALPA. 29 lial capsule. This pit is the perithoracic tube or spiracular tube, and its external opening, which soon closes, is the spiracle. Before the two peri- thoracic tubes lose their external openings they elongate, and pushing across the body cavity into the substance of the visceral layer of follicle cells, they meet and unite on the middle line to form the cloaca. A reconstruction in a transverse vertical plane, from the horizontal section shown in Plate XVII, Fig. 5, and in Plate XII. This stage in the history of the perithoracic tubes is shown in Plate XII, which is a series of horizontal sections of a young specimen of Salpa pinnata, and in cut B, which is a vertical transverse section constructed from the series of horizontal sections. The reader who wishes to understand the structure of the embryo must compare these figures with each other. The double fold of the embryo sac, 21 and 22, and the epithelial capsule, b' or B', appear in all the sections, but as they have already been described, it only remains to point out that while these membranes are shown in the cut separated from the embryo by an empty space, as they are in the living embryo and in unshrunken specimens, they are represented in the horizontal sections as close to the surface of the embryo, as they are in specimens which have been imbedded in melted paraffine, which causes these deli- cate unsupported folds to shrink. The plane of section 1, Plate XII, cuts the right spiracle g" close to its external opening, while it cuts the left one below its opening and just above the point where it communicates with the cloaca which is shown at g'" in section 2. The epithelium which lines the cloaca and perithoracic tubes is derived from the somatic layer of follicle cells, but in the plates it is colored orange like the blastomeres, as I did not obtain proof of its folli- 30 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. cular origin until the plate was finished. The orange color is used in the same way for the follicular epithelium of the cloaca and perithoracic tubes in Plate XXII. Below the level of the cloaca the perithoracic tubes run downwards, one on each side of the middle line as shown at g 1 in Fig. 3 and at g v in the cut, but they end blindly at this stage, and in Fig. 4 the left is cut at its end below the cavity, while the right, gr, is cut close to the bottom of the cavity. The next section, Fig. 6, does not cut any part of the perithoracic tubes, although their positions are outlined on each side of the middle line in visceral follicle cells with scattered blastomeres. The next stage in the development of the perithoracic system is shown in Plate XIII and in cut C. CUT C. A reconstruction in a vertical transverse plane from the horizontal sections shown in Plate XIII. The cloaca, Fig. 7, g'", and the two prolongations of the tubes down- wards, Fig. 8, g lv , are about as before, except that these latter are a little longer. The spiracular tubes, however, have undergone a great change, as they have lost their external openings and have moved towards the middle line, where they lie side by side at g", in Fig. 6, above the cloaca, imbedded in the visceral follicle cells. In Plate XIV and in cut D, the pharynx, c, is shown, as well as the two gill-slits, gf lv , which have been formed out of the descending portions of the perithoracic tubes, Fig. 6, g lv , which now open above into the cloaca, Fig. 5, g'", and below into the pharynx, Figs. 8, c and 9, c. In Figs. 3 and 4, the two spiracular tubes g" are shown side by side, above the cloaca, W. K. BROOKS ON THE GENUS SALPA. 31 imbedded in visceral follicle cells. Plate XXII shows a series of trans- verse sections at the same stage, but as these are not perfectly vertical but in planes which make an acute angle in front with the horizontal axis, they are less easily intelligible than the cut. Fig. 2 passes through the pharynx, c ; Fig. 3, through the two gill-tubes, g", and Fig. 4, through the cloaca, g'", and the two spiracular tubes, g", g". The details of the formation of the pharynx and gill-tubes and gill, o, are shown in Plate XLII, Figs. 6, 7 and 8. The cavity of the pharynx, Fig. 8, c, and Plate XIV, Figs. 8 and 9, c, and Plate XXII, Fig. 2, c, is hollowed out in the mass of visceral follicle cells, below the cloaca, by the degeneration of the follicle cells. These become amoeboid and are set free in the cavity, where they persist for some time. They are easily recognizable by the transparency of their nuclei, and by the fact that these are usually in pairs. 3 Plate 14 CUT D. A reconstruction in a vertical transverse plane from the horizontal section shown in Plate XIV. The communication between the perithoracic tube g" and the pharynx is formed in essentially the same way. The somatic follicle cells, 7, of the perithoracic tube, Fig. 6, Plate XLII, and Fig. 8, and the visceral follicle cells between it and the pharynx, Fig. 8, 8, become amoeboid, and wander out into the cavity which is thus formed. The rod-like mass of cells, Fig. 8, o, which is left between the cloaca g, above, and the pharynx c, below, and the gill-slits g", on the sides, is the so-called gill. It consists of a mass of visceral follicle cells, 8, which contains 32 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. scattered blastomeres, &Z, and is covered above and at the sides by a layer of somatic follicle cells, 7. The degeneration of the follicular lining of the cloaca begins before the gill-slits are formed. Fig. 7 of Plate XLII is part of Plate XIII, Fig. 8, showing part of a perithoracic tube at the stage when it ends blindly. The visceral follicle cells are shown on the left ; then the follicular epithe- lium of somatic cells, and then three of these cells, with double nuclei, which have become amoeboid, and have wandered into the lumen of the tube. Fig. 6 is a section through the axis of the gill-slit, from a specimen at the same stage as Plate XXII, showing the follicle cells breaking apart to form the channel of communication with the pharynx. As the embryo grows the pharynx increases in size, at first, very much faster than the cloaca, and at the stage shown in Plates XVI and XVII, the pharynx, Fig. 2, c, is very capacious, while the cloaca, XVI, Figs. 1 and 2, g" and g lv , is very small. The relative sizes of the two structures are also shown at a somewhat later stage in Plate XVIII, Figs. 4 and 8, c and g'", and also in the surface view in Plate XLI, Fig. 2, c and f/". The two spiracular tubes seem to fuse into one by the disintegration of the partition, as shown in Plate XVI, Fig. 1, g" ; and their chamber becomes part of the cloaca. All the somatic follicle cells of the cloaca ultimately fall into its cavity and degenerate, although this process is not completed until the other organs of the body are well advanced in their development, and they are shown in Figs. 1, 2 and 3 of Plate XVI, and at x in Fig. 9 of Plate XLII. SECTION 5. The Blastodermic Tissties of the Embryo. We have now to consider the way in which the blastodermic epithe- lium of the pharynx and cloaca replaces this temporary scaffolding of follicle cells. While the changes which we have described are taking place, the blastomeres gradually become smaller and more numerous, as shown in Plate XLII, Figs. 1, 2, 3, 4, 5, and those which are to give rise to the epithelium of the pharynx and cloaca become distributed through the mass of visceral follicle cells under the region of the perithoracic tubes and cloaca, as shown in Plate XII, Figs. 6 and 7, 9, and in Plate XIII, Fig. 9. As the cavity of the pharynx is formed they become arranged between the visceral follicle cells, Plate XLII, Fig. 8, 8, and the degenerating somatic cells, 7, as is shown at bl in Fig. 8 and in Fig. 5. In this way the W. K. BROOKS ON THE GENUS SALPA. 33 epithelial lining of the pharynx is gradually completed and extended until at last it becomes continuous, as shown in Fig. 9, in which 6 is the cavity of the pharynx and 15 the body cavity. Even after this epithelium is well denned, it is here and there interrupted by a follicle cell, with its two transparent nuclei, as is shown at 8 in Fig. 10. This figure is part of Fig. 2 of Plate XVII. Other blastomeres migrate upwards along the gill-slits, under the somatic cells, as shown in Fig. 6 of Plate XLII, which is from an embryo at the stage of Plate XXII. These cells multiply, and finally build up a continuous epithelium in the gill-slits, as shown in Plate XLII, Fig. 9, which is part of the section shown in Plate XVI, Fig. 3. In Fig. 9, gr' v is the cavity of the gill-slit, a; is a group of degenerating somatic follicle cells, 6' is the blastodermic epithelium, 75 is the body cavity, 8 the visceral follicle cells, A mesoderm cells, a the ectoderm, and &' the cells of the degenerating epithelial capsule. The oesophagus, stomach and intestine are formed from a diverticulum from the posterior wall of the pharynx, a little to the right of the middle line, as shown at a very early stage at q in Plate XVII, Fig. 2, and more magnified in Plate XLII, Fig. 10, q. The opening of this diverticulum becomes the oesophagus, shown at q in Plate XVIII, Figs. 6, q and 8, q, and it elongates to form the stomach and intestine, but as it soon becomes twisted it cannot be described in detail without a greater number of figures than the subject seems to merit, as the only point which seems noteworthy is that the digestive tract is a secondary outgrowth from the pharynx. As the blastodermic epithelium of the gill is formed, the intestine extends forwards into it, as is shown in Plate XLI, Figs. 3 and 5, and at p in Plate XXXV. The anus is not formed until the embryo is well advanced, when it breaks through into the cloaca at the point marked p" in Plate XXXV, on the middle line at the anterior end of the cloaca. Plate XIX, Fig. 1, is a horizontal section, reversed in the drawing, through the gill of an embryo a little older than Plate XXXV, showing the intestine p in the gill o, and the opening of the oesophagus q, on the right side at the base of the gill. At a very early stage, Plate XVII, Figs. 1 and 2, a stomodaeal invo- lution of the ectoderm z grows inwards to meet a corresponding out- growth from the pharynx, although the aperture of the mouth is not formed until the embryo is well advanced. In Plate XXXV the stomo- daeum z is shown with its inner end covered up by the unbroken epithe- lium of the pharynx. Plate XLV, Fig. 8, is a vertical section through the mouth at the time of its appearance, from an embryo like Plate XLI, Fig. 5. 34 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. In Plate XLV, Fig. 8, v is the cellulose mantle which is not yet per- forated, and c is the cavity of the pharynx. The ectoderm and endoderm are united at the edge of the lower lip, but they are as yet separate in the upper lip, which is rounded and thick and protuberant, with three trans- verse muscles, w, and at a later stage the thin lower lip is tucked inwards under the rounded upper lip, to form the oral valve of the branchial sac. The gill of salpa, Plate XLII, Fig. 8, o; Plate XVIII, Fig. 4 and Fig. 8, o, Plate XXXV, o, and Plate XLV, Fig. 4, o, is simply the space bounded by the pharynx below, the gill-slits at the sides and the cloaca above. It is at first a solid mass of follicle cells with blastomeres which give rise to its lower or endodermal epithelium, but its sides are only slowly covered with blastodermic epithelium, and its upper surface consists of follicle cells in embryos which have acquired most of their organs. The follicle cells in its cavity degenerate as the intestine extends into the gill. The cloaca is at first lined throughout by somatic follicle cells. Some of these begin to migrate into its cavity very early, and in some speci- mens the whole chamber is so choked up with them that it is difficult to trace it. The blastodermic epithelium gradually extends over its whole surface, and the follicle cells degenerate and disappear. Its cavity then enlarges very rapidly, as will be seen by comparing Fig. 8 of Plate XVIII with Plate XXXV, or by comparing the embryos on Plate XLI. After its epithelial lining is complete, the ectoderm, Plate XVII, Fig. 6, a, bends downwards around a circular area to meet it, in such a way that a lump of visceral follicle cells is shut in under an arched cover of ectoderm. These cells then become vacuolated and finally disappear, as does also the cap of ectoderm. The ectoderm around the edge of the circle now bends inwards upon itself as shown in Fig. 7, and becomes continuous with the cloacal epithelium, and all of this inside the circular line of adhesion degenerates as shown in the figure, and finally disappears, to form the cloacal aperture. Figure 6 of Plate XVII is from the embryo shown in Plate XXXV, and Fig. 7 from one like Plate XLI, Fig. 3. It will be seen from this account that the cloacal aperture is a new formation, and that it is not the two spiracles united into a single aperture, although I believe that the history of salpa is quite reconcilable with the view that it is phylogenetically a pair of spiracles. The spiracles which are formed in the somatic layer of the follicle lose their external openings, as we have seen, and the spiracular tubes move towards the middle line and unite at the spot, above the cloaca, where the aperture is W. K. BROOKS ON THE GENUS SALPA. 35 afterwards formed. They are not blastodermic but follicular, and they therefore do not repeat the ancestral history in all details, but their changes of position are quite intelligible on the assumption that they are a record which has been preserved from a time when the spiracles them- selves moved up to the middle line of the back and fused to form the cloacal aperture. The changes which take place in the position of the aperture during the growth of the embryo are most interesting. The first trace of it, Plate XLI, Fig. 2, g v , is at the upper end of the long vertical axis of the embryo, and the space between it and the ganglion, s, is about equal to the space between the ganglion and the mouth, z ; and the axis of the mouth and that of the cloacal aperture make an angle of about 90. As the embryo grows, Figs. 3 and 5, the mouth and the ganglion preserve essentially their original relations to each other, but the space between the ganglion and the cloacal aperture gradually increases until, at last, mouth and cloacal aperture come, in the adult Salpa pinnata, to lie in the same axis at opposite ends of the body, as is shown in Plate I, Fig. 1. In the fixed ascidians the mouth and the cloacal aperture are close together, with the ganglion between them, and in this, as well as in other respects, the young salpa embryo is much more like a fixed ascidian than the adult, and I think we must see, in the primary position of the cloacal aperture, evidence that the salpae are descended from fixed ascidians, or, at least, from ancestors very similar to the ascidians in structure and habits. We have now traced the broad outlines of the history of the digestive organs, and of the perithoracic tubes and their derivatives, and we will pass to other systems of organs. Returning to the stage shown in Plate X, Fig. 8, we have seen that the outer wall consists at this time of a somatic layer of follicle cells, 7, which is continuous, over the area 10, with the central mass of follicle cells, 8, and blastomeres, 9. The area, 10, where the two layers are continuous, marks what is to be the middle line of the dorsal surface of the embryo, and some of the blastomeres soon move outwards along this line until they pass entirely out of the follicle and lie directly under the epithelial capsule. These cells are the ectodermal blastomeres, and they are shown at a" in cuts A and B and in Plate XXII, and at 9' in Plate XVII, Fig. 5, and in Plate XII, Figs. 1 and 2. The epithelial capsule, which at first passes over them without interruption, as shown in cut A, soon becomes 36 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. folded down on each side of them, so as to form a median dorsal ridge, shown in cut B and in Plate XVII, Fig. 5. At first, as is shown in cut A, these ectodermal blastomeres are not abruptly separated from those at deeper levels, but as the perithoracic tubes and their derivatives are formed and grow inwards, they separate the ectodermal blastomeres, in the middle region of the body, from those of the visceral mass, as is shown in cuts B and C. The series of sections on Plate XII shows, however, that there is no such interruption before or behind the cloaca. At the posterior end of the body, the end which is at the top in the figures, a continuous series of blastomeres may be traced through all the sections, from 9' in Plate XVII, Fig. 5, through 9' in Plate XII, Figs. 1 and 2 ; and 18 in Figs. 4 and 5 to 18 in Fig. 6. At the anterior end a similar series of blastomeres may be traced from 9 in Plate XVII, Fig. 5, through 9' in Plate XII, Fig. 1 ; 9 in Fig. 2, and s in Figs. 4, 5 and 6, to Fig. 7. The mass of visceral follicle cells and blastomeres which makes up the embryonic region is at first nearly spherical, as shown in the diagram in Plate XII, Fig. 10. As the embryo grows and the body cavity becomes more capacious, the visceral mass becomes folded into two vertical plates, intersecting at right angles in such a way that a horizontal section shows it as a cross, Plate XII, Fig. 5, with its long arm in the middle plane of the embryo, and crossed near its anterior end by the short arm. In Fig. 1 the ends of the short arm are continuous around the spiracular openings of the perithoracic tubes, with the somatic layer, as they are also in Fig. 2, where it contains the cloaea, which, with its lining of somatic cells, runs across the middle line and perforates the long arm of the cross. In Fig. 4, the enlarged rounded ends of the short arm contain the blind ends of the perithoracic tubes, as also in Fig. 5, while in Fig. 6 and in Fig. 7 the short arm contains the blastomeres which are to form the pharynx or branchial sac, into which, at a later stage, the perithoracic tubes open through a single gill-slit on each side. The long arm of the cross is formed by what is shown by the series of sections to be a thin verti- cal plate of visceral follicle cells and blastomeres, hanging down into the body cavity from an area on the middle line of the dorsal surface where it is suspended from the layer of somatic follicle cells. In the middle region of the body this vertical plate is interrupted by the peri- thoracic organs, and it is perforated by the cloaca, but both below and above the cloaca it is a continuous plate. W. K. BROOKS ON THE GENUS SALPA. 37 At the level of Figs. 5 and 6 which are in the planes indicated by the lines 5 and 6 in cut B, the whole cross lies free in the body cavity. At the level of Figs. 4 and 2 the ends of the short arm become continuous with the somatic layer, and the long arm is interrupted by the cloaca. In Fig. 2 the posterior end of the long arm becomes continuous with the somatic layer, as does its anterior end also in Fig. 1, where the long arm is continuous above the cloaca, as it is below it in Fig. 4. Most of the organs of the body except the perithoracic system are outlined in the visceral follicle cells of the median plate, and the blasto- meres are grouped with reference to this outline. At the level of Fig. 4 there are three well marked enlargements of the median plate, s, 19 and 18. One of these, s, lies anterior to the trans- verse plate, and is the rudiment of the ganglion. In Fig. 2 it is about as in Fig. 4, but in Fig. 1 its follicle cells become continuous with those of the somatic layer, and in Plate XVII, Fig. 5, its blastomeres, s, become continuous with the extra-follicular blastomeres of the ectodermal ridge which is shown at A" in cut B. The enlargement of the median plate at its posterior end, 18, is the rudimentary nervous system of the caudal region, and in Figs. 2, 1, and XVII, Fig. 5, its blastomeres can be followed up into the posterior end of the ectodermal ridge, just as those of the ganglionic rudiment can be followed up into its anterior end. The third thickening, which is marked 19 in Plate XII, Fig. 2, is the rudiment of the notochord. At the level of Fig. 2 it contains no blastomeres ; at the level of Fig. 3 it contains two blastomeres, but there are none in Fig. 4, while they are numerous at lower levels, as is shown in Fig. 5 and also in Fig. 6, where the notochord is erroneously marked 18 instead of 19. At the level of Figs. 6 and 7 the median plate, with its blastomeres, swells out into a pair of lateral lobes which lie under the blind ends of the perithoracic tubes, and give rise to the pharynx, as already described. All these structures are shown, in essentially the same relations, in the older embryo in Plate XIII, except that the anterior end of the body is at the top of the figures, and the caudal nervous system is marked 30 instead of 18, in both Plate XIII and Plate XIV. The most important changes in the visceral mass at this stage con- cern the perithoracic system, and have already been described. The f ollicular rudiment of the caudal nervous system is also begin- ning to break down and disappear. The degeneration begins at the upper end, where it is continuous with the ectodermal ridge, and at this stage, as shown in Plate XIII, Figs. 6 and 7, 30, it is now represented only 38 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. by scattered cells, although it is still sharply defined at a lower level, as is shown in Figs. 8 and 9, 30. The next older embryo, Plate XIV, shows it in about the same condition, degenerated and with no distinct boundary in Figs. 4, 5 and 6, 30, but sharply defined in Figs. 7, 8 and 9, 30. I do not know what becomes of its blastomeres, but as they gradually become unrecognizable, I see no reason for supposing that they persist, and they probably degenerate and disappear. The structure which is marked 19 in Plates XII, XIII and XIV becomes the larval organ which is generally known in salpa as the eleoblast. Its successive stages of development in Salpa pinnata are shown at k in Plates XVI, XVII, XXXV, XIX, and in Plate XLI, Figs. 2, 3 and 5, k. As these plates show, it grows with the growth of the embryo, and is in the older larvas a prominent organ of considerable size. Salensky suggests that it may be a rudimentary tail, and while he does not describe its structure minutely nor present much proof, his view is unquestionably the true one. In Salpa pinnata degenerative changes begin in it very early, at the stage of Plate XVI, and go on as it grows, so that its internal structure is always vague and indefinite, but it is very much less rudimentary in the embryos. of other species, especially those of the cordiformis group, and sections of it, in advanced embryos of these species, show that it is unquestionably the embryonic and degenerated representative of the locomotor tail of the tadpole larvae of other ascidians. In Plate III, Fig. 4, I have drawn an advanced embryo of Salpa hexagona, and in Plate VIII, Fig. 3, I have copied for comparison one of Uljanin's figures of the tailed larva of Doliolum, showing the chorda ch, and the caudal vesicle k, which is formed by its degeneration. Comparison of these figures shows clearly that the position and the anatomical relations of the eleoblast, k, of Plate III, Fig. 4, are identical with those of the tail-vesicle k of the Doliolum larva. Fig. 1 of Plate XLIV is a transverse section through the eleoblast of the embryo of Salpa hexagona which is shown in Plate III, Fig. 4. This figure shows that it consists, first, of the outer sheath or cellulose mantle, v ; secondly, of a layer of very thin flat ectoderm cells, a ; third, of a circular tract of the body cavity, 15, filled with blood corpuscles, bl, and migratory follicle cells, 29; and fourth, of a great mass of wedge-shaped or subconical chorda cells, k, radially arranged, with all their protoplasm, and their nuclei, at their outer ends, while the empty bodies of the cells converge towards the center. These conical cells are wedge-shaped in section, but W. K. BROOKS ON THE GENUS SALPA. 39 as their position is not perfectly radial, but inclined to the axis of the chorda, a section does not lay open the whole length of any one cell from periphery to center. The axis of the chorda is occupied by a mass of granular protoplasm with scattered nuclei. In an ordinary ascidian the tail degenerates before the definitive structure of the adult is acquired, but in Doliolum, as Uljanin's figures show, it is retained for a time after the little animal has ceased to be, in other respects, a larva, and has completely acquired all the characteristic structures of the adult Doliolum. In this respect Salpa agrees with Doliolum, for the young Salpa hexagon a, in Plate III, Fig. 4, is not a larva, but a perfectly formed young salpa, although it still retains its notochord k and its placenta pi. Other species of salpa retain the rudimentary tail still longer, and there is a trace of it in the adult Salpa cordif ormis. Plate IV, Fig. 3, is a young specimen of the solitary form of this species, some time after birth, showing the tail as a distinctly marked outgrowth of the cellulose mantle, with a cavity which is part of the body cavity. There is no trace of the chorda, although comparison with embryos of this species and of Salpa hexagona shows that this process is the same as the tail of Fig. 4 of Plate III. It grows smaller compared with the body, as the animal grows up, but it is easily seen even in the adult Salpa cordiformis, Plate IV, Fig. 5. In Salpa pinnata the tail reaches considerable size, as is shown in Plate XLI, Fig. 5, but degenerative changes occur in it so early that its internal structure is vague and indefinite. It consists, as is shown at k in Plate XXXV and in Fig. 9 of Plate XIX, of an outer wall of flattened ectoderm cells, a, which is covered by a thin layer of cellulose, and forms the wall of a chamber which has groups of blood corpuscles around its periphery, while its central portion is occupied by a vacuolated mass of irregular cells in process of degeneration, and this species, studied alone, would give little evidence as to its nature, although other species show clearly that it is a tail, and that its central mass is a somewhat degener- ated notochord. It now remains to trace its origin from the body which is marked 30 in Plates XIII and XIV, and 19 in Plate XII. Plates XVI and XVII show it, in a transitional stage, at k. In this embryo all traces of the caudal nerve have disappeared, and the notochord is represented by a sharply bounded mass of cells filled with large vacuoles. I was not able to trace the history of the blastomeres, but as the circular vacuoles make their appearance at about the time when the round blastomeres 40 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. disappear, they are possibly the spaces which they occupied. If this is true, the rudiment of the true blastodermic chorda degenerates in Salpa pinnata before its follicular case, but both structures soon break up, and are used as food. We have traced the migration of the ectodermal blastomeres from the interior of the visceral mass to their extra-follicular position in the ectodermal ridge, Plate XVII, Fig. 5, 9, and cuts B and C, a". In this position they are covered up by the epithelial capsule, but are on the out- side of the embryo. At a stage a little older than Plate XIV and Plate XXII, they begin to multiply and to spread out over the embryo on both sides of the middle line to form the ectoderm. Plate XLII, Fig. 11, is a transverse section of the ectodermal area of an embryo a little older than Plate XIV. 21 is the outer fold of the embryo sac, 22 its inner folds, and 6' is the epithelial capsule ; a is the ectoderm spreading out at the sides between the epithelial capsule and the somatic layer, 8, of follicle cells. At this early stage in the development of the ectoderm its cells and nuclei are so much larger than those of the follicle that they can be distin- guished clearly, and the nuclei of the blastoderm cells are rich in chro- matin and have a well marked reticulum with nucleoli and large granules, and are in this quite different from the vesicular nuclei of the follicle cells. The ectoderm has a growing edge, like that of a meroblastic embryo, and it gradually spreads on all sides, and pushing under the separated cells of the epithelial capsule, forces them off, and thus finally becomes the outer covering of the embryo. The embryo, which is shown in Plates XVI and XVII, is almost covered by the ectoderm, and Figs. 6 and 7 of this plate and 9 of Plate XLII are parts of this embryo, more highly magnified to show the details of its structure. In all these figures a is the ectoderm, and &' the epithelial capsule. The cellulose mantle makes its appearance as soon as the ectoderm is fully formed, as a trans- parent layer, which is shown at v in Plate XVII, Fig. 6, and in Plate XLV, Figs. 3, 4 and 6. A few small cells are sometimes included in it, as in Plate XVII, Figs. 5 and 6, v, but the cells of the epithelial capsule are on the outside of it, as is shown at v in Plate XLV, Fig. 3. We have seen that the blastomeres of the ganglionic rudiment, s, are at first continuous, above with the extra-follicular blastomeres of the ectodermal ridge, as shown in Fig. 5 of Plate XVI, where s marks the ganglionic blastomeres. At a lower level, Plate XII, Fig. 1, s, they are imbedded in follicle cells at the point on the middle line of the body W. K. BROOKS ON THE GENUS SALPA. 41 where the visceral follicle cells are continuous with those of the somatic layer. In Fig. 2, s, they are shut into the visceral layer, and the gang- lionic rudiment lies in the body cavity, 15. At a still lower level, Figs. 6 and 7, the blastomeres pass without any line of demarcation into those of the pharynx. In an embryo a little older, Plate XIII, we find essentially the same condition. In Fig. 3, s, the ganglion cells are extra-follicular; in Figs. 4 and 5, s, they gradually pass into the visceral layer of the follicle; in Figs. 6, 7 and 8, the ganglionic rudiment is within the body cavity, and in Fig. 9 it becomes continuous with the rudiment of the pharynx. In Plate XIV, Fig. 4, s, it begins to be shut in dorsally and separated from the somatic layer, while a cavity appears within it, as shown in Figs. 5 and 6, and in Plate XXII, Fig. 2, s. The cavity of the pharynx, c, is formed at the same time, and at first consists of a broad chamber, Plate XIV, Figs. 8 and 9, and a much narrower anterior portion, which is shown in Fig. 8 and, at an older stage, in Plate XVII, Fig. 1. This narrow portion, which is the stomodaeal portion of the pharynx, soon loses its distinctness, as the whole pharynx widens out directly up to the mouth, as shown in Plate XIX, Figs. 4 and 5, but before this takes place the stomodaeal diverticulum lies directly under the ganglion, as will be seen by comparing Figs. 6, 7 and 8 in Plate XIV, although the cavities of the two structures are at first separate, as shown in the intermediate section, Fig. 7. The follicle cells which separate them soon disappear, and the cavity of the ganglion opens into the cavity of the stomodaeal diverticulum of the branchial sac. This is the case at the stage which is shown in Plates XVI and XVII, where the cavity of the ganglion is shown at s in Figs. 2 and 3 of Plate XVI, while the stomodaeal diverticulum is shown in the same place in Fig. 1 of Plate XVII. The study of the intermediate sections, which are not figured, shows that there is now no partition between the two, and the relations between the two structures are as shown in a longitudinal section of an embryo a little older, in Plate XVIII, Fig. 8, s and z. The opening of the ganglion into the pharynx is also shown in Plate XVIII, Fig. 2, where s is the cavity of the ganglion and c the cavity of the pharynx. Plate XXXV, s, shows the ganglion at a still older stage, and its opening into the pharynx at t, in longitudinal section. The pericardial rudiment first makes its appearance in the longitu- dinal plate of the visceral mass, at/, in Plate XIV, Figs. 8 and 9, between 42 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. the notochord and the pharynx. It quickly becomes a large hollow vesicle, Plate XVIII, Figs. 1, 2, 3 and 4, /, which runs vertically behind the pharynx. On the side next the pharynx the heart, e, is formed as a vertical groove or fold in the wall of the pericardium, which ultimately becomes converted into a tube by the union of its edges, as shown at c in Plate XIX; the union first taking place in the middle, and extending towards the ends, which are permanently open and communicate with the space of the body cavity in which the blood circulates. As the heart becomes shut in, the follicle cells which lie between it and the wall of the pharynx, as shown in Plate XVII, become folded into it, as shown at e in Plate XIX, Figs. 8 and 9, and they then become vacuolated, and ulti- mately disappear. At the stage shown in Plate XIX the heart has essentially its adult form, and the series of sections illustrates its position and anatomical structure with sufficient clearness. Figure 1 is near the top of the heart, and shows its opening into the body cavity, which is shown in Fig. 6 more enlarged and filled with blood corpuscles, which are formed in the body cavity and are drawn into the heart by its pulsations. Fig. 2, which is shown more magnified in Fig. 8, and Fig. 3, which is shown more magnified in Fig. 9, show the structure of the middle region of the heart, while Fig. 5 shows its inferior opening into the body cavity. The heart changes its position with the growth of the embryo. It is at first behind the pharynx, as shown in Plate XLI, Fig. 2, /, but as the pharynx elongates it pushes the heart down and grows over it, as shown in Plate III, Fig. 4, e and /, where the heart is in its adult position under the posterior end of the pharynx. I have not been able to trace the origin of the mesoderm, and have first found it at the stage shown in Plate XLII, Fig. 8. At the lower part of this figure a number of small cells, with deeply stained nuclei, much smaller than those of the follicle cells, are shown in the body cavity, y, outside the visceral follicle cells, and other similar cells, mes, are shown arranged in an epithelium on the surface of the visceral cells, 8. These are mesoderm cells, some of which are shown more magnified at A in Fig. 9, and in Plate XLII, Fig. 7. Their position seems to indicate that they are derived from the endodermal blastomeres of the pharynx. SECTION 6. The Degeneration of the Follicle. We must now turn back from the history of the embryonic cells and their derivatives and trace the fate of the follicle cells. At a very early W. K. BROOKS ON THE GENUS SALPA. 43 period in the history of the embryo some of the visceral follicular nuclei wander in among the blastomeres, or push into their substance, as is shown in Plate XLII, Fig. 1, and there degenerate, as has already been described. The stage which is shown in Plate XII and in cut B is the starting point for a description of the fate of the others, since the follicle is at this time most fully developed, and consists, as already described, of a somatic layer, 7, which forms the outer wall of the embryo ; a visceral mass, 8, and the somatic lining of the perithoracic structures, g, g', g", g'" and g". The somatic layer of the follicle is divisible into two parts, the upper portion, cut A, 7, which is covered by the epithelial capsule, B', and the lower part, 10, which is exposed to the body cavity, y, of the chain-salpa. These two regions are so different in their history that they must be treated separately. The lower portion, 10, forms the roof of the placenta, cuts A and B, 10, and, as is shown in cuts C and D, it soon loses its continuity with the upper portion, and establishes a union with the upper edge of the supporting ring, 23, which, as I have shown, itself loses its continuity with the epithelial capsule, B', and bends inwards, as shown in cut C, to unite with the follicular roof of the placenta. In Plate XII, the cavity of the placenta, colored yellow, is shown at y" in Figs. 2, 4, 5, 6, 7, 8 and 9, and the supporting ring, 23, which is colored red, is shown in the same figures, where the part of the somatic layer of the follicle which is to enter into the formation of the roof of the placenta is marked 10. The relations between these various structures are so well shown in the figures and in the reconstructed cross section in cut B that description seems unnecessary, but we must note that the upper edge of the placenta is not a flat circle, but that it reaches much farther up on the sides of the embryo than it does on the middle line, and that the line which separates the placental portion of the somatic layer from the embryonic portion follows the same course, as does also the zone of transition from the epithelial capsule to the supporting ring. Comparison of cuts A, B, C and D also shows that as the embryo grows, the roof of the placenta becomes flat, and that the embryo itself pushes upwards from the surface which is thus formed. The fate of the follicular cells of the roof of the placenta will be described in the account of the placenta, and we have now to consider the fate of those follicle cells which are more intimately bound up in the structure of the embryo. 44 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. At the stage shown in Plate XII, the somatic follicle on and near the middle line, just above the placenta, at 7, in Figs. 4 and 5, begins to lose its cell outlines and to become thickened and indefinite, and, as is shown at 7 in Plate XIII, Fig. 9, it very soon breaks up into its con- stituent cells, which separate from each other and become amoeboid, although most of them remain in the places which they occupied at an earlier stage. This separation of the cells is accompanied by the formation of a transparent gelatinous substance between them, so that they become converted into a cartilage-like substance which lies around the circumference of the body cavity of the embryo just above the placenta, and most abundant near the middle axis, as is shown at fc in Plate XVIII, Figs. 1 and 8, and also in Plate XXXV. The disintegration of the somatic layer gradually extends upwards, as is shown in Plates XIII and XIV and in cuts C and D, but as the cells break apart and become free they wander into all parts of the body cavity, and all traces of the somatic layer as a distinct tissue soon disappear, although, as shown in Plate XIII, Figs. 3, 4, 5 and 6, its outline is preserved for a short time after the cells are completely separated. The body cavity, which appears to be empty, is undoubtedly filled with a fluid or semi-fluid serum, of greater density than water, and sufficiently firm to hold the amoeboid cells and to keep them floating. By the time that the pharynx appears, Plate XIV and Plate XXII, the somatic layer is completely broken down. Plate XLII, Fig. 11, is part of a transverse section across the middle line of the dorsal surface of an embryo at the time when the last remnant of the somatic layer is going to pieces, and it shows the way this takes place. In this figure 21 and 22 are the folds of the embryo sac, &' is the epithelial capsule, a is the ectodermal rudiment, 15 is the body cavity, and 8 the follicle cells. These are multiplying rapidly by direct division of the nuclei, and their exposed ends are irregular and amoeboid. Many of the cells break away bodily, while others divide into a portion which retains its position, and a portion which splits off as a free amoeboid cell. Multiplication of the nuclei goes on in the detached cells, as well as in those which are fixed, and two nuclei are usually present in most of the wandering cells. Thus the embryonic portion of the somatic layer of follicle cells dis- appears as a distinct layer, and at the stage shown in Plates XVI and XVII, and in Fig. 2 of Plate XLI, it is represented only by migratory amoeboid cells scattered through the body cavity of the embryo. W. K. BROOKS ON THE GENUS SALPA. 45 The next part of the follicle to disappear is the somatic lining of the perithoracic system. As shown in Plate XLII, Figs. 6 and 7, the cells of this layer become detached and acquire amoeboid outlines before the gill- slits acquire their openings into the pharynx, and as I have already stated, all of them ultimately separate and pass into the cavity of the cloaca, which becomes so well filled with them that at the stage shown in Plate XVIII, Fig. 8, the outline of the cloaca is hard to trace in the sections. The details of the process of disintegration, which are shown in Plate XLII, Fig. 8, g", are so much like the history of the somatic layer just described, that no further account seems necessary. The disintegration of the visceral mass of follicle cells begins with the formation of the cavity of the pharynx, as already described and figured at c in Fig. 8. After the blastodermic epi- thelium of the pharynx is formed, most of the visceral follicle cells are left outside it in the body cavity, as shown at 8 in Plate XVII, Figs. 1, 2 and 3, where they form a secondary wall outside the blastodermic epithe- lium, as they do also around the gill-slits, Plate XVI, Fig. 3, and the cloaca, Fig. 2. All these cells ultimately degenerate and disappear, but while most of them first become migratory and amoeboid, as shown at 8 in Fig. 9 of Plate XLII, others become vacuolated and break down and disappear without any migration, as for example those which are shown in Plate XVII, Fig. 6, included between the epithelium of the cloaca, g'" and the ectoderm a, at the point where the cloacal aperture is to be formed. The history of the process of degeneration is as follows : The large irregular amoeboid follicle cells, usually with two nuclei, Plate XX, Fig. 5, wander into all parts of the body cavity and become very much vacuolated, and while this is going on the small amoeboid mesoderm cells of the embryo lodge upon their surfaces, as they do upon the surfaces of the embryonic cells, and give rise to a fibrous capsule around each follicle cell or group of cells, as is shown in Plate XX, Figs. 5 and 6, Plate XXI, Figs. 1 and 2, and Plate XIX, Figs. 9 and 10. Mesoderm cells or blood corpuscles are often found also in the vacuoles of degenerating follicle cells, as in Plate XX, Fig. 5. As the cells disappear they leave behind them an empty meshwork of fibers, as is shown in Plate XXI, Fig. 2. While it is of course impossible to trace the history of each follicle cell, I think there is sufficient evidence to warrant the statement that, while the organization of the embryo is at first blocked out in follicle cells, all of this scaffolding is afterwards torn down ; for I have found follicle cells in the act of detachment in all the regions of the embryo and in all the f ollicular structures. 46 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. SECTION 7. The Placenta. We have now to trace the history of the placenta, and as this organ is much simpler in Salpa hexagona than it is in Salpa pinnata, I shall first describe the origin and fate of the placenta of Salpa hexagona. The part of the follicle which is bathed by the blood of the chain- salpa begins to grow, Plate XI, Fig. 3, as soon as the rest of the surface of the embryo is covered by the epithelial capsule, and it soon becomes many cells thick, and irregular folds and spaces appear in it. As soon as the walls of the placental chamber, Plate XLV, Fig. 1, are formed by the supporting ring, 23, the thickened portion of the follicle grows down into it as a ring, 31, around its sides, and as a free pendant mass, 24, which Barrois has compared to a bell-clapper. As the embryo grows, and the cavity of the placenta becomes larger, Plate XLV, Figs. 2, 3 and 4, cell multiplication goes on very rapidly in these structures, until the chamber is completely filled by convoluted inosculating strings of cells, so arranged as to break up the blood space into a number of tortuous channels, in which the circulation is retarded, so that the plasma and the blood cor- puscles are almost brought to rest, as they find their way through these obstructed passages. Figs. 2, 3 and 4 of Plate XLV are sections through the placenta and embryo of Salpa hexagona at three successive stages of development, but they will also serve to illustrate the structure of the placenta, since they cut it in three different planes. Figure 4 is from an embryo at the stage of Plate III, Fig. 4, passing, through the neck of the placenta and through the gill, o. Comparison with Plate III, Fig. 4, will show that it is a vertical transverse section in the vertical axis of the placenta. Figure 3 of Plate XLV passes through the neck of the placenta and through the ganglion, s, and while it is from an embryo con- siderably younger than Fig. 4 of Plate III, comparison with this figure will show that it must cut the greater part of the placenta in front of the median plane of Fig. 4. Fig. 2 is from a still younger embryo, passing through the neck of the placenta, and through the place of the cloacal aperture, g\ At this stage this point is very much farther forward than it is in the embryo shown in the figure on Plate III, and nearly where it is shown at g v in the embryo of Salpa pinnata shown in Fig. 3 of Plate XLI, but examination will show that the section must cut the greater part of the mass of the placenta about as much behind the plane of Fig. 4 as the section in Fig. 3 is in front of it. While all these sections pass through the neck of the placenta, the upper part of Fig. 3 is in effect a vertical W. K. BROOKS ON THE GENUS SALPA. 47 transverse section through the anterior portion, Fig. 4 through the middle portion, and Fig. 2 through the posterior portion of the body of the placenta. The cellular strings become more and more numerous and crowded as the placenta grows, as is shown by the series of stages, but they are always most developed in the middle plane, as is shown in Fig. 4, while in front and behind the spaces are much larger, and the cellular strings more independent. At the stage shown in Fig. 1 the blood circulates vaguely back and forth into and out of the placenta, but hori- zontal sections show that at the stages of Figs. 2, 3 and 4 the neck of the placenta is divided into two openings, an anterior and a posterior, by a median transverse partition. This partition is not flat, but folded in such a way that no single vertical section can show the whole of it, but each series shows it at all positions between that of Fig. 2, where it is united below the neck of the placenta to the wall of the cloaca on the left, to Fig. 3 where it joins it on the right. The partition is continuous with the " blood-bud," 24, and is formed from a substance which appears to be the same as the cellulose mantle. This gelatinous substance is formed in many parts of the body of salpa, between the mesodermic endothelium of the body cavity and the endo- dermal structures. It is shown, for example, at 32 in Figs. 3 and 5 of Plate XXXIV. This partition divides the cavity of the placenta into an anterior blood chamber, Plate XLV, Fig. 3, bl, 1, and a posterior chamber, Fig. 2, bl, 2, which communicate with each other through the spongy mass formed by the plexus of strings of cells which fills the middle portion, as shown in Fig. 4. In Salpa pinnata the partition is formed by the "blood-bud," 23, itself, which in advanced embryos fits like a plug into the neck of the placenta in transverse section, Plate XVIII, Fig. 4, while in longitudinal section, Fig. 8, or in a transverse section in front of the plug, Fig. 2, or behind it, Fig. 5, there is a free channel for the entrance or exit of blood, as is shown also in Plate XXXV, and in the series of horizontal sections from the same stage shown in Plate XLVI, Figs. 2, 3 and 4. The blood which enters the placenta behind this partition and passes into the chamber marked bl, 2, must make its way slowly through the spongy mass of cells before it can gain access to the anterior chamber, bl, 1, and escape from the placenta, and the conditions are of course essentially the same when the circulation is reversed. The general anatomical structure of the placenta of Salpa pinnata can be understood by comparing the 48 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. longitudinal section in Plate XXXV with the transverse sections in Plate XVIII, and the horizontal sections in Plate XLVI, Figs. 2, 3 and 4. In a horizontal section near the top, Fig. 2, the cavity is cut up by strings and clumps of cells which have no constant arrangement, although the study of sections at a lower level shows that certain channels, bl, 1 and bl, 2, communicate with one or the other of the two chambers which open into the blood system of the chain-salpa, while others are channels of communication between one of these chambers and the other. At a somewhat lower level, Fig. 3, the strings of cells gradually unite to form a transverse partition which separates the anterior chamber, bl, 7, from the posterior, bl, 2, and these chambers gradually become more and more capacious, and the blood channels in the partition less and less numerous, until, near the neck of the placenta, Fig. 4, the partition becomes, in longitudinal sections, a straight rod, running across the placenta like the handle of a basket. In a section just above the neck of the placenta the ends of this rod are separated from the supporting ring by a space, 33, which is also shown at 33 in Plate XXXV. This space runs around the whole circum- ference of the placenta, as shown in Fig. 4, and it is bounded internally by the endothelium of the placenta and externally by the supporting ring. It is interrupted at short intervals by fibers which cross it, and it is lined by very small cells, very much smaller than the blood corpuscles. This space acts, perhaps, as a valve to close or expand the opening of the placenta, although sections of preserved specimens throw little light on its function. The strings of cells are nourished by the plasma of the blood which is retarded in the meshes of the spongy mesh work, and also by the blood corpuscles; and in both Salpa pinnata and Salpa hexagona, blood corpuscles may be found settling upon the surfaces of the strings and sinking into their substance, as is shown in Fig. 5 of Plate XLVI, which is a highly magnified drawing of part of Fig. 3. SECTION 8. TJie Nutrition of the Embryo. As the mammalian placenta nourishes and aerates the blood of the foetus by the diffusion of gases and food in solution through the walls of the blood-vessels, it has been generally taken for granted that the placenta of salpa performs its function in the same way, and it has been described as divided into a foetal chamber and a maternal chamber, although its cavity is in reality part of the body cavity of the chain-salpa, and the blood which circulates in it that of the chain-salpa. The salpa W. K. BROOKS ON THE GENUS SALPA. 49 embryo is bathed by the water which is constantly flowing past it, and it is therefore in very much closer relation to the external world than a mammalian embryo, shut up in the interior of a large thick-walled body. There does not seem to be any need in salpa for a respiratory placenta, and its thick spongy walls seem to indicate that it is not respir- atory. We find in its structure nothing like the interlacing villi of the mammalian chorion, and the sections show that the embryo is nourished in a way quite unlike anything which has been described in the mam- malia. The subject is a very interesting one. The rapid growth of the salpa embryo is one of its most conspicuous characteristics, and the nutrition which this rapid growth demands is secured by two very peculiar organs, the follicle and the placenta. While the egg at the time of fertilization is very minute, the embryo at the time of birth is enormous, as compared with the size of the chain- salpa which carries it, and it certainly increases many thousandfold during development. The growth is only partially due to cell multipli- cation, and it is in part a result of the growth of the individual cells, for instead of growing smaller with repeated division, they actually increase in size in all parts of the body. As the older stages are less magnified in the figures than the younger ones, this growth of the cells is not conspicuous in the figures, but it is one of the most notable peculiarities of the salpa embryo, and in many parts of its body cells as large as the original ovum are found. The growth sets in very early, and it goes on uninterruptedly throughout the whole foetal life, so that the embryo becomes gigantic as compared with the body of the chain-salpa which contains it. Quoy and Gaimard describe an embryo, two inches long at birth, in a salpa (S. forskalii) a foot long, and Leuckart says that the embryo of S. democrat] ca at birth is two-fifths as long as the chain-salpa which carries it. The fully grown embryo of S. hexagona is almost as long in comparison with the chain form of the same species. It is not unusual for the embryos of viviparous animals to gain slightly in size and weight before birth, but, as Leuckart points out, the mammals are the only animals which exhibit anything comparable to the rapid growth of the salpa embryo from a minute egg, and the history of the salpa embryo at once calls to mind the growth of the embryo in the placental mammals; nor is this resemblance entirely superficial, for in both the mammal and in salpa we find an especial foetal organ, the 50 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. placenta, for the purpose of affording to the growing embryo an abundant supply of nutriment. The resemblance between the fostal life of salpa and that of a mam mal is most remarkable, and it is all the more noteworthy since we may be absolutely confident that the placenta of salpa is an independent acquisition, entirely without genetic relation to that of mammals. No modern writer except Todarro has ventured to regard the two structures as homologous, and their phylogenetic independence is so obvious that it is not necessary to discuss it, although a greater physio- logical and anatomical resemblance than the facts warrant has usually been assumed. We should hardly expect fundamental similarity in structures of diverse origin. On the contrary, we might reasonably look for profound differences between the placenta of salpa and that of the mammals. The various writers on salpa, while recognizing this fact, and while pointing out the great differences in the way in which the placenta is formed in the two cases, have nevertheless assumed, either explicitly or by implication, a much greater resemblance to the mammalian placenta, in structure and in function, than actually exists. The later writers say very little about the function of the placenta of salpa, but they assume a fundamental similarity to its function in mammals. So far as it is in both cases an organ for supplying the embryo with nutritive matter, derived from the blood of the supporting organism, the resemblance is real, but it goes no farther than this, and the way in which the nourishment is conveyed to the embryo is totally unlike ; a fact which has never been described nor even noted. In the mammalian placenta the blood of the embryo, as it circulates through the villi of the chorion, is brought into such close contact with the blood of the mother, that diffusion takes place through the separating walls, and thus the blood of the fostus is oxidized, relieved of its waste products, and supplied by diffusion with nutritive matter in solution. Notwithstanding the very intimate union between the blood-vessels of the foetus and those of the mother, there is no direct communication between them, and nothing except gases and liquids can pass from the body of the parent to the body of the child, without the violent rupture or perforation of the walls of the vessels, unless, perhaps, some very minute bacteria are an exception. It has been generally assumed that this must be true of salpa also. Thus Barrois says, incidentally and very briefly (4) p. 495, that the func- W. K. BROOKS ON THE GENUS SALPA. 51 tion of the placenta of salpa is to bring about by osmosis an interchange of fluids between the blood of the parent and that of the embryo, as in the placenta of a mammal. The subject has received very little attention, but as no one has ever commented upon the view set forth at considerable length by Leuckart (1) pp. 61 and 62, this may be regarded as the accepted view. He says: "The histological differentiation of the organs and tissues of the embryo is accelerated, to a high degree, by the circulation in the body of the young salpa, which is completely separated from the circulation of the mother. At no time does the blood of the mother pass through the wall of the placenta into the body of the embryo. The transfusion between the mother and the foetus is, as in the mammals, purely endosmotic, through the substance of the placenta, and it is most essentially facili- tated by the movement of the blood, both in the embryo and in the chain- salpa. " The upper wall of the placenta, which is the peculiar seat of the process of diffusion, projects into the body of the embryo, and is sur- rounded by the median ventral blood sinus. As the blood corpuscles of the embryo are much smaller than those of the chain-salpa, it is easy to see that no mingling takes place." It is probably true that no transfusion of blood corpuscles takes place, and it is difficult to show from the study of sections of hardened specimens that no serum from the blood of the chain-salpa is diffused through the wall of the placenta, although its great thickness seems to be a very unfavorable condition for this purpose, and I shall show farther on that the mechanism of nutrition is very different from that of mammals ; that this is effected by the actual migration of great placenta cells, Plate XLV, Fig. 4, 20, into the body cavity of the embryo. The pla- centa is an organ for the nourishment of the placenta cells by the blood of the chain-salpa ; and the subsequent degeneration of these cells, after they have migrated into the body of the embryo, supplies the material for the growth of the embryo. This is in all probability the only function of the placenta, for there does not seem to be any need for an especial apparatus for oxidation, or for the removal of waste products. The salpa embryo stands in much more direct relation to the external world than the mammalian embryo. It projects into the cloaca of the chain- salpa, and is freely exposed to the constant current of fresh sea-water which flows around it, and its thin surface seems to be much more favorable than the thick wall of the placenta for the diffusion of gases. 52 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. During the later stages of foetal life its own mouth is open, its muscles contract, and there is no reason why it should not breathe for itself exactly like an adult. I therefore regard the placenta as a nutritive organ; pure and simple, and it serves its purpose not by the diffusion of a fluid, but by the transportation of solid food into the body of the embryo. From this point of view it is clear that those investigators who have described it as divided into a foetal chamber and a maternal chamber have been misled by an erroneous notion of its function. The detachment of the placenta cells has been observed and noted by both Salensky and Barrois, but it has been regarded as a destructive change and as a sign that the organ has served its purpose and has become superfluous. It has been assumed that it reaches its perfect form and serves its purpose, and that it then degenerates and breaks down, and no import- ance has been attached to the process of degeneration, as it has not been regarded as significant. No note has been made of the very early stage at which degeneration begins, nor of the fact that it is initiated as soon as the embryo begins to grow, and long before it has reached half or a quarter of the size which it is to have at birth. This is hard to explain so long as the disintegration of the placenta is regarded as its destruction, but it becomes quite intelligible as soon as we learn that the detachment of the placenta cells, instead of marking the end of its functional life, is actually a manifestation of its useful activity. As the figures in Plate XLV show, the strings of cells multiply at their lower ends by direct division of their nuclei, and as the new cells which are thus formed push up towards the top, they grow very large, while their nuclei become filled with diffused chromatin granules. In Salpa hexagona these cells ultimately reach the top of the placenta, where they gradually become elongated and irregular, and then break through into the body cavity of the embryo as the migratory follicle cells which are shown at 29 in the figures. While the details are slightly different in Salpa pinnata, placenta cells migrate bodily into the embryo in the same way, and they are shown in many of the figures, as in Plate XVIII, 29, for example. The rapid growth of the embryo seems to be most important to salpa, and while we know almost nothing of its birth rate, the quickness with which the surface of the ocean becomes covered with salpse of all W. K. BROOKS ON THE GENUS SALPA. 53 ages in a long calm, shows that the animals are most prolific, and the complicated structure of the organs for nourishing the embryo shows that every provision is made for rapid growth. The placenta is not the only nutritive organ, for, as we have seen, the follicle also makes most important contributions to the supply of material which is available for the construction and rapid completion of the body of the embryo, and while I have spoken of the segmentation and the formation of the blastodermic germ layers as retarded, the retardation is probably not actual, but only relative, and the process of development is, on the whole, accelerated by the presence of the follicle, and by its share in the growth of the embryo. I have now shown that the ultimate fate of all the follicle cells is the same, and that they may be found, in the sections, detaching themselves and degenerating, first, in the somatic layer of the embryo ; secondly, in the somatic follicular lining of the perithoracic structures ; third, in the cavity of the pharynx ; fourth, in the visceral mass outside the digestive cavity, and last, in that part of the placenta which is derived from the somatic layer of the follicle. While it is not possible to trace the history of every cell from first to last, we have as ample evidence as we could hope from sections, that the function of the follicle of salpa is exclusively nutritive; that it is transitory and embryonic, and that the tissues of the embryo are not built up out of follicle cells, but from blastomeres, after the analogy of all the rest of the animal kingdom. CHAPTER III. THE MORPHOLOGICAL SIGNIFICANCE OF THE SALPA EMBRYO. A basis for the comparison of the salpa embryo with even its closest allies is hard to find, for although it is still a true embryo and not a bud, its early stages have been profoundly modified by secondary changes. Salensky holds indeed, (5) 396, that a knowledge of the develop- ment of other animals does not help to clear up the obscurity which involves the salpa embryo ; that its peculiarities are so different from all that we know of other animals that we must not hope to bring it into the general scheme of animal embryology, and that, while it begins its development by the sexual method, this soon gives place to budding from the wall of the follicle. I have shown that this view is untenable, and that the embryology of salpa is not totally and fundamentally irreconcilable with the princi- ples of general embryology, although it is quite true that the nature and origin of the secondary changes are most perplexing subjects. As our starting-point in making comparisons, we may safely assume that the fundamental plan of development was originally that of the Tunicates in general, but we have very few facts to show the way in which the complications were introduced. Salensky believes that the embryology of Salpa bicaudata is less modified than that of the other species, but only as regards accessory structures. As regards the peculiar history of the blastomeres and migratory follicle cells this species is like the others, and there is, so far as we know, no species which presents a transitional stage in this history. Nor can we get much help from Pyrosoma or Doliolum, the two nearest allies of Salpa, for while the life-history of Doliolum, with its invaginated gastrula and its tailed larva, is possibly more primitive than that of Salpa or Pyro- soma, it presents no trace of the distinctive peculiarities of either of them, and it therefore affords no better basis for comparison than the ordinary Tunicates. The embryos of Salpa and Pyrosoma represent two distinct lines of secondary modification, and neither serves as an explanation of the other. W. K. BROOKS ON THE GENUS SALPA. 55 We therefore have only a slight basis for a phylogenetic history of the modifications which the salpa embryo has undergone in reaching its present form. We must remember, though, that it is possible to make instructive and valuable comparisons even when they do not lead to exact or definite phylogenetic conclusions, as when we compare adult echinoderms with each other without committing ourselves to any view of their ancestral relationship ; and I think we may give clearness and definiteness to our conception of the salpa embryo by comparing it with the embryos of other Tunicates, although we may not be able to mark out the path it has followed in reaching its present structure. We cannot have a clear notion of structure without comparison, although we may make note- worthy progress without historical data. Even if we were totally without evidence as to the history of the secondary modifications which the salpa embryo has undergone, their morphological nature could be studied by comparing it with the embryos of other Tunicates, and I shall show that we do have some evidence, although it is true that this is scanty. The Embryology of Primitive Tunicates. The embryology of Pyrosoma, the nearest relative of Salpa, is com- plicated by the early degeneration of the embryo, and by the early appearance of asexual multiplication by buds. Both these peculiarities are secondary, and they have been acquired, or at least very much accented since Pyrosoma and Salpa diverged from their common ancestor. We may therefore leave the four Ascidiozooids of the Pyro- soma embryo out of consideration, and take the Cyathozooid as a basis for comparison with the salpa embryo, but we must do away in imagi- nation with the degeneracy of the Cyathozooid, and must picture it as an embryo with the potency of an adult, like the embryo which becomes a solitary salpa. We may also assume that, at some time in the past, these embryos, like the embryo of Doliolum, passed through an Appendicularia-like larval stage, corresponding to the tadpole larva of other Tunicates. Still further back, we must believe that at some remote period the ancestral embryo of all the Tunicates and of all other chordata passed through an invaginated gastrula stage, formed by the total regular segmentation of a holoblastic egg. We are also forced, by all the facts of embryology, to 56 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. believe that the whole organization of the body, in the most complete form which it had then attained, was latent or potential in the fertilized egg, and was formed from it by cell multiplication. Furthermore, we may be sure that, for a time, all the chordata origi- nally followed the same line in their ontogenetic development from the egg ; that all the Tunicates retained a common life-history for a longer time ; that the embryos of Salpa, Pyrosoma and Doliolum were originally alike for a little longer, and that the differences between the embryos of these three forms are the results of more recent modification. These statements involve no opinion on the exact character of the relationship between the ordinary Tunicates on the one hand, and Salpa, Pyrosoma and Doliolum on the other, and no opinion on this point is necessary for our purpose, which is to study the morphological signi- ficance of the salpa embryo. Has the Egg of Salpa passed through a Stage with a Large Food- Yolk? Pyrosoma has a big food-yolk and a disk-shaped blastoderm ; and the arrangement of the germ layers and the anatomical relations of the embryo are profoundly modified by its presence. Was this yolk acquired before or after Pyrosoma and Salpa diverged from each other? The salpa egg has nothing of the sort now, and it undergoes total segmentation, but if the yolk was acquired before the divergence of the two life-histories, we must, in our comparative study of the salpa embryo, take into consideration the inherited results of its existence in the ancestors of this embryo. The question cannot be definitely answered, but the incompleteness of the ectoderm and the slow closure of the floor of the pharynx of salpa are what we might expect if a food-yolk has once been present. In an ordinary gastrula the continuity of the germ layers is complete, but it will be seen by comparing the figures in Plates XVI, XVII, XVIII and Figure 11 of Plate XLII, that the ectoderm of salpa has a growing edge, and that it gradually spreads out on all sides over the embryo until its growth is stopped, in the older stages, by the placenta. The starting- point from which the ectoderm spreads is on the middle line of the dorsal surface of the embryo around the region which I shall soon give my reasons for regarding as the place of the blastopore. In meroblastic eggs, like those, for example, of Teleosts and Birds, it is well known that even after the germ layers are established they have W. K. BEOOKS ON THE GENUS SALPA. 57 an external or peripheral boundary and a growing edge, and that the inclusion of the yolk is a very slow process. From Salensky's statement (17), p. 455, that "outside the limits of the germinal area the yolk is perfectly naked," I infer that in Pyrosoma it never becomes completely inclosed by the ectoderm of the cyathozooid, but that this degenerates before the yolk is covered. This secondary adaptation to the presence of a food-yolk is so well understood that we need not dwell on it. So far as I am aware, the salpa embryo is the only one without a food-yolk which is known to have an incomplete ectoderm with a growing edge, and the incompleteness of the floor of the digestive tract is another fact of exactly the same kind. There is therefore good reason for believing that salpa is descended from a form with a food- yolk, like that of Pyrosoma, which has afterwards been lost. The acqui- sition of new methods of nourishing the embryo by means of migratory follicle cells and a placenta is a satisfactory explanation of the way in which the need for a food-yolk was done away with, and its disappear- ance is therefore quite intelligible. In considering the influence of a food-yolk upon the structure of the embryo, we must have a clear idea of what the books call its "morpho- logical position." To my mind, the way this term is used by writers on the embryology of vertebrates is open to criticism. The fact that the unconsumed remnant of the yolk of vertebrates is surrounded by endo- derm or inclosed in endoderm cells, is no evidence that it was laid down in the egg in any relation to the regions of the body of the future embryo, or in any "morphological position" whatever. The series of stages in the phylogenetic history of the acquisition of a food-yolk is a series of eggs, not embryos. In secreting the yolk the egg functions as a cell, not as a potential embryo, and there is no reason to believe that the yolk material is laid down in any definite relation to the structure of the latent embryo. Its distribution is probably determined by the structure of the egg as a cell, and the place which the unassimilated remnant occupies in the body of the embryo depends upon the physiological activity of cells of the embryo itself. In fact, as all the cells of a young amphibian embryo are packed with yolk, the yolk of an amphibian egg has no " morphological position." The yolk of Pyrosoma is not inherited from a common source with that of true vertebrates, and there is, of course, no reason why it should be assimilated in exactly the same way. I believe that a careful study of Salensky's figures will show that it is in the body cavity, and not in the digestive cavity of the embryo, for no layer of endoderm ever extends over any considerable part of it. 58 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. If the salpa embryo has ever had a food-yolk it undoubtedly agreed in all respects with that of Pyrosoma and lay in the body cavity. It may, perhaps, seem to some that the incompleteness of the germ layers of salpa may have been acquired as an adaptation to the presence of the follicle or the placenta, and that it may therefore be quite inde- pendent of the same phenomenon in Pyrosoma, but this view cannot be seriously urged, for while we may believe that the ectoderm is potentially present, in an undifferentiated state, in the surface of the yolk outside the growing edge, as it certainly is in young amphibian embryos, we cannot believe that it is potentially present in the follicle of salpa. I therefore think that we may safely assume, from the incomplete- ness of the germ layers of Salpa, that its ancestors had a food-yolk like that of Pyrosoma. The Primitive Salpa Embryo. Whatever view of the food-yolk we take, there can be no doubt whatever that, far back in the past, the ancestor of both Salpa and Pyrosoma had simple holoblastic eggs like those of Clavelina. This is a necessary deduction from the principles of comparative embryology, and it is also supported by the fact already pointed out that the salpa egg shows the same type of segmentation as that of Clavelina. We may therefore safely assume, as the point of departure for our comparative study of the embryo, a life-history like that of the primitive chordata, where the holoblastic egg undergoes total regular segmenta- tion and gives rise to a hollow blastula, from which a gastrula is formed by invagination. From the dorsal part of the endodermal wall of the primitive diges- tive cavity the notochord was formed, and the nerve tube arose from the dorsal median ectoderm. At least one pair of ectodermal ingrowths united with paired endodermal outgrowths from the pharynx, to form the pharyngeal clifts through which the digestive cavity opened to the exterior. The region of the tail elongated and the embryo became an active locomotor chordate animal. So far the characters of the primitive salpa embryo are common to the chordata. Furthermore we must assume that in the larval stage of all known Tunicates, except Appeiidicularia, the ectodermal portions of the gill- tubes moved towards the middle line, and united to form a common dorsal cloaca, into which the digestive and reproductive organs came to open. W. K. BROOKS ON THE GENUS SALPA. 59 The embryology of Salpa furnishes, as I shall show in another sec- tion, still further evidence regarding the phylogeny of the adult, but we are here concerned only with the comparative study of the embryo. The Origin and Significance of the Follicle of Salpa. The most peculiar and anomalous feature in the life-history of Salpa is the follicle, but even here we are not absolutely without means for comparative study. In numerous, widely separated members of the animal kingdom, follicle cells migrate into the substance of the ovarian eggs, and supply them with food. Many instances are found among the Tunicates ; Cla- velina for example ; and, as we have seen, Salpa must be added to the list. Salpa undoubtedly inherits this peculiarity, which is also found in Doliolum, from their common ancestor, and very probably from a still more remote ancestor, common to the other Tunicates and to Appendicu- laria. Kowalevsky (Arch. f. Mikros. Anat., Band XI) was the first to show that the migration and degeneration of follicle cells goes on in Pyrosoma after the egg is fertilized. He says that great numbers of follicle cells separate from the follicular epithelium and wander into the space between it and the developing egg, as free inner follicle cells. Some of them gather in groups external to the germinal disk, while others work their way in between the segments and are for a long time distin- guishable from the blastomeres. Finally, he says (p. 607), these cells as well as the yolk are surrounded and inclosed by the blastoderm and turned to nutritive material or to blood corpuscles. While Salensky believes that instead of nourishing the embryo these migratory follicle cells take part in its construction, his account of their origin is essentially like Kowalevsky's, except that he shows that besides those which make their way between the blastomeres, many others pass under the edge of the blastoderm and accumulate in the segmentation cavity, or the space between the blastoderm and the yolk, and that others wander into the substance of the yolk. Cuts E and F are two early stages in the segmentation of Pyrosoma, copied from Salensky's memoir. They are sections through the blasto- derm, and the food-yolk is not represented. It should occupy the space below the figures. In my pen-copy of his figures I have dotted the migratory follicle cells to make them more conspicuous. In cut E they 60 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. are shown migrating into the segmentation cavity under the edge of the blastoderm, and filling the space between it and the yolk ; while in cut F they are shown pushing in between the blastomeres, and some of them are embedded in their substance. CUT E. CUT F. In Salpa there are two periods of migration. One, shown in Plate XXXI, Figs. 5 and 6, which has never before been described, is very early in the history of the ovarian eggs, when the migratory cells are assimilated by the egg cell, and thus furnish the material for its growth. A second period of migration, which was discovered and minutely described by Salensky (5), begins immediately after fertilization, and the migration goes on for a long time with great energy and rapidity on an extensive scale, as shown in my Plates IX and X. Pyrosoma shows this migration of the follicle cells into the substance of the embryo in a simpler form than Salpa, and while there may be no conclusive evidence that the life-history of Salpa has come about by the modification of that of Pyrosoma, there can be no doubt that the two have had a common starting-point, and that, so far as the migration of follicle cells goes, the Pyrosoma embryo is nearer this starting-point than the Salpa embryo. It may therefore be used to interpret the structure of the salpa embryo, and I shall attempt this without assuming that the Pyrosoma embryo is in the direct line of modification. We have earlier stages in the evolution of the migratory follicle than that exhibited by the Pyrosoma embryo. In cuts Gr and H, I have copied from Davidoff two figures of early stages in the segmentation of Dis- taplia magni larva. Cut Gr is an egg which has divided into two blasto- meres, and cut H is one in which the ectoderm and endoderm are differ- entiated. The egg is surrounded by a capsule of follicle cells, 6, which are equivalent in ultimate origin to the germ cells, as they are in Salpa. In Distaplia, according to Davidoff, the layer of cells by which the growing egg cell becomes surrounded are so directly derived from cells which are destined to become eggs, that this author calls them abortive eggs. W. K. BROOKS ON THE GENUS SALPA. 61 During the early stages of segmentation some of the cells which surround the egg of Distaplia pass into the fissures between the blasto- meres (p. 536) in much the same way that they do in Pyrosoma, but on a much less extensive scale. Ultimately, after segmentation is well advanced, (p. 548), these cells enter into the substance of the endodermal blastomeres, and Davidoff has no doubt (p. 549) that they serve as food for the endoderm cells. COT G. CUT H. In cut G the outer follicle, 6, is separated from the surface of the egg by an intervening space, which is filled with free follicle cells, 7, or, as the author calls them, abortive eggs, which have migrated into it from the capsule. As the blastomeres are marked off by segmentation these cells wander into the spaces between them, where some of them are shown at 5. Except for these spaces filled by migrating cells, the blasto- meres are in contact, and there is no empty segmentation cavity. As development goes on the migrating cells penetrate the substance of the blastomeres, as shown in cut H, and are used up as food, so that at last all traces of the follicle disappear. In Distaplia the blastomeres are so much larger than the migratory cells, at the latest stage when these are found, that the two sorts of cells are easily distinguishable, and it is certain that the follicle cells take only a nutritive part in the construc- tion of the embryo. . In Pyrosoma and in Salpa, most of the migratory cells retain their individuality until the blastomeres have become so small by repeated division that they are no larger than the follicle cells, and it becomes very hard to follow the course of each cell and to trace its fate. I have shown that while this is difficult in Salpa pinnata, it is not impossible, and that the ultimate fate of all the follicle cells is to supply food for the cells of the embryo. In the embryo of Pyrosoma, Salensky has traced the history of the two sorts of cells until they are equal in size and no 62 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. longer distinguishable from each other, and as he finds no cells in the act of degenerating, he believes that they all help to build up the embryo ; but there is every reason to believe that the follicle cells have the same fate in Pyrosoma that they have in Distaplia and Salpa. While the Distaplia egg undergoes total segmentation, it is well filled with yolk, and the segmentation cavity is thus obliterated; but this is a secondary change, and we may picture the egg without it, as we may also, in imagination, divest the Pyrosoma embryo and the Salpa embryo of all secondary peculiarities except those which engage our attention ; and the most satisfactory way to study the history of the follicle in these three embryos is to strip them mentally of their irrelevant peculiarities and picture them as reduced to their simplest expression, and to imagine them as developing according to the primitive chordate type. I have attempted to do this in the following diagrams. If we picture the Distaplia egg without its yolk, and with a spacious segmentation cavity, it will be like cut I, in which 15 is the segmentation cavity and 2 is the epithelial capsule of follicle cells. The space between this and the embryo is filled with migratory cells, and these are repre- sented as wandering into the substance of the blastomeres, and also pushing their way between them into the segmentation cavity. In Distaplia the follicle cells are used up very early, but I have rep- resented them in the diagram as persisting in great abundance at an advanced stage of segmentation, as is the case in Pyrosoma and Salpa. The blastula in the diagram is figured as about to undergo invagination to form a gastrula, and the cells in the left hemisphere, 3, are ectodermal, while those in the right, 7, are endodermal. For reasons drawn from the embryology of Salpa, I have represented the migration into the sub- stance of the blastoderm cells as most active in the endodermal hemi- sphere, and the migration into the segmentation cavity as most active in the equatorial zone where the two germ layers meet, and as more active between the endoderm cells than between the ectoderm cells. If now we picture the invagination of a blastosphere like cut I, we have a gastrula like cut J. As this stage is more advanced than the first, the epithelial capsule of the follicle is represented as entirely broken up into distinct cells, and while a few of these are still outside the embryo, most of those which have not migrated into its substance are folded into the digestive cavity as this is formed. A few of the migrating cells are in the ectoderm, more in the endoderm, and others are pushing in between the cells, especially around the edge of the blastopore, and wandering W. K. BEOOKS ON THE GENUS SALPA. 63 into the segmentation cavity or body cavity. In Salpa these latter become arranged in an epithelium or continuous layer, and in cut J, I have represented them as beginning to assume this structure by settling down over the inner ends of the blastoderm cells. The embryo of Pyrosoma is a disk-shaped blastoderm on a surface of food-yolk ; but so far as the migratory follicle cells are concerned, I regard it as comparable to cuts I and J ; and if we imagine the gastrula in cut J converted into a disk-shaped embryo on the surface of a big yolk, we shall have something like the Pyrosoma embryo, so far at least as the follicle cells are concerned. CUT I. CUT J. We do not know whether the migratory habit of the follicle cells and their peculiar share in the development of many Tunicates were or were not acquired before the embryos departed from the primitive type which is shown in these diagrams; but however this may be, we know that they were acquired by embryos which were, at least, modifications of this type, and we may therefore refer them back to it in imagination and picture them to ourselves as added on to it. Between the Pyrosoma embryo and Salpa embryo there is a gap which may be filled, in imagination, in this way. Suppose the follicle cells, which, in cut J, lie in the body cavity of the gastrula, to become arranged over the inner ends of the cells of the ectoderm and endoderm, so as to form a continuous epithelium of elongated cells, as shown in cut K, where 3 is the ectoderm, 9 the endoderm, 15 the body cavity, and 7 and 8 the two layers of follicle cells ; and suppose also that besides these follicle cells there are others, in the substance of the endoderm cells, and others in the digestive cavity, and a few outside the embryo in the vicinity of the blastopore. In Plate X, Fig. 8, the follicle cells, which are colored blue, have very nearly the arrangement that they have in 64 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGEAPHS. cut K, although the salpa embryo exhibits other peculiarities which have not yet been discussed. In the diagram the arrangement of the follicle cells inside the body cavity is so much like the arrangement of a meso- derm that I have called its outer and inner layers somatic and visceral, after the analogy of the mesoderm. At the stage shown in the diagram the embryo has a body wall made up of an outer layer of ectoderm, 3, and an inner layer of somatic follicle cells, 7 ; and a digestive cavity whose wall is made up of a layer of endoderm, 9, and a visceral layer of follicle cells, 8, while its lumen is filled with follicle cells. The body cavity is bounded on all sides by follicular epithelium. CUT K. Cur L. Now if we picture this gastrula as gradually shaping itself into a Tunicate embryo, it is plain that, as the various organs are blocked out by foldings and ingrowths and outgrowths of the germ layers, the folli- cular epithelium of the body cavity will take part in all these changes, and will thus conform to the shape of all the organs which are formed before it degenerates. Thus all the organs of the embryo will become outlined in both blastomeres and follicle cells, so that the follicle will become a sort of mold or cast of the embryo. These changes are so hard to draw and so easy to imagine that diagrams of them seem uncalled for. The most remarkable peculiarities of the Salpa embryo are now to be described. They are due to another set of secondary changes which are quite anomalous. After the follicle had become developed in the way which I have sketched, so that it formed a cast of the embryo, segmentation and the formation of the germ layers became retarded gradually, step by step, as compared with the development of the follicular layers, until the salpa embryo as we now know it was evolved. Cut L is a diagram of a salpa gastrula, and the character of its secondary modifications will be under- stood by comparing it with cut K. W. K. BKOOKS ON THE GENUS SALPA. 65 The segmentation of the salpa egg is so much retarded that, in the gastrula stage, the germ layers are represented only by the scattered blastomeres, 3 and 9. There is no digestive cavity, as the space between the scattered endodermal blastomeres, 9, is completely filled with migra- tory follicle cells, as is the blastopore also ; while the ectoderm is repre- sented by a few scattered ectodermal blastomeres, 3, external to the follicle in the region of the blastopore. While the development of the true germ layers is thus retarded, the follicle cells follow, in their migration, the paths which were established for them before the retardation of the germ layers was initiated, and they now shape themselves into the cast of a gastrula, and this pseudo gastrula becomes converted into the simulacrum of a tunicate embryo, carrying with it the retarded blastomeres, in the way which has already been fully illustrated and described. The blastomeres thus reach their final positions in the body of the embryo before they become differ- entiated and arrange themselves in organs and tissues. Finally, the follicular layers break up. Many of their cells are taken into the sub- stance of the blastomeres and digested, while others are left in the body cavity as free wandering cells, destined to degenerate at later stages and to serve as food for the tissues of the body. In this section I have not been able to avoid the language of phy- logeny, and I therefore wish to say once more that the account which is here given is both hypothetical and diagrammatic. The actual salpa embryo is complicated by the possession of a placenta, and by peculiari- ties which may be due to the former presence of a food-yolk which has disappeared. These features have been left out of consideration, and, even with this qualification, I have no desire to assert that, during the actual history of the evolution of the salpa embryo, it has moved along the path which I have indicated. We have no history of its origin, and it is quite possible that the course of development has been very different from this imaginary recon- struction. I believe, however, that the comparison throws light upon the funda- mental nature of the embryo, and upon its relation to the embryos of other tunicates, and I claim that, so far, it is valuable, although I should be the last to assert its definite phylogenetic significance. I have spoken of the history of the blastomeres as retarded, but it is probable that the retardation is not actual but only relative, and that the embryo is actually produced more quickly than it was before the folli- cular migration became established. CHAPTER IV. THE ORIGIN OF THE PROLIFEROUS STOLON. My own observations on the origin of the stolon of Salpa are by no means in complete harmony with the accounts which have been pub- lished, but I shall adhere to the plan which I have followed so far, and shall try to describe my own observations briefly and simply, leaving literary comment for a later chapter. SECTION 1. Outline Sketch. The aggregated salpae are produced from a proliferous stolon, which grows out from the body of the solitary salpa. It makes its appearance while the solitary salpa is an embryo, and as it begins to become con- verted into chain-salpae before this is born, its early stages must of course be studied in the embryo. The stolon is a tube which is joined, at its proximal end, to the body of the solitary salpa, while distally it ends blindly. Its structure and its anatomical relations are well shown in the following figures. Plate XXXV shows at a' a longitudinal section of a very young stolon, and its position is intelligible, since the entire figure is a longitudinal section through the body of an embryo. This stolon is shown more magnified in Plate XX, Fig. 7, and a series of transverse sections through another stolon at the same stage is shown in Plate XX, Figs. 1, 2, 3 and 4. Plate XLI, Fig. 3, is an embryo a little older than the one in Plate XXXV, and its stolon is seen at st. In Fig. 4 of the same plate the same stolon is shown more enlarged, and it is shown in longitudinal section in Plate XVI, Fig. 5. The stolon, st, of the embryo, in Fig. 5 of Plate XLI, is shown more enlarged in Fig. 6, and a series of transverse sections of the same stolon is shown in Plate XXI, Figs. 1-7. In this series, Fig. 1 is the most proximal section, at the root of the stolon, and Fig. 7 the most distal one, near its tip. A transverse section of an older stolon is shown in Plate XVI, Fig. 4; another, of a still older one, in Plate XLV, Fig. 5, and a section through the root or proximal end of the stolon of a fully grown solitary Salpa pinnata is shown in Plate XXXIV, Fig. 1. W. K. BROOKS ON THE GENUS SALPA. 67 As this series of figures shows, the stolon of Salpa pinnata lies below the middle line of the ventral surface of the solitary salpa, with its free end pointing forward. In several of the figures, as in Plate XVI, Fig. 4, Plate XXI, and Plate XLV, Fig. 5, the two folds d-d which compose the endostyle of the solitary salpa are shown, and it will be seen that the stolon lies under the furrow in the middle line of the ventral surface of the pharynx between these folds. The figures in Plate XXI also show that the vertical longitudinal plane, between the halves of the endostyle, which cuts the solitary salpa into symmetrical halves, also cuts the stolon symmetrically, so that Plate XXXV cuts both the embryonic solitary salpa and the stolon in the plane of bilateral symmetry. When in its true or morphological position, then, the stolon is symmetrical with reference to the middle plane of the solitary salpa, although, as the stolon grows older, it is usually somewhat twisted on its own axis, as Plate XVI, Fig. 4, and Plate XLV, Fig. 5, show. In many species of Salpa, the stolon, as it grows, becomes wrapped in a spiral around the digestive organs, as is shown at st, in the figure of the solitary Salpa africana, in Plate IV, Fig. 2, and when this is the case, the stolon begins to curve to one side almost as soon as it makes its appearance. There can be no doubt, however, that in all cases it is, morphologically, a median structure, symmetrical in the same plane as the solitary salpa ; that Salpa pinnata shows its true or primary position, and that its position in species like Salpa africana is secondary. When fully formed it consists, in cross section, Plate XXXIV, Fig. 1, of the following structures: 1st, a tubular sheath of ectoderm, a; 2d, a nerve tube, /; 3d, an endodermal tube, d'; 4th, two perithoracic tabes, (/ and li ; 5th, two blood-tubes, i and j, and 6th, a genital rod, m-n. In addition to these well-defined constituents, there are isolated cells between the cloacal tube and the adjacent structures, shown, uncolored, in Plate XXXIV, Fig. 3. Before the chain-salpae make their appearance, all these structures run from the base of the stolon nearly to the tip, so that every section is like the figure, but at the tip the nerve tube, genital rod and perithoracic tubes disappear, and the endodermal tube d' ends blindly, as the longi- tudinal section, in Plate XVI, Fig. 4, shows, so that the two blood-tubes communicate with each other. They do not communicate with the exterior, however, or with the cavity of the endodermal tube. 68 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. SECTION 2. The Orientation of the Stolon. As the regions of the stolon undergo very great changes of position during its conversion into a series of salpae, it is necessary to fix upon definite terms for its various regions. I shall call the tip of the stolon the distal end. Morphologically this end is anterior, and the end where it joins the body of the solitary salpa is posterior; but as in many species the stolon is not straight, but coiled, the terms distal and proximal are better, and I shall call the root of the stolon its proximal end. As the region along which the nerve tube lies is above, or towards the body of the solitary salpa, I shall call it the top of the stolon, and the opposite surface, where the genital rod lies, I shall call the bottom. The side which is on the right, when the solitary salpa is pictured as placed in front of the reader with its oral end above and its dorsal surface towards the reader, I shall call the right side of the stolon. SECTION 3. The Ectoderm of the Stolon. This, like the ectoderm of the solitary salpa? and chain-salpae, is colored purple in all the sections. As Plate XVI, Fig. 5, and other figures show, it is continuous at the proximal end of the stolon with the ectoderm of the solitary salpa. The multiplication of the ectoderm cells is the most efficient agent in the production of the tubular stolon. In Plate XLI, Fig. 2, x marks the place where the stolon is to be developed, although there is as yet no trace of it. Plate XX, Fig. 6, is a median longitudinal section through this region of the body of an embryo a little older, and the youngest in which I have found traces of the stolon. Plate XXXV will serve to show what part of the body is included in this figure, although it is from an embryo younger than Plate XXXV. In Plate XX, Fig. 6, /, colored yellow, is the pericardium ; k is the eleoblast ; n is the germinal mass, or aggregation of embryonic germ cells, in the body cavity of the solitary salpa embryo ; a is the unmodified ectoderm of the embryo, and a' the tract of ectoderm which is to become the ectoderrnal tube of the stolon. The ordinary ectoderm cells of this region of the body are flat, but those which form this tract are cylindrical, and while none in active multiplication are shown in this figure, other sections show that they do multiply, and the ectodermal area of the stolon thus extends farther and farther backwards, as shown at a' in Figs. 5 and 7, until the germinal mass, Fig. 7, n, and the endodermal tube, d', are shut in, or covered by a W. K. BROOKS ON THE GENUS SALPA. 69 dome of ectoderm, as is shown at a' in Figs. 1, 2, 3 and 4, which are a series of sections through a stolon a very little older than Fig. 7, parallel to a line drawn from the letter c to the letter n. A stolon a little older is shown in surface view in Plate XLI, Fig. 4, and in longitudinal section in Plate XVI, Fig. 5. The dome of specialized ectoderm has here become a deep bowl, and its growing edge pushes farther and farther inwards, so as to form a double fold of ectoderm around the proximal end of the stolon. In Salpa pinnata this fold soon results in the forma- tion, around the base of the stolon, of a prepuce-like structure, shown in Plate XLI, Fig. 6, so that, while the distal end of the stolon is covered by only one layer of ectoderm, its proximal end has, as Plate XLV, Fig. 5 shows, three layers : first, the ectoderm, A, of the solitary salpa ; second, the reflected layer, A", and third, the ectoderm, A', of the stolon. The portion of the stolon which is covered by these three layers of ectoderm is often so much compressed that the structures included in it are crowded together, as Plate XVI, Fig. 4, shows, so that the space between the endodermal tube and the ectoderm appears at first sight to be filled up with a mass of undifferentiated cells, although more careful examination shows that all the structures which are shown in Plate XXI, Fig. 7, are really present. The effect of this compression will be understood by comparing this last figure with Fig. 4 of Plate XVI. The presence of three layers of ectoderm around the proximal end of the growing stolon gives great complexity to sections like those in Plate XXI. In these figures, a is the unmodified ectoderm of the solitary salpa, a' that of the stolon, and a" the reflected layer, and the relations of the three layers are well shown in Fig. G, which will serve to interpret the other figures. I do not know whether these three prepuce-like layers of ectoderm are found in other species than Salpa pinnata. So far as I am aware they have never been described. Seeliger gives (6), Taf. XII, Fig. 7, a section of a young stolon of Salpa democratica at a stage in which he says that the space between the endodermal tube and the ectoderm is filled up by a solid mass of undifferentiated mesoderm. There is certainly no such stage in Salpa pinnata, and it is possible that his figure represents a compressed section like my Fig. 4 in Plate XVI. I shall, however, discuss his view of the origin of the stolon further on. 70 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. SECTION 4. Tlie Nerve Tube. This is colored violet in the sections, like the ectoderm, as it is ecto- dermal in origin, although very young stolons must be studied to show this. In all the older stolons it is distinct from the ectoderm from end to end, and appears in all the sections, as it is shown in Plate XXXIV, Fig. 1, at Z, as a closed tube, with a lumen, in contact with the inner surface of the ectoderm, on the middle line of the upper surface of the stolon. Even in a stolon as young as the one shown in Plate XXI, sections throw no light on its origin, for Fig. 7 shows that it has the structure that it has in the figure just referred to. In the stolon, a little older than Plate XX, Fig. 7, w r hich is shown in transverse section in Figs. 1, 2, 3 and 4, I found the clearest evidence of its ectodermal origin. In the more proximal sections 1, 2 and 3, the ectoderm covers only the bottom and sides of the stolon, but the tip of the distal end, Fig. 4, is completely shut in, although its ectoderm, a', has not yet separated from the ectoderm, a, of the solitary embryo, and at the point where the two layers are continuous, the abundance of nuclear figures shows active cell multiplication, and a number of the cells have pushed in to the cavity of the stolon, where they form a projecting knob, I, the rudimentary nerve tube. At this early stage it is solid, and its lumen, which appears later, never has any communication with the exterior. At least I have found no opening, although Salensky, who has recently shown (17) that the nerve tube of the ascidiozooid of Pyrosoma is ectodermal, finds a dis- tinct neural invagination. A longitudinal section of the somewhat older stolon of Plate XLI, Fig. 3, is shown in Plate XVI, Fig. 5. Here the nerve-rudiment, I, stretches for some distance along the middle line of the stolon, and at its distal end the cells are arranged around its axis, although the lumen has not yet made its appearance. Plate XXXV shows that the stolon is about as far away from the ganglion of the solitary embryo as it could well be, and it is difficult to believe that there is any phylogenetic connection between the nerve tube of the stolon and the ganglion of the embryo. It is of course possible that ontogenetically the two structures may be common descendants of the same cells, but there is no indication of any such ontogenetic relation- ship. Uljanin believes (7) that the budding of salpa is to be traced back, phylogenetically, to fission in a double embryo. If this has been its history we certainly should expect to find corresponding structures arising in positions which are more consistent with their supposed W. K. BROOKS ON THE GENUS SALPA. 71 fundamental identity of origin. I shall, however, discuss the subject further on. SECTION 5. Tlte Endodermal Tube. This is colored red in the sections, as are also its derivatives. As d' in Figs. 2, 3, 4 and 5 of Plate XIX shows, it arises as a tubular outgrowth from the ventral wall of the pharynx on the middle line between the two folds of the endostyle, which are marked d--d in these and other figures; and in Salpa pinnata and Salpa cylindrica it per- manently retains its communication with the cavity of the pharynx. Seeliger states that in Salpa democratica it soon loses its communica- tion with the pharynx, but this species is a difficult one to study, and the opening is easy to find in the straight stolons of the two species which I have studied most thoroughly, even in the oldest specimens. I have not found in any of my sections any food or foreign organisms in the en do- dermal tube of the stolon at any stage, nor have I ever found in the stomachs of the young chain-salpae any evidence that they are nourished through it, although their digestive organs are derived from it, and retain for a long time their communication, through it, with the pharynx of the solitary salpa. It is formed, as Plate XXXV shows, before the mouth of the solitary embryo communicates with the exterior, and there is no reason for believing that it has a nutritive function. As the longitudinal sections in Figs. 5, 6 and 7 of Plate XX show, the inner ends of its cells have a ragged or indefinite outline, as have also the cells of the endostyle, Fig. 2, d, and it is probably ciliated when alive, and, at later stages, when the pharynx of the solitary salpa is filled with fresh sea-water, it may, perhaps, convey to the young chain-salpa3 a respiratory current. In a mature stolon, Plate XXXIV, Fig. 1, d', its upper and lower walls are formed of flattened cells, with clearly defined outlines, shown more magnified in Fig. 3, while along each side of it there is a thick strip of elongated cells, with their inner edges indefinite and ragged. At the distal end of the tube these two strips unite, and as the longitudinal section in Plate XVI, Fig. 5, shows, its tip consists of elongated cells. Before we examine its structure and origin in detail, it will be neces- sary to briefly describe the endostyle of the solitary embryo. Plate XXI, Fig. 5, shows in red at b-d-d-b a transverse section across the middle line of the pharynx of the solitary embryo. The ordinary endoderm of the pharynx is shown at b. It consists of flattened cells with sharp outlines. The endostyle consists of two parallel ridges of greatly elongated cells 72 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. with indefinite outlines, separated from each other on the middle line by a deep furrow, the floor of which consists of flat, sharply defined cells, like those of the pharynx in general. At the stage shown in Plate XX, Fig. 7, where the ectoderm of the stolon forms a shallow bowl or dome, the endoderm grows down into the cavity of the bowl, as shown in the transverse sections in Figs. 1, 2 and 3, to form the endodermal tube, which is at first widely open above into the cavity, c, of the pharynx, and consists of a floor of flattened, sharply defined cells, which is part of the floor of the endostylic furrow of the solitary embryo, and which crosses the middle line of the stolon to join at each end a vertical wall of thickened cells, with indistinct and irregular inner ends, like the cells in the endostylic ridges. As Fig. 1 shows, these side walls of the endodermal tube are actually part of the endostylic ridges, into which they can be traced in Fig. 1. Soon, however, as the older or more distal section in Fig. 2 shows, the endostylic ridge proper becomes separated from the endostylic side wall of the stolon by an inter- vening area at d-d in Fig. 2, where the cells become flat and sharply defined. In a still more distal or older section, Fig. 3, these two belts of flattened cells are pushed towards each other until they meet on the middle line to form the roof of the endodermal tube. The way in which this tube is formed shows that it is actually a section of the endostyle which drops down out of the solitary embryo into the stolon. As the solitary embryo develops, its two endostylic ridges come nearer to each other, and finally meet on the middle line, as the succes- sive stages, in Plate XXI, Plate XVI, Fig. 4 and Plate XLV, Fig. 5 show, and the channel which connects the cavity of the endodermal tube with that of the pharynx of the solitary salpa is thus reduced in size until it becomes a small slit, d' of Plate XXI, Fig. 1. Within the stolon the endodermal tube assumes the shape shown in the figures in Plate XXI. In cross-section it is a little like a letter H , and is made up of a horizontal cross-bar and two vertical ends. In young stolons the floor and roof meet and come into contact with each other on the middle line, so that the cavity is divided into two lateral chambers which communicate with each other only at the distal end, Plate XX, Fig. 4, but as the stolon grows older the roof and the floor separate from each other, as Plate XLV, Fig. 5 shows, until in mature stolons the cavity assumes the shape which is shown in Plate XXXIV, Fig. 1, d'. W. K. BROOKS ON THE GENUS SALPA. 73 SECTION 6. The Blood-Tubes. There are two of these, the upper one, j, and the lower one, ?', both colored yellow in the sections. They communicate with each other at the tip of the stolon, around the tip of the endodermal tube. Elsewhere they are separated from each other by the endodermal tube and the perithoracic tubes, which are so placed as to form a horizontal longi- tudinal partition. In their origin the tubes are part of the cavity of the body of the solitary embryo, and they are present in the stolon at all stages. When the ectoderm and endoderm which are to enter into the structure of the stolon first become differentiated from the tissues of the embryo, Plate XX, Fig. 6, these two layers are in contact with each other, and, as Plate XLI, Fig. 7, shows, both the ectoderm, a, and the endoderm, 6, are in direct contact with the germinal mass, n, but as soon as the rudimentary stolon begins to elongate, Plate XX, Fig. 7, the endoderm separates a little from the other structures, so that there is space for the blood corpuscles of the solitary salpa to circulate between them, and these spaces are present at all subsequent stages, as the transverse sections in Plates XX and XXI, and the longitudinal section in Plate XVI, Fig. 5, show. The lower blood space, , is the first one to be folded into the stolon, as shown in Plate XXI, Figs. 1 and 2, i ; and in Plate XXXV it is contin- uous with that part of the body of the solitary salpa which contains the eleoblast, k. I shall show farther on that the degenerating cells of the eleoblast furnish food for the blood corpuscles and migratory mesoderm cells of the embryo, and as the sections of the stolon show that these latter pass into it in great numbers, they no doubt play an important part in the nutrition of the growing stolon and the chain-salpse which are formed from it. The upper blood space, j, is in intimate relation with the heart, e, and the great ventral blood-vessel of the solitary embryo, and it will be necessary to say a few words here about these structures. The pericardium, / of the sections, and the heart, e, are shown in horizontal sections of the solitary embryo, in Plate XIX, and in vertical section in Plate XXXV. The pericardium is a closed vesicle, colored yellow in the figures, behind the pharynx, c, and to the left of the middle line, and the heart is formed by the infolding of its walls on the side next the pharynx, as shown at e in Plate XIX, Figs. 1, 2, 3, 4 and 5. In its origin the chamber of the heart is thus part of the body cavity, external to the pericardium. In the young embryo it is behind the pharynx, c, and 74 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. vertical. It is at first simply a furrow, and its upper and lower ends remain permanently open, but in its middle region, Fig. 3, more magni- fied in Fig. 9, its lips fold in and meet so as to convert it into a tube. Its lower opening is shown at e in Fig. 5, and more enlarged in Fig. 10, and its upper opening at e in Fig. 1, more enlarged in Fig. 6. The upper end of the heart, Plate XXXV, is situated at the base of the gill, o, which is a tubular rod bathed on all sides by the water which circulates through the pharynx, c, and cloaca, g'". In the mature salpa the lower end of the heart communicates with a large blood space or vessel which runs along the middle line of the ventral surface under the endostyle. This vessel is shown in section, not marked by a letter, in Plate XXI, Fig. 7, and also in Plate XVI, Fig. 4. It is also shown in a young chain-salpa, in Plate XXXVIII, Fig. 29, atj. In the young solitary embryo it is very short, and it opens into that part of the body cavity which lies over and around the placenta. Plate XXI, Fig. 1, cuts the heart, e, and the pericardium, /, of the solitary embryo. Fig. 2 shows on the side next the stolon the epithe- lium of the pericardium and that of the heart in contact with each other. Figs. 3, 4, 5 and 6 pass through the inferior opening of the heart, and Fig. 7 cuts the ventral blood-vessel. If the upper blood space of the stolon, j, of Fig. 7 be traced back through sections 7, 6, 5, 4 and 3, it will be seen that it communicates with the opening of the heart of the embryo, and through the ventral blood- vessel, with the placental portion of its body cavity, and it is therefore clear that it stands in very intimate relation to the two sources of nutri- tion, the degenerating eleoblast and the placenta, as well as to the gill. When the heart of the chain-sa-lpa is beating in one direction, blood flows from the eleoblast into the lower tube of the stolon out to its tip, and back to the heart through the upper tube. In the reversed circula- tion, blood from the gill of the embryo is driven by its heart into the upper tube, and out to the tip of the stolon and back to the eleoblast. In the mature stolon, Plate XXXIV, Fig. 1, each blood tube has its own endothelial lining, which lies in such close contact with the other organs of the stolon that it easily escapes observation at all points except the places where it bridges over the gap between the ectoderm and the endodermal tube, Fig. 3, or in the angles at the sides of the nerve tube. It is shown in Plate XXIII, Fig. 10, j, and in Plates XXIV and XXX. W. K. BEOOKS ON THE GENUS SALPA. 75 It is difficult to determine whether this endothelium is formed in place, as is probably the case, or derived from the endothelium of the blood spaces of the embryo, for its great delicacy in young stolons renders it unfavorable for study in sections. A fragment of it is shown in Plate XXI, Fig. 6, above the letter g, and it is also shown in Figs. 2, 3, 4 and 5. SECTION 7. The Perithoracic Tiibes. These are colored green in the plates, and the right one is marked gr, and the left one h. They run along each side of the stolon, between the ectoderm and the thickened side walls of the endodermal tube. In mature stolons, each of them has a distinct tubular lumen, as is shown in transverse section in Plate XXXIV, Fig. 3, h, and in longitudinal section in Fig. 6, h. In very young stolons, Plate XXI, Fig. 7, g and h, the lumen is absent. I have devoted especial attention to the question of the origin of these tubes, but I have not been able to obtain any evidence which I regard as conclusive, although my observations indicate that the tubes arise in the ectoderm of the young stolon, at the points where this joins, at the sides of the stolon, the ordinary ectoderm of the embryo. In old stolons, like the one shown in Plate XXXIV, Fig. 1, the tubes can be traced to the proximal end of the stolon, and at the point where the ecto- derm bends outwards to join that of the solitary salpa they come to an end, although I have not been able to find, in old stolons, any union between them and the ectoderm. The stolon shown in the transverse sections in Plate XX had no perithoracic tubes, although, on the left side of Fig. 3, the fold where the ectoderm, a', of the stolon joins that of the embryo, a, runs down for some distance into the stolon, alongside the endoderm. This may possibly be the rudiment of a perithoracic tube, the right one, as these sections were drawn from inverted specimens, but I have not been able to prove to my own satisfaction that it is. In Plate XXI, Fig. 2, the left perithoracic tube is shown to lie in very intimate relation to the ectoderm at the root of the stolon close to the point where, as section 1 shows, the ectoderm folds upon itself. This section, like the one before it, seems to show that the perithoracic tubes arise from the ectodermal fold at the proximal end of the very young stolon, and I know of no theoretical ground for doubting this evidence, although none of my sections show any vegetative activity in the cells 76 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. at this point. A horizontal longitudinal section through a very young stolon would probably furnish more decisive evidence, but all my own specimens of the proper stage were used for other purposes. SECTION 8. The Mesoderm of the Stolon. In addition to the blood corpuscle and the cells which form the walls of the blood spaces, a number of cells become shut into the stolon on each side, between the ectoderm and the perithoracic tube, as shown in Plate XXI, and in Plate XXXIV, Fig. 3, while others are found, as Plate XXXIV, Fig. 3 shows, between the endoderm and the perithoracic tube. Some of these cells appear to enter the stolon as detached mesoderm cells, while it is possible that others are derived from the pericardium, for this unquestionably enters into the structure of the stolon. On the left side of the inverted section in Plate XX, Fig. 1, it will be seen that a diverti- culum from the pericardium, /, pushes in between the ectoderm and the right side of the endodermal tube, and Figs. 4 and 5 of Plate XXI show that the end of this process is constricted by the growing ectoderm, and a part of it, at least, probably becomes included in the stolon, although at an older stage there is nothing which can be identified as its derivative, and if it is represented at all it is probably represented only by inde- pendent cells. Another constituent of the stolon may be mentioned here, the gela- tinous substance which fills the spaces in the angles between the endo- thelium of the blood spaces and the other organs. This substance, which is homogeneous and transparent, is shown at 32 in Plate XXXIV, Figs. 3 and 5. SECTION 9. The Genital Rod. The origin and history of the genital rod will be treated at length in another chapter. We must note here, however, that its rudiment, Plate XLI, Fig. 7, is present in the body cavity of the embryo before the stolon is formed, and that it is shut into the stolon, as the figures on Plate XX show, by the growth of the ectoderm. ft SECTION 10. The Derivatives from the Parts of the Stolon. The ectoderm of the stolon gives rise to the ectoderm of the chain- salpse, to the organs by which they are fastened to each other after birth, W. K. BROOKS ON THE GENUS SALPA. 77 and to their cellulose mantles. The nerve tube gives rise to the ganglia. The lateral portions of the endodermal tubes give rise to the corres- ponding halves of the pharynx ; and the oesophagus, stomach and intes- tine are derived from the one on the right side. The perithoracic tubes give rise to the cloaca and to the cloacal portions of the two gill-slits by which it opens into the pharynx. The body cavity consists, in part at least, of diverticula from the blood spaces, and it is lined by their endothelium. The muscles and the stoloblast, or the equivalent of the eleoblast, are derived from mesoderm cells from both sides of the stolon. The heart and pericardium are probably formed from some of the mesoderm cells on the right side. The eggs and their follicles and fertilizing ducts, and the testes, are derived from the genital rod. CHAPTER V. THE TRANSFORMATION OF THE STOLON INTO THE SERIES OF AGGREGATED SALP^E. SECTION 1. Outline Sketch. The origin of the aggregated salpge is complicated by secondary changes, but in its essential features it is a very simple history. I shall therefore preface my account by a brief outline of the process, stated in its simplest form, and divested of all secondary complications. I. The Proliferous Stolon. As already described, this, when fully formed, consists, 1st, of a tubular sheath of ectoderm, Plate XXXIV, Fig. 1, a, which is derived from the ectoderm of the solitary salpa; and which contains, 2d, an endodermal tube, d', which is derived, as Plate XXI, Fig. 1, d' shows, from the pharynx of the solitary salpa; 3d, a nerve tube, Plate XXXIV, Fig. 1, Z, which is derived, as Plate XX, Fig. 4, I, shows, from the ectoderm of the stolon ; 4th, of two perithoracic tubes, g and 7^, which are probably derived from the ectoderm of the stolon, as shown in Plate XX, Fig. 3 ; 5th, an upper haemal tube, /, which communicates at the base of the stolon with the cavity of the heart of the solitary salpa, as is shown at j in Plate XXI, Fig. 3 ; 6th, a lower ha3mal tube, i, which communicates with the body cavity of the solitary salpa, as is shown in Plate XXI, Fig. 2, i ; and 7th, of a genital string which consists of a series of eggs, m, inclosed in a follicular sheath, n, and which is derived from the germinal mass, Plate XX, Fig. 6, n, of the solitary salpa. II. The Segmentation of the Stolon. The first indication of the seg- mentation of the stolon is a series of ectodermal folds, Plate XXXIV, Fig. 11, a, which first appear at its sides, but soon extend up and down and completely encircle it, and, pushing inwards, mark out the body cavities of the salpas, and also cut up the tubular structures inside the stolon into segments. The active agent in this process of segmentation is the growth of the ectodermal folds, and the other structures are actually cut by these folds, W. K. BROOKS ON THE GENUS SALPA. 79 as Plate XXXIV, Fig. 10 and Plate XXIII, Fig. 7 show. As the result of this process the nerve tube becomes cut up into a series of ganglia, one for each salpa, Plate XV, Fig. 10, s ; the perithoracic tubes become cut up into a series of perithoracic vesicles, two for each salpa, Plate XXXIV, Figs. 9, 10 and 11, and Plate XV, Figs. 11 and 12, g and h ; the genital string becomes cut up into a series of eggs, Plate XV, Fig. 2, n, one for each salpa, inclosed in a follicle, m; and the thickened endodermal epithelium at the sides of the endodermal tube, d', becomes cut up into a series of vertical pouches or pockets, two for each salpa, the rudiments of the right half of the pharynx, 27, and of its left half, 28, as is shown in Plate XXIII, Figs. 4, 7, 8 and 9, and Plate XV, Figs. 6, 7, 8, 11 and 12, where these pouches are colored red. III. The Rudimentary Chain-Salpa. The structures which I have enumerated form the rudiments of a single salpa. They are shown in Plate V, Fig. 1, which is a reconstruction from a series of tran verse sec- tions through the stolon of Salpa pinnata. At this stage each salpa is bilaterally symmetrical, and its plane of symmetry is the same as that of the stolon, while its long axis is at right angles to that of the stolon, which becomes converted into a single row of salpae, so placed that, as is shown in cut M, the dorsal surfaces of all of CUT M. them are towards the base of the stolon, their ventral surfaces towards its tip, their right and left sides on its right and left respectively, their oral ends at its top or neural side, and their aboral ends at its bottom or genital side. IV. The Secondary Changes. The single row of salpae becomes con- verted into a double row, Plate IV, Fig. 1, which consists of a series of right-handed salpae and a series of left-handed ones, placed with their dorsal surfaces out, their ventral surfaces towards the ventral surfaces of those in the opposite row, and the left sides of those on the right and the right sides of those on the left towards the base of the stolon. In 80 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. order to illustrate these secondary changes of position let us represent the series of salpse by a file of soldiers, all facing the same way. Now imagine that each alternate soldier moves to the right, and the others to the left, to form two files still facing the same way. Now let them face about so that the backs of those in one row are turned towards the backs of those in the other row. They will now represent two rows of salpae like those shown in Plate IV, Fig. 1. To make the illustration more perfect suppose that, instead of step- ping into their new places the soldiers grow until they are pushed out by mutual pressure, and suppose that their heads, growing fastest, form two rows while their feet still form one row, and suppose furthermore that as each soldier rotates his feet turn first, and that the twist runs slowly up his body to his head, which turns last. We must also imagine that these various changes all go on together, and that while they are taking place each soldier not only grows larger, but also develops from a simple germ to his complete structure. As thus outlined these secondary changes are simple, but as growth, development, pushing to the side, and rotation are all going on together, and as they all take place gradually, the interpretation of sections is very puzzling ; although the history of their development becomes very simple when it is illustrated by diagrams from which the secondary changes are omitted, instead of by figures from actual sections. V. The Development of the Chain-Salpa. (1) The body cavity. At first this is directly continuous, through the blood spaces i andj, with those of the adjacent salpae, but it is gradually shut in by the growth of the ectodermal folds, of which the inner edges is shown in the diagrams and in the figures on Plate V. (2) The pharynx. The lateral pouches, Plate V, Fig. 1, 27 and 28, from the sides of the endodermal tube, d', grow forwards towards the oral end of the body, cut N, 27 and 28, o, and backwards towards the aboral end, cut 0, 27 and 28. They are the rudiments of the right and left halves of the branchial sac or pharynx, and they may therefore be called the pharyngeal pouches. Their oral ends soon bend in towards each other, cut 0, and finally meet and unite on the middle line, cut P, ventral to the ganglion, s, to form the oral end of the pharynx, which is shown at 28-O-27 in cuts P to X. Finally, the oral end of the pharynx unites with the ectoderm and becomes perforated to form the mouth, cut S, z. W. K. BROOKS ON THE GENUS SALPA. 81 CUT N. CUT 0. CUT P. CUT Q. 82 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. The aboral ends of the pharyngeal pouches elongate, as is shown in cuts O and P, 27 and 28, and, bending towards each other, cut Q, finally meet on the middle line and unite with each other, as shown in cut R, 27-28, to form the aboral portion of the pharynx. They are shown in contact but not yet united, in Plate XXXIII, L~L', Figs. 3, 4, 5, 27 and 28, while in M-M' and N-N', Figs. 3 and 4, they are united on the middle line of the ventral surface. Both the oral and the aboral ends of the pouches ultimately unite on the middle line throughout their whole length, so that, as is shown in cut S and cut X, the pharynx becomes a large, unobstructed chamber, CUT R. OUT S. opening to the exterior at the mouth, and communicating with those of adjacent salpa3 through the opening which is marked d' in the figures. (3) The perithoracic tubes and cloaca. As the pharyngeal pouches grow backwards they carry with them, on their dorsal sides, the peri- thoracic vesicles, g and h, as is shown in cut N. At about the stage shown in cut 0, each of these vesicles unites with the dorsal surface of the W. K. BROOKS ON THE GENUS SALPA. 83 pharyngeal pouch and opens into it, as is shown in section in Plate XXVI, Fig. 3, E-E', where the perithoracic vesicles, g and h, colored green, are cut through their openings into the pharyngeal pouches, 27 and 28, which are colored red. These two openings are the two gill-slits, and in cuts P, Q and R their edges are indicated by dotted lines. Com- parison of the figures will show that they soon undergo a very great increase in size, until at the stage of cut S they include all the dorsal sur- face of the aboral end of the pharynx except a strip, o, on the middle line. Each perithoracic vesicle, after it has established its communication with the pharyngeal tube, gives rise to a diverticulum, cut P, which grows inwards towards the middle line of the dorsal surface, as is shown in Plate XXV, Fig. 7, H-H', g and h, and these diverticula ultimately meet and unite on the middle line, as is shown in cut Q, to form the cloaca, g'". Plate XXXIII, Fig. 3, K-K' shows, colored green, these two diverticula meeting on the middle line of the dorsal surface, which is the lower surface in the sections. Soon after the cloaca is thus formed its dorsal wall unites with the ectoderm on the middle line, and the cloacal aperture, G* of cut S, is formed at the point where this union has taken place. Plate XXXVI, Fig. 5, g v shows the union between the dorsal wall of the cloaca and the ectoderm, and it is shown again more advanced in Plate XXXVIII, Figs. 88 and 97, g\ and also in Plate VIII, Figs. 1 and 2, g\ (4) The gill. The gill of salpa, cut S, o, and Plate XXXVI, Figs. 5 and 6, o, is simply that part of the body cavity which is bounded ventrally by the dorsal wall of the pharynx, dorsally by the ventral wall of the cloaca, and at the sides by the gill-slits. (5) The digestive organs. The oesophagus, stomach and intestine are formed by gradual specialization in the course of the tubular diverti- culum which is shown at q in cut P, growing out from the posterior end of the right pharyngeal pouch, 27. In cut P, its blind end is dilated to form the stomach, and in cut Q, the intestine, p, is shown growing out from the ventral side of the stomach and bending upwards towards the dorsal surface, where in cut S it opens into the cloaca. (6) The pericardium and the heart. The pericardium makes its appearance very early, cut 0, as a closed vesicle, F, external to the aboral end of the right pharyngeal pouch, 27. As this pouch lengthens it carries the pericardium back with it, cuts P and Q, F, and Plate XXIV, Fig. 2, e, and as the digestive tract is formed it pushes the heart towards the ventral surface, cut S, F, and Plate XXV, Fig. 4, H, e, and Fig. 7, F, 84 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. e, and it finally comes to lie under the ventral surface of the right side of the pharynx, Plate XXXVI, Figs. 3, 4 and 5, /. As shown in these figures, the heart arises as a furrow which is formed by the involution of the dorsal surface of the vesicular pericardium. This outline of the history of the larger organs of the chain-salpa will give enough insight into the mode of development of its body to enable the reader to understand the more minute and detailed descrip- tion which follows. SECTION 2. The General Characteristics of Aggregated Salpce. The way in which the proliferous stolon grows out from the body of the salpa embryo and gradually acquires its complicated organization may be spoken of with perfect propriety as budding; but inasmuch as the stolon itself contains the rudiments of all the important systems of organs, its transformation into a series of aggregated salpae by cell multiplication and by the folding of its various parts, is more like the development of an embryo with germ layers into the body of a compli- cated adult than it is like ordinary budding, and it may perhaps be more proper to speak of it as the strobilization of the stolon. It is of course a process of asexual multiplication, but as the stolon contains the potency of all the following generations of chain-salpae, it stands, as Seeliger has pointed out (11, p. 583), in somewhat the same relation to the chain-salpae as that which a young embryo, with its future structure latent in its germ layers, bears to the perfect adult, since, in each case, the process of development consists in the unfolding of its rudimentary organization. As a knowledge of the final result will aid us in tracing the minute details of the process, I shall give a short outline of the most conspicuous features before I describe the process of strobilization. The growing stolon lies in a chamber which is hollowed out in the cellulose mantle of the solitary salpa, and as the aggregated salpae are set free they escape through an opening which connects this chamber with the exterior. In Salpa pinnata, Plate I, Fig. 6 ; Salpa chamissonis, Plate I, Fig. 7 ; Salpa cylindrica, Plate III, Fig. 7, and in a few other species, the stolon lies underneath the middle line of the ventral surface of the body of the solitary salpa, and it is symmetrically placed, with its free or distal end pointing forward, and with its right and left sides symmetrically placed with reference to the plane of symmetry of the solitary salpa. W. K. BROOKS ON THE GENUS SALPA. 85 In Salpa democratica, Plate II, and in most of the species of salpa, it becomes asymmetrical almost as soon as it is recognizable, and it grows around the nucleus in a spiral, as is well shown by Seeliger's excellent figure of the stolon of Salpa democratica (11, Taf. X, Fig. 5), and by his description (11, p. 593), and also in my figure of the solitary form of Salpa africana, Plate IV, Fig. 2, st. In Salpa pinnata, Salpa chamissonis, and probably in all the pinnata- like species, the stolon presents a graduated series of stages of develop- ment; each successive salpa, from the root of the stolon to its tip, being a little larger and more developed than the one behind it, as is shown in Plate XLVI, and also in the series of figures on Plates XXIII to XXXIII. In these species the salpa at the tip of the stolon is largest and most developed, and new ones are continually being marked out at the base of the stolon as those at the tip are set free. In all the other species the chain-salpae are developed in sets, as is shown in the cut M, and all the individuals in a set are in essentially the same stage, although there is gradation among the members of the youngest set at the root of the stolon. The diagram does not show the number of individuals in each set. They are always very numerous, and in some species each set contains a hundred or more. We know nothing of the birth-rate of salpa, but the solitary salpa begins to set free chain-salpae soon after it is born. I have never found a specimen with an exhausted stolon, and there is no evidence of any fixed limit to the process of asexual multiplication. The number of buds on the stolon at one time is very great. Salpa democratica usually has three or four sets at one time, and Leuckart (1, p. 67) found forty in one set and sixty-five in another. Seeliger says (11, p. 593) that he counted sixty-one in a single set in this species, and the average is probably about sixty in each set, or between two hundred and two hundred and fifty in all on the stolon at one time. The stolon of Salpa pinnata has about the same number, from two hundred to two hundred and fifty. The number is very much greater in Salpa cylindrica, and I have counted two hun- dred in a single set from this species. As its stolon carries three or four sets, the total number of buds at one time is from six hundred to eight hundred. The position which the bodies of the aggregated salpae occupy when they are first marked out in the stolon has been much misunderstood. I shall show soon that, morphologically, they form only a single series, and that they all arise in exactly the same position, with the neural or dorsal 86 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. surfaces of all turned towards the root of the stolon, with all their right sides on the right of its plane of symmetry and their left sides on its left, and with their long axes at right angles to the long axis of the stolon, and their oral ends above, as they are shown in the diagram, cut M. As they increase in size, however, and become crowded, they push out of the line to the right and left alternately, and thus form two ranks instead of one. At the same time the body of each salpa rotates ninety degrees upon its own axis, so that the neural or dorsal surfaces come to face out- wards, while the left sides of those on the right and the right sides of those on the left become turned towards the base of the stolon, and the planes of symmetry of the salpae, instead of coinciding with the plane of symmetry of the stolon, make right angles with it. It is most important to grasp clearly the fact that this position is a secondary one, and that, morphologically, there is only a single series of animals, all placed in the same position and all facing the same way like a single file of soldiers ; for the change of position takes place at a very early stage, and all the published accounts of the budding of salpa are so vitiated by a failure to discover it, or else to understand it, that they are almost worthless. The chain-salpae of Salpa scutigefa, Plate IV, Fig. 1, retain this secondary position after they are born, but in most species this arrange- ment, which persists in Salpa scutigera, is transitory, and still other changes of position soon take place. The position which is shown in cut M may be called position one, and the position which persists in Salpa scutigera, position two. In every species the salpse arise in the stolon in position one, which may therefore be called the true, or morphological, position, and in every species they quickly assume position two. Plate VIII, Fig. 1, shows two of the aggre- gated salpae of Salpa pinnata in this position, and Fig. 2 two of Salpa cylindrica. This position is also shown in sections of Salpa pinnata in Plates XXXVI, XXXVII and XXXVIII, and in sections of Salpa cylindrica in Plates XXXIX and XL. These plates, and Plate XV and Plates XXIII to XXXIII, represent sections which are parallel to the long axis of the stolon, and transverse to the bodies of the salpae, and in all of them the right side of the stolon is on the right side of the figure, and its proximal end towards the bottom of the page. In Salpa pinnata, Plate I, Fig. 2, and Salpa chamissonis, Plate XLI, Fig. 9, only a few aggregated salpse, about eight in Salpa pinnata and twelve in Salpa chamissonis, are set free at one time, and these, just before they escape, arrange themselves in a wheel or rosette with their W. K. BROOKS ON THE GENUS SALPA. 87 dorsal surfaces outwards, and their long axes parallel to the axis of the wheel. In all my preserved specimens the tip of the stolon had been so much flattened by contact with the side of the bottle, in transportation, that I have not been able to study in detail the way in which this wheel-like arrangement is acquired, and the subject should receive the attention of those who are able to study living specimens. There is an obvious resemblance between the wheel-like arrange- ment of the first four ascidiozooids which in Pyrosoma form the basis for the cylindrical community, and the wheel-like aggregation of the Salp in species of the pinnata group, as may be seen by comparing Salensky's figures of the young Pyrosoma community (17, Taf. II) with Plate XLI, Fig. 9. In each case the animals are arranged in a circle with their long axes parallel to the central axis, their dorsal surfaces out- ward and their oral ends above. I shall show that there are other reasons for believing that the pinnata-like species of Salpa are most primitive and most closely related to Pyrosoma, and it is not improbable that the wheel-like arrangement has been inherited in both cases from a common source, and that it is the primitive arrangement for the Salpae. In most species, however, the aggregated salpae are set free in the well-known floating clusters which have long been called chains. Part of a chain of Salpa cordiformis is shown in Plate IV, Fig. 6, and part of one of Salpa scutigera is shown in Plate IV, Fig. 1. As is shown in these figures, a chain consists of two parallel longitudinal rows of individuals, so placed that those in one row alternate with those in the other, while the neural or dorsal surfaces of all are external, and their ventral surfaces in contact with the ventral surfaces of those on the other side of the chain. The members of the community are united to each other by process from the walls of their bodies, which are hollow and contain diverticula from their body cavities, although there is no communication between the body cavities of adjacent salpae. In the pinnata group, Salpa pinnata, Plate I, Fig. 3 ; Salpa chamis- sonis, Plate VIII, Fig. 6 ; Salpa affinis and Salpa dolichosoma, there is only one of these processes, situated on the middle line of the ventral surface in front of the heart. The way in which it arises is well shown in the figures in Plate VII, Figs. 4 and 5, and Plate VIII, Fig. 1. Plate I, Fig. 3, shows it in Salpa pinnata in its perfect form, while the other figures show it at earlier stages. It is also shown in section, at succes- sive stages, in Plate XXXVI, Fig. 9, Plate XXXVII, Figs. 10 and 21, and Plate XXXVIII, Figs. 52 and 61. 88 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. These figures show that the process is ectodermal, hollow, and that the ectoderm of adjacent salpae comes into contact at the end of the pro- cess. After the cellulose mantle is formed the processes come to consist almost entirely of cellulose, although the actual contact between adjacent salpae is ectodermal. Plate XXXVII, Fig. 21, shows that in Salpa pinnata each individual is united by it to four others, the two which are diago- nally opposite it on the other side of the series, and the two adjacent to it on its own side. As the wheel-shaped colonies, Plate I, Fig. 2, are formed, the processes from all the members meet in the center and bind them together. In all the other salpae, each individual in the chain is joined on to four others, the adjacent ones on its own side of the chain, and the alter- nating ones on the opposite side, but instead of being effected by a single process as it is in Salpa pinnata, the union is usually brought about by eight, as is shown in the figure of Salpa scutigera, Plate IV, Fig. 1, and Salpa cordiformis, Plate IV, Fig. 6. We know of no species which stand midway between those of the pinnata group and the ordinary salpae, and we therefore have no phylogenetic evidence, but it seems probable that Salpa pinnata gives us the primitive method, and that originally a single process joined each salpa on to four others, and that this single process has been gradually converted into eight separate ones. In all cases the processes are primarily ectodermal, and they are shown at an early stage in Salpa cylindrica, in section in Plate XXXVII, Fig. 26. A chain of salpae may be compared to two trains of cars on two parallel tracks, placed so that the middle of each car on one track is opposite the ends of two cars on the other track, and each joined by two couplings to the car in front of it on its own track, and in the same way to the one behind it, and also to those diagonally in front of it and behind it on the other track. In young chains, of all species, on the stolon, the long axes of the salpae are at right angles to the long axis of the stolon, as if the cars in the two trains were set on end, and this primitive position is, as I have said, persistent in Salpa scutigera and Salpa bicaudata. Salpa democratica and Salpa tilesii pass through this stage, and before their chains are set free the bodies of all the individuals become inclined in the same direction, as, if the cars in the train were pushed over till each one rests against the one in front of it, Plate XLIII, Fig. 1. This change takes place in such a way that the oral end of the body of each salpa is thrown towards the distal end of the stolon, and in the W. K. BROOKS ON THE GENUS SALPA. 89 species which have just been named this inclined arrangement is per- sistent. In still other species, such as Salpa cordiformis, Plate IV, Fig. 6, Salpa runcinata, Salpa africana and Salpa hexagona, the axes of the bodies rotate until they become nearly or quite horizontal and parallel to the axis of the chain, and we thus have two series of salpae with each one joined at its anterior or oral end to the posterior end of the one next in front of it in the same row, and with the two rows facing each other by their ventral surfaces, and with the middle of the body of each opposite the joint where two in the opposite row are joined together. While the union between the salpa3 in the series is ectodermal, the cellulose mantles, as they grow, usually come into contact, and flattening against each other to help to maintain the integrity of the chain. In chains like those of Salpa scutigera, Plate IV, Fig. 1, and Salpa demo- cratica, Plate XLIII, Fig. 1, the cellulose mantle is in contact, on the sides of the body, with that of adjacent salpae in the same row, while in all chains the ventral surfaces of the mantles of the salpa3 on opposite sides of the chain come into contact. In chains like that of Salpa cordi- formis, Plate IV, Fig. 6, where the salpae in each row are placed end to end, the area of contact between their mantles is increased, as is shown in the figure, by pyramidal processes at the ends of the body. As long as the chain is intact these processes are bent at right angles to the long axis, of the body of the salpa, but when the chains are broken up by storms or other accidents the processes gradually straighten out into the long axis of the body, as is shown in Plate III, Figs. 2 and 3, which are dorsal and lateral views of a detached specimen of the aggregated form of Salpa cordiformis. This brief sketch of the characteristics of salpa chains should be followed by a discussion of their comparative history and of the phylogeny of the chain, but it will be best to postpone this for the present and to treat it in a separate section together with other questions of phylogeny, and we are now prepared to study the details of the process by which the stolon becomes converted into a series of salpae. SECTION 3. The Segmentation of the Stolon. The statement that the stolon becomes converted into, or is mor- phologically equivalent to, a single row of salpae placed like a file of soldiers with their long axes vertical or at right angles to the stolon and 90 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. with all their dorsal or neural surfaces turned towards its base, and their ventral surfaces towards its tip ; that this single row is converted into a double row by the passage of all the odd salpae to one side and all the even salpae to the other, and that each salpa rotates on its own axis so that all their dorsal surfaces come to face outwards, and all their ventral surfaces towards those of the opposite salpa3, sounds very simple. In reality the changes are far from simplicity, and they are extremely difficult to study or to describe in detail. I have enumerated them in succession and they must be so described, but they all go on simultaneously, and they begin at a very early stage, so that all the space relations of the body of each salpa are changing continually during its development ; nor do the changes affect all parts of the body alike, the oral ends with the ganglia being the first to move out into two ranks and the last to rotate. The oral ends of the bodies, and the ganglia, which are marked s in the figures, move from their primitive position on the middle line of the stolon at a very early stage, as is shown in Plate V, Fig. 2, although in Salpa cylindrica, Plate VIII, Fig. 2, long after the aboral or nuclear end of the body has assumed its secondary position, with the dorsal surface and cloacal aperture, g v , turned outwards or away from the axis of the chain, the ganglion s still lies on the proximal surface of each salpa, that is, on the surface which is turned towards the base or prox- imal end of the stolon, as is shown in the sections of the same stage in Plate XL. In this plate, as in all the others which represent sections of chain-salpae, the base or proximal end of the stolon is towards the bottom of the page, and the right side of the stolon is on the right side of the figure. The ganglion of a salpa which has moved to the right is shown at s in Figs. 14, 15 and 16, and the ganglion of one which has moved to the left at s, in Figs. 19 and 20, and it will be seen that even at this late stage the ganglia on both sides of the series are proximal, although the aboral ends of the bodies have rotated into their secondary position, as is shown in Plate XXXIX, Figs. 10, 11 and 12. The fact that these changes take place gradually and simultaneously and affect different parts of the body in different ways, renders a clear conception of their character an indispensable preliminary to the study of sections of the stolon, but, unfortunately, the converse is also true, and a clear conception of the character of the secondary changes can only be gained by the study of the details of the process of development. W. K. BEOOKS ON THE GENUS SALPA. 91 As two accounts are therefore necessary, I shall first describe the history of the chain-salpa as it would be if there were no secondary changes, and I shall illustrate this first account by imaginary diagrams. I shall also refer from time to time to figures of actual sections which show the points which are referred to, as this will gradually familiarize the reader with the secondary changes, although it will be as well for him to refrain from any attempt to analyze these secondary changes until they are taken up and discussed in due course. We left the stolon at the stage which is shown in Plate XXXIV, Fig. 1, in which it consists of a tube of ectoderm, a, colored violet in the figures ; inclosing a nerve tube, I, also colored violet, and running along the middle line of the upper surface of the stolon ; an endodermal tube, d', colored red ; a right perithoracic tube, /*, and a left one, g, both colored green ; a string of ova, n, colored orange, inclosed in a tubular follicle, w, colored blue ; and a lower ha3mal tube, i, and an upper onB, j, colored yellow. All these structures must be pictured as running along the stolon from its base to the point where it begins to strobilize into salpa3. Three longitudinal sections, with the same colors and letters, are shown in Figs. 4, 5 and 6. Fig. 4, which is near the bottom of the stolon, cuts the ectoderm, a, the lower ha3mal tube, /, and the string of eggs, n, in its follicle, m. Fig. 5 cuts first, on the right the ectoderm, a, then the right perithoracic tube below its lumen, then a number of scattered mesoderm cells, left uncolored, between the perithoracic tube and the area of thickened endodermal epithelium on the right side of the endodermal tube, d', then the ordinary flattened endodermal epithelium on the left, then, in the upper part of the figure, the lower blood tube, i, with its endothelium, colored blue, and in the lower part the layer of cellulose, 32, which fills up all the unoccupied spaces outside the haemal tubes, as is shown in Fig. 3, and finally, on the left the section again cuts the ecto- derm, which, in the section which was drawn, shows by its undulating outline the first trace which I have found of the segmentation of the stolon into salpae. Fig. 6 is a little higher up, and it passes through the lumen of the right perithoracic tube, h, and through mesoderm cells between it and the ectoderm, as well as between it and the thickened endoderm on the right of the endodermal tube. The section passes through the thickened endoderm on the left side of the endodermal tube, but it passes below the left perithoracic tube. I have not figured a longitudinal section through the nerve tube, Fig. 1, 1, at this stage, as it would be shown as a simple continuous tube. 92 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. Fig. 3 shows the details of a part of the right side of Fig. 1 more highly magnified, but it does not require explanation, since comparison with Fig. 1 will be enough. As already stated, the first trace which I have found of the strobiliza- tion of the stolon into a chain of salpae is the undulatory outline of the ectoderm, which is shown on the left of Figs. 5 and 6. The peculiar arrangement of the cells seems to indicate that each of the vertical ridges, which are cut transversely by this longitudinal section of the stolon, may be produced by the multiplication of a single row of ecto- derm cells, but I have not been able to prove this. In the stolon from which the sections were made the ridges appear on the left side before they do on the right, but the difference is very slight and is perhaps accidental. They first appear in the ectoderm of the middle region of the stolon, and gradually extend up and down towards its neural and germinal surfaces. Figures 8 and 9 are more highly magnified sections through the left half of the middle region of a slightly more advanced part of the same stolon, and 10 and 11 are through the right half. They are longitudinal sections like 4, 5 and 6, and are, of course, at right angles to the section shown in Fig. 3. They show that the ectodermal ridges are from the first almost exactly equal to each other in width, and that between the ridges the ectoderm grows inwards towards the axis of the stolon, in double folds, which thus form deep vertical furrows separating the ridges from each other. As these folds grow inwards they press upon the right and left perithoracic tubes, h and g, in such a way as to con- strict them, and at last to cut them up into series of closed cloacal vesicles, which soon become completely separated from each other, as is shown in the lower part of Fig. 11 at h. My sections indicate that the perithoracic tubes are passive, and that the active agency which divides them up into vesicles is the growth of the ectodermal folds, which, after passing across the perithoracic tubes, begin to push their way in to the lateral masses of thickened endodermal epithelium, as is shown in Fig. 11 and, further advanced, in Plate XXIII, Figs. 7 and 8. The ingrowth of the ectodermal folds goes on, carrying the endoderm before it, as shown in Fig. 9 of Plate XXIII, until the lateral masses of endoderm are folded into a series of vertical pockets, Fig. 9, 28, which open into the endodermal tube of the stolon, d'. The endodermal pouch on the right side of the stolon, Plate XXIII, Fig. 8, 27, is the rudiment of the right half of the branchial sac of the chain-salpa ; and that on the W. K. BROOKS ON THE GENUS SALPA. 93 left side of the stolon, Fig. 9, 28, is the rudiment of its left half. The right perithoracic vesicle, Fig. 8, g, is the rudiment of the right half of the cloaca ; and the one on the left, Fig. 7, h, the rudiment of its left half. Plate XV, Fig. 11, is part of a horizontal section on the left side of the stolon, showing these changes in a still more advanced stage. The ectodermal folds have now penetrated to a considerable depth, and they appear from the sections to be the active agents in the formation of the endodermal pockets as well as the cloacal vesicles. At the stage shown in Plate XXXIV the perithoracic tubes lie outside the endodermal thickenings, and the cloacal vesicles have for a time the same positions, as is shown in Plate XXIII, Figs. 7 and 8, g and h ; but as the endodermal pockets, Plate XV, Fig. 11, 28, grow deeper, their blind ends push past the cloacal vesicles, h, on their distal sides, so that the cloacal vesicles themselves come to lie nearer the base of the stolon than the endodermal pockets. Fig. 11 shows this for the left side of the stolon, and the arrangement of the parts is exactly the same on the right side. In this figure, as in all the others, the bottom is proximal or towards the base of the stolon, and the top distal or towards its tip. The changes which we have described are accompanied by important changes in other parts of the stolon, which must now be noticed. They are shown in Plate XV, and also in Plate V, Fig. 1. Like all the subsequent changes, they are of such a character that they cannot be understood or described with- out sections in at least two planes. I have therefore figured sections at each successive stage from two stolons, of which one was cut parallel to the long axis of the stolon, and transverse to the bodies of the salpse. A series of these sections is shown in Plate XV, Figs. 1 to 10. Fig. 1 is close to the germinal surface of the stolon; Fig. 2, a little higher up, through the chain of eggs; Figs. 3, 4 and 5, through the lower blood tube ; Figs. 6, 7 and 8, through the endodermal tube ; Fig. 9, through the upper blood-tube, and Fig. 10, through the neural surface of the stolon. In all these figures the top is distal and the bottom proximal, and the right side of the stolon on the right. Another stolon was cut into sections transverse to the long axis of the stolon, or parallel to the long axes of the salpa3, but instead of making separate drawings of these sections, I have superimposed the drawings and have thus constructed solid figures. Plate V, Fig. 1, is one of these reconstructions, to show the stage which is shown in Plate XV. It is a dorsal or proximal view and shows the surface which is below in the sections in Plate XV. 94 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. When the ectodermal ridges and furrows first appear on the stolon they are restricted to the regions of the perithoracic tubes, but as the folds grow deeper they also extend up and down, and soon completely encircle the stolon, dividing its surface up into a series of complete rings, each of which marks out the body of a salpa. At the stage shown in Plate XV the rings encircle the stolon, as the figures show, although the ectodermal folds penetrate much deeper into the stolon in some places than they do in others. In Plate V, Fig. 1, the shaded area around the periphery shows the extent of the infolded ectoderm, and comparison of this figure with the sections in Plate XV will show its relations much better than words. On the neural surface, Fig. 10, it has grown inwards so far that it has cut up the nerve-tube into a series of ganglia, s, and has pushed down around the ganglion in such a way as to shut it into a pocket of ectoderm, open towards the upper haemal tube. The active agent in the transformation of the nerve tube into a series of hollow vesicular ganglia seems to be the growing ectodermal fold. Below the ganglion there is a region, cut by section 9, where the ectodermal folds are very superficial and faintly marked ; while still lower down in the region of the endodermal tube, d', they are very deep, as shown in Figs. 8, 7, 6 and 5 ; still lower down, in the region of the lower blood-tube, i, they are slight and superficial, Figs. 4 and 3 ; while upon the germinal surface of the stolon, Fig. 1, they are very deep, so that they divide the genital string into a series of partially separated segments, which are about half shut in to ectodermal pockets. In general, the ectodermal folds are, as Plate V, Fig. 1 shows, deepest in those regions where the stolon contains solid structures, and most superficial where it is hollow, although each fold forms a complete ring. The structures shown in Plate V, Fig. 1, are the rudiments of a single salpa seen in proximal view, with a circular groove-like body cavity, which opens on all sides into the upper and lower blood spaces, / and j, and communicates through them with the body cavities of the salpae before and behind it in the series, as is shown in Plate XV, Fig. 4. This body cavity con- tains a closed vesicular ganglion, Fig. 10, s ; two closed cloacal vesicles, a right one, Figs. 7 and 8, g, and a left one, Figs. 5 and 6, h ; and two vertical endodermal pockets, which form the rudiments of the right half, Fig. 8, 27, and the left half, Figs. 7 and 6, 28, of the branchial sac. These pockets open into the endodermal tube, Plate V, Fig. 1, d', of the stolon, and through this communicate with each other, and also with the branchial sacs of the salpae before and behind them in the W. K. BROOKS ON THE GENUS SALPA. 95 series, as is shown in Plate XV, Fig. 7. At the lower end of the body is an egg, n, which lies partly in the body cavity and partly in the lower blood-tube, i. As Plate XV, Fig. 2, shows, it is not yet com- pletely shut off from the other eggs in the series, although the follicle is beginning to grow in between the eggs. Still lower down, in a pocket of the body cavity, the rudiment of the testis is represented by two folds of the follicle, Fig. I, m, m. Plate XXIII, Fig. 5, shows that the active agency in the segmentation of the genital string is the growth of the fold of ectoderm, a, and that this presses in to the genital string and cuts it up just as it cuts up the nerve tube and the peri thoracic tubes. The body cavity also contains scattered mesoderm cells, shown in Plate XV, Fig. 11, and the rudiment of the pericardium, Fig. 7, e. At this stage this is external to the outer surface of the right endodermal pocket, 27, and it is probably formed from the cells which are shown in Plate XXXIV, Fig. 3 and Fig. 11, between the right perithoracic tube and the ectoderm. They are shown at a more advanced stage at e in Plate XXIII, Fig. 8. I do not know whether there is a left pericardial rudiment or not. In Fig. 5 of Plate XV, shown more enlarged in Fig. 12, there is a rudiment on the left side, exactly like the one shown on the right side in Fig. 7, e. Salensky states (2, p. 44) that there is a left pericardial rudiment as well as a right one in the rudimentary ascidio- zooid of Pyrosoma, although the definitive pericardium and heart are formed from the right one alone. In Salpa the heart and pericar- dium are certainly formed from the right one, and if there is a left one it disappears very early, for I found no trace of it in older sections, and after I had drawn Figs. 5 and 111 discovered reasons for suspecting that I may possibly have drawn an inverted and misplaced section, and that the pericardial rudiment may possibly be only another figure of the right one, which is shown in Fig. 7. SECTION 4. The Development of the Chain- Sdlpa. The vertical endodermal pockets, 27 and 28, at first open, along their whole length, into the endodermal tube, d', of the stolon, but their upper or oral ends soon begin to grow up inside the body cavity of the salpa towards the ganglion, as is shown in Plate V, Fig. 2, so that sections above the level of the endodermal tube show them as closed tubes, Plate XV, Fig. 8, 28. In Fig. 9 the right side of the section cuts their upper blind ends, while the section passes above them on the left. They 06 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. are shown at a younger stage in Plate XXIII, Fig. 10, which is just a little above the level of Fig. 9. In Fig. 9 the pockets, 28, open into the cavity, d', of the endodermal tube, while in Fig. 10, 28, they are closed, and separated by the upper blood space, j, from the wall, d", of the endo- dermal tube, d'. These blind pouches grow rapidly and soon reach the top of the stolon, as is shown in cut N. ajid their slightly dilated oral ends, 27, o and 28, o, lie at the sides of the ganglion, s, a"s is also shown in section in Plate XXIV, Fig. 5, 27, 28 and s. This figure is the uppermost of the series shown in Plates XXIII and XXIV, and in these plates, as well as in those which follow, the capital letters A- A', B-B', etc., indicate the same individual in the series of sections on Plates XXV-XXXIII, the capital letter without an accent marking its right side, and the one with an accent its left side. A-A' is the youngest and most proximal salpa, B-B' the next, and so on to N-N', which is the oldest and most distal one shown in these plates. A-A', C-O, F-F', H-H', J-J', L-L' and N-N' are right-hand salpse, and the alternate ones are left-hand salpae. As the body of an older salpa is longer than that of a younger one, it is cut by more sections, and is shown in more figures. In all the figures 27 is the right half of the branchial sac, and 28 its left side. It will be seen from the series of sections in Plate XXIV that the upper or oral ends of the halves of the branchial sac, Fig. 5, 27 and 28, are dilated, and that, as we follow them down, Fig. 4, they become smaller, until they enlarge again at the level of the endodermal tube, Fig. 2, d', into which they open. Below the level of the endodermal tube they run down for a short distance as closed tubes, Fig. 1, C, and these tubes soon lengthen, like the oral ones, and become so long that they reach the genital surface of the stolon ; but before this stage the bodies of the salpae begin to push out of the line of the stolon towards the right and left alternately, and they also begin to rotate on their own axes. The omission of all reference at present to these secondary changes will simplify the description so much that I shall from this point on describe the development of the chain-salpa as if these changes did not occur, and I shall illustrate it by imaginary diagrams. We shall then be the better fitted for going over the subject a second time, as it is actually exhibited in nature, and as it is illustrated by the figures of real sections. Cut represents a proximal or dorsal view of a single salpa at the stage just described, in which the endodermal pouches have reached the W. K. BROOKS ON THE GENUS SALPA. 97 neural surface of the stolon or the oral end of the salpa, and have also begun to lengthen towards the genital surface of the stolon or the posterior end of the salpa, carrying with them the two perithoracic vesicles, g and h, and the rudiment of the pericardium, e. The dotted line, i and j, shows the inner edge of the ectodermal fold, and it also of course marks the boundary of the aperture by which the body cavity of the salpa communicates with those of the salpae before and behind it, while the circle, d', is the boundary of the opening which connects the rudi- mentary pharynx of the salpa with those of the adjacent salpae. The aboral ends of the endodermal pouches, 27 and 28, soon begin to grow backwards, as is shown in cut 0, and they carry with them the right perithoracic vesicle, . In Salpa cylindrica the aboral ends of the pouches remain distinct from each other until an advanced stage of development is reached, Plate XXXIX, 27 and 28, although they come together on the middle line and are separated only by a thin flat mesentery, which is shown in the sections as a double layer of flattened endoderm cells, running vertically from the gill, o, to the middle line of the ventral surface of the pharynx between the folds of the endostyle. The connection between the axial portion of the body and the lateral portion also persists in Salpa cylindrica very much longer than it does in any other species which I have studied. The position of the endostyle is most instructive in Salpa cylindrica. In Salpa pinnata it does not make its appearance until about the stage shown in Plate VIII, Fig. 1, after the pharynx has assumed its adult form and has entirely lost its connection with the endodermal tube, so that this species tells us nothing whatever about its morphological position, but in Salpa cylindrica it makes its appearance at the stage shown in diagram Q as a thickening or ridge in the ventral wall of each pharyngeal pouch, involving both its aboral and oral portions. As these pouches approach each other and unite, the two folds which are to form the endostyle approach and become parallel at their anterior and posterior ends, while they diverge from each other in the middle region of the body and pass around the endodermal tube. As they are differentiated at such an early stage in this species, they are of course involved in the secondary changes of position, and they furnish excellent landmarks for studying the character of these changes. Thus, in Plates XXXIX and XL if we fix our attention on the right half of the pharynx, 27, and the right half of the endostyle, in Fig. 10, we can trace them through Figs. 11, 12, 14, 15, 16, 17, 18, 19, 20 and 21. This last section is near the oral end of the body, anterior to the ganglion, Figs. 19 and 20, s, and we see that the right half of the pharynx of a right-hand salpa is a continuous structure, as it is represented at 27 in cut W. In a section, which has not been drawn, between Fig. 20 and Fig. 21, it gives rise to the small tube which is shown at 27 in Fig. 21. This tube, which is also shown at 27 in Plate VIII, Fig. 2, is the connection between the right half of the pharynx and the right side of the endo- 112 JOHNS HOPKINS UNIVEESITY MOEPHOLOGICAL MONOGEAPHS. dermal tube, d'. The relations of the left half of the pharynx of a left- hand salpa and the left half of its endostyle are, of course, the same; but as the sections which are figured in the plates do not cut the two salpse at the same points, it will be best to trace these structures also. Starting as before with Fig. 10 of Plate XXXIX, we can follow the left half of the pharynx, 28, and the left half of the endostyle of the left- hand salpa through Figs. 1 1, 12, 13, 14, 15, 16, 17, 18 and 19, and we find that like the structures on the right of the right-hand salpa they are uninterrupted, and we also notice that in section 15 the left half of the pharynx becomes separated from a small tube, 28, which can be followed through sections 16, 17, 18, 19, 20 and 21 until in section 23 it opens into the endodermal tube, d'. The relations of these parts are thus seen to be exactly as they are represented in cut W 27. The relations of the left halves of the endostyle and pharynx of a right-hand salpa are more complicated, as are also of course those of the right half of a left-hand salpa. Starting as before with Fig. 10 of Plate XXXIX, we will follow the left half, 28, of the right-hand salpa. Figs. 11, 12 and 13 show little change. In Fig. 14 the left half of the endostyle d 2 becomes stretched out towards the far side of the stolon. In Fig. 15, the left pharyngeal pouch 28 has divided into two tubes, one on the right side of the stolon, which comes to an end in Figs. 17 and 18, and one on the left side of the stolon, also marked 28, which can be traced through all the figures, until "in Fig. 23 it opens into the endodermal tube, and then goes on, as shown at 28 in Plate VIII, Fig. 2, over the top of the stolon to expand again, in Fig. 21, into the left half, 28, of the oral end of the pharynx. Tracing the right half, 27, of the pharynx of the left-hand salpa in the same way, we find it, in Figs. 14, 15 and 16, passing with the right half of the endostyle on to the right side of the stolon, and after passing over it, reappearing, at 27 in Fig. 18, at the oral end of the pharynx. These connecting tubes are shown in Plate VIII, Fig. 2, for the right- hand salpa, and it will be seen that they are exactly as they are repre- sented in cut W at 28. The right half of the pharynx is joined by a short tube, 27, to the right side of the endodermal tube, d', while the oral and aboral ends of the left half of the pharynx have no communication with each other except through the tube, 28, which passes around the stolon, and con- nects with the left side of the endodermal tube. The left half of the endostyle d 2 bends over onto the far side of the axial tube, as is shown at d 2 in Plate XL, Fig. 14, while the right half d' remains parallel to the W. K. BKOOKS ON THE GENUS SALPA. 113 axis of the body, as shown also in the figures on Plates XXXIX and XL. (Through an oversight in the lettering of Fig. 15 of Plate XL, the letter d l which is elsewhere used for the right half of the endostyle is used here for the left, and d, 2 for the right.) Soon after the stage which is shown in these figures, the axial por- tion of the body of Salpa cylindrica becomes entirely shut off from the lateral portion, which becomes converted into a complete salpa, while the axial portion which is morphologically a section taken out of the middle region of its body degenerates and disappears, the endodermal portion being the first to lose its connection with the salpa proper. In Salpa pinnata these changes take place at a very much younger stage, and for this reason they are difficult to follow without the guid- ance of Salpa cylindrica. SECTION 7. The Development of the Aggregated Form of Salpa pinnata. The direction of the movement outwards by which the single row of salpae becomes two rows differs in different species. In Salpa africana the movement is almost directly outwards, and Seeliger's figures show that this is true of Salpa democratica also. In Salpa cylindrica the movement is outwards and downwards in the direction indicated by the arrows in cut N, as is also shown by the figure of Salpa cylin- drica in Plate VIII, Fig. 2. In Salpa pinnata the movement is more downwards than outwards, as the figures in Plates V, VI and VII show. So far as the secondary changes are concerned, Salpa africana and Salpa democratica are the simplest, and they would be the easiest to understand if the stolon were straight. Salpa cylindrica comes next in intelligibility, but as its stolon is straight it is actually a more favorable subject for study. In Salpa pinnata the secondary changes are most obscure and difficult to trace, but the fact that the series of animals in its straight stolon develop in succession instead of in sets, renders it in many respects the most favorable species to study, and as its secondary changes are fundamentally like those of the other species, they should present no great difficulty after the history of Salpa africana and Salpa cylindrica is understood. I have selected this species, Salpa pinnata, for the most extended illustration, since, on the whole, it is the most instructive species, and the advantages it affords by its straight stolon and by the gradual development of the salpse more than compensate for the obscurity which comes from the very early stage at which the secondary changes occur. 114 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. A series of sections of this species, parallel to the long axis of the stolon, and transverse to the bodies of the salpae, is shown in Plates XV, XXIII-XXXIII and XXXVI, XXXVII and XXXVIII. Another stolon was cut into a series of sections at right angles to those in these plates, but, in order to diminish the number of figures, I have, instead of drawing all these sections, combined them to form the solid pictures shown in Plates V, VI, VII, and Fig. 1 of Plate VIII. In these figures the numbered cross-lines indicate sections which correspond, or nearly correspond, to the part of the figure which is crossed by the line, but it is very difficult to secure exact correspondence in all cases. As far as possible, I have shown corresponding stages in the two sets of plates, but the salpa3 change so rapidly during their development that the successive stages which are exhibited by one stolon may correspond to the unrepresented intervals between the successive salpae in another stolon, and I have not been so fortunate as to find two well preserved and perfect stolons which are exactly alike. The departures from exact agreement are not very great, nor are they of such a character as to perplex the reader, as they relate to slight differences in the shape of the bodies or to slight variations in the relative positions of the organs. Thus, for example, the line 41 in Plate VIII, Fig. 1, is represented as passing through the process which unites each salpa in one series with the ones diagonally opposite it in the other series. In Plate XXXVIII, how- ever, section 41 actually passes through the bottom of the lower blood space, i, and the connection between the salpa3 is cut in sections 50 and 61. This difference is partially due, perhaps, to distortion caused by hardening and imbedding, but it is chiefly due to the fact that the two specimens are not at exactly the same stage of development, although they are so nearly alike that they can be compared without difficulty. I shall now describe the two sets of figures in detail. Plate V, Fig. 1, shows a single salpa in the symmetrical position before the secondary changes of position begin. The shaded area shows the extent of the fold of ectoderm which separates this salpa from those adjacent to it, and it will be seen that this fold is most developed where the stolon contains internal organs, and least developed where it is empty. Thus the testicular folds of the genital string, Plate XV, Fig. 1, m, and the lower portion of the egg, n, are shut into an ectodermal pocket, while a little higher up, Figs. 2 and 3, the ectodermal folds are little more W. K. BEOOKS ON THE GENUS SALPA. 115 than undulations in the outline of the ectoderm. In the region of the endodermal pouches, 27 and 28, the body cavity is very much more shut in, and section 4 of Plate XV cuts the bottom of the left body cavity just above the point where it begins to deepen, while the right half of the section is at a lower level and shows the ectoderm as an undulating line. Section 6 of Plate XV shows the left pharyngeal pouch, 28, and the left perithoracic tube, h, shut in to a deep ectodermal fold, while on the right side of the section the ectodermal fold is cut below the level of the pharyngeal pouch. This section also shows the opening by which the left pharyngeal pouch, 28, opens into the endodermal tube, d', and in section 7 the opening of the right pharyngeal pouch, 27, as well as that of the left, 28, is shown. Fig. 7 also cuts the pericardium, e, and the right perithoracic tube, g, above its lumen. Fig. 8 is like Fig. 7 on the right side, except that it passes above the pericardium, but on the left it passes through part of the upper blood space, .;', and cuts the oral pro- longation of the left pharyngeal tube above its connection with the endo- dermal tube, d'. Fig. 9 passes above the left pharyngeal pouch, but it cuts the right one close to its blind end. Finally, section 10 cuts the ganglia, s, which are entirely shut in by the folds of ectoderm. The cuts N and 0, on page 80, and the series of sections in Plates XXIII and XXIV, show the gradual extension of the pharyngeal pouches towards the oral and aboral ends of the body. The oral ends, Plate XXIV, Fig. 5, 27 and 28, have reached the level of the ganglion, and are dilated and much larger than the tubes, Figs. 4 and 3, which connect them with the region of each pouch which opens into the endodermal tube, d', of Fig. 2. At this stage, as in the one before, all the salpae are exactly alike and symmetrically placed with reference to the bilateral plane of the stolon, but in the next stage, Plate V, Fig. 2, and in all the following stages, the salpa3 which move to the right are different from those which move to the left. Plate V, Fig. 2, shows a salpa which is to pass to the right, at the stage in which the first traces of the secondary changes make their appearance. Like all the other figures in this plate, it is a proximal or dorsal view of a single salpa. The lower part of the figure is like Fig. 1 in all essential particulars, but great changes have taken place in its upper part. As shown in Plate XXXI, Fig. 3, B-B', the ganglion, s, has moved outwards and downwards, and the oral ends of the two pharyngeal pouches have met and united behind or distal to the ganglion, or on what is to become the 116 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. ventral surface of the body, to form the oral end of the pharynx. This communicates directly with the aboral end of the right pharyngeal pouch, 27, as will be seen by examination of the series of sections of B in Plates XXXI and XXX. At the stage which is shown in Plate V, Fig. 2, the oral end of the pharynx communicates, through a long connecting tube over the upper surface of the stolon, with the aboral end of the left pharyngeal pouch, 28, but this connecting tube, which, as we have seen, persists in Salpa africana and Salpa cylindrica, Plate VIII, Fig. 2, 28, until the chain-salpa is perfectly developed, is very transitory in Salpa pinnata, and in salpa B-B' in the sections it has begun to degenerate and disappear. At the level of sections 3 and 2 of Plate XXXI it is shown at B, but in section 1 of this plate there is no trace of it and the body cavity of B' is empty. It reappears again at B in Plate XXX, Fig. 3, and in Fig. 2 the opening of the left pharyngeal pouch, 28, into the endodermal tube, d', is shown, and still lower down, at B' in Plate XXVIII, the communication between the left perithoracic vesicle, h, and the left pharyngeal pouch, 28, is shown. Plate V, Fig. 3, is also a right-hand salpa, a little older than Fig. 2. Both halves of the body have begun to move downwards, or towards the genital side of the stolon, and the only trace of the connecting tube of Fig. 2 is a short prolongation, 28', from the oral end of the pharynx. This prolongation can be traced up towards the top of the stolon for a short distance, but it soon disappears, so that the only communication between the oral and the aboral ends of the left half of the pharynx, 28, is now through the endodermal tube, d'. Plate V, Fig. 4, shows a left-hand salpa in which the connecting tube has completely disappeared, and the halves of the body have moved down so far that the ganglion, s, is on the level of the ectodermal tube, d, as is also shown in the section of the salpa E-E' in Plate XXX, Fig. 2. Above the endodermal tube the body cavity is now empty, as shown at E-E' in Figs. 1, 2 and 3 of Plate XXXI, and there is no trace of the connecting tube. The greater part of the right pharyngeal pouch, 27, now lies below the level of the endodermal tube, d', and its aboral dilated end is joined to the right side of the endodermal tube by a constricted tubular prolongation, which is shown, in section at E, in Figs. 1, 2 and 3 of Plate XXX. Fig. 3 cuts it at E below the level of the endodermal tube, while Figs. 1 and 2 show its opening into this tube. In Plate XXIX, Fig. 2, it is cut at C, as it begins to expand, and at C in Fig. 1 its communication with the right perithoracic vesicle, g, is shown, as it is W. K. BROOKS ON THE GENUS SALPA. 117 also at C in the figures in Plate XXVII. In Plate XXVII, C, the peri- cardium, e, is shown in Figs. 1 and 2, and also the diverticulum, g, which is to give rise to the digestive tract. Through an oversight, this is colored green in the figures instead of red. It will be seen from Figs. 3 and 4 of Plate V that the left pharyngeal pouch of a salpa which goes to the right, Fig. 3, like the right one in a salpa which moves to the left, moves much further from its primary position than the other pouch, and becomes correspondingly reduced in cross section, as is shown by the sections which we have just examined of the right pharyngeal pouch of a left-hand salpa. The very small tubular upper part of this pouch, shown in section in Plate XXX, Figs. 1 and 2, E, is the same as the connecting tube, which is shown, at a very much older stage, in Salpa cylindrica, at 27, in Plate VIII, Fig. 2. In Salpa pinnata, as in Salpa cylindrica, it connects the dilated aboral end of the right-hand pouch of the left-hand salpa, Plate V, Fig. 4 (or the left-hand pouch of the right- hand salpa, Plate VIII, Fig. 2), with the corresponding side of the endodermal tube, but in Salpa pinnata it soon degenerates and disap- pears, as will be shown soon, while it is present in advanced embryos of Salpa cylindrica. We have now to examine the sections through the left half of the left-hand salpa, shown in Plate V, Fig. 4. Plate XXX, Fig. 3, E' shows the union of the oral ends of the two pharyngeal pouches. Figs. 2 and 1, E', show the communication between the left one, 28, and the endodermal tube, d', and they also show the ganglion s, proximal or dorsal to the endodermal pouch and at the level of the connecting tube. In Plate XXIX, Fig. 2, c', the section cuts the left perithoracic vesicle, h, and the left pharyngeal pouch, 28, just below the endodermal tube. C' in Fig. 1 shows the communication between these two organs, as does also c' in Plate XXVIII, Figs. 1 and 2; d, in Fig. 3 of Plate XXVII, cuts the aboral ends of the pouch and vesicle, and d in Figs. 2 and 1 passes below these structures. It will be seen by comparing Plate V, Fig. 3, with Fig. 4, and by the examination of the sections, that the pericardium, e, is always in relation to the aboral end of the right pharyngeal pouch, and that the digestive organs always arise from this pouch, whether it is the larger, as it is in a right-hand salpa, Fig. 3, or the smaller, as it is in a left-hand salpa, Fig. 4. The next figure, Plate VI, Fig. 1, is a dorsal or proximal view of a right-hand salpa at the stage which is shown at H-H' in the sections on 118 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. Plates XXV-XXXI. The structures which are involved in the formation of the body proper have moved down onto the lower surface of the axial tube, and are beginning to approach each other on the middle plane of the body, which lies pretty nearly in the imaginary line between the letter n and the Fig. 27. This is a most important stage, and its careful examination is essential to a comprehension of the development of Salpa pinnata. The course which the ganglion has taken in its migration from its primitive position to the one which it occupies at this stage will be understood from the study of the figures on Plate V. In Fig. 1 all the ganglia are on the middle line of the stolon and in a row, as is shown in Plate XV, Fig. 10. In Fig. 2 those which belong to right-hand salpse have moved to the right, and those which belong to left-hand salpae to the left, although they are still near the top of the stolon, as is shown in Plate XXXI. In the next stage, Fig. 3, they have begun to move down towards the lower surface of the stolon, although they are still above the level of the endodermal tube^ while in Fig. 4 they have reached the level of the endodermal tube, as is shown in Plate XXX, Fig. 2, at F for a right-hand salpa, and at E' for a left-hand one. Finally, in the stage shown in Plate VI, Fig. 1, they are near the level of the bottom of the stolon, as is shown for a right-hand salpa in Plate XXVIII, Fig. 1, H, and for a left-hand salpa in Plate XXVI, Fig. 2, G'. As the salpa grows older the ganglion moves further and further from its primitive position, as is shown by comparing the figures in Plate VI, 1, 2 and 4, Plate VII, 3, 4 and 5, and Plate VIII, Fig. 1. Returning now to Plate VI, Fig. 1, it will be noticed that the extension of the two pharyngeal pouches downwards has resulted in the separation of each pouch into a small connecting tube and an expanded end within the body of the salpa. One of these connecting tubes, 27, joins the right pharyngeal pouch to the right side of the endodermal tube, while the other, 28, joins the left one to the left side of the endodermal tube. The latter is the same as the one shown at an earlier stage for a left-hand salpa at 27 in Plate V, Fig. 4, and is also the same as the lower part of tube 28 in Plate VIII, Fig. 2, while the former, 27, is the same as tube 27 of Plate VIII, Fig. 2. The three upper sections through Plate VI, Fig. 1, hardly call for explanation, although examination of the sections shows that an impor- tant change, the obliteration of the folds of ectoderm between the salpae, has already begun in this region, and at a later stage the ectoderm of all W. K. BROOKS ON THE GENUS SALPA. 119 the upper part of the axial tube becomes thin and flat and loses all traces of segmentation. Section H-H' of Plate XXX, Fig. 1, cuts the left connecting tube, H', at its opening into the endodermal tube. The section also cuts the lower wall, d", of the endodermal tube, crosses the upper part of the lower blood space, i, and cuts the right connecting tube, H, 27, just below its opening into the endodermal tube. Section H of Plate XXIX, Fig. 2, cuts the oral end of the pharynx and of the ganglion, s, but the left half of the section is not shown in the figure, although F' shows what it would be if it were present. In Plate XXVIII, H, sections 1 and 2 pass through the ganglion, s, and the oral end of the pharynx, 27, and the sections in Plate XXVII, H, cut the right pharyngeal tube below the ganglion. The section H-H', in Plate XXVI, Fig. 2, cuts the left connecting tube at H 1 just before it begins to expand in the body of the salpa, and it also passes through the lower blood space, i, close to its floor. In Fig. 1, H-H', the right pharyngeal pouch, 27, and the right perithoracic tube, g 1 . are cut through the aperture or gill-slit which unites these cavities. The left pharyngeal tube, 28, is much flattened dorso-ventrally, as is also the left perithoracic vesicle, and this latter is much elongated towards the middle line of the dorsal surface. In Plate VI, Fig. 1, this elongated portion of the left perithoracic vesicle is marked tf". It is the cloacal diverticulum shown at G'" in cut O. In the diagram the cloacal out- growths from the two perithoracic vesicles are represented as equal, but in Salpa pinnata the one which is derived, in the right-hand salpa, from the left vesicle, Plate XXVI, Fig. 1, H', h, and in the left-hand salpa, Fig. 1, G, from the right vesicle, g, appears much larger than the other in sections. This section also shows on the dorsal surface, between the two perithoracic vesicles, the two folds of the follicle which are to give rise to the fertilizing duct of the egg, and, nearer to the ventral surface, between the pharyngeal pouches, the single fold which is to give rise to the testis. Plate XXV, Fig. 6, H-H', shows the egg, m, in its follicle on the middle line of the dorsal surface between the perithoracic tubes g and h, and Fig. 4, H-H', shows on the left the blind end of the left pharyngeal tube and the aboral surface of the left perithoracic tube. On the right the section passes through the digestive tract, g 1 , and the pericardium, e, which is on the ventral surface of the right end of the body, and so directly behind the digestive tract that it is completely hidden behind it in a dorsal view like Plate VI, Fig. 1. 120 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. Fig. 2 of Plate VI is a dorsal view of a right-hand salpa and the tubes which connect it with that part of the axial tube which pertains to its body, while beyond it and partially hidden behind it is the body of a left-hand salpa. Fig. 3 is a ventral view of the same two salpa3, but as this second figure was reconstructed from the same set of sections, it is not reversed, but the left-hand salpa is on the left side as it is in Fig. 2. In effect, Fig. 3 is Fig. 2 as it would appear under a microscope when focused down to its far side. Sections of a right-hand salpa at nearly the same stage are shown in Plates XXXII and XXXIII at J-J', and sections of a left-hand salpa in K-K'. The planes of these sections are shown by the cross lines in the figures. The salpa? shown in the sections are a little younger than those from which the solid figures were reconstructed, but the difference is not very great. In Plate XXXIII, Fig. 9, cuts, on the right side below the level of the endodermal tube, the tubes which connect it with the right halves of the pharyngeal pouches ; all the tubes on the right side of this figure belong to the right halves of the bodies, but they belong alternately to right- hand and left-hand salpaB, while all those on the left, cut at the level of their openings into the endodermal tube, belong to the left halves of the bodies. Fig. 8, K-K', cuts the right and left connecting tubes of the left- hand salpa, as is shown by the line K, 8, in Fig. 3, while Fig. 8, J-J', cuts, as the line J, 8, in Fig. 2 shows, the left connecting tube of the right-hand salpa at J', while on the right it cuts the oral end of the pharynx and the ganglion. Fig. 7, K-K cuts, on the left, the left-hand connecting tube of the left-hand salpa at 28 just before it expands into the pharynx, while on the right at K it cuts the right connecting tube of the same salpa. The right-hand salpa, J-J', is not shown in Fig. 7, but L-L' is essentially like it, as is J-J' of Fig. 6. As the line J, 7, in Fig. 2 shows, this section cuts the left pharyngeal tube of the right-hand salpa twice, once at the extreme left of section L', and again near the middle of the section. In Fig. 6, J-J', however, the left pharyngeal tube, 28, is cut only once. The right end of section 7, ./, cuts the oral end of the pharynx at 27 and the ganglion at s. PART TWO. THE SYSTEMATIC AFFINITY OF SALPA IN ITS RELATION TO THE CONDI- TIONS OF PRIMITIVE PELAGIC LIFE ; THE PHYLOGENY OF THE TUNIC ATA : AND THE ANCESTRY OF THE CHORDATA. CHAPTERS VI, VII, AND VIII. CHAPTER VI. THE SYSTEMATIC POSITION OF SALPA. SECTION 1. The Evidence that Salpa is descended from a Fixed Form. I formerly believed that Salpa is the modern 'representative of an ancient Tunicate stem which has been pelagic throughout its whole history, and has been evolved at the surface of the ocean from an ancestor something like the modern Appendicularia ; and that the group has nothing in common with the fixed Ascidians except a common descent from this ancient form. While I still regard Appendicularia as the starting-point, I now feel confident that in all other respects this view is wrong, and that the facts force us to believe that Salpa is a modified descendant from a fixed form ; that it owes nearly all the distinctive peculiarities of its structure to a sedentary life ; that its adaptation to a free life at the surface is secondary and comparatively recent ; and that its only connection with Appendicularia is through this fixed ancestral form. It is possible, and I think probable, that this fixed form was not identical with any Ascidian which now exists, and it may have lacked some peculiarities which are shared in common by all modern Ascidians, but it must have been either a fixed Ascidian, or else an ancestor of the fixed Ascidians, with their habit of life and with essentially their structure. The facts which have forced me to abandon my original opinion and to substitute the view which has just been outlined are these. In the first place, comparative anatomy forces us to believe that the atrium of Salpa is identical with the perithoracic and atrial chamber of ordinary Ascidians, and the facts of embryology show beyond question that this is a real homology. In the ordinary Ascidians this system arises in the embryo as two ectodermal invaginations, one on each side of the body at some distance from the middle line, which grow inwards towards the pharynx, and become the perithoracic vesicles, which ultimately establish a communi- cation with the pharynx through the gill-slits. The two perithoracic vesicles or lateral atria approach each other on the middle line, dorsal 124 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. to the pharynx, and unite with each other to form the median atrium or cloaca, which opens to the exterior on the dorsal middle line through the atrial aperture. This aperture is, morphologically, the two original spiracular openings of the perithoracic invaginations, and in most cases these openings move towards each other until they meet and unite to form the single median atrial aperture. In some cases, however, they close up and disappear, while the atrial aperture breaks through as a new opening on the middle line. In the latter case, however, it is plain that we have to do with secondary changes, and the replacement of a circuitous ancestral history by a more direct mode of development. This secondary history is exhibited by Salpa, as I have shown already. To recapitulate briefly, we have in the embryo of the solitary form, cut A, p. 29, first, a pair of lateral perithoracic involutions from the surface of the body. Then, as Plate XII shows, these involutions, Fig. 1, g", extend towards the middle line and meet to form the median atrium, Fig. 2, g"', and also extend downwards towards the rudiment of the pharynx as a pair of perithoracic tubes, Fig. 4, g' and Fig. 5, g, which, at the stage shown in Plate XII, end blindly. Cut B on p. 29 shows these structures at this stage in a vertical transverse section. In the embryo which is shown in Plate XIII, the spiracular openings of the perithoracic tubes have closed, and the external portions of these tubes have moved towards the middle line, Fig. 6, g", where they meet above the median atrium, Fig. 7, g'", from which the perithoracic tubes, Fig. 8, gr iy , are continued down towards the pharynx, although they still end blindly, as the vertical section of the same embryo, on p. 30, shows. In the embryo shown in Plate XIV they open into the pharynx by a single large aperture or gill-slit, g", on each side of the middle line. In this plate, Figs. 3 and 4 cut the perithoracic tubes above the median atrium ; Fig. 5 cuts the median atrium ; Figs. 6 and 7 cut the tubes at lower levels, and Figs. 8 and 9 pass through the pharynx, c. Cut D on p. 31 is a vertical section of the same embryo. Finally the median atrial aperture, Plate XXXV, g", is formed as a new opening on the dorsal surface in the way which is shown in Plate XVII, Figs. 6 and 7. In the aggregated salpa the atrial structures are formed in essen- tially the same way. On each side of a cross-section of the stolon, Plate XXXIV, Fig. 1, there is a perithoracic tube, g and h, which probably arises at the base of the very young stolon, Plate XX, Fig. 3, as an involution of the ectoderm, although as Plate XXI, Fig. 7 shows, the W. K. BROOKS ON THE GENUS SALPA. 125 external opening is soon lost. Folds of the ectoderm of the stolon, Plate XXXIV, Figs. 9, 10 and 11, g and h, soon divide each tube up into a series of vesicles, one on each side of the body of each salpa. Each perithoracic vesicle, Plate XXVI, Fig. 3, E-E', g and h, now acquires an opening or gill-slit by which it communicates with its own half, 27 and 28, of the pharjmx, and they also become produced towards the middle line of the dorsal surface, Fig. 1, H-fT, g and h, where they meet, as shown at g and h in Plate XXXIII, Fig. 3, K-K and N-N', and ultimately unite to form the median atrium, Plate XXXVI, Fig. 6, g"', which afterwards acquires a median dorsal aperture, Fig. 5, g\ Writers on the embryology of Salpa and allied animals have involved the history of the atrial system in unnatural obscurity, for its origin in the salpa embryo and in the aggregated salpa is in perfect accordance with the teaching of comparative anatomy, and quite irreconcilable with any view except the one which regards these structures in salpa as strictly homologous with the median and lateral atria of ordinary Ascidians. It will be necessary to discuss in a later chapter the various views of the writers on the origin and homology of the atrial structures of Salpa, but at present this would lead us too far from our subject. I have shown in another place, p. 35, that, as Leuckart pointed out long ago, the atrial aperture of salpa is much nearer the mouth when it first appears than it is later, and that in this respect the ontogeny of salpa exhibits evidence of an Ascidian-like stage in its ancestry. The compactness of the ganglion of salpa, as contrasted with the elongated central nervous system of primitive chordata, and its position between the two apertures of the body, are also features of resemblance to the Ascidians ; and while there are now no traces, at any stage of its develop- ment, of numerous stigmatic gill-slits, like those of Pyrosoma, I shall show soon that there is indirect evidence that they at one time existed in the ancestors of salpa, which are, in this respect, Pyrosoma-like. The muscle bands of Salpa are easily intelligible as modified oral and atrial sphincters, and they are distinctly more irregular in the young than they are in the adult. In the young aggregated Salpa cylindrica, Plate VIII, Fig. 2, the fourth and fifth body muscles are clearly shown to arise as branches from an atrial sphincter, and Fig. 1 shows one of the body muscles arising in the same way in the aggregated Salpa pinnata. The peculiar anatomical relations of the pharynx and atrium of Ascidians are generally and justly regarded as modifications which were 126 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. gradually added on to the primitive Tunicate type, as adaptations to a sedentary life. If Salpa has been evolved from a swimming ancestor like Appendicularia through an uninterrupted series of free pelagic stages, we can give no explanation whatever of its Ascidian type of structure, while this is perfectly intelligible on the view that it is a modified Ascidian. SECTION 2. Views on the Relationships of the Swimming Tunicates. We must now discuss its systematic relation to the other swimming Tunicates, and the nature of its relation to the Ascidians. The Tunicates which are most like salpa are Pyrosoma, Anchinia, Dolchinia, Doliolum, and Octacnemius. We know too little of Octacne- mius to make much use of it, and if it is related to salpa at all it is not a stem form, but a salpa which has been secondarily modified, so we need give it no more attention at present. If Dolchinia is not a Dolio- lum, it is so near to Doliolum that it throws no new light on the origin of Salpa. We do not know the whole life-history of Anchinia, but our know- ledge of it is sufficient to show the justice of the opinion which, so far as I am aware, is universally accepted, that it is very closely related to Doliolum, and our comparison may therefore be narrowed down to Pyrosoma, Salpa, and Doliolum, including with the latter Anchinia and Dolchinia. The student of the recent literature on the systematic relationships of these three groups finds total confusion and irreconcilable contradic- tion. I shall not attempt to discuss all the opinions on record, as three selections are enough to show how little is definitely established. I shall therefore confine myself to the views of Grobben (Doliolum. Arbeiten Zool. Inst., Wien, IV, 2, 1882), Uljanin (7), and Herdman (14), and before I discuss the statements of these authors I will state as briefly as pos- sible the distinctive characteristics of each. Grobben holds (p. 67) that Pyrosoma, Doliolum and Salpa are very closely related ; that they form a natural group, and that they have been derived from the Compound Ascidians, through a Pyrosoma-like ancestor. So far as they are contained in this outline, I myself accept Grobben's views, except that I doubt whether the Ascidian ancestor had the dis- tinctive characteristic of any modern group of Ascidians. W. K. BROOKS ON THE GENUS SALPA. 127 Uljanin believes (p. 123) that we have in Salpa, Doliolum, and Pyro- soma, representatives of three distinct and independent lines of descent from the Simple Ascidians, from which Salpa and Doliolum are directly although independently descended, while Pyrosoma is a modified Com- pound Ascidian, and is, together with all the Compound Ascidians, descended from the Simple Ascidians. Uljanin gives a genealogical tree of the Tunicates, which, so far as the forms which we are now examining are concerned, is like this- Pyrosoma Doliolum Salpa Compound Ascidians Simple Ascidians Appendicularia Grobben and Uljanin agree in the opinion that Salpa, Doliolum, and Pyrosoma are all of them descended from Ascidians, but here the agree- ment ends, while Herdman, p. 124, rejects this very point, and holds that Salpa and Doliolum are descended directly from Appendicularia, and that they have nothing else in common with the Ascidians. He believes that Salpa and Doliolum are closely related, and that they form together a natural group, while Pyrosoma belongs to another line entirely, and is descended from a Compound Ascidian ancestor. While these views do not exhaust all the permutations and combina- tions of the three factors, they are enough to show the absence of a common ground for comparison, although it should not be difficult to place the subject upon a sound and permanent basis, as the forms which are to be considered are so very few and our knowledge of them so complete. 128 JOHNS HOPKINS UNIVEESITY MOEPHOLOGICAL MONOGEAPHS. SECTION 3. Salpa and Doliolum. First, as regards the comparison between Salpa and Doliolum, are we to believe with Uljanin that they are widely separated, or with Herdman and Grobben that they are closely related ? To my mind there is no room for doubt. Unquestionably Doliolum, Anchinia, and Salpa are more closely related to each other than they are to any other Tunicate except Pyrosoma. I have already shown, page 9, that the contrast in the muscle bands upon which the groups Cyclomyaria and Desmomyaria are based has no existence. In all Doliolums some of the muscle bands are imperfect rings ; in many species of Salpa the oral and atrial muscles are perfect rings, and in the most common and best known species of Salpa, the solitary Salpa democratica, most of the body muscles are as perfectly closed dorsally and ventrally as the rings of Doliolum. Anchinia, at least, is only by courtesy a Cyclomyarian, for it has no circular muscles except the oral and atrial sphincters, as the figure of the sexual animal given by Kowalevsky and Barrios (4), Plate III, Fig. 8, clearly shows. The groups Cyclomyaria and Desmomyaria are then purely artificial and without scientific value. Uljanin lays stress upon the presence of a tailed larval stage in the embryo of Doliolum and its absence in Salpa, but a comparison of the Doliolum larva which he shows in his Plate Y, Fig. 1, and which I have copied in my Plate VIII, Fig. 3, with the embryo of Salpa hexagona which is shown in my Plate III, Fig. 4, will show that the eleoblast k of the salpa embryo is a true tail, bearing exactly the same anatomical relations to the body as the tail of the doliolum larva. In my account of its minute structure, page 38, I have shown that sections prove it to be, without question, a degenerating larval tail. Uljanin says that he regards Salpa as standing alone among the Tunicates, and that its resemblance to Doliolum is superficial and due to secondary adaptation, but he gives no valid reason for this opinion. He says that the anomalous foetal development of Salpa, and the part which, according to Salensky, the tissues of the mother-organism take in the construction of the embryo, show that it is very different from all other Tunicates and render it difficult to place in the system. I have shown that the development of the salpa embryo, while very remarkable indeed, is by no means totally anomalous, and while we know nothing of the way its foetal mode of development and its placenta were acquired, they W. K. BROOKS ON THE GENUS SALPA. 129 must have been acquired at some period in its history, and I do not see that Uljanin comes any nearer to an explanation of its peculiarities by tracing it back to the Ascidians along an independent line. So far as I know, nothing in the history of the Ascidians helps us to understand these peculiarities of Salpa, and Uljanin's view accomplishes nothing except to force him to seek for a secondary explanation of the con- spicuous and undeniable resemblances between Doliolum and Salpa. As Grobben says, pa,ge 67, the shape of the body in Salpa and Doliolum, the situation of the mouth and atrial aperture at opposite ends, the arrangement of the muscle bands around the barrel-shaped body, and the free-swimming habit, are in themselves very conclusive evidence of their affinity, and while it is quite true that, as Uljanin points out, these adaptations to similar conditions of life might have been independently acquired, there is no good reason for thinking that this has happened, for they exhibit both superficial and fundamental similarity of structure. Uljanin indeed holds that the gills of Salpa are not homologous with those of Doliolum. If by this statement he means that the rod which usually, in Salpa, goes by the name of " gill " is not the same thing as the branchial slits or stigmata of Doliolum, his objec- tion is unworthy of consideration, for no one has ever seriously proposed any such homology, although the " gill " of Salpa has its homologue in Doliolum, as in Pyrosoma and the Ascidians, in the dorsal lamella. If he means that the two apertures by which the pharynx of Salpa com- municates with the atrium are not homologous with the branchial slits of Doliolum, I can only quote my observations, already detailed, which show that the gill-slits of Salpa are strictly homologous with those of the Ascidians. I shall soon examine Uljanin's statement that the atrium of Doliolum is not homologous with the atrium of Ascidians. SECTION 4. Salpa and Pyrosoma. I think that we may safely assume, as the first step in our com- parison, that Doliolum and Salpa are closely related, and we come now to the question whether Pyrosoma is closely related to Doliolum and Salpa as Grobben believes, or is a swimming Tunicate of very different origin, as Herdman and Uljanin believe. Since it was first pointed out by Huxley (Remarks upon Appendi- cularia and Doliolum, p. 602), the affinity between Pyrosoma and Doliolum has received general acceptance, and Grobben, p. 68, has shown that there is a very close anatomical agreement between them. 130 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. Uljanin (7), p. 124, characterizes the method of comparison which Grobben employs as irrational, inasmuch as he bases it upon purely imaginary changes in the anatomy of the fully developed animals, and makes no use of embryology. He says that Grobben overlooks the fact that the atrium or cloaca of Doliolum does not correspond to that of Pyrosoma and the other Tuni- cates, inasmuch as there are, in the Doliolum embryo, (p. 67) no peri- thoracic tubes like those of the Ascidians, since the cloaca of Doliolum is formed as a single median unpaired invagination of the ectoderm. It is not impossible that there may be a pair of lateral perithoracic tubes in Doliolum before the median cloaca and its aperture are formed, for his observations were made upon entire embryos, and no sections of the structures in question are figured. In Anchinia, which Uljanin justly regards as the nearest relation of Doliolum, Barrois has shown (10), pp. 226, 230, 236 and 242, that the atrial structures of the sexual animals arise in the buds as paired lateral ectodermal invaginations, and that their history is exactly the same as the primitive history in the Ascid- ians, so that Uljanin must either deny the homology between the atrium of Doliolum and that of Anchinia, or else he must recognize its homology with that of the Ascidians and Pyrosoma. Even if his observations are accepted as final, his deduction by no means follows. In some echinoderm Iarva3 the coelomic pouches separate from the gut before they separate from each other, while in most cases they are distinct from each other before they become separated from the gut, yet all embryologists regard them as homologous, and it is vastly more probable that the ectodermal rudiments of the perithoracic tubes of Doliolum meet on the middle line before the invagination takes place than that the atrium of Doliolum is a new structure. While history gives ample reason for his statement, p. 124, that the facts of comparative anatomy may be distorted or misrepresented, all naturalists know that anatomy often proves homology and furnishes a key to embryology. Thus mammalian teeth and the flat bones of the mammalian cranium are held to be dermal scales, although mammalian ontogeny gives no record of their phylogeny. So too the mammalian body cavity is held to be a series of coelomic pouches from the gut, and the mouth of a starfish is held to be strictly homologous with the mouth of a sea-urchin, although in the one case it is the same as the larval mouth, while in the other it breaks through on the left side of the larva. In all these cases we reconstruct the primitive ontogeny from the W. Z. BROOKS ON THE GENUS SALPA. 131 evidence of comparative anatomy, and explain modern ontogeny as the result of secondary modification. Comparative anatomy shows clearly that the atrium of Doliolum is homologous with that of Pyrosoma and the Ascidians, and the onto- genetic history in Doliolum does not present any great difficulty. I shall have to refer again further on to the nature of the evidence from embryology, but I think all morphologists agree that when organs or animals which are shown by their anatomy to be homologous, differ in their ontogeny, we have good ground for expecting to find evidence that the ontogeny has undergone secondary modification, and that very considerable embryological diversity is quite compatible with close syste- matic affinity. On the other hand, when two animals whose anatomy does not for- bid comparison exhibit striking ontpgenetic resemblances, these must be held to be evidence of phylogenetic relationship. The strongest evidence of the affinity of Salpa and Pyrosoma is of this sort, and every student who has concerned himself with either the embryology or the asexual multiplication of these animals has expressed or implied his strong conviction of their relationship. My own studies have forced me to differ from Salensky most essen- tially regarding the part which the follicle cells take in the construction of the salpa embryo, but I fully agree with him (17), p. 84, that the embryological phenomena prove that Salpa and Pyrosoma are closely related. The embryological evidence of their affinity is so fundamental that their whole history must be studied before its weight can be fully appreciated. I have shown in the chapter on the significance of the Salpa embryo, that the history of its development can be explained only on the hypothesis that it exhibits, in a high degree, secondary complications which are shown at a much more primitive stage by Pyrosoma, which is, so far as our knowledge goes, the only animal which does give us any material aid in the interpretation of the Salpa embryo. I have also shown that there is some reason for believing that the egg of Salpa has at some time in the past been furnished with a food-yolk, and has had a meroblastic mode of development like Pyrosoma, although it may be that these characteristics never became as highly evolved in the ances- tors of Salpa as they are in Pyrosoma. We must not forget, however, that while there is no other animal known to us with an embryology as much like that of Salpa as Pyrosoma, the differences between the two are very considerable. 132 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. The most characteristic peculiarities of the Salpa embryo are its foetal development, and the presence of a placenta and foetal envelopes. The Pyrosoma embryo does not present the slightest trace of these pecu- liarities, nor is there any indication whatever in the life-history of Salpa of the degeneration which is so remarkably manifested by the Pyrosoma embryo, and we cannot believe that the life-history of either of them is like that of their common ancestor. In fact the differences are so great that if the resemblance between the embryos were the only thing which these animals have in common, we might well doubt whether this alone is sufficient to prove their affinity, but their whole organization testifies to their very close relation- ship, and if we make a comparison, not between Salpa and Pyrosoma alone, but between Pyrosoma on the one hand and Salpa, Doliolum and Anchinia on the other, the anatomical resemblance is most impressive. It is exhibited not only by the fundamental plan of their structure, but also in superficial details. Thus the primary colony in Pyrosoma consists, as it does in Salpa pinnata, Plate I, Fig. 2, of a wheel or rosette made up of a number of zooids placed parallel to each other with their dorsal surfaces outwards, their ventral surfaces towards the axis of the colony, and their mouths all pointing the same way. It is also interesting to find in both Salpa and Pyrosoma peculiar luminous organs with the same structure, and in positions which, while not strictly homologous, are sufficiently alike to show that they have in all probability been inherited from a common source. The solitary Salpa pinnata, Plate I, Fig. 6, has five pairs, and the aggregated form of the same species, Plate I, Fig. 3, one pair of longi- tudinal rod-like organs in the intermuscular spaces at the sides of the dorsal surface. These organs, which were noted by Forskal, have given rise to much speculation, although no one could examine a living Salpa pinnata with- out discovering that they are intensely luminous. I am not aware that this interesting fact has ever been recorded. They have been regarded as ovaries (Meyer), or renal organs (H. Muller), but recent writers have refrained from conjectures as to their function. My first living specimens of this species were examined on deck under the full blaze of the noonday sun, but in spite of the bright day- light my eye was instantly caught by these longitudinal rods which glowed with a light of their own as the animals floated in a dish of W. K. BROOKS ON THE GENUS SALPA. 133 water. In sunlight the luminous organs have a tinge of purple, but at night their light is as white as the glow of an incandescent wire. Sections show that each organ consists of an aggregation of blood corpuscles in a sinus or dilation of the blood spaces of the body cavity. As most of the corpuscles are swollen, irregular, granular, and in process of degeneration, the sections seem to indicate that the degeneration of the corpuscles is the source of the light, although we cannot look for much information on this point from dead specimens. Since the time of Savigny, Pyrosoma has been known to have a pair of elongated organs on the sides of the pharynx near the mouth, and as in the case of Salpa, there has been much vague speculation as to their nature. Savigny thought they were ovaries ; Huxley suggested that they were kidneys, and Keferstein and Ehlers believed that they might be organs for the production of buds. According to Herdman (p. 22), Panceri first showed that they are really organs for the production of the light for which Pyrosoma is so famous and from which it gets its name. Salensky's account of the development of these organs (17), p. 48, and his figure, Taf . 7, Fig. 59, Lzgr, seem to show clearly that, in Pyro- soma, as in Salpa, the organs are simply spaces in the body cavity filled with swollen and degenerating cells. This point of agreement between Salpa pinnata and Pyrosoma, while in itself of little weight, must be regarded as a part of the evidence of their relationship. Ussow discovered and Bolles Lee has more fully described in the aggregated form of Salpa, a pair of sensory ectodermal tentacles situated near the mouth, and Salensky (17), p. 28, finds in Pyrosoma a pair of tentacles which exhibit a general similarity to those of Salpa, and thus add a little to the weight of the evidence from other sources ; although, inasmuch as Salensky's figures and description show that they are at the posterior end of the body in Pyrosoma, we cannot give unqualified assent to his statement that they are identical with the sense tentacles of Salpa in form and structure as well as position. The opinion that Salpa and Pyrosoma are closely related does not, however, rest upon these superficial resemblances, but upon their funda- mental identity of structure, although one of the details, the resemblance in their asexual multiplication, is so complete as to be almost enough in itself to establish their affinity. Kowalevsky's and Huxley's accounts of the proliferous stolon of 134 JOHNS HOPKINS UNIVEESITY MORPHOLOGIC A.L MONOGRAPHS. Pyrosoma, and the more recent accounts by Seeliger (15) and Salensky (17), would, so far as the anatomical structure of the stolon is concerned, serve as a description of the salpa-stolon. In each case the stolon is bilateral in the same plane as the animal which carries it ; in each case it grows out on the ventral middle line near the heart, and in each case it consists of, 1st, an ectodermal tube continuous with the ectoderm of the parent ; 2d, an endodermal tube which arises on the ventral side of the pharynx between the folds of the endostyle ; 3d, two perithoracic tubes, which, in the primary stolon at least of Pyrosoma (Salensky, 17, p. 475), are ectodermal in ultimate origin, as they are in Sal pa; 4th, a nerve tube, or, in the primary stolon of Pyrosoma, a series of nerve vesicles (Salensky, 17, p. 475),which are ectodermal in origin like the nerve tube of Salpa ; 5th, two blood tubes, and, 6th, a genital string. The organs which are derived from these various structures are alike in both cases, and my account of the formation of the aggregated salpaB shows that, morpho- logically, they form a single series placed belly to back, and developing in succession from the tip of the stolon to its base exactly as they do in Pyrosoma, and that each one in the series is joined by its dorsal surface to the ventral surface of the next younger, and by its ventral surface to the dorsal surface of the next older in the series, and that the right and left sides of each one, as well as its dorsal and ventral surfaces, lie, mor- phologically, in the same relation to space as the corresponding parts of the parent. In order to appreciate this resemblance in all its details it is neces- sary to master the minute history in each case, but the phenomena are so complicated and their points of agreement so numerous that the deduction seems to be irresistible that the method of asexual multipli- cation has been inherited in both Salpa and Pyrosoma from a common source. I believe, therefore, that the present condition of our knowledge enables us to state with confidence that Pyrosoma, Doliolum and Salpa form a natural group, or great branch from the Tunicate stem. I have spoken of the various members of this group as closely related, but I have not intended to imply that the relationship is in all cases equally close. Anchinia, Dolchinia and Doliolum are clearly more closely related to each other than to any other Tunicate, and the gen- erally accepted view that, in this group, Anchinia is nearest to Pyrosoma seems to have much to commend it. The various species of Salpa are more closely allied to each other than to any other Tunicate, and Salpa W. K. BROOKS ON THE GENUS SALPA. 135 pinnata seems to have more features of resemblance to Pyrosoma than any of the others, but I think this is all we can state with any confi- dence. I am quite unable to satisfy myself as to the exact nature of their relationship, or to formulate it either in definitions or in a genealogical tree. SECTION 5. The Nature of the Relationship of Salpa, Doliolum, and Pyrosoma to the Ascidians. This subject has been discussed by both Uljanin and Herdman, but there is little harmony in their results, although they agree that Pyro- soma must be regarded as a descendant from the Compound Ascidians. Uljanin says that we are forced to believe that Doliolum has branched off directly from the Simple Ascidians, since its '' ausserordentlich grosse " relationship to them is proved by its embryology. His account of its embryology is very peculiar, however, as he says that the archen- teron of the gastrula disappears, and that the entire digestive system of the adult is formed subsequently from an involution of the ectoderm at the oral end of the larva. As nothing of this sort has ever been described in the Simple Ascidians, the embryology of Doliolum can hardly be stated to bear any " unusually great " resemblance to that of the Simple Ascidians. Uljanin, basing his discussion of the relationships among the Tuni- cates upon their chordate affinities, and upon the ancestral significance of Appendicularia and the Ascidian tadpole, assumes, as his fundamental principle, that those Tunicates which have most perfectly retained their primitive or ancestral ontogeny, as this is exhibited by primitive chor- data, must themselves be most primitive. Even if we admit this, we see that the ontogeny of Doliolum by no means proves its primitive character, for we are forced by his own account to believe that there has been very great secondary modification in its very early stages of development. The principle cannot be accepted, however, as all embryologists know. No one would hold that a starfish, in which the ontogeny has been simplified by development in brood pouches, is more modern than one with a free larval life. The modified ontogeny is more modern than the unabridged ontogeny, of course, but it by no means follows that the adult animal is also modern. As the ontogeny of the Simple Ascidians adheres more closely than that of the Compound Ascidians to the primitive chordate type, as this 136 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. is exhibited by the anatomy of Appendicularia and by the ontogeny of Amphioxus, he assumes that the simple Ascidians themselves are more primitive than the Compound Ascidians, but his reasoning seems to me to rest upon an erroneous conception of the value and significance of the evidence from embryology. The resemblance between the larvas of the Simple Ascidians and the primitive chordata is unquestionably phylogenetic, but it does not by any means follow that the adult animals are older than the Compound Ascidians, and Pyrosoma, Salpa, and Doliolum. The locomotor chordate tadpole larva serves to distribute the species and to establish new sessile animals at a distance from their fixed parents, and these parents might attain to almost any degree of evolution without losing this phylogenetic larval stage so long as its retention continued to be important. There is no reason for believing that an adult which has retained it is itself primitive, and the great size, complexity and independent indi- viduality of the Simple Ascidians seem to be marks of great specialization rather than of low rank. The most characteristic and dominant organs of a Tunicate are the branchial sac and the atrial system, and these structures are peculiarly complex and highly evolved in the Simple Ascidians. The leathery, lumpish "sea-squirts" are much less attractive than the delicate, transparent, pelagic Tunicates, whose whole organization is at once seen to be beautifully adapted to the conditions of their life, and it is natural to assume that these latter are the most improved and modern forms, and that the Simple Ascidians are much older, but the muscular mantle and richly ciliated pharynx of the Simple Ascidians are well adapted for protection and nutrition, and the number and diversity of the families and genera and species of Simple Ascidians, and their abundance and wide distribution, prove that their organization has been brought into very complete harmony with their conditions of life, and that, in this respect, they are highly specialized. The colonial Ascidians spread by budding ; and the accidents which break up the colonies scatter the fragments and distribute the species. Small colonies of Perophora and Botryllus are often taken in the tow-net far from land, and the ancestral locomotor stage is much less essential to these animals than to the Simple Ascidians. The adult Pyrosoma, Doliolum, and Salpa are themselves specialized for locomotion, and as they are not dependent upon local conditions, but are at home all over the ocean, they do not need locomotor larvaB, while W. K. BEOOKS ON THE GENUS SALPA. 137 their habit of life seems to be peculiarly favorable for the development of the young upon or within the parent. While the large food-yolk of Pyrosoma, and the rudimentary con- dition of its Cyathozooid, and the placenta and foetal life of Salpa, and the differentiation of a set of Ascidiozoids to carry the sexual animals in Doliolum and Anchinia, are obviously secondary, they do not prove that the adults are any more modern than the Simple Ascidians, nor is the reduction of the tail of the larva to a food reserve anything anomalous, for in all ascidians it is used up as food after it has served its purpose, and we might expect to find it directly devoted to this new use when its locomotor function becomes unnecessary. It therefore seems to me that the reasons which Uljanin draws from the facts of embryology for holding the view that the Simple Ascidians are more primitive than the other Tunicates, and for regarding Pyrosoma and Salpa as more divergent from the primitive type than Doliolum and the Compound Ascidians, are perfectly consistent with a different inter- pretation, and we find that Herdman's view is very different. Briefly summarized, his opinion is that Pyrosoma is not at all closely related to Salpa and Doliolum, and that the two latter forms, with Anchinia and Octacnemius, form a natural group, the Thaliacea, which has been pelagic throughout its whole history, and has nothing in com- mon with the Ascidians except its common descent from an ancestor like Appendicularia, while he regards Pyrosoma as a fixed Compound Ascidian which has secondarily become adapted to a swimming life. He says (p. 149), " It seems to me that the passage from Appendi- cularia mossi through Anchinia rubra to Doliolum, and through the ancestral Doliolidae to Salpa, is so natural and simple that it becomes very improbable that the Thaliacea have ever been fixed forms. It is extremely unlikely that they are, as Uljanin supposes, a group of Simple Ascidians which, after being fixed, betook themselves again to a free swimming mode of life." On page 124 he says that Appendicularia mossi "is perhaps the nearest form known to the ancestral Tunicates, from which the two great lines of degeneration (?) diverged, the one leading to the Doliolidse and Salpida3, and the other to the Simple and Compound Ascidians." Herdman's introduction of Appendicularia mossi into his discussion is unfortunate, and he also fails to give due weight to the evidence of the affinity of Salpa and Pyrosoma which has been pointed out so often by Huxley, Grobben, Salensky, and many others, and in view of all the facts 138 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. of the case we must reject the first of his two lines of descent from Appendicularia. The second line leading to Pyrosoma is perhaps pictured more minutely than the facts warrant, but I think we can safely follow him : 1st, in his statement of the way in which the Ascidian line arose from an ancestor like Appendicularia, and second, in his derivation of Pyro- soma from an Ascidian-like ancestor which is not descended from the Simple Ascidians. Finally I think we are fully justified in believing with G-robben, in opposition to Uljanin and Herdman, that Salpa (including Octacnemius), Doliolum (including Anchinia and Dolchinia), and Pyrosoma are more closely related to each other than to any other known organism, but I do not see how we can safely commit ourselves to a more minute phylogeny. Gastrang, in a recent paper on the gill-slits of Tunicates (Proc. Eoyal Soc. London, May 19, 1892, pp. 505-513), holds that the transverse stig- mata of Pyrosoma are protostigmata; that they present conditions which are embryonic or larval in the Ascidians, and that all the secondary stig- mata in a transverse row, in the pharynx of an Ascidian, correspond to or are homologous with a single one of the transverse protostigmata of Pyrosoma. He therefore believes that the pharynx of Pyrosoma approaches nearer to the ancestral type than that of the Ascidians. The literature of the subject is not all of it in harmony with his view, but I see no valid ground for rejecting his conclusion, which is certainly worthy of a provisional acceptance. Speculations upon the ancestry of Pyrosoma must then conform to the conditions which it imposes, and we cannot derive the pharynx of Pyrosoma from that of any fixed Ascidian which we know. The facts seem to me to prove that Salpa and Pyrosoma are descended from a fixed form, although Gastrang's studies seem to force us to believe that this fixed form resembled Pyrosoma rather than the Ascidians in the structure of its pharynx, and that it was in this respect more primitive than any Ascidian which we know, although I see no reason why it may not have been the parent of the fixed Ascidians, or why it may not have resembled them in habit of life and in general structure. In this discussion I have accepted without comment the opinion which seems to be shared by all recent writers on the Tunicata, that W. K. BROOKS ON THE GENUS SALPA. 139 the starting-point for the Tunicate phylum is to be found in a form which was essentially like the modern Appendicularia, and that the line which joins the Tunicata to the Vertebrate phylum must pass back- wards from the Ascidians through the ancestors of Appendicularia to its point of union with the Vertebrate line. In this opinion I fully concur, but before I discuss the remote ances- try of the Tunicata, I wish to approach the subject from another point of view, and to speak of the life-history of Salpa in certain broad rela- tions to the question of the origin of the Metazoa. I shall then in another chapter discuss the question of the degeneracy of the Tunicata and the significance of the Appendicularia stage in their development. CHAPTER VII. SALPA IN ITS RELATION TO THE EVOLUTION OF LIFE. Salpa is distinctively a pelagic animal, adapted by its whole structure for a free existence, and for life at the expense of the micro-organisms in the water of the ocean. To understand its position and significance in the economy of nature, we must have before us the broad outlines, at least, of a picture of the conditions under which oceanic life has been evolved. I believe that the history of the evolution of Salpa, as told by its embryology, is most suggestive and important, and that it contributes to the solution of some of the most profound and fundamental problems of biology, and brings us into conflict with some of the most favorite dicta of modern morphology. I shall therefore devote considerable space to a review of certain familiar features of ocean life, in order that I may pre- sent in this way my view of the significance of the phylogeny of Salpa, in its bearing upon the first principles of morphology. Contrast between Terrestrial Life and Marine Life. In a picture of the land, the mind calls up a vast expanse of verdure, broken only by water, and stretching through forest and meadow from high up on the mountains, over hills and valleys and plains, down to the sea. Our picture of the ocean is an empty waste, stretching on and on with no break in the monotony, except, at long intervals, a floating tuft of sargassum, or a flying fish, or a wandering sea-bird, and we never think of the ocean as the home of vegetable life. It contains plant-like animals, "zoophytes," in abundance, but while they resemble plants or flowers in form and color, and in their mode of growth, they are true animals and not plants. At Nassau, in the Bahama Islands, the visitor is taken in a small boat, with windows of plate glass set in the bottom, to visit the "sea- gardens" at the inner end of a channel, through which the pure water from the open sea flows between two coral islands, into the lagoon. W. K. BROOKS ON THE GENUS SALPA. 141 Here the true reef corals grow in quiet water where they may be visited and examined. The bottom of the boat is below the surface ripples and reflections. When illuminated by the vertical sun of the tropics, and by the light which is reflected back from the white bottom, the pure transparent ocean water is as clear as air, and the smallest object, forty or fifty feet down, is seen distinctly. As the boat glides over the great mushroom-shaped coral domes which arch up from the depths, the dark grottoes between them, and the caves under their overhanging tops, are lighted up by the sun far down among the flower-animals or anthozoa and the animal plants or zoophytes which are seen through the waving thickets of brown and purple sea fans and sea feathers as they toss before the swell from the ocean. There are miles of these "sea-gardens" in the lagoons of the Bahamas, and it has been my good fortune to spend many months studying their wonders, but no description can convey any conception of their beauty and luxuriance, and I never spent a day among the reefs without longing, at every turn, for the skill to copy with a brush the new beauties which never ceased to present themselves. The general effect is very garden-like, and the beautiful fishes of black and golden yellow and iridescent cobalt blue hover like birds among the thickets of yellow and lilac gorgonias. The parrot fishes (Diodon and Ballistes) seem to be cropping the plants like rabbits, but more careful examination shows that they are biting off the tips of the gorgonias and branching madrepores, or hunting for the small Crustacea which hide in the thicket, and that all the apparent plants are really animals. The delicate star-like flowers are the vermilion heads of boring annelids, or the scarlet tentacles of actinias, and the thicket is made up of pale lavender bushes of branching madrepores and green and yellow and olive masses of brain coral, of alcyonarians of all shades of yellow and lilac and purple and red, and of red and brown and black sponges. Even the lichens which incrust the rocks are hydroid corals, and the whole sea-garden is a dense jungle of animals where plant life is represented only by a few calcareous alga3, so strange in shape and texture that they are much less plant-like than the true animals. The scarcity of vegetation becomes still more noticeable when we study the ocean as a whole. On land, herbivorous animals are always much more abundant and 142 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. prolific than the carnivora, as they must be to keep up the supply of food. Insectivorous birds are very abundant, but they are not numerous enough to keep the plant-eating molluscs and insects in check, and the devastation which is caused every year by the armies of grasshoppers and locusts and herbivorous beetles and by other less conspicuous insects, shows that their natural enemies are not numerous enough to overtax their productive power. The birds which feed upon grain and seeds and fruit are very abundant indeed, and they sometimes gather at their breeding grounds, or places of assembly, in innumerable multitudes, but the hawks and owls which prey upon them are never numerous. The small rodents, such as the rats, mice, squirrels and rabbits, are the most abundant and prolific of animals ; but the small carnivora are so rare that their very existence is known to few except naturalists and trappers. The homes of the wild sheep and goats, deer, antelopes, cattle and horses support these large mammalia in incredible numbers, but their carnivorous enemies are never abundant. It is clear that if the destruc- tion of the plant-eaters exceeded their productive power, both herbivora and carnivora would disappear, and terrestrial life would come to an end. The animal life of the ocean shows a most remarkable difference, for marine animals are almost exclusively carnivorous. The birds which live upon the ocean, the terns, gulls, petrels, divers, cormorants, tropic birds and albatrosses, are very numerous indeed ; so numerous that in many parts of the ocean some are always visible in calm weather around the vessel, wherever it may be. The only parallel to the pigeon-roosts and rookeries of the land is found in the dense clouds of sea-birds around their breeding places, but these sea-birds are all car- nivorous; most of them are fishers, and others, such as the petrels, scoop up the copepods and pteropods from the surface. Even the birds of the sea-shore subsist almost exclusively upon animals such as mol- luscs, Crustacea and annelids. The seals pursue and destroy fishes ; the sea-elephants and walruses live upon lamellibranchs ; the whales, dolphins and porpoises, and the marine reptiles, all feed upon animals, and most of them are fierce beasts of prey. The manatee is a vegetable feeder, but it is not strictly a marine animal, since its home is in the mouths of great rivers. There are a few fishes which pasture in the fringe of seaweed which grows in the littoral zone of the ocean, and there are some which browse W. K. BROOKS ON THE GENUS SALPA. 143 among the floating tufts of algae upon its surface, but most of them frequent these places in search of the small animals which live among the plants. All the floating fishes whose home is the floating sargassum ; the file fishes and trigger fishes (Ballistida) ; the trunk fishes (Ostracion) ; the frog fishes (Antennarius) ; and the puffing fishes (Tetradon and Diodon) are carnivorous, living upon the barnacles and molluscs and hydroids which grow upon the sargassum, or upon the Crustacea, young fishes and the floating larvae which seek its shelter. In the Chesapeake Bay, the sheepshead (Diplodus probatocephalus) browses among the algae upon the submerged rocks and piles like a marine sheep, but its food is exclusively animal, and I have lain upon the edge of a wharf watching it crush the barnacles and young oysters until the juices of their bodies streamed out of the angles of its mouth and gathered a host of small fishes to snatch the fragments as they drifted away with the tide. Many important fishes, like the cod, pasture on the bottom, but their pasturage consists of molluscs and annelids and Crustacea, instead of plants. The vast majority of marine fishes are fierce hunters, pursuing and destroying smaller fishes, and often exhibiting an insatiable love of slaughter, as in the case of our own blue-fish and the tropical albacore and barracuda. Others, such as the herring, feed upon smaller fishes and the pelagic pteropods and copepods ; and others, like the shad, upon the minute organisms of the ocean, but all, with few exceptions, are carnivorous. In the other great groups of marine animals we find some scaven- gers, some which feed upon micro-organisms, and others which hunt and destroy each other, but there is no group of marine animals which corresponds to the herbivora and rodents and plant-eating birds and insects of the land. The pelagic copepods are, of all the marine Meta- zoa, the ones whose place in the economy of nature is most like that of the terrestrial plant-eaters. They swarm in innumerable multitudes at the surface of the ocean, and also below it down to a depth of a mile or more, and they furnish the chief food for most young fishes, and for great armies of herrings and pteropods and jelly-fishes and siphono- phores, and for most pelagic larvae. There are plant-eating molluscs and echinoderms and annelids in the ocean, but not in sufficient numbers to play any conspicuous part in its economy, and the copepods are the only plant-eaters which exist 144 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. in sufficient numbers to be compared with those of the land, and the food of the copepods is only partially vegetable, for they devour micro- scopic animals as well as microscopic plants, and probably to an equal amount. The group Crustacea as a whole is a carnivorous one, however, for while a few subsist on algas, their number is inconsiderable. Others chew the mud of the bottom and extract its organic matter, but this is chiefly animal and consists of foraminifera and rhizopods and infusoria. The molluscs as a whole are carnivorous, and while there are many exceptions, such as the nudibranchs for example, many nudibranchs feed on hydroids. The cephalopods and pteropods and heteropods and many of the gasteropods pursue and destroy their prey, and other gasteropods are scavengers, while the lamellibranchs gather up the microscopic organ- isms which are drawn into their gills with the water. The majority of the worms and echinoderms are animal-feeders. Some of them, like the common starfish, are actively predaceous; others, like the crinoids, gather up microscopic organisms from the water; others, such as most holothurians, eat the mud of the bottom and digest out of it the foraminifera and small molluscs and annelids and crus- tacea which it contains, while others, such as the sea-urchins of the coral reefs, grind away and swallow the living coral. The universal presence of a poisoning apparatus in the coelenterates shows that the food of this great and important group of marine animals must consist, in the main, of animals which are able to resist or to escape, and observa- tion shows that this is true. Floating jelly-fishes and siphonophores are often found fastened to the half-digested carcasses of sagittas or hetero- pods or fishes larger than their captors, and they consume enormous numbers of copepods, pteropods, young fish, and pelagic larvae of all sorts. So far as we know, all the sea-anemones and coral polyps and alcyonarians and hydroids are carnivorous. Some of the discomedusa3, the rhizostomes, feed upon microscopic organisms, but this mode of life is exceptional, and some recent observations, as yet unpublished, by Dr. R. P. Bigelow, show that the food of the rhizostomes consists of cope- pods. Except for a few plant-eating fishes and molluscs and worms and echinoderms, all the animals of the ocean fall into two classes, those which subsist on microscopic organisms, and those which prey upon each other and correspond to the rapacious animals of the land. W. K. BROOKS ON THE GENUS SALPA. 145 There is practically nothing in the ocean corresponding to the terres- trial herbivora, and nothing like terrestrial vegetation, except the fringe of seaweeds in the shallow water along the coast, and a few floating islands of algae like the Sargasso Sea. While these tracts of vegetation are pretty extensive, they are totally inadequate to support the animal life of the ocean, and as the whole animal world is dependent directly or indirectly upon plants, we must ask what takes the place of terrestrial vegetation. The Fauna of Mid-ocean. There is so much room in the vast spaces of the ocean, and the part which is open to our direct observation is such an inconsiderable part of the whole, that it is only when great multitudes of pelagic animals are gathered together at the surface that the abundance of marine life becomes visible and impressive ; but some faint conception of the bound- less wealth of the ocean may be gained by observing the quickness with which marine animals become crowded at the surface in favorable weather. On a cruise of more than two weeks from Cape Hatteras to the Bahama Islands I was surrounded continually, night and day, by a vast army of dark-brown jelly-fishes (Linerges mercutia), whose dark color made them very conspicuous in the clear water. They were not densely crowded, although they were so abundant that nearly every bucketful of water we dipped up contained some of them. We could see them at a distance from the vessel, and at noon, when the sun was overhead, we could look down into the water to a great depth through a well in the middle of the vessel where the centerboard hung, and as far down as the eye could penetrate, fifty or sixty feet at least, we could see the brown spots drifting by like motes in the sunbeam. We cruised through them for more than five hundred miles, and we tacked back and forth over a breadth of almost a hundred miles, and they were everywhere in equal abundance. The recent literature of pelagic exploration, which has been sum- marized by Haeckel (Plankton Studien : von Ernst Haeckel, Jena, 1890), is full of references to great accumulations of pelagic animals, from which I have selected those which follow. Chiercha says that during a cruise of forty days between Peru and Hawaii the net brought in from the surface and from all depths down to 146 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. about two miles, a multitude of pelagic animals which would be incred- ible to those who have not witnessed it. The naturalists of the Challenger found the waters of the equatorial Pacific swarming with life, not at the surface alone, but in its deeper layers, and the ship often sailed through great banks of pelagic animals. The equatorial Atlantic is like the Pacific, and Chiercha says that its zone of equatorial calms is rich beyond all measure in animal life, and that the water often looks and feels like coagulated jelly. Of the Indian Ocean, Haeckel says that in his voyage to and from Ceylon he was wonderstruck with the wealth of pelagic life day after day on the mirror-like surface. At night it was an unbroken sheet of sparkling light as far as the eye could reach, and the water which was dipped up at random held such a thick swarm of densely crowded luminous animals (Ostracods, Salpa, Pyrosomas, and Medusae) that a printed book could be read distinctly in a dark night by this pelagic light. In temperate and arctic waters there is less diversity, but, as Haeckel shows, there is no evidence of any decrease in individuals, and banks of pteropods (Clio and Limacina), so dense that they seem almost solid, are met even beyond the arctic circle. Haeckel says that in a cruise to the northwest of Scotland he met with such enormous masses of Limacina that each bucket of water which was dipped up contained thousands. The tendency to gather in crowds is not restricted to the smaller pelagic animals, and many species of raptorial fishes are found in densely packed banks. The fishes in a school of mackerel are as numerous as the birds in a flight of wild pigeons. Goode, in his History of Aquatic Animals, tells of one school of mackerel which was estimated to contain a million barrels, and of another which was a windrow of fish half a mile wide and at least twenty miles long ; but while the pigeons are plant-eaters, the mackerel are rapacious hunters, pursuing and devouring the herrings, as well as the pteropods and pelagic Crustacea. Herring swarm like locusts, and a herring bank is almost a solid wall. In 1879 three hundred thousand river herring were landed in a single haul of the seine in Albemarle Sound ; but the herrings are also carniv- orous, each one consuming myriads of copepods every day. In spite of this destruction and the ravages of armies of medusas and siphonophores and pteropods, the fertility of the copepods is so great that they are abundant in all parts of the ocean, and they are met with in numbers which exceed our powers of comprehension. W. K. BROOKS ON THE GENUS SALPA. 147 On one occasion the Challenger steamed for two days through a dense cloud formed of a single species, and they are found in all lati- tudes from the arctic regions to the equator, in masses which discolor the water for miles. We know, too, that they are not restricted to the surface, and that the banks of copepods are sometimes a mile thick. When we reflect that thousands would find ample room and food in a pint of water, we can form some faint conception of their universal abundance. * TJie Primary Food-supply. As the result of our review, we find that the organisms which are visible without a microscope in the water of the ocean and on the sea bottom are almost universally engaged in devouring each other, and many of them, like the blue-fish and the albacore, are never satisfied with slaughter, but kill from mere sport. Insatiable rapacity must end in extermination unless there is some unfailing supply, and as we find no visible supply in the water of the ocean we must seek it with a microscope. By its aid we find a wonder- fully rich and diversified fauna made up of innumerable Iarva3 of all sorts of marine animals, together with a few minute and simple metazoa, but these things cannot form the food-supply of the ocean. It is clear that a single carnivorous animal could not exist very long by devouring its own children, and the result must be the same however great the number of individuals or species. The total amount of these organisms is inconsiderable, however, when compared with the abundance of a few forms of protozoa and protophytes, and both observation and deduction force us to recognize that the most important element in the total amount of marine life consists of some half-a-dozen types of protozoa and unicellular plants, of giobigerina and radiolarians, and of trichodesmium, pyrocystis, pro- tococcus, and the coccospheres, rhabdospheres and diatomes. Modern microscopic research has shown that these simple plants, and the globigerina3 and radiolarians which feed upon them, are so abundant and prolific that they meet all the demands made upon them and supply the food for all the animals of the ocean. This is the fundamental conception of marine biology. The basis of all the life in the modern ocean is to be sought in the micro-organisms of the surface. This is not all. The simplicity and abundance of the microscopic 148 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. forms and their importance in the economy of nature show that the organic world has gradually shaped itself around and has been controlled by them. They are not only the fundamental food-supply, but the primeval supply, which has determined the whole course of the evolution of marine life. The pelagic plant-life of the ocean has retained its primitive sim- plicity on account of the very favorable character of its environment, and the higher rank of the littoral vegetation and that of the land is the result of hardship. On the land the mineral elements of plant-food are slowly supplied as the rains dissolve them ; limited space brings crowding and com- petition for this scanty supply; growth is arrested for a great part of each year by drought or cold; the diversity of the earth's surface demands diversity of structure and habit, and the great size and compli- cated structure of terrestrial plants are adaptations to these conditions of hardship. The conditions of the surface of the ocean ; the abundance and uniform distribution of mineral food in solution ; the area which is available for plants ; the volume of sunlight and the uniformity of the temperature are all favorable to the growth of plants, and as each plant is bathed on all sides by a nutritive fluid, it is advantageous for the new plant-cells which are formed by cell multiplication to separate from each other as soon as possible in order to expose the whole of their surface to the water. Cell aggregation, the first step towards higher evolution, is therefore disadvantageous to the pelagic plants, and as the environment is so homogeneous at the surface of the ocean that there is little oppor- tunity for an aggregation of cells to gain a compensating advantage by seizing upon a more favorable habitat, the pelagic plants have retained their primitive simplicity. The list of pelagic micro-organisms is a long one, but a few forms are so predominant that the others have little significance at the present day in comparison, and we may regard the great primary food-supply as made up of two simple protozoa, Globigerina and the Radiolarians, and some five or six unicellular plants. Of these only two, the Radiolarians and the Diatomes, show any great diversity of species, and while the Radiolarians are so diversified that the Challenger collection alone furnished more than four thousand species, this variety does not obscure the primitive simplicity of the type, W. K. BROOKS ON THE GENUS SALPA. 149 and the most distinctive peculiarity of the microscopic food-supply of the ocean is the very small number of the forms which go to make up the enormous mass of individuals. The Origin of Pelagic Animals. All the animals of the ocean are dependent upon the microscopic food-supply, and many of them are adapted for preying upon it directly. Among these Salpa is one of the most conspicuous examples. It passes its whole life in the open water, and it has no sessile stage in its onto- geny, as many floating animals have. It abounds in all parts of the ocean, and over some great seas it is always present at the surface. As the result of three years' observation, Schminkewitch says that the Salpas are perennial pelagic animals, and Chun has shown that they are also found in abundance at great depths. As long as it is alive and breathing a steady stream of micro- organisms is slipping along its pharynx and down through its oesophagus into its stomach, and sections of the intestine of Salpa afford most beau- tiful preparations of radiolarians and diatomes. The pelagic food-supply is very ancient, and we have, in Salpa, an animal which has been especially evolved to pass its life swimming through the living broth of the mid-ocean. If we were to select the typical pelagic animal we should probably choose Salpa, and it is therefore most surprising to find that Salpa itself has not been produced at the surface of the ocean by gradual evolution from a simple pelagic ancestor. The structure which fits it so well for its mode of life has come to it by the inheritance of peculiarities which were originally acquired by bottom animals in adaptation to the needs of a sessile life. This is all the more remarkable since both Salpa and its fixed allies show by their embryology that still more remotely they are descended from a pelagic form like Appendicularia. The place in the pelagic world which Salpa fills so well has been ready for it from primeval times. Why then has not the simple pelagic Appendicularia given rise, in the open sea, to series of more and more perfected pelagic descendants culminating in Salpa ? Why should the descendants of a pelagic ancestor have passed through a sessile stage before they acquired their improved pelagic structure ? 150 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. If this were a solitary case it would not deserve notice ; but exam- ination will show that no highly organized animal has ever been evolved at the surface, although all depend on the pelagic food-supply. The animals which now find their home in the open waters of the ocean are, almost without exception, the descendants of forms which live upon or near the bottom or along the sea-shore or upon the land, and the exceptions are all simple animals of minute size. The metazoa which are primitively pelagic, that is, those which have been pelagic throughout their whole history and do not owe their structure to competition with improved forms from the bottom or the shore, are astonishingly few, and these few are among the smallest and simplest of the metazoa. It is only necessary to review the chief groups of metazoa in order to perceive that most of their pelagic representatives exhibit the clearest evidence of descent from forms which lived upon or near the bottom or the shore. Many indeed have no pelagic members, but are restricted to the bottom. The sponges are obviously a bottom group ; most of them are fixed, all are sedentary, and their whole organization is an adaptation for life in the bottom. The coral polyps, actinias and alcy on arias, are among the most characteristic bottom forms, and the abundance of the fossil remains of polyp skeletons proves that these animals became established on the bottom very early, and that the whole history of their evolution has taken place at the bottom. The acraspedote medusae are universally and justly regarded as the descendants of fixed polyp-like ancestors, and we may state with confidence that they are not primitively pelagic, but that a fixed period in their history has come between the modern swimming jelly-fish and its remote and unknown primitive pelagic ancestor. The veiled medusae are usually held to have had a similar history, but I shall soon give my reasons for holding that some of these at least are primitively pelagic. There can, however, be no doubt that the evolution of hydroid cormi has taken place at the bottom. The siphon- ophores are descended from ancestors like the anthomedusae, and the various families and genera and species of siphonophores have most certainly been produced by divergent specialization among pelagic forms, and the greater part of their history, if not the whole of it, is therefore pelagic. The echinoderms are most characteristic inhabitants of the bottom, as they have been from palaeozoic times, and while synapta is sometimes W. K. BROOKS ON THE GENUS SALPA. 151 found at the surface of the open ocean, this is exceptional, and we may state without hesitation that the evolution of the echinoderms has taken place at the bottom. This is equally true of the brachiopods and of most of the animals classed as vermes, the gephyreans, bryozoa, nemertians, and so forth. The pelagic annelids, such as Tomopteris, are secondary modifications of bottom forms, and while some of the more primitive annelids may possibly be originally pelagic, the group as a whole is as characteristic of the bottom as the echinoderms. Many groups of Crustacea have pelagic representatives, and the pelagic crustacean fauna is rich and varied, but in most cases the pelagic forms show unmistakable evidence of secondary change of habit, and all the higher Crustacea have been evolved at the bottom in adapta- tion to a bottom life. I shall soon give my reasons for believing that there is one important exception to this rule, however, and I shall try to show that there is good ground for holding that the copepods are primitively pelagic, and that while the greater part of the history of the Crustacea is bottom history, the characteristics of the crustacean type were outlined in pelagic animals at a very early period in the history of the metazoa. The heavy calcareous shells of the mollusca could not have been acquired at the surface, and that most characteristic molluscan organ, the lingual ribbon, is adapted for attacking more solid bodies than the delicate primitive pelagic animals. The classes and orders of mollusca must have been evolved at the bottom, and there is ample evidence that the swimming shelless gasteropods and cephalopods have, like those great pelagic groups the pteropods and heteropods, been secondarily adapted for a pelagic life. Many of the marine fishes are strictly pelagic, and the structure and habits of fishes are in all respects so well fitted for a wandering life in the open water that the pelagic habit of fishes seems at first sight to be their most distinctive peculiarity, although a little examination will show that there is ample evidence that it is secondary, and not primitive. The perfection of their adjustment to a free life in the open sea is no evidence that this life is primitive, for the highest marine animals and those whose adaptation to a pelagic life is most complete, the sea-birds and cetacea and marine reptiles, are air-breathing terrestrial animals which have gone back into the ocean. The most primitive groups of living fishes are the cyclostomes, elas- mobranchs and ganoids. The cyclostomes are too small a group, and 152 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. the living forms are too aberrant in habit, to contribute much informa- tion regarding the nature of the primitive vertebrates, but they exhibit no evidence of adaptation to a pelagic life, and our scanty knowledge of them is quite in harmony with the view that their remote ancestors were bottom animals. The case is very different as regards the great groups of modern fishes for which the term palaeichthyes is often used; the sharks, rays and ganoids. The living representatives of these great and ancient groups are of peculiar interest to naturalists on account of their close affinity to the oldest vertebrate fossils which have been discovered. These points of resemblance to the more modern, but still ancient, amphibia and teleosts show that the modern palaeichthyes have preserved their ancient struc- ture with very slight modification, and that we have in them one of the most remarkable stem forms in the whole animal kingdom. This is shown still more conclusively by the fact that some of the palaeozoic families of elasmobranchs have lived through period after period of geological history and have held their ground up to our own times. The abundance and variety of the remains of elasmobranchs in the palaeozoic rocks prove the great development of the group at this remote and early period, and the silurian sharks probably differed but little from those of the present day, although we are forced to see in them the ancestors of the ganoids and of all the divergent groups of extinct and living vertebrates. Of the three groups of modern elasmobranchs, two, the chimaeras and the rays, are bottom-feeders. The whole organization of the ray is as obviously adapted for life upon or near the bottom as that of a bird is for life in the air, and the flat pavement teeth are adapted for crush- ing and grinding the hard-shelled molluscs and Crustacea and echino- derms of the bottom. It is true that the sawfish is not confined to the bottom, and the devil-fishes often capture their prey at the surface. In the West Indies they are often found very far from land, but these cases are exceptional, and the true rays rarely leave the bottom, nor are they adapted for rapid movement through the water. The rays are undoubtedly much more modern than the true sharks, but there is ample evidence that they have retained habits of life which are common to all the primitive elasmobranchs. Many of the modern sharks live on or near the bottom, where they W. K. BROOKS ON THE GENUS SALPA. 153 are found in immense numbers and at considerable depths. In 1888 I was invited by Marshall McDonald, the Superintendent of the IT. S. Fish Commission, to make use of the opportunity for surface collecting which was afforded by an expedition which was sent out to fish with hook and line on the bottom and along the edge of the Gulf Stream. The fishing commenced at the 500 fathom line, and every time the line was taken in we found numbers of dogfish (Scyllium) on the hooks, even when the water was considerably more than half a mile deep. Many genera of sharks, such as the houndfish (Mustelus) and the dogfish (Scyllium), are known to feed upon the molluscs and Crustacea and worms of the bottom, and the flat pavement-teeth of other genera whose habits are less known show that their mode of life is the same. Some of the bottom-feeding sharks (Cestracion for example) are the oldest of living vertebrates. The mailed ganoids were undoubtedly derived from a shark-like ancestor, and the structure of the oldest ones, such as perichthys, cocco- steus and cephalaspis, shows that they were not very rapid swimmers. They were, undoubtedly, bottom-feeders like the modern sturgeon, and like many large and important families of modern teleosts, such as the cod, the siluroids and the pleuronectidae. So far as we know the palaeozoic waters from fossils, there were no active locomotor animals of large size to furnish prey for raptorial fishes, and the existence at the present day of so many species and genera and families of bottom-feeders, and the fact that the most archaic forms have this habit, are all grounds for believing that the fishes are secondarily adapted to a pelagic life, like the sea-birds and the cetacea. So far as amphioxus furnishes evidence, this bears in the same direction, for its home is in the sand of the bottom. In fact it may almost be called a subterranean animal, for when it is placed in an aquarium it sinks into the sand at the bottom and disappears at once, and it makes its way through the sand with great ease and rapidity. All the evidence shows that the primitive vertebrates lived upon or near the bottom, and that the early steps in the evolution of the classes of vertebrated animals were made at the bottom. As the result of this review we see that the evidence from palaeon- tology, from embryology, and from the structure and habits of living- animals all bears in the same direction, and shows that there are no large or highly organized animals which have been pelagic through all the stages of their evolution, and that, in this particular, the life-history of Salpa is not exceptional, but typical. 154 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. In its descent from an inhabitant of the bottom and in its secondary adaptation to a pelagic life, its history resembles that of all the highly organized pelagic animals. Embryology also gives us good ground for believing that Salpa follows the analogy of all the metazoa in its still more remote descent from a small and simple pelagic ancestor, and there is good ground for believing that the earliest metazoa were all pelagic, and that they were represented at a very early period in the history of life by floating or swimming animals of minute size and simple structure. We may see in the free larval forms of many marine metazoa, such as the tornaria of balanoglossus, the swimming echinoderm larva, the ascidian tadpole, the floating ciliated larvae of annelids, brachiopods and molluscs, in the coelenterate planula, and, as I believe, in the crustacean nauplius, traces of this primitive mode of life ; often obscured or complicated by more recent adaptation and sometimes almost obliterated by secondary changes. When this fact is seen in all its bearings and its full significance is grasped, it is certainly one of the most noteworthy and instructive features of the history of evolution. The food-supply of the- ocean consists of a few species of unicellular microscopic plants, and of a few simple protozoa which feed upon them. This supply is inexhaustible, and it is the only source of food for all the inhabitants of the ocean, except a few which live upon floating sargas- sum and the -littoral algae, and the drainage from the land. Many marine animals are adapted for direct subsistence upon these organisms, and some' of them, like Salpa, are universally distributed and are found in enormous numbers in all parts of the ocean. The food-supply is not only inexhaustible, it is also primeval, and all the life of the ocean has gradually taken shape in direct dependence upon it during the history of its evolution. In view of all these facts we cannot but be profoundly impressed by the thought that all the highly organized marine animals are products 'of the bottom, or of the shore, or of the land, and that while the largest animals on earth are pelagic, the few which are primitively pelagic are very small and very simple. The reason is obvious. The conditions of pelagic life are so easy that there is no fierce competition, and the inorganic environment is so simple that there is little chance for diversity of habits. The growth of terrestrial plants is limited by the scarcity of food, W. K. BROOKS ON THE GENUS SALPA. 155 but there is no such limit to the growth of pelagic plants or the animals which feed upon them, and while the balance of life is undoubtedly adjusted, competition for food is never very fierce even at the present day, when the ocean swarms with highly organized animals which have become secondarily adapted for a pelagic life. Even now the destruction or escape of a microscopic pelagic organism depends upon the accidental proximity or remoteness of an enemy rather than upon defense or pro- tection, and survival is determined by space relations rather than by a struggle for existence. The abundance of food is shown by the ease with which wanderers from the land, like birds, find places for themselves in the ocean, and the rapidity with which they spread over its whole extent. As a marine animal, the insect, halobates, must be very modern as compared with most pelagic forms, yet it has spread over all tropical and subtropical seas, and it may always be found skimming over the surface of the water as much at home as a gerris in a pond. I never found it absent in the Gulf Stream when conditions were favorable for collecting. The easy character of pelagic life is also shown by the fact that the larvas of innumerable animals from the bottom and the shore have retained their pelagic habit, and I shall soon refer to facts which prove that the larva of a shore animal is safer at sea than it is near the shore. The absence of fierce competition in the open ocean is well shown by the simultaneous existence in the modern ocean of graded stages in the evolution of a type, such as the series of Pelagidae; and also by the persistency of a stem form like the elasmobranch, side by side with, and often in competition with, various improved lines of divergent descendants. In the primitive pelagic fauna and flora there was little opportunity for an organism to gain superiority by seizing upon an advantageous site or by acquiring peculiar habits, for one place was like another, and peculiar habits could count for little in comparison with accidental space relations. After the pelagic fauna had been enriched by the addition of all the marine animals which are secondarily pelagic, competition with these improved forms from the bottom brought about improvements in those which were strictly pelagic in their origin, and through this competition, complicated animals of considerable size, like the siphonophores, have been evolved at the surface, but while their whole history has thus been pelagic they are not primitively pelagic ; that is, they are not the out- 156 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. come of purely pelagic influence. The wanderers from the bottom have introduced another factor in the evolution of pelagic life, for their bodies have been utilized for purposes of protection or concealment or on account of other advantages, and we now have fishes which shelter themselves in the poisoned curtain of physalia ; Crustacea which live in the pharynx of salpa; barnacles and sucking fishes fastened to whales and turtles, besides a host of external and internal parasites. The primi- tive ocean furnished no such opportunity, and the conditions of pelagic life must, at first, have been extremely simple. Among the higher metazoa and the higher plants size is, in itself, an important factor in evolution. Variations in the constituent cells of a large organism are continually being seized upon and fixed by natural selection, on account of their value in the functions of relation to other parts. Primitive pelagic organisms are all minute, and it is easy to understand why. To plants which are bathed on all sides by food, like the pelagic protophytes, small size is advantageous, since a small body has a larger surface in proportion to its bulk than a large one ; and the pelagic plants are, as I have shown, most favorably placed for rapid growth when new cells separate as soon as they are formed, and thus expose all their surface. The same ratio between bulk and nutritive surface tends to limit in the same way, if not to the same degree, the growth of the pelagic animals which live in the midst of an abundant supply of vegetable food. Competition was not entirely absent among the primitive pelagic organisms, for the conditions of life are never absolutely uniform, although the possibilities of evolution must have been extremely limited and the progress of divergent modification very slow, so long as life was restricted to the waters of the ocean. There can be no doubt that pelagic Ife was abundant for a long period during which the bottom was uninhabited. The history of the slow process of geological change by which the earth gradually assumed its present character, presents a boundless field for speculation, but there can be no doubt that the surface of the primeval ocean became fit for life long before the deeper waters or the sea-floor. The early steps in the evolution of plants must have been taken in the transparent surface water under the influence of sunlight, and as both animals and plants are dependent upon oxygen, the primal flora and fauna must have lived in aerated water. The oxygen which is W. K. BROOKS ON THE GENUS SALPA. 157 diffused through the ocean from the surface, where it is absorbed from the air, is gradually exhausted by oxidizable substance, both inorganic and organic, and it diminishes with the distance from the source of supply at the surface. The oceanic circulation tends to equalize its dis- tribution, and no part of the ocean now seems to be totally without oxygen. Oxygen has been shown to be reduced to a minimum at the bottom of some of the great depressions of the sea-floor, and it is clear that a slight change in the conditions which influence it might render the sea-bottom unfit for life. In early palaeozoic times the sea-floor was perhaps more level than it is now, and there may have been no deep hollows like those in which the oxygen is now found to be deficient, but the average depth must have been considerably greater, when all the water which is now locked up in the sedimentary rocks of the bottom and of the shores was still free in the ocean. The circulation may also have been less active when geo- graphical conditions were more simple, and the air was undoubtedly less rich in oxygen in early palaeozoic times than it is at present. It is therefore easy to understand that long after the crust of the earth had acquired essentially its present character, there may have been a period when the supply of oxygen was so scanty that the activities of pelagic organisms and the products of their decomposition used it up in the surface water, so that life on the bottom was impossible at a time when the superficial water supported a luxuriant fauna and flora. During this period the proper conditions for the production of large and complicated organisms did not exist, and while the total volume of life was probably very great, it consisted of the organisms of minute size and simple structure which I have termed the primitive pelagic fauna and flora. The Primitive Pelagic Fauna. In using this term I do not, of course, intend to imply that these organisms are the beginning of life, or to express any opinion as to the way in which life first came into existence. I use it merely as a con- venient designation for the total sum of the organisms which have been evolved by purely pelagic influences from a starting-point which is absolutely unknown at present. The attempt to reconstruct in imagination the primitive pelagic fauna and flora is most fascinating, but all the available evidence is indirect, and as we can have little hope of finding any record of it in the rocks, we must trust to deduction rather than observation. 158 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. The modern pelagic protophytes have probably retained nearly their ancient form, but the modern radiolarians and pelagic foraminifera exhibit indications of secondary adaptation, and they have undoubtedly been modified by competition with improved organisms from the bottom. All the metazoa have pelagic larvae, or else larval or embryonic stages, which must be regarded as the degenerated vestiges of a pelagic habit ; but in most cases these larvae have been so much changed by the accelerated development of adult features, or by the acquisition of habits or structures to fit them for the conditions of modern pelagic life, that we can deduce little more from them than the former existence of pelagic ancestors. When a pelagic larva is still represented by a modern pelagic adult of minute size and simple structure, as the tadpole larva of ascidians is represented by Appendicularia, we may be confident that it is a pelagic production, and that it existed in the primitive pelagic fauna, although this view is directly opposed to accepted dogmas regarding the origin of the Chordata. When all the members of a great group have a definite pelagic larval stage which adheres to the same plan of structure in all of them, we may be pretty confident that this larva is the repre- sentative of a primitive pelagic adult animal, even if this ancestor has now no unmodified descendants. To my mind the best example of the retention, by all the members of a great group, of a larval stage which represents an extinct primitively pelagic ancestor is to be found in the Crustacea nauplius, and notwith- standing the popular verdict against it, I do not hesitate to regard the nauplius as a pure pelagic product, and to include it in the primitive pelagic fauna, although I shall discuss this question further on. In cases like that of the echinoderms, where the pelagic larvae of the various classes and orders are very different from each other in the details of their organization, we are hardly safe in assuming more than the primi- tive existence of an unknown pelagic organism, from which they have been derived. This is true to even a greater degree of the trochic larvae of annelids, molluscs, etc., but while there is little ground for regarding the forms of these modern larvae as ancestral, we must regard their pelagic habit as an inheritance from unknown ancestors in the primitive pelagic fauna, in which we must therefore include representatives of such larvae as the molluscan veliger, the nemertian pilidium, the acti- notrocha of phoronis, the brachiopod larva, the coelenterate planula, and so forth, although we are quite unable to say how many independent starting-points these various metozoan lines had in the primitive pelagic W. K. BROOKS ON THE GENUS SALPA. 159 fauna, or what these starting-points were like. Our inability to describe or picture these ancestral forms is no reason for doubting their reality, for in biology the weight and certainty of a deduction are often indepen- dent of its definiteness. We may, for example, feel sure that the cetacea are descended from terrestrial animals and yet find it impossible to picture their ancestor, or even to decide whether their ancestral lines converge into one stem before or after the pelagic habit was acquired. We may in the same way feel sure, even in the absence of sufficient evidence to trace their direct paths, that all the great groups of metazoa ran back to minute pelagic ancestors, and we must, therefore, include in the primitive pelagic fauna a great, but indefinite, number of distinct and somewhat widely separated ancestral forms, and together with them, no doubt, an equal or greater number of somewhat similar forms which have been exterminated and have left no descendants. In these extinct forms we should, if we could study them, find the connecting links between divergent groups, and we would thus be able to complete the genealogical tree of the metazoa by bringing together the great divergent branches of the metazoan stem whose primary relationships now seem beyond discovery. In addition to the primitive pelagic animals which are known to us only by the traces of their former existence which they have left in the structure and habits of. modern larvae and embryos, there are a few modern pelagic adult animals which show by their minute size and simple structure and by their systematic affinities that they are primi- tive pelagic animals, owing their structure to purely pelagic influences. Appendicularia is a good example of this class, and I believe that the copepods are the most important group of the primitively pelagic metazoa. The Origin of the Crustacea. The view that the copepods are degenerated descendants from Crustacea like the phyllopods, and that the Crustacea were evolved on the bottom, and that the pelagic habit of the copepods is secondary, is so generally accepted that it is hardly worth while to advance a different view in this place where there is no room for its exhaustive treatment. The consideration which seems to have the greatest weight with morphologists is the supposed necessity of a phylogenetic explanation of metamerism, but a little reflection will show the persistent existence of an influence which tends to metamerism at the present day. 160 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. For this influence, which is shown by such phenomena as the inheritance by the child of polydactylous feet from the polydactylous hands of the parent, or the development of wing-feathers on the toes of fantail pigeons; the influence which has carried the feet of the horse family along the same line of evolution with the hands, I have, in another place, proposed the term ontogenetic inheritance. Among the arthropods, examples of this sort of modern metamerization are very common, both as normal features of their structure, in the movable body-rings of the ocular and antennary somites of stomatopods, for example, and as monstrosities, as in the twelve-legged coleoptera. I believe that a thorough study of this most interesting and instruc- tive class of facts will convince any one that there is no philosophical necessity for assuming that the primitive crustacean had a highly meta- merized body like that'of a phyllopod, and that all the common features in the structure of arthropods may have been derived from a common ancestor as simple as a nauplius. The analogy between the parapodia of annelids and the limbs of Crustacea has been held to prove that the primitive crustacean limb was not a rowing organ fitted for a pelagic life, like the limbs of the nauplius and the copepod, but flat and leaf -like and adapted for movement over the bottom. It is hardly possible, however, to believe that the arthropods have been derived from the higher polychaetous annelids, and as the simpler and more primitive annelids have no parapodia, the resemblance, which is not actually very noteworthy, can be nothing more than an analogy. There are plenty of degenerated copepods, and we have in their structure abundant proof of the degeneracy, and an adequate explanation of it in their parasitic habits, but they are degenerated descendants of ordinary swimming copepods, and not of phyllopods, and there is no reason for holding that the copepodan type itself is degenerate, except the supposed exigencies of morphological philosophy. The active locomotor habits of the eucopepods of the open ocean would seem to be conducive to advancement rather than to degenera- tion, and the occurrence of phyllopods in the lower Cambrian is, of course, no more evidence that they are primitive Crustacea than the occurrence of pteropods and gasteropods is that they are primitive molluscs. I am unable to see any valid objection to the view that the copepods are primitively pelagic ; that they have been evolved at the surface of W. K. BROOKS ON THE GENUS SALPA. 161 the ocean from pelagic nauplii, and that the great group Crustacea has been derived from them. We have already seen that the eucopepods are the chief intermediary between the micro-organisms of the ocean and the larger and higher marine animals ; that they prey upon the protophytes and protozoa, and in their turn supply either directly or indirectly most of the food for the large inhabitants of the water ; that most pelagic larva? feed upon them ; that they are the food of the great pelagic banks of pteropods and hetero- pods, of many coelenterates, of the young of most fishes, and of some of the most abundant and important adult fishes, like the herring, and that the sea-birds, the cetacea, and in fact almost all of the larger pelagic animals, prey upon animals which in their turn prey upon copepods. The animals which are most important at one period in the earth's history are often replaced by others at another period, and it is, of course, possible that the modern copepods now fill a place which was in former times filled by something else ; but as their organization, as com- pared with that of the other Crustacea, exhibits all the characteristics of a primitive pelagic stem-form, and inasmuch as the remains of animals, like the pteropods, which now live almost entirely upon copepods, are found in the oldest fossiliferous rocks, there is every reason to believe that the group formed an important constituent of the primitive pelagic fauna. No one who advocates at our time the morphological heresies which are involved in the view that appendicularia is a stem-form which is pelagic in its origin ; that the nauplius is a persistent representative of the primitive Crustacea, and that the whole history of the copepods has been pelagic ; and that the veiled medusa? have been evolved in direct relation to pelagic influences ; no one who makes these statements can hope to escape the charge that his view "ist die unwahrscheinlichste von unwahrscheinlichkeiten." The books all tell us that the free active appendicularia is the "degenerated" descendant of an ancestor which crept over the bottom; that the nauplius is a secondary larval form ; that the active free-swim- ming copepods are degenerated phyllopods; and that the locomotor hydro-medusa is, in its origin, a specialized member of a sessile, poly- morphic, hydroid cormus. The first of these opinions, that appendicularia is a degenerated form, rests upon a supposed necessity for deriving the body of a verte- brate, which consists of a series of segments homologous with each other, 162 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. 9 but highly differentiated among themselves, from an ancestral aggrega- tion of similar, but less differentiated, segments. The second opinion, that the copepods are degenerated and that the nauplius is a secondary larval form, is the result of a supposed necessity for explaining the segmentation of the arthropods in the same way, while the third view has its origin in the belief that the polymorphic members of a hydroid cormus must have arisen through specialization and division of labor from an ancestral undifferentiated aggregation. These are a few, from among many, illustrations of the general acceptance among morphologists of a dogma which, while it is often refined and qualified until its character is almost lost, may be broadly stated as a belief that the homology between different parts of the same organism is always to be explained, like the homology between corres- ponding parts of different animals, as the result of phylogenetic inherit- ance ; or, to state it in a different way, that the vegetative duplication of parts in animals has a phylogenetic significance, and implies descent from a duplicated ancestor. The dogma is not the dictum of any one teacher, and it has grown almost imperceptibly from its starting-point in the discovery that the body of a metazoon is an aggregation of cells, each with an individuality of its own, specialized and differentiated by polymorphism and division of labor, and each one homologous with an unicellular organism. The dogma has been a most useful and suggestive working hypoth- esis when well controlled, but when uncontrolled it has led to the most fantastic and grotesque unscientific speculation. The climax of incon- sistency into which its blind adherents have been led was well shown by the simultaneous appearance, in a recent morphological journal, of two memoirs, one an essay on "The Origin of the Vertebrates from the Arachnids," and the other on " The Origin of Vertebrates from a Crus- tacean-like Ancestor." After my first examination of the second of these memoirs I laid it down, much distressed in mind by the thought that this author had unkindly descended from the sphere of experimental research in physi- ology, to expose the unscientific methods of the morphologists by a severe and well merited, if somewhat ponderous, satire. In my next chapter on the morphological significance of appendi- cularia I shall try to show that there is no philosophical necessity for a phylogenetic explanation of duplicated structures in animals, whether they are radial, bilateral, metameric or indefinite, and I must refer the W. K. BEOOKS ON THE GENUS SALPA. 163 reader to that chapter for my reasons for including appendicularia, the copepods and the veiled medusae among the primitive pelagic animals. The Phytogeny of the Metazoa. The primitive pelagic fauna, before the influence of the bottom and of the shore had been brought to bear upon it, consisted of small animals of simple structure; but we are forced, by the facts of comparative anatomy and embryology, to believe that a number of distinct types of structure were found among them. Most of the great metazoic stems show by their embryology that they run back to simple and minute pelagic ancestors, and that their common meeting-point must be projected back to a still more remote time, before the differentiation of their pelagic ancestors had been effected. After we have traced each great line of metazoa as far back as we can from the study of fossils and by the aid of comparative mor- phology, we still find these lines distinctly laid down. The lower Cam- brian Crustacea, for example, are as distinct from the lower Cambrian echinoderms or pteropods or brachiopods or lamellibranchs, as they are from those of the present day. The efforts of anatomists and embry- ologists to reconstruct the primary phylogeny of the metazoa have so far yielded few trustworthy results, and the results which are most trustworthy are usually those which are the most indefinite. We are therefore forced to believe that the early steps in the estab- lishment of the various types of metazoa were taken under conditions which had some essential difference from those which have prevailed, without any fundamental changes, from the time of the oldest fossil to the present day ; and we are also forced to believe that most of the great lines of descent were represented at some time in the remote past by ancestors which, living a different sort of life, differed essentially in structure as well as habits, from the representatives of the same types which are known to us. Furthermore, embryology teaches that each great group still bears internal evidence of descent from pelagic ances- tors, and while the characteristics of these ancestors are in most cases unknown, a few, like appendicularia, are still found alive. Our knowledge of the evolution of the metazoic types has certain general features which are essentially the same for all, but each group has also in its history much that is individual, and any general state- ment requires so much qualification that the history of an illustrative group is more instructive than a general summary. 164 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. In the echinoderms we have a well-defined type represented by abundant fossils, very rich in living forms, very diversified in its modifi- cations, and therefore well fitted for use as an illustration. This great stem contains many classes and orders, all constructed on the same plan, which is sharply isolated and quite unlike the plan of structure in any other group of animals. All through the series of fossiliferous rocks echinoderms are found, and the plan of structure is always the same. Palaeontology gives us most valuable evidence regard- ing the course of evolution within the limits of a class as in the cri- noids and in the echinoids; but we appeal to it in vain for light upon the organization of the primitive echinoderm, or for connecting links between the classes. To our questions on these subjects and on the rela- tion of the echinoderms to other animals, paleontology is silent, and throws them back upon us as unsolved riddles. The morphologist unhesitatingly projects his imagination, held in check only by the laws of scientific thought, into the dark period before the times of the oldest fossils, and feels absolutely certain of the past existence of a stem form, from which the classes of echinoderms have inherited the fundamental plan of their structure, and he affirms with equal confidence that the structural changes which have separated this ancient type from the classes which we know were very much more profound and extensive than all the changes which each class has under- gone from the earliest paleozoic times to the present day. He is also disposed to assume, but, as I shall show, with much less reason, that the amount of change which structure has undergone is an index to the length of time which the change has required, and that the period which is covered by the fossiliferous rocks is only an inconsid- erable part of that which has been consumed in the evolution of the echinoderms. The morphologist does not check the flight of his scientific imagina- tion here, however, for he trusts implicitly to the embryological evidence which teaches him that, still further back in the past, all the echinoderms were represented by a minute pelagic animal which was not an echino- derm at all in any sense except the ancestral one, although it was distinguished by features which natural selection has converted, under the influence of more modern conditions, into the structure of echino- derms. He finds, in the embryology of modern echinoderms, phenomena which can bear no interpretation but this, and he unhesitatingly assumes that they are an inheritance which has been handed down from genera- W. K. BROOKS ON THE GENUS SALPA. 165 tion to generation through all the ages from the prehistoric times of zoology. Other groups tell the same story with equal clearness. Who can look at a living lingula without being overwhelmed by the effort to grasp its immeasurable antiquity, and by the thought that, while it has passed through all the chances and changes of geological history, the structure which fitted it for life on the earliest palaeozoic bottom is still adapted for a life in the sands of the modern sea-floor ? The everlasting hills are the type of venerable antiquity; but lin- gula has seen the continents grow up, and has maintained its integrity unmoved by the convulsions of nature which have given to the crust of the earth its present form. As measured by the time-standards of the morphologist, lingula itself is modern, for its life-history still holds, locked up within it, the record of a structure and of a habit of life which were lost in the unknown past at the time of the lower Cambrian, and it tells us, vaguely but unmistakably, of a life at the surface of the primitive ocean at a time when the brachiopod stem was represented by minute and simple pelagic animals. Broadly stated, the history of each great line of metazoa has been like that of the echinoderms or brachiopods, for while the brachiopods are certainly much more closely related to the polyzoa or the gephyreans than to the echinoderms, and while these latter are nearer to the chordata than to the brachiopods, yet each great line stands sharply by itself. The oldest pteropod or lamellibranch or crustacean or echinoderm or vertebrate which we know from fossils exhibits its own type of structure with perfect distinctness, and later influences have done no more than to expand and diversify the type, while anatomy fails to guide us back to the point where these various lines met each other in a common source, although it forces us to believe that this common source once had an individual existence. Embryology teaches that each line once had its own pelagic repre- sentatives, and that the early stages in the evolution of each type have passed away and left no record. The palaeontological side of the subject has recently been ably summed up by Walcott in an interesting memoir on the oldest fauna which is known to us from fossils (The Fauna of the Lower Cambrian or Olenellus Zone, by Charles Doolittle Walcott, U. S. Geological Survey, 10th Annual Report, Washington, 1890). 166 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. The fossils of the lower Cambrian are not absolutely the oldest known, but it is the oldest fauna which is represented with sufficient completeness for a general view, and is, therefore, interesting to biologists. Walcott says that no plants are known in the rocks of the lower Cambrian, and that he has satisfied himself, after a study of all the reputed species of algae, that they are not plants, but the trails of worms or molluscs. The number of species is small, but their diversity is most note- worthy and remarkable. Walcott' s collection of 141 American species from the lower Cam- brian is distributed over most of the marine groups of the animal kingdom, and, except for the absence of the remains of vertebrates, the whole province of animal life is almost as completely covered by these 141 species as it could be by a collection from the bottom of the modern ocean. Four of the American species are sponges, two are hydrozoa, nine are actinozoa, one an echinoderm, twenty-nine are brachiopods, three are lamellibranchs, thirteen are gasteropods, fifteen are pteropods, eight are Crustacea, fifty-one are trilobites, and the trails and burrows show the existence of at least six species of bottom forms, probably worms or Crustacea. The most noteworthy characteristic is the completeness with which these new species outline the whole fauna of the modern sea-floor. Nothing brings home more vividly to the zoologist a picture of the diversity of the lower Cambrian fauna and of its intimate relation to the bottom fauna of to-day than the thought that he would have found, on the old Cambrian shore, about the same opportunity to study the embry- ology and anatomy of pteropods, gasteropods and lamellibranchs and Crustacea and medusa3 that he now has at a marine laboratory, and that his studies in phylogeny would have had about the same form then that they have now. Biological evidence based on embryology and anatomy and on the habits and affinities of animals is justly regarded, by zoologists at least, as a more perfect record of the early history of life than palaeontology, and we accept, without question, proofs of phylogeny which refer to a time very much more remote than the age of the oldest fossils. We must not forget, however, that our generalizations in primitive phylogeny rest for the most part on the study of swimming or floating larvae of minute size and simple structure, which we can have little hope of finding as fossils. W. K. BROOKS ON THE GENUS SALPA. 167 In the formations which follow the lower Cambrian, species grad- ually become more numerous, but this is due to divergent specialization, and Walcott says that if a comparison be made between the Olenellus Zone (lower Cambrian) and the Silurian fauna, the superiority of the latter in number of species, genera and families is at once apparent. "If the comparison be extended to class characters, the disparity between the two is very much reduced, and it is made evident that the evolution of life between the epoch of the Olenellus fauna and the epoch of the Ordvician fauna has been, with one or two exceptions, in the direction of differentiating the class types that existed in the earlier fauna." The ground which we have covered in our review of these various broad aspects of the animal kingdom brings us, then, to the following point of view : There are no highly organized animals which have been pelagic through all the stages of their evolution. The metazoa, which have been pelagic through their whole history, are either small and simply organ- ized, as compared with the higher representatives of the group to which they belong, like appendicularia, or else, like the siphonophores, they have been perfected through competition with higher types. Marine life is older than terrestrial life, and as all marine life has shaped itself in relation to the pelagic food-supply, this itself is the only form of life which is independent, and it must therefore be the oldest. There must have been a long period in primeval times during which- there was a pelagic flora and fauna, rich beyond limit in individuals, but made up of only a few small simple types. During this time the pelagic ancestors of all the great groups of metazoa were slowly evolved, as well as others which have no living descendants. So long as life was restricted to the surface, no great or rapid advancement through the influences which now modify species was possible, and we know of no other influence which might have replaced these. We are, therefore, forced to believe that the differentiation and improvement of the primi- tive flora and fauna was slow, and that for a vast period of time life consisted of an innumerable multitude of pelagic organisms made up of a few forms. During the time which it took to form the thick beds of older sedimentary rocks the physical conditions of the ocean gradually took their present form, and during a part, at least, of this period, the total amount of life in the ocean may have been about as great as it is now without leaving any permanent record of its existence, for no rapid 168 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. advancement took place until the advantages of a life on the bottom were discovered. The Discovery of the Bottom, and its Effect on Evolution. We must not think of the populating of the bottom as a physical problem, but as colonization, very much like the colonization of oceanic islands. Physical conditions for a long time made it impossible, but its initiation was the result of biological influences, and there is no reason why the starting-point should be the point where the physical obstacles were first removed. It is useless to speculate upon the character of the physical obstacles ; there is reason to believe that one of them, probably a very important one, was the deficiency of oxygen in deep water. Whatever their character may have been they were all, no doubt, of such a nature that they first disappeared in the most shallow water around the coast, but it is not probable that bottom life was first estab- lished in shallow water, or before the physical conditions had become favorable at considerable depths. The sediment near the shore is destructive to most pelagic animals, and recent explorations have shown that a stratum of water of very great thickness is necessary for the complete development of the pelagic flora and fauna. It is a mistake to picture pelagic life as confined to a thin surface stratum. Pelagic plants probably flourish as far down as the light penetrates, and pelagic animals are abundant at very great depths. As the earliest bottom animals must have depended directly upon the floating organisms for food, it is not probable that they first established themselves in shallow water, where the food-supply is not only scanty in amount but also mixed with sediment ; nor is it probable that their establishment on the bottom wag delayed until the great depths had become favorable to life. The belts around elevated areas which are far enough from shore to be free from sediment and to have above them a sufficient depth of water to permit the pelagic fauna to reach its full development, are the most favorable spots, and I shall soon show that there is palaeontological evidence which indicates that they were seized upon very early in the history of bottom life. It is very probable that colony after colony was established on the bottom, and afterwards swept away, like clouds before the wind, by geological changes, and that the bottom fauna which we know was not the first. W. K. BEOOKS ON THE GENUS SALPA. 169 Colonies which started in shallow water were exposed to accidents from which those in great depths were free, and in view of our present knowledge of the permanency of the sea-floor and of the broad outlines of the continents, it is not impossible that the first fauna which settled in the deep zone around the continents may have persisted and given rise to our modern life. However this may be, we must regard this deep zone as the birthplace of the fauna which has survived ; as the ancestral home of all the improved metazoa. The effect of life upon the bottom is more interesting than the place where it began, and we have now to consider its influence in the evolution of animals. The effect of the secondary acquisition of a sedentary life by modern animals has been fully discussed by many writers, but no one, so far as I am aware, has ever considered the effect of the first settlement of the bottom by pelagic animals, all whose competitors and enemies had previously been pelagic. It is doubtful whether the animals which first settled on the bottom secured any more food than the floating ones, but they undoubtedly obtained it with less effort, and were able to devote their superfluous energy to growth and to multiplication, and thus to become larger and to increase in numbers faster than pelagic animals. Their sedentary life must have been favorable to both sexual and asexual multiplication, and the tendency to multiply by budding must have been quickly rendered more active. It is sometimes stated that the capacity for budding has been acquired among the metazoa as the result of a sedentary life, but this view hardly seems to be the true one. Capacity for asexual multiplication is very old, older in all probability than sexual reproduction, and there is no reason to believe that it has ever been lost even by the highest animals, for it must be regarded as nothing more, in ultimate analysis, than discontinuous growth. The tissues of all animals have vegetative power, and external influences determine whether this shall result in continuous or discontinuous growth, and a trace of the power to multiply asexually is retained even among the embryos of mammals. It is therefore wrong to speak of the acquisition of a capacity for budding, and it is not at all improbable that the primitive pelagic metazoa multiplied by buds ; although the tendency to form connected cormi, and to retain the connection between the parent and the bud until the latter was able to obtain its own food and to care for itself, was a result, and probably one of the first results, of life on the bottom. 170 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. The animals which first acquired the habit of resting upon the bottom therefore soon began to multiply faster, both sexually and asexually, than their swimming allies; and their asexual progeny remaining for a longer time attached to and nourished by the parent stock, were much more favorably placed for rapid growth. As bottom animals live on a surface, or at least a thin stratum, while swimming animals are distributed through solid space, the rapid multiplication of bottom animals must soon have led to crowding and to competition, and it soon became harder and harder for new forms from the open water to force themselves in among the old ones, and colonization soon came to an end. The great antiquity of all the types of structure which are repre- sented among the modern metazoa is therefore what we should expect, for after the foundation for the fauna of the bottom was laid it became, and ever afterwards remained, difficult for new forms to establish them- selves. Our knowledge of the sea-bottom is for the most part from three sources : from dredgings and other methods of exploration ; from rocks which were originally laid down beyond the immediate influence of the continents, and from the patches of the bottom fauna which have been gradually brought near its surface by the growth of coral reefs; and from all these sources we find testimony to the density of the crowd of animals on favorable spots. Deep-sea exploration can give only the most scanty and fragmentary basis for a picture of the sea-bottom, but it shows that its animal life may thrive with the dense luxuriance of tropical vegetation, and Sir William Thomson says that he once brought up at one time on a "tangle," which was fastened to a dredge, over 20,000 specimens of a single species of sea-urchin. While cruising on the U. S. Fish Commission schooner Grampus, I was interested to find that when a ground-line with baited fish-hooks had been sunk to the bottom in nearly a mile of water, several of the hooks dropped into the mouths of large sea anemones, so that they were brought up uninjured, and were carried more than three hundred miles to the laboratory, where they lived for some time in an aquarium. The number of remains of palaeozoic crinoids and brachiopods and trilobites which are crowded into a slab of fine-grained limestone is most astounding, and it testifies most vividly and forcibly to the wealth of life on the old sea-floor. W. K. BROOKS ON THE GENUS SALPA. 171 No description can convey an adequate conception of the boundless luxuriance of a coral island, but nothing else affords such a vivid picture of the capacity of the sea- floor for supporting life. The marine plants are not abundant on coral islands, and the animals depend either directly or indirectly upon the pelagic food-supply, so that in this respect their life is like that of animals in the deep sea far from land. The abundant life is not restricted to the growing edge of the reef, and the inner lagoons are often like crowded aquaria. At Nassau, my party of eight persons found so much to study in a little reef in a lagoon, close to our laboratory, that for four months and more we found new things every day, and our explorations seldom carried us beyond this little tract of bottom. Every inch of the surface was carpeted with living animals, while others were darting about among the corals and gorgonias in all directions ; but this was not all, for the solid coral was honeycombed everywhere by tubes and burrows; and, when broken to pieces with a hammer, each mass of coral gave us specimens of nearly every great group in the animal kingdom. Fishes, Crustacea, annelids, molluscs, echinoderms, hydroids and sponges could be picked out of every fragment, and the abundance of life inside the solid rock was most wonderful. The absence of pelagic life in the landlocked waters of coral islands is as impressive and noteworthy as the luxuriance of life upon and near the bottom. On my first visit to the Bahama Islands I was sadly disappointed by the absence of pelagic animals where all the conditions seemed to be peculiarly favorable. The deep ocean is so near that, as one cruises through the inner sounds past the openings between the islets which form the outer barrier, the deep-blue water of mid-ocean is seen to meet the white sand of the beach, and soundings show that the outer edge is a precipice as high as the side of Chimborazo and much steeper. Nowhere else in the world is the pure water of the deep sea found nearer land or more free from sediment, and on the days when the weather was favorable for towing outside, we found siphonophores and pteropods and pelagic mol- luscs, Crustacea, salpae, and all sorts of pelagic Iarva3 in great abundance in the open sea just outside the inlets. Inside the barrier the water was always calm, and day after day it was as smooth as the surface of an inland lake. When I first entered 172 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. one of these beautiful sounds where the calm, transparent water stretches as far as the eye can reach, and new beauties of islets and winding channels open before one, as those which are passed fade away on the horizon, I felt sure that I had at last found a place where the pelagic fauna of mid-ocean could be taken home alive and studied on shore. The water proved to be not only as pure as air, but also as empty. At high water we sometimes captured a few pelagic animals near the inlets, but we dragged our surface-nets through the sounds day after day only to find them as clean as if they had been hung out in the wind to dry. The water in which we washed them usually remained as pure and empty as if it had been filtered, and we often returned from our towing expeditions in the sounds without even a copepod or a zoea or a pluteus. The absence of floating Iarva3 is most remarkable, for the sounds swarm with bottom animals which give birth every day to millions of swimming larvas. The mangrove swamps and the rocky shores are fairly alive with crabs carrying eggs at all stages of development, and the boat passes over great black patches of sea-urchins crowded together by thousands, and the number of animals which are engaged in laying their eggs or in hatching their young is infinite, yet we rarely captured any larvae in the tow-net, and most of those which we did find were old and nearly through their larval life. It is often said that the water of the coral sounds is too full of lime to be inhabited by the animals of the open ocean, but this is a mistake, for the water is perfectly fitted for supporting the most delicate and sensitive animals, and we had no difficulty in keeping alive, in water taken from the sounds, the surface animals which we caught outside. Even trachomedusas and doliolums, which are extremely sensitive to impurities in the water, could be kept alive in the house very much better than in any other place where I have ever tried to keep them, and instead of being injurious, the pure water of the sounds is peculiarly favorable for use in aquaria for surface animals. The scarcity of floating organisms can have only one explanation. They are eaten up, and competition for food is so fierce that nearly every organism which is swept in by the tide, and nearly every larva which is born in the sounds, is snatched by the tentacles around some hungry mouth. Nothing could illustrate the fierceness of the struggle for food among the animals on a crowded sea-bottom more vividly than the W. K. BROOKS ON THE GENUS SALPA. 173 emptiness of the water in coral sounds. The only larvae which have much chance of establishing themselves for life are those which are so fortunate as to be swept out into the open ocean, where they can com- plete their larval life under the milder competition of the pelagic fauna, and while it is usually stated that the pelagic habit has been retained by the larvae of bottom animals for the purpose of distributing the species, it is more probable that it has been retained on account of its com- parative safety. There can be no doubt, in view of these facts, that competition came swiftly after the establishment of the first bottom fauna, and that it soon became very rigorous and led to rapid evolution ; and we must also remember that life on the bottom introduced many new opportunities for divergent modification and for the perfecting of animals. The increase in size, which came with the economy of energy, increased the possibilities of variation, and led to the natural selection of those peculiarities which improved the efficiency of various parts of the body in their functions of relations to each other, and this has certainly been an important factor in the evolution of complicated organisms. The new mode of life also permitted the acquisition of protective shells, hard supporting skeletons, and other imperishable structures, and it is therefore probable that the history of evolution in later times gives us no index as to the time which was required to evolve, from pelagic ancestors, the oldest animals which were likely to be preserved as fossils. Life on the bottom also introduced another most important influence in evolution competition between blood relations. In the animals which we know most intimately, divergent modification, with the extinc- tion of connecting forms, results from the fact that the fiercest com- petitors of each animal are its closest allies, which, having the same habits, living upon the same food, and avoiding enemies in the same way, are constantly striving to hold exclusive possession of all the essen- tials to their life. When a stock gives rise to two divergent branches, each of them escapes competition with the other, so far as they differ in structure and habits, while the parent stock, competing with both at a disadvantage, is exterminated. Among the animals which we know best, evolution leads to a branching tree-like phylogeny with the topmost twigs represented by living animals, while the rest of the tree is buried in the dead past. The connecting form between two species must, therefore, be constructed in imagination or sought in the records of the past. 174 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. Even at the present day things are somewhat different in the open ocean, and they must have been very different in the primitive ocean, for a pelagic animal has no fixed home, one locality is like another, and the competitors and enemies of each individual are determined, in great part, by accidents. We accordingly find, even now, that the evolution of pelagic animals is often linear instead of divergent, and the early steps in the series often live on side by side with the later and more evolved forms. The radiolarians and the medus* and the siphonophores furnish many well-known illustrations of this feature of pelagic life. No one is much surprised to find in the South Pacific or in the Indian Ocean a salpa, or a pelagic crustacean, or a surface fish which has pre- viously been known only in the North Atlantic, and the list of species of marine animals which are found in all seas is a very long one. The fact that pelagic animals are so independent of those laws of geo- graphical distribution which limit land animals is additional evidence of the easy character of the conditions of pelagic life. We have seen that one of the first results of life upon the bottom was to increase asexual multiplication and to lengthen the time during which buds remained united to and nourished by their parents. One result of this is the crowding together of individuals of the same species, and competition between relations. We have in this and in other obvious peculiarities of life on the bottom a sufficient explanation of the fact that, since the first establishment of the bottom fauna, evolution has resulted in the elabo- ration and divergent specialization of the types of structure which were already established, rather than in the production of new types. Another result of the struggle for existence on the bottom was the escape of varieties from competition with their allies by flight from the crowded spots and a return to the open water above; just as in later times the cetacea and sea-birds have gone back from the land to the ocean. These emigrants, like the civilized men who invade the homes of peaceful islanders, brought with them the improvements which had come from fierce competition, and they carried everything before them and produced a great and rapid change in the character of the pelagic fauna. The rapid intellectual improvement which has taken place among the mammalia since the middle tertiaries, and the rapid structural development which took place in animals and plants when the land fauna and flora were first established, are well known, but the fact that the discovery of the bottom initiated a much earlier, and probably much W. K. BROOKS ON THE GENUS SALPA. 175 more important era of rapid development in the forms of animal life has never received the attention which it so well merits. If the views which I have advanced are correct, the primitive bottom fauna must have had the following characteristics : 1. It was entirely animal, without plants, and it at first depended directly upon the pelagic food-supply. 2. It was established around elevated areas in water deep enough to be beyond the influence of the shore. 3. The great groups of metazoa were rapidly established from pelagic ancestors. 4. There was a rapid increase in the size of the bottom animals and hard parts were quickly acquired. 5. The bottom fauna soon produced progressive development among pelagic animals. 6. After the establishment of the bottom fauna, elaboration and differentiation among the representatives of each primitive type soon set in and led to the extinction of the connecting forms. There is no reason to suppose that the first animals which were adapted for preservation as fossils have been discovered, and many of the oldest fossils, like the pteropods, are most certainly the modified descendants of simpler ancestors with hard parts, but it is interesting to note that the oldest fossil fauna which is known to us is an unmistakable approximation to the primitive bottom fauna as I have outlined it. Walcott has given the following sketch of the broad general charac- teristics of the lower Cambrian fauna : The lower cambrian fossils are distributed through strata which, in "Washington and Rensselaer counties in New York, are nearly two miles thick, and some of them, at least, were deposited in water of considerable depth. This is shown by the fineness of the sediment and by the perfect preservation of tracks and burrows in soft mud and of soft animals like jelly-fishes. These show that the sediment was laid down slowly and gently, in water so deep as to be free from disturbance, and under con- ditions so favorable that it contains the remains of some animals which are not found again until we reach a very much more modern period. The fossil medusae of the lower cambrian are so perfect that their identity is unquestionable, yet it is not until the Solenhofen lithographic slate of the Jura is reached in ascending the geological scale, that medusas are again met with ; and corals and lamellibranchs are found in the lower cambrian, although as they are not found again until the 176 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. Silurian rocks are reached, we have no record of their existence through the long period covered by the middle and upper cambrian. The fauna of the lower cambrian, while it undoubtedly lived in water of very considerable depth, was not oceanic but continental, and Walcott says that "one of the most important conclusions is, that the fauna of the lower cambrian lived on the eastern and western shores of a continent that in its general configuration outlines the American conti- nent of to-day. Strictly speaking, the fauna did not live upon the outer shore, facing the ocean, but on the shores of interior seas, straits, or lagoons that occupied the intervals between the several ridges that ran from the central platform east and west of the main continental land- surface of the time." The lower cambrian fauna was rich and varied, but it was not self- supporting, for no fossil plants are found, and the primary food-supply was pelagic. Animals adapted for a rapacious life at the surface, such as the pteropods, were abundant, and they prove the existence of a rich supply of pelagic animals. All the forms are either carnivorous animals, such as medusa3, corals, Crustacea and trilobites, or they are adapted, like the sponges, brachiopods and lamellibranchs, for straining minute organ- isms out of the water, or for gathering up those which rained down from above, and the conditions under which they lived were obviously very similar to those on the bottom at the present day. Walcott's studies show that the earliest known fauna had the follow- ing characteristics : 1. So far as the record goes it consisted of animals alone, and these animals were dependent upon the pelagic food-supply for support. 2. While small in comparison with many modern animals, they were gigantic in size as compared with primitive pelagic animals. 3. The species were few, but they represented a very wide range of types. 4. All the types have modern representatives, and most of the modern types are represented in the lower cambrian. 5. The habitat was not the bottom of the deep ocean, but the sub- merged surface of a sinking continent, under water of considerable depth. Remains of bottom animals are found in rocks below the cambrian, and Walcott believes that while the olenellan fauna adds a little more to our knowledge of the rate of convergence backwards in geological time of the lines representing the evolution of animal life, it also proves, at W. K. BROOKS ON THE GENUS SALPA. 177 the same time, that an immense interval has elapsed between the begin- ning of life and the epoch represented by the olenellan fauna. He says: "That the life in the pre-olenellus seas was large and varied, there can be little, if any, doubt. The few traces known of it prove little of its character, but they prove that life existed in a period far preceding lower cambrian times, and they foster the hope that it is only a question of search and favorable conditions to discover it." No one can question the validity of the basis for Walcott's hope, for pelagic animals have undoubtedly established themselves on the shores of elevated tracts again and again, during the oscillations of the sea- bottom, and we have every reason to expect and look for their remains. If, however, it is true that the primitive stem-forms were pelagic and minute, there is little hope of finding their delicate microscopic remains in the sedimentary rocks of the shore. The cambrian fauna is usually regarded as a half-way station in a series of organisms which reaches back into the past for an immeasur- able period, and it is even stated that the history of life before the cambrian is longer, by many fold, than its history since. So far as this opinion rests on the diversity of types in cambrian and silurian times it has no good basis, for if the view which I have advocated is correct, the evolution of the ancestral stem-forms took place at the surface, and all the necessary conditions for the rapid production of types were present when the bottom fauna first became established. As we pass backwards towards the lower cambrian we find closer and closer agreement with the biological conception of the primitive life at "the bottom. We cannot regard the olenellan fauna as the first bottom fauna, for it contains forms which have been secondarily adapted for a pelagic life, such as the pteropods. We may, however, feel confident that the first bottom fauna resem- bled that of the lower cambrian in its physical conditions, and in its most distinctive peculiarity, the abundance of types and the slight amount of differentiation among the representatives of these types. Far from seeing in the lower cambrian fauna a half-way station in a long series of bottom animals, the biologist must regard it as an unmistakable and decided approximation to the primitive fauna of the bottom, beyond which life was represented only by simple and minute pelagic organisms. CHAPTER VIII. THE ORIGIN OF THE CHORDATA, CONSIDERED IN ITS RELATION TO PELAGIC INFLUENCES. SECTION 1. The Ancestral Chordata. I shall now attempt to study the origin and significance of the structure of appendicularia in accordance with those conditions which must, as Dohrn has pointed out (Studien, etc., VIII, p. 79), direct all inquiry into the genealogy of animals. All biologists will agree with Dohrn that no amount of morphological information, or of exhaustive microscopical study of the structure and development of animals, can suffice, in the absence of comprehensive knowledge of their mode of life and of the conditions of their existence, for the institution of inquiries into their phylogenetic relationship. Unquestionably the first condition for genealogical inquiry is, as Dohrn says, the establishment of a direct connection between our mor- phological studies and the facts of physiology and biology. "The homologies which are established by comparative anatomy, and the primititive identities which are established by comparative embryology, are only the means for this end. They are in themselves valuable in phylogenetic inquiry only so far as they furnish us the opportunity to pass from the consideration of the structure of organs as they now exist, and of the functions of these organs at the present time, to the consideration of conditions which have passed away ; to the study of the history of the modifications which have come between these struc- tures and functions and those which we must attribute to the same organs at an earlier genealogical stage." Keeping these conditions of genealogical inquiry in view, let us try to study the structure of appendicularia in relation to the conditions of its life, so far as these are known to us, and let us see what functions we must, according to the principle of change of function, attribute to the organs of the remote ancestors of the tunicates, and what are the paths these organs have traversed in reaching their modern structure. If the reader of the following pages should think that I wander too W. K. BROOKS ON THE GENUS SALPA. 179 far from the beaten paths of observation, I must plead as my excuse that the study of phylogeny is impossible without the use of the imagination, and that the field is already occupied by a phylogeny of the tunicata which cannot be set aside until a more satisfactory one has been found. Appendicularia is a very simple organism, and while much ingenuity has been expended in the negative task of accounting for the absence of all the structures which it lacks, I hope that the more positive attempt to account for its actual structure will not lead us into any great difficulties. In the belief that the sequel will justify the assumption, I shall, as my starting-point, picture the ancestor of appendicular-ia as a simple, minute, unsegmented, chordate animal, leading a free, locomotor, pelagic life and subsisting upon the micro-organisms of the ocean. I shall also assume that this ancestor had an elongated, unsegmented body stiffened by an axial, unpaired, unsegmented notochord, like that of amphioxus, appendicularia, and the ascidian larva ; that it had a simple, elongated, dorsal, nervous system, and an elongated, ventral, digestive tube, without pharyngeal clefts ; that this tube was nearly straight ; that it had a capacious lumen, and that, as in amphioxus and the tunicates, this was permanently distended and ciliated, and that the water, with the micro- organisms that float in it, was swept through it by endodermal cilia and not by muscular contractions. In order to entangle the floating particles of food and to hold them while the water swept on through the intestine and out of the anus, gland-cells for the excretion of slime were scattered among the ordinary ciliated endoderm cells of the digestive tract. In origin, these slime-cells may have been modified or specialized digestive gland-cells. As particles which are entangled and held captive near the oral end of the gut are more perfectly exposed to its digestive action than those which continue to float with the stream, the most anterior slime-cells are most efficient and valuable, and as each variation in this direction gave its possessor an advantage, the slime-cells gradually, through the action of natural selection, became localized in the pharyngeal region, and this region gradually became enlarged and was thus set apart, at a very early period, as a specialized tract of the gut. It is also probable that, at a very early stage in the phylogeny of these primitive chordata, a blind pouch was developed, behind the pharynx, to catch the food-particles as they were hurried past with the stream of water and to retain them long enough for perfect digestion, and that the rudiment of the organ which has in the higher vertebrates become the liver was thus established. 180 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. In these primitive animals the current of water through the diges- tive organs was most useful as the vehicle for floating food, but while necessary, it was a necessary evil, for the large distended lumen which furnished it a channel also permitted undigested food to be swept away and lost. The immovable, permanently distended, ciliated digestive tract of a modern lamellibranch is very similar to that of these primitive chordata, but the lamellibranchs have acquired an apparatus for straining off the water from the captured food, so that the digestive tract is relieved from this disadvantageous current. If, after the pharynx had been established, a secondary opening from it to the exterior were to be formed, this opening would permit the water to escape without passing through the intestine, and as the advantage of this new arrangement is obvious, there can be no doubt that after an opening of this sort was once formed, it would be preserved and perfected by natural selection, as a channel for the escape of the water after the food has been strained out and entangled by the excretion of the pharyn- geal slime-glands. I shall show, further on, that if an useful opening of this sort were to be fixed and preserved by natural selection on one side of the body, the laws of growth would soon cause it to be duplicated on the other side. These two openings are the so-called gill-slits of appendicularia, although they are beyond question much older than the modern appendicularia, dating back to a time before this animal had acquired the features which distinguish it from its more primitive chordata ancestors. I am not able to suggest what led to the first establishment a of secondary opening into the pharynx ; but, once formed, its preservation and gradual improvement, by natural selection, as a channel for the escape of superfluous water, and its duplication on opposite sides of the body, are easily intelligible. If we accept the view that the chordata type was evolved under purely pelagic influences, we are forced to believe that the first chordata were minute, and that their small bodies were soft, and unprotected by a hard covering. If we also admit that their digestive tract was a channel for a current of water, we can hardly believe that they needed respiratory organs, or, for that matter, excretory organs, for all the tissues of a minute soft animal, bathed within and without by pure water, must have been sufficiently aerated and purified without any organs for this purpose. W. K. BROOKS ON THE GENUS SALPA. 181 It is not at all probable, then, that the pharyngeal clefts were origin- ally either gills or renal organs, and we have seen that the conditions of pelagic life furnish a much more simple explanation of their advantage, and I believe that the view that they were originally concerned in nutri- tion rather than in respiration will commend itself to all who approach the subject without any philosophical preconception. After they were once established they gradually effected a rearrange- ment of the slime-cells and ciliated cells of the pharynx, for as it now became important that all the food particles should be entangled by the product of the slime-cells before it reached the pharyngeal clefts, the slime-cells were gradually restricted to the anterior part of the pharynx, while the ciliated cells gradually became specialized to carry the entangled food past the openings and to convey it safely into the ossophagus. All the parts of the pharynx of appendicularia are beautifully con- structed for this purpose. The pharyngeal clefts are situated far back in the pharynx, and are separated by nearly its whole width from the oesophagus. They are fringed by large cilia to expel the water, and they are separated from each other by a vertical shelf or velum on the ventral floor of the pharynx, so placed as to prevent cross-currents. In front of this shelf the slime-cells are brought together in two rows, near the middle line, just inside the mouth, to form the hypo- pharyngeal band or endostyle. Between these two rows of slime-cells there is a median row of large ciliated cells, so placed that they drive the slime forwards to the point where a ciliated peripharyngeal band receives it and carries it up each side of the pharynx just behind the mouth, into the most favorable place for entangling the food, as this enters with the current of fresh water. On the dorsal middle line the -threads of slime are gathered up and guided along the epipharyngeal band or dorsal lamella, beyond the influ- ence of the current of water which sets backwards, on each side of the ventral velum, to the pharyngeal clefts, and the food is thus safely con- ducted into the oesophagus while the water escapes. Up to this point I believe that the ancestral history of the tunicates was identical with that of the vertebrates, for the hepatic ca3cum, the dilated pharynx, the pharyngeal clefts, the hypopharyngeal gland and the peripharyngeal bands have been inherited by all the chordata, and have impressed themselves so firmly in their organization that even the highest vertebrates still retain them, either as vestiges, or as organs which have been fitted to new functions. 182 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. I believe, however, that while they were acquired before the tuni- cates diverged from the chordata stem, they were acquired by an organ- ism whose environment and habits of life were essentially like those of the modern appendicularia. All the parts of the pharynx of appendicularia are so beautifully co- ordinated for effecting a purpose so useful and so well adapted to the conditions of its simple pelagic life, that we find it difficult to resist the belief that its ancestors had essentially the same habits, and that they lived under essentially the same conditions, and that this simple organi- zation was directly acquired in adaptation to these conditions. If this view involved any great or unusual difficulties we might well distrust it, notwithstanding its simplicity ; but I shall try to show that it does not. In the preceding chapter I have shown that it accords with our knowledge of the fundamental principles of the general biology of the ocean, and further on I shall try to show that it is equally in accord with the principles of morphology. At present we must devote our attention to the history of the evolu- tion of the tunicates from this primitive chordata stem. SECTION 2. The Origin of the Tunicates. Like most recent students of the tunicates, I believe that we have in appendicularia a persistent representative of the primitive tunicata; but, unlike many of them, I fail to find in its structure any evidence of degeneracy, or in its habits any basis for the assumption that it is degenerated. In most respects its structure is like that of the hypo- thetical ancestor whose evolution we have traced. It has an unseg- mented notochord, and a capacious lumen throughout the whole course of the digestive tract from mouth to anus. This lumen is permanently distended and food is carried through it by cilia. It has a blind diverti- culum from the stomach, and the greatly expanded pharynx opens laterally through two ciliated pharyngeal clefts, through which the water escapes while the food passes into the oesophagus. There is a ventral slime-gland just inside the mouth, and its excretion is conveyed upwards around the pharynx by the cilia of the peripharyngeal bands, and is then swept into the oesophagus with the entangled food. This increasing complexity and perfection of the pharynx is accom- panied by an increase in its size, so that in the primitive tunicates it soon comes to be the most important and dominant organ of the body, and W. K. BROOKS ON THE GENUS SALPA. 183 brings about adaptive changes in other parts. One of these is the differ- entiation of a stomach for the retention and digestion of the food, in the direct course of the gut. As long as the food was mixed with great quantities of water, digestion and assimilation probably went on simul- taneously in all parts of the post-pharyngeal gut, but as the water found another exit and the food thus became more compact and solid, the stomach of appendicularia became established and thus divided the gut into an oesophageal, a gastric, and an intestinal region. Our knowledge of the primitive vertebrates seems to me to be too scanty to show whether this differentiation occurred before or after the tunicates diverged from the ancestors of the vertebrates. We are now concerned with the history of the tunicata line alone, and the fact that the differentiation now exists in all tunicates shows that it was brought about very early in their history. Another most important change in the relations of the gut also took place very early in their history. The intestinal portion became bent upon the enlarged pharynx so as to form a cj with the intestinal bar of the cj ventral to the pharyngeal portion, and with the anus on the ventral middle line under the pharynx. Herdman represents the primitive con- dition of the digestive tract of tunicates as a d, with the intestine and anus dorsal instead of ventral (page 128) ; but I shall show further on that the relations exhibited by appendicularia are the primitive ones, from which we must derive those which are exhibited by other tunicates. By this change the tail was freed from the gut and was made much more efficient as an organ of locomotion, while the fa3ces were discharged from the anus into the current of water which set out through the pharyngeal clefts. This latter feature may not have been of any value so long as habits of active locomotion were retained, but, as we shall see, it became very important at a later stage. The embryology of the ascidians shows that this arrangement of the digestive tract was secondary; that at one time it was straight, extending into that region of the body which is now specialized in appendicularia as a tail. The advantage to an active pelagic animal of this change is obvious, since it permits the tail to become purely loco- motor. As each slight variation in this direction must have given a slight increase in the freedom of movement, the shape of the body of appendicularia is easily intelligible as the result of natural selection, and while the change is complete in this, the most primitive tunicate which we know, so that we can only Conjecture the transitional stages, the 184 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. change itself is not a complicated one. It presents little difficulty, although the resulting differentiation of appendicularia into two regions or " segments," a body and a tail, has been made the basis of much speculation. The great development of the pharynx and the reduction of the tail to an organ of locomotion soon resulted in a pronounced change, of the sort for which Dana long ago proposed the term cephalization. As the functions of the pharynx, and of its oral end in particular, became more and more complicated and more and more exactly co-ordi- nated, while those of the tail became simplified, the elongated nervous system became differentiated in a corresponding way, and its caudal portion became reduced to a caudal nerve, while its oral extremity became evolved into a cerebral vesicle with sense-organs and nerves in relation with the co-ordinated structures of the pharynx. All the characteristics of appendicularia, except the structure of the heart and the structure and position of the reproductive organ, are thus seen to be intelligible as direct adaptations to a pelagic life; for its distinctive features, as compared with other primitive chordata, are the U-shaped folds of the digestive cavity, the sharp separation of the tail from the body, and the differentiation of the nervous system into a caudal nerve and anterior vesicle. We have little basis for speculation as to the path by which the reproductive organ acquired its present position, and it is by no means certain whether the tunicate heart is homologous with that of the other chordata. The conditions of pelagic life are so permanent that we may safely make use of the structure and habits of the modern pelagic forms to reconstruct this part of the ancestral history of the tunicates, for time writes no wrinkles on the azure brow of the ocean. As regards the later history the case is different. Between appendi- cularia and the ascidians there is a great gap which we can bridge only in imagination. The transitional animals are totally unknown, and the conditions of life on the bottom of the modern ocean may, possibly, be very different from those which prevailed when the fixed ascidians were first evolved. It is easy to imagine changes which might have gradually converted an ancestor like appendicularia into a descendant like the fixed ascidians, through successive adaptations to a sedentary life, but in the absence of all evidence we cannot feel implicit confidence that the imaginary picture bears any minute and detailed resemblance to the actual history. W. K. BROOKS ON THE GENUS SALPA. 185 It seems probable that after the bottom of the ocean became fit for life, some of the descendants of the primitive pelagic tunicates gradually acquired the habit of sometimes swimming upon or near it in an inclined position with the mouth downwards to suck up the organic sediment, and that they also acquired the habit of resting upon the bottom in this position. We may well doubt whether these animals obtained any more food than their pelagic ancestors, but it is well known that it is not the amount of food, but the ratio between the supply and the amount of expended energy which affects size. As this new habit economized energy both during rest and during activity, it permitted an increase in size, and it is interesting in this connection to note that Chun has found at great depths appendicularias which may well be called gigantic as compared with all which are known to exist at the surface. With each increase in size, the habit of visiting the bottom must itself have become more and more fixed, until the life upon the bottom, which may have been at first only intermittent and more or less acci- dental, at last became established in the ancestors of the ascidians as a constant characteristic peculiarity. As this new mode of life was gradually acquired, some method of aerating the fluids of the body must also have been gradually evolved ; for without it, a minute animal adapted for a free active life in the highly aerated surface-water, could not, at the same time that it grew larger, acquire a less active habit of life in the bottom strata where the water is less perfectly aerated, the products of decomposition of organ- isms more concentrated, and the capacity for passing from exhausted and impure water to a fresh environment, restricted both by the more stationary habit and by the fact that life in space has been exchanged for a home which is limited by a surface. Undoubtedly the change of habit was accompanied by the gradual perfecting of the system of blood-spaces around the pharynx, which, at first indefinite and irregular, became constant on the margins of the pharyngeal clefts, which thus gradually acquired a new function and became gill-slits, and also became duplicated as the animals grew larger and the need for more perfect respiration increased with their change of habits. I hope that no one will interpret the last sentence as an expression of the belief that the need for respiration caused the gill-slits to multiply. I believe, and shall try to show further on, that the tendency to dupli- 186 JOHNS HOPKINS UNIVEKSITY MORPHOLOGICAL MONOGRAPHS. cate a structure, either radially, bilaterally or serially, is a result of the method of growth by cell multiplication, and that in the case in question the serial reduplication has been fixed and preserved by natural selection on account of its value in respiration. The context shows that I also regard the gill-slits of vertebrates and those of tunicates as homologous structures inherited from a common source, the primary pharyngeal clefts ; but that I regard the increase in their number as a secondary change which has occurred in both lines after their genealogical paths had diverged. It does not seem necessary to defend the thesis that the number of gill-slits in the ascidians is the result of secondary multiplication, since, as I shall show further on, it is accepted by Dohrn (Studien, etc., IX, 417), who has proved himself a most rigorous critic of the logic of morphology. There is reason to believe that the multiplication of gill-slits in the tunicates has not only taken place independently, but that it has taken place in a peculiar way. Anatomy and embryology give evidence that while the perforations of the tunicate pharynx multiplied, the perfora- tions of the outer wall of the body did not ; and that the external por- tions of the two primary clefts became distended into a pair of spacious perithoracic chambers, each with numerous ciliated openings into the pharynx, and a single opening to the exterior which perhaps became enlarged as the gill-slits multiplied. So long as the primary function of the first pair of pharyngeal clefts, the discharge of the superfluous water, was the only one, they probably remained circular like those of appendicularia ; but as they became con- cerned in respiration and increased in number, and were furnished with definite blood-vessels, they became elongated vertically and, forming a series side by side over a considerable area on each side of the pharynx, they thus became much more efficient organs for the aeration of the blood. In this simple way metamerism, that fetish of the morphologists, was established among the tunicates, and there is no evidence that it has ever involved any of their organs except the gill-slits and the pharyngeal blood-vessels. A vertical series of slits, elongated longitudinally, would undoubtedly have permitted the water to escape just as well as a longitudinal series elongated vertically, but it is possible that, during the gradual establish- ment of the respiratory circulation, those of the irregular and variable W. K. BROOKS ON THE GENUS SALPA. 187 blood-spaces which were most nearly transverse to the current of water from the mouth to the primary clefts, were the ones which were first made definite by natural selection, and that" the arrangement of the gill- slits was thus determined. We can only conjecture how this unknown ancestral swimming organism first became fixed, but the discovery of its descendants on the modern sea-floor is among the possibilities of future explanation. The sedentary habit undoubtedly came gradually, and at first it may have been temporary, confined perhaps to the breeding season, when, loaded down with eggs, the animal may have learned to rest upon the fragments of crinoids, or the shells of trilobites or brachiopods or mol- luscs, to avoid clogging its delicate ciliated and vascular pharynx with sediment. At the point where the heavy anterior end of the tadpole- shaped body rested, the ectoderm cells, just below the mouth, probably became modified for the excretion of an adhesive cement. The sedentary habit, which must have resulted in a still greater economy of energy and a corresponding increase of size, undoubtedly became more and more firmly established, and the changes which followed and resulted in the evolution of the ascidian type are easily intelligible as adaptations to a fixed home, although we have little to show the sequence of their acquisition. So long as the animal led a free life the fate of the deoxidized water after it left the gill-slits had no meaning, but with the fixed habit came the need for driving it away as far as possible, and the external apertures of the perithoracic chambers became small, moved towards each other, and finally united to give to the exhaled current the strength of concen- tration. The attitude of the animal upon the bottom undoubtedly determined the dorsal instead of ventral location of the common aper- ture and of the median atrium or cloaca. As each step in this process of concentration must have been advantageous, its evolution by natural selection is easily intelligible. The accumulation of faeces from the intestine, around a fixed animal, is so unsanitary that the anus has disappeared in many sedentary metazoa, while in others, such as the crinoids and the lamellibranchs for example, secondary adaptations for sweeping away the refuse matter have been acquired. The folding of the originally straight digestive tract of the primitive chordata into a U with the anus and intestine ventral to the pharynx, took place in the ancestral tunicates as an adaptation to locomotion, but, as appendicularia shows, it incidentally brought the anus into the 188 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. region of the pharyngeal clefts. As the sedentary habit became slowly established the anus became shifted from the middle line into the exhaled current from the left perithoracic chamber, and finally into the margin of its aperture, so that, during the migration of the exhalent openings, the U of the digestive tract became twisted into an 8 in such a way that, as Plate VIII, Fig. 2 shows, the intestine p passed on the left side of the oesophagus, q, to open dorsally into the atrium, near the middle line, but a little to the left. This arrangement of the digestive organs is very characteristic of the tunicates, and the few exceptions are clearly due to later changes. Thus in doliolum the atrium has moved backwards as an adaptation to locomotion, and the anus has followed it until the gut has become nearly straight. The intestine and anus of the adult aggregated Salpa pinnata, Plate I, Fig. 1, are ventral; but I have shown that in the young the intestine crosses to the left of the oesophagus to open dorsally, as it does in the adults of all ordinary salpae. In the Polyclinida3 the loop of the intestine has been elongated, with the elongation of the body, until the bend of the 8 has been obliterated, and the presence of the characteristic 8 in more primitive ascidians such as clavelina shows that the Poly- clinidas have been more recently modified. All sedentary animals which take their food by means of cilia have their apertures raised in some way above the reach of sediment. In the crinoids this end is reached by a stalk ; in the lamellibranchs it is attained either by siphons, or by the vertical elongation of the shell as in the oyster ; and the shifting of the area of attachment of the ascidians from the oral end to the aboral end, the elongation and approximation of the mouth and the atrial aperture, the acquisition of oral and atrial sphincter muscles, the degeneration and disappearance of the locomotor tail, and the simplification of the nervous system, are such obvious adaptations to a sedentary life that it is not necessary to discuss them. SECTION 3. The Annelidian Hypothesis. I believe that the structure of the tunicates has been acquired as an adaptation to the biological conditions which prevailed at the surface of the primitive ocean, and that it has been evolved by the gradual addition of successive complications on to the body of a still more primitive and simple ancestor. This involves the total rejection of the dogma that the vertebrates are modified annelids, and that the tunicates are degenerated vertebrates. W. K. BROOKS ON THE GENUS SALPA. 189 While it is not my purpose to discuss the ancestral history of the vertebrates, the remote phylogeny of the tunicates is unquestionably identical with that of the other chordata, and I cannot ignore the general acceptance of an opinion which is absolutely irreconcilable with the one which I have presented. This prevailing opinion has interwoven itself with the literature in such a complicated way that one may well shrink from the interminable labor which the critical revision of the whole of it would involve. I myself decline to undertake what I regard as an unprofitable and useless task ; unprofitable, as the literature rests on an untenable and false basis, and useless, since I do not hope to induce those who have stored their minds with the endless details of morphology docketed and pigeon- holded according to a false system, to unload all this rubbish and to build again on a new foundation. I shall therefore restrict myself to a discussion of the origin of the two most characteristic systems of tunicata organs, the gill-slits, and the pharyngeal ciliated cells and gland cells; and I shall here confine myself to the observations and reflections of a single writer, Dr. Dohrn. I make this selection the more willingly, as Dohrn's name is most intimately associated with the annelidian hypothesis, and because his writings are not only the ones which have been most influential, but also the ones which are most comprehensive and most attractive to the reader. The "Ursprung der Wirbelthiere " is a most fascinating book. Soon after it appeared I placed it in the list of works which my students are advised to read, and for many years an acquaintance with it has been expected of all who have been examined for the degree of Ph. D. in the Johns Hopkins University. My students have even prepared for their own use an English trans- lation of it, and I have read it with them several times with interest and pleasure. At the first reading my pleasure was almost that of convic- tion, but as the ingenious details became familiar, and the essay was more sharply focused in its completeness, and was held, as it were, at arm's length, so that the whole picture could be seen at one view, I have read it, as I have read Gulliver's Travels, with admiration for the skill which has elaborated it in such logical minuteness from a fundamental assumption which is purely imaginary. The story, as told by Dohrn in the "Ursprung," is so consistent and logical that I see no reason why animals like the tunicates might not have been evolved in the way which he pictures so vividly, although I 190 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. believe that the actual tunicates have been produced in a very different way. I shall therefore examine the account of the origin of the gill-slits which Dohrn gives in the "Ursprung," and the view of the ciliated and glandular structures of the pharynx which is developed in his "Studien," especially in Parts VII, VIII and IX, in order to determine how far the origin of these structures is accounted for by the annelidian hypothesis, and what superiority, if any, this has over the much simpler hypothesis which is here advanced. Dohrn says (Ursprung, p. 10) that the branchial apparatus of the tunicates and that of balanoglossus are so much more complicated than that of the selachians, and their origin is so much more difficult to understand, that they are of no help to us in our attempt to trace the origin of gill-slits. I am quite at a loss for the meaning of this passage, for no secondary perforation of the pharynx could possibly be less complicated than the gill-slits of appendicularia, nor could it be developed in a simpler way than by the involution of a pit on the side of the body. It is quite true that we do not know how the gill-slits of appendicu- laria first came into existence, or what influence led to their formation, but their usefulness as channels for the escape of the water which, before they were formed, must have passed through the intestine, is clear, and we can understand why they have been preserved, by natural selection, on account of this advantage. We are forced to believe that the pharynx did, in some way, acquire a secondary communication with the exterior, although we are not able to say how it was acquired. Dohrn' s view of the origin of gill-slits is based upon the need for an explanation of the original formation of the perforation. He says (Ursprung, p. 10) : "What is a gill-slit? Perforations of the body-wall do not take place directly, and still less do they form connections with corresponding perforations of the digestive tract," and he therefore undertakes to study the origin and primitive function of gill-slits by the aid of the law of the change of function, and to find in a more primitive function an explanation of their present function as channels for water. As his point of departure is the need for an explanation of the origin of the perforation, we feel a natural hope that we are to be led to this explanation, but this hope ends in disappointment. He regards the gill-slits as modified segmental organs, but he tells us W. K. BROOKS ON THE GENUS SALPA. 191 explicitly, on page 10, that "we are not able to assign any reason why segmental organs should unite with the gut," and his explanation of the origin of the perforations is no explanation at all, since it simply assumes, but does not account for, the very phenomenon which it is supposed to make clear. His inability to understand the direct origin of the secondary per- forations of the gut has one most remarkable result, for the view that the gill-slits are segmental organs involves the view (Ursprung, p. 57) that the anus of the tunicates is not a primary anus nor a secondary one, but a tertiary one, and that the ancestors of the tunicates have not only acquired two new secondary anal apertures, but that they have lost one mouth and acquired a second, and that they have lost this and acquired a third. As these mouths are supposed to be modified segmental organs, we are, according to the acknowledgment on page 10, "unable to assign any reason why they should have united with the gut." The original mouth of the ancestors of the chordata was, according to Dohrn (page 3), on what is now the dorsal surface, and the primitive ossophagus passed through what is now known as the fossa rhomboidea of the brain. This ancestral mouth degenerated and disappeared as it was gradually superseded in the remote progenitors of the vertebrates by a second mouth (page 5), which is the mouth of the vertebrates of the present day, and of the ancestors of the tunicates (page 57) as well, although it was gradually converted first into a sucker, and finally into an organ for fastening the tunicata to foreign bodies, while these animals gradually acquired a tertiary mouth (page 58) by the formation of a secondary communication between the nasal chamber and the gut. Dohrn says (page 60) that these assumptions "set the relation between the fishes and the ascidians in the right light," although the perforation of the gut, which the hypothesis is to explain, is not only left unaccounted for, but is multiplied so many times that, like the man with an unclean spirit, its last state is worse than the first. Dohrn says that the secondary nature of the mouth of the verte- brates is proved by its very late appearance in the young vertebrate after its embryonic body and its great systems of organs are fully formed, and by the fact that, when it does make its appearance, it does not lie at the anterior end of the body, in the place which it finally occupies in the great majority of vertebrates, but at a spot some distance behind this place. 192 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. It is not possible to attach much weight to either of these arguments, for slight changes in the position of organs are not unusual, and it is well known that the ontogenetic acceleration or retardation in the relative time of appearance of structures is by no means exceptional, and it would be as safe to assume that the change in the pitch of the voice of man is phylogenetically older than the sexual maturity of the ancestors of man, as it is to assume, from the same sort of evidence, that the aortic system of vertebrates is older than the mouth. The vertebrate mouth unquestionably bears a great morphological resemblance to a pair of gill-slits. As Dohrn points out, it is bordered, like the gill-slits, by a pair of visceral arches, it lies in front of the first pair of true gill-slits, it arises at the same time with them in the embryo, and like them it opens into a section of the gut. A ventral view of a shark shows the resemblance between the mouth and the true gill-slits in the most impressive way, and if any pair of them were to be united with each other at their ventral ends, they would become perfectly equivalent to the mouth. The armature of the mouth is repeated on the gills, and there is reason to believe that the jaw-arches have at one time carried gills like the gill-arches. This resemblance is not imaginary. Beyond all question it is real, and it is certainly most remarkable and suggestive, but does it prove that the vertebrate mouth is phylogenetically a pair of gill-slits ? When, in my student days, my instructor held before me the skull of a turtle and called upon me to observe the centrum, the transverse processes and the neural arch of the occipital vertebra, I was, for the time, convinced that the occipital bone had arisen by the differentiation and specialization of a bony vertebra, like those in the neck of a turtle, and that its history had been identical with that of the thoracic vertebrae, which have been differentiated and specialized in the same way into con- stituent parts of the bony box which covers the body of the turtle, as the skull covers the brain. In all these cases the morphological resemblance is undeniable, but our opinion of its phylogenetic significance depends upon our view of the nature and origin of the metamerism of vertebrates, a question which will soon be discussed. At present we must confine ourselves to a narrower point of view, and learn where we are led by Dohrn's opinion that the vertebrate mouth is actually a pair of gill-slits. If the present mouth of the vertebrates was once a pair of gill-slits, W. K. BEOOKS ON THE GENUS SALPA. 193 the ancestors of the vertebrates must have had at that time another mouth, and during the long series of stages of development, while the gill-slits were gradually assuming the function of a mouth, food must have been taken in through both openings ; for the new function of the gill-slits must have been acquired slowly alongside their old function, until the new mouth finally became so perfectly adapted for its new function that it supplanted and replaced the old one. According to Dohrn, these considerations force us to believe that the primitive mouth of the ancestors of the vertebrates and of the tuni- cates was situated in the fossa rhomboidea, where an oesophagus pushed inwards to join the mid-gut, in the same way that it is joined in insect embryos by the fore-gut. This primitive mouth and its oesophagus were homologous with the corresponding organs of modern arthropods and annelids. The mouth of the modern vertebrates is then to be regarded as a secondary mouth, which has gradually supplanted and replaced the old one on account of its greater efficiency. It follows from this, according to Dohrn (p. 56), that the "so-called larva" of the ascidians is a degenerated fish, and that all the features which show the derivation of the cyclostomes from the fishes show also that the process of degeneration has reached its extreme in the tunicates. The cyclostomes are held to owe their degeneracy to parasitism, and the most important element in the more advanced process of degeneration is that the ascidians no longer fasten themselves to fishes nor make use of their bodies as food, but that they fasten themselves to stones, to ships, or to the bodies of other animals which do not serve as food, such as the shells of crabs or the tubes of annelids. The mouth (p. 57) which in the cyclostomes serves both as an organ for attachment to the skin of fishes, and also as a sucker for extracting their blood, has become converted in the ascidians into an organ for attachment; and these animals have thus lost their old mouth, which was homologous with that of the true vertebrates, and have acquired a new one which is homologous with the vertebrate nasal chamber. The process, Dohrn says, must be represented as follows : The fishes take in the water for respiration through the mouth, but as this is used by the parasitic cyclostomes as a sucker, they have acquired another arrangement, and the water is not only discharged through the gill-slits, but is also inhaled through them, and, in the myxenoids, through the nasal passage also, which has in the tunicates become the functional mouth. The vertebrate mouth has lost its old function in the cycle- 194 JOHNS HOPKINS UNIVERSITY MOEPHOLOGICAL MONOGRAPHS. stome-like ancestors of the tunicates, as these have gradually lost their parasitic habit, and have established themselves on lifeless bodies ; but the original lips have remained, and they are to be recognized in the so-called sucking knobs of the ascidian larva, while the teeth of the cyclostomes are supposed to be represented by "bristle-carrying end knobs" upon the suckers. The "so-called larva" of the ascidian s is represented in almost every feature of its organization by the adult, sexually mature, appendicularia. No better example of the correspondence between an adult animal and an ontogenetic stage in the history of another can be desired, and we may feel confident that, whatever the phylogenetic history of appendi- cularia has been, that of the ascidian larva has been the same. Nearly all of the students who have devoted themselves to the study of the tunicates agree in regarding appendicularia as a persistent repre- sentative of their primitive condition; but appendicularia is an active swimming organism, and I have shown that its simple structure is so well adapted to the needs of its pelagic life, that there can be no inherent improbability in the view that it owes its origin to simple pelagic influ- ences. Nothing whatever in its habit of life or in its structure lends the least support to the view that it is a degenerated animal, and if we accept it as evidence, we are forced to believe that, far from being the fixed and degenerated descendants of parasitic vertebrates, the tunicates are descended from free, active, pelagic animals of very simple structure and minute size. Even Dohrn seems to admit that the ancestors of the tunicates were swimming animals, for he tells us in support of his view of the homology of the endostyle (Studien, etc., VIII, p. 62) that the ancestors of the tunicates were "obviously " free swimming animals, and therefore in the position to seize their food by hunting. "Offenbar waren sie frei schwimmende Geschopfe und damit in der Lage, ihre Nahrung durch Jagd selbst zu packen." If the tunicates are, as their embryology and comparative anatomy indicate, the descendants of an ancestor which was obviously a free swim- ming animal, it is surely simpler, in view of all the facts, to regard the gill-slits as perforations which were originally retained and fixed by natural selection as channels for the exit of the water which was taken into the mouth with the food, than to refer them back to imaginary segmental organs which have left no other trace of their existence in the body of any known tunicata. W. K. BROOKS ON THE GENUS SALPA. 195 Minute pelagic animals, with soft bodies bathed on all sides by pure water, do not need special organs of excretion or respiration, and it is not at all probable that the pharyngeal clefts were originally respiratory ; but it is easy to understand how the channels through which the water flowed became converted into gill-slits, in accordance with the law of change of function, as the descendants of the primitive tunicates grew larger and became sedentary, and thus came to need respiratory organs. It may be argued that the thing to be explained is not the existence of gill-slits, but their serial reduplication or metamerism. It may be held that the metameric repetition of the gill-slits of ascidians forces us to regard the ascidian pharynx as the primitive form, from which that of appendicularia has been produced by " degeneration." As we are told, however, by no less an authority than Dohrn (Studien, IX, p. 417, and VIII, p. 61) that the great number of gill-slits in the ascidians is due to secondary multiplication, "nachtragliche Vermehrung," this considera- tion need not detain us. If the logical conditions of sound morphological philosophy admit the possibility of "nachtragliche Vermehrung," and permit us to believe that the twenty or thirty pairs of gill-slits which are found in ascidians are to be traced back to the eight pairs which the primitive fishes are said to have possessed, the same logic will surely permit us to believe, on sufficient evidence, that they have arisen not from eight but from a single pair like those of appendicularia. All the vertebrates have a peculiar organ known as the thyroid gland, and while it holds no prominent place in our general conception of a vertebrate, this gland is actually one of their most constant and characteristic organs. In all the jawed vertebrates, from the sharks up to man, its typical structure is adhered to so closely as to prove that the gland as it exists in man is an organ of vast antiquity. In all these animals it is a ductless gland, situated far back in the throat, behind the hyoid skeleton ; but at an early stage in its ontogeny it is a part of the endodermal epithelium of the pharynx, and it arises on the middle line just within the mouth. Its function in the jawed vertebrates is problematical, but these two features in its ontogeny seem to show that far back in the remote past, before it had assumed its characteristic form, it had another function which stood in some direct relation to the mouth. The tunicate endostyle is a conspicuous organ which attracts the eye of all observers, but its true structure was first demonstrated by Pol, 196 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. who proved that it is a pharyngeal gland with excretory cells to produce slime, and with ciliated cells to drive the slime out through the long, narrow, slit-like duct into the pharynx. Fol also showed its true relation to the ciliated peripharyngeal bands and dorsal lamella, and proved by simple but conclusive experiments that these organs are co-ordinated parts of a single system, which has for its function the capture of the microscopic floating food which enters the mouth with the water. W. Miiller was the first to point out the homology of the tunicata endostyle with the vertebrate thyroid gland, and this homology has been established beyond the possibility of doubt by Schneider's discovery that the thyroid body of ammocoetes is a slime gland with an opening into the pharynx near the mouth, and that on each side of this opening a ciliated furrow or peripharyngeal band runs Tipwards on the inner wall of the pharynx, just in front of the first gill-slit, to its dorsal middle line, where the two unite to form an epipharyngeal band or dorsal lamella which runs backwards to the oesophagus. Even more conclusive proof of this homology is afforded by Dohrn's account (Studien, VIII) of the histological structure of the pharyngeal gland of ammocoetes, ' for his studies show on the one hand a most complete fundamental identity with the very peculiar and characteristic histological structure of the tunicata endostyle, and they also, on the other hand, prove its identity with the vertebrate thyroid gland, by showing that, as development pro- gresses, it is cut up by ingrowths of connective tissue into the isolated follicles which are so characteristic of the thyroid gland. Still further confirmation is furnished by Dohrn's discovery in the torpedo embryo (Studien, VIII, p. 60) of two endodermal grooves which run from the ventral margins of the spiracles to the ventral middle line of the pharynx, to end at the median unpaired thyroid invagination in such a way as to prove that they are rudimentary peripharyngeal grooves. This most remarkable homology can no longer be questioned. The simplest explanation, and the one which first presents itself, is the one which Miiller advances, that the common ancestor of the tunicates and of the other chordata, possessed this system of organs in the form in which we now find it in the tunicates, and that while all the jawed verte- brates have inherited the ventral pharyngeal gland, it has been turned in them to some new use, as yet undiscovered by the physiologists, and has lost its primitive connection with the pharynx and its functional relation to the mouth, and has become a ductless aggregation of follicles far back in the throat. W. K. BEOOES ON THE GENUS SALPA. 197 I have tried to show that the structure and anatomical relations of this system of organs in the tunicates are quite consistent with the view that it was originally acquired for the purpose which it now serves, the capture of food. The simplest explanation of its origin is that which attributes it to the preservation by natural selection of a long series of slight changes, each of which improved the adaptation to the simple conditions of primi- tive pelagic life. Dohrn disputes this position, and says that "many persons would have great difficulty in believing that this simple mechanism is primi- tive" (Studien, VIII, p. 62). The future must show how many of these persons there are, but I shall now lay before them Dohrn's own explana- tion, that they make comparisons for themselves. "We ask," he says (p. 62), "how the ancestors of the tunicates obtained their food before the endostyle was formed. Obviously they were free swimming animals, and therefore in the position to seize their food by hunting. It is as certain that they needed other contrivances than the ciliated furrows and the slime-gland, as it is that the ancestors of the cirripeds sought their food in some other way than by the forma- tion of little vortices to sweep into their mouths everything within their influence. The limbs of the swimming forefathers of the cirripeds were certainly different from the cirri of modern barnacles ; even so were the ancestors of the tunicates differently constructed from the modern ones, and before the slime-gland and the ciliated grooves became the exclusive means of nutrition, they must have been the accessory aids to some more primitive mode of capturing food "... "Ammocoetes lives in the sand, into which even the youngest larvaB bore. Although direct observations fail, it must be assumed that the excretion of slime and the ciliation have some advantage in the nutritive or respiratory functions of organisms which live in the mud. May we not believe that, in spite of all the sifting through the oral tentacles and the velum, the hard particles of sand would be injurious to the delicate epithelium of the gut, if this were not protected by a thick coating of slime ; that the ciliated furrows are adapted for conveying this slime to the most exposed parts, and that, in this function, they have their start- ing-point ? Once brought into existence, it is not remarkable to see these useful structures further evolved until the whole mass of food is invested with a slimy admixture to facilitate its passage through the gut. It is not impossible that besides acting mechanically as an investment, the 198 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. slime also acts chemically as an aid to digestion. If this is the case, it is easy to understand how a peculiarity so useful to sedentary animals like the ascidians, or to floating ones like the salpae, gradually assumed the whole function of nutrition. Thus the problem of the change of func- tion is solved." Although it seems as if the delicate walls of the gut of a burrowing animal would be more effectively protected if slime were directly excreted " upon the most exposed spots," than by this highly specialized system of organs, we might yet believe that the system " has its starting-point " in the habits of ammocoetes, if we did not find in the structure and embry- ology of every chordata animal which is known to exist evidence of descent from an ancestor in which it had attained, not a starting-point merely, but its full development. The ontogenetic evidence that the vertebrate thyroid body was at one time a pharyngeal gland opening just within the mouth, and the dis- covery by Dohrn of rudimentary peripharyngeal grooves in the torpedo embryo (Studien, etc., VIII, Plate H, Figs. 7f, 7g, 7h and 7i), seem to me to be convincing proofs that the organs did not have their starting-point in the habits of ammocoetes nor in any degenerated fish, but that they arose in a lineal ancestor of the selachians and of the higher vertebrates, which was also an ancestor of the tunicates and cyclostomes. Passing now from the biological relations of the system of the endo- style to its homologies, we are told by Dohrn that it is equivalent to two pairs of gill-slits ; that these gill-slits were present and functional in the fish-like ancestors of the cyclostomes and tunicates, and that two of them, the mandibular clefts, moved downwards and met on the ventral middle line to form the thyroid gland or endostyle, while the endodermal portions of the others, the spiracular clefts, lost their connection with the exterior and became converted into the peripharyngeal grooves (Studien, etc., VII and VIII). Homologies are expressions of genetic relationship, and Dohrn tells us (p. 79) that they are valuable in phylogeny only as they furnish us with the opportunity to pass from the consideration of the structure of organs as they now exist, and of the functions of these organs at the present time, to the consideration of conditions which have passed away ; to the study of the history of the modifications which have come between these structures and functions, and those which we must attribute to the same organs at an earlier genealogical stage. I regard the structures which we find in the tunicates and in ammo- W. K. BROOKS ON THE GENUS SALPA. 199 coetes as primitive, and as homologous with those which we find in the jawed vertebrates ; and I have tried to trace the history of the modifica- tions which have come between these structures of modern vertebrates and those which we must attribute to the same organs at an earlier genealogical stage in the primitive history of the ancestral pelagic chor- data. The reader must judge of my success. Let us now see what light Dohrn's homology throws on the history of these primitive modifications. He tells us (Studien, etc., VII, p. 47) that he will point out, further on, the significance of the changes which have led to the fusion, on the middle line, of structures which were originally paired ; but I have been able to find nothing more upon this point except the acknowledgment, on page 63, that "I frankly admit that I have at present no available argument to bring the peculiar organization (of the ciliated grooves) of ammoccetes from a pair of imperforated (spiracular) gill-slits, into accordance with the concept of change of function ; and that the origin of the slime-gland of ammo- ccetes from two ventrally fused (mandibular) gill-slits must for the present remain an unsolved problem." Whatever may be thought of my own view, it must be admitted that Dohrn's homology of the endostylic system with two pairs of gill-slits has very little phylogenetic value, even when measured by his own test : the opportunity it furnishes for passing from the structure and functions of modern organs to the history of earlier genealogical stages. Dohrn's memoirs upon the thyroid body are full of interesting anatomical details, such as the similarity between the thyroid body of the shark embryo and the true gill-slits, in their relations to the cartil- ages, to the muscles and to the blood-supply (VII, p. 44) ; and the resemblance between the peripharyngeal grooves of ammoccetes and the spiracular gills of selachians (VIII, p. 55) ; but as he admits that the annelidian hypothesis leaves the origin of the endostylic structures of tunicates an unsolved problem, our subject, the history of the tuni- cates, does not require us to enter into the discussion of these complicated details of vertebrate morphology. The considerations which I have presented will undoubtedly be met by the assertion that while the simple and direct origin of the tunicates seems plausible so long as we confine ourselves to these animals alone, such a restricted view is unscientific. I shall no doubt be told that we are forced by more fundamental evidence to believe that the body cavity of the chordata is, in ultimate analysis, a segmented enteroccel formed 200 JOHNS HOPKINS UNIVERSITY MORPHOLOGICAL MONOGRAPHS. from a series of pairs of gut-pouches, and that the simplicity of appendi- cularia cannot be primitive, inasmuch as the ancestors of the tunicates once possessed these complicated structures. The first step to take in discussing this objection is to learn whether there are any traces of gut-pouches in the tunicates. Seeliger (p. 9) has given us a very minute and detailed account of the history of the mesoderm in the clavelina embryo, and has shown that it arises from two rows of endoderm cells which give origin, in the tail, to the caudal muscles and, in the body, to free mesoderm cells which multiply with great rapidity and wander everywhere through the body cavity, which is bounded on one side by the endodermal wall of the gut, and on the other by the ectoderm. He says emphatically (p. 128) that the mesoderm arises as two totally unsegmented rows of cells, each forming a single layer ; that the body cavity is not an enterocosl, but a primary body cavity; and that the ontogeny of the tunicate mesoderm gives no evidence of derivation from paired pouches comparable to the coelomic pouches of amphioxus. It is a rare thing for students of tunicate morphology to agree, but in this case the phenomena are simple, and Davidoff (p. 16) completely confirms Seeliger's observations, so far as they bear upon the question, by his own studies of clavelina and distaplia. His account of the origin of the mesoderm differs from Seeliger's in only one minor point, which has no bearing upon the question under consideration. Like Seeliger, he derives the mesoderm from two rows of endoderm cells, but he says that these cells remain as endoderm cells after they have given rise to the mesoderm, while Seeliger states that they become converted into the mesoderm. In all other respects Davidoff s observations are a complete confir- mation of Seeliger's, for he says (600) that in distaplia the mesoderm of the caudal region persists as a solid rudiment and becomes the muscular layer of the tail, while elsewhere it breaks up into wandering mesenchyma cells. "It is to be particularly emphasized that in no part of the mesoderm is any trace of segmentation to be discovered, and that there is not the least indication of any cavity comparable to a myoccel. The embryonic history of the mesoderm of distaplia cannot be referred back in any way whatever to anything comparable to Hertwig's concep- tion of the enterocoelomata." Of clavelina he says (607) : " There is not even a transitory division of the mesoderm into a somatopleur and a splanchnopleur. Even where W. K. BROOKS ON THE GENUS SALPA. 201 the mesoderm is two-layered, so that a parietal and a visceral layer may be distinguished, there is no homology between these layers and the bounding walls of the co3lom of the enterocoelomata." After reviewing all the literature on the subject, he gives as the general result of his studies the statement (p. 622) that " the body cavity of the ascidians lies between the two primary germ layers and must be regarded as a blastocoel, which would be identical with the segmentation cavity if this were not temporarily obliterated during gastrulation by the contact of the ectoderm cells and endoderm cells." "While the salpa-embryo is very complicated and unfavorable for studying this question, my own observations, which have already been described, seem to show that the body cavity of salpa is, like that of clavelina and distaplia, a primary one, fundamentally identical with the segmentation cavity, and that the mesoderm arises as free mesen- chyma cells derived from the endodermal blastomeres. The body cavity of the salpa-embryo, Plate XXXV, 75, is identical with the space between the somatic and visceral layers of follicle cells, Plate XII, 15, and while there is a stage in which these two layers are in contact, Plate X, Fig. 9, the follicular cavity is undoubtedly the same as the cavity shown at 75 in Plate XI, Fig. 3, and this is the same as the space which is shown in Plate X, Fig. 3, between the segmenting egg and the follicle. In the chapter on the significance of the salpa-embryo I have given my reasons for believing that this space is homologous with the segmen- tation cavity of more normal tunicate embryos, and if this view be correct the body cavity of salpa is not an enterocoel but a primary body cavity or blastocoel. The mesoderm of salpa consists of free migrating cells, and the chamber of the heart is part of the body cavity, so that these cells pass through it ; and while salpa is a peculiarly unfavorable subject, my observations are in complete accord with those which Seeliger and Davidoff have made under simpler and more favorable conditions. No student of the embryology of tunicates has ever described any trace of a series of body cavities, and Kowalevsky, the discoverer of the coelomic pouches of amphioxus, failed to find anything comparable to them in the tunicates, although the existence of a single pair of entero- coels has been claimed by certain observers. Van Beneden and Julin (Zool. Anzeiger, 4, 1881 ; Bull. Acad. Belg. (3) 7. 1884 ; Arch. Biol. 6, 1884) believe that the anterior portion of the body cavity of ascidians arises as a pair of gut-pouches, and that its mesoderm consists of a somatopleur 202 JOHNS HOPKINS UNIVEESITY MORPHOLOGICAL MONOGRAPHS. and a splanchnopleur, but Davidoff has shown by careful serial sections that this statement is probably based upon erroneous observations. Salensky holds (17, 460) that the mesoderm of the blastoderm of pyrosoma consists of two symmetrically placed ccelomic pouches, and that pyrosoma is, therefore, to be placed among the true enterocoelomata. The space between the vertebrate blastoderm and the yolk is undoubtedly homologous with the enteron, but it is by no means certain that this is the case in pyrosoma, where the food-yolk is an independent acquisition ; nor do Salensky's figures show, as clearly as we might wish, that the two coslomic vesicles open into this space, and even if this is the case, we must remember that the pyrosoma-embryo is very aberrant, and that the structure of its body cavity may be a secondary adaptation to the presence of the yolk. Taken alone it certainly is not enough to prove, without corroboration from other sources, that the body cavity of the tunicate is an enterocoel. The ontogeny and homology of the tunicate mesoderm have been recently discussed at very great length by Seeliger (11, pp. 85-104 and pp. 126-131), by Davidoff (16, pp. 592-628), and by Salensky (17, pp. 456-462 and pp. 468-470 and 36^6), and as those who wish can find in these papers an extended presentation of the complicated and perplexing theory (?) of the mesoderm, I have attempted to treat it very briefly. The literature shows that there is no direct evidence whatever of the existence, at any time in the history of the tunicates, of a metameric series of coelomic pouches, and the supposed necessity for believing that such a series existed in the primitive chordata is only another aspect of the dogma that the metamerism of the vertebrates must have been inherited from a primitive metameric ancestor. If, as I believe, the metamerism of vertebrates is secondary, the metamerism of the mesoderm and body cavity may have resulted from the duplication of a single pair of coelomic pouches similar to those of echinoderm larvae, and it is quite conceivable that these may have been acquired by the ancestors of the vertebrates after the divergence of the tunicates. If, however, future research should show that there is a pair of gut- pouches in the embryo of appendicularia, or should prove in some other way that the structures which Salensky describes are true enterocosls inherited from an ancestral tunicate, such a discovery, which is certainly among the possibilities, would be no evidence that the primitive tunicate was the degenerated descendant of an ancestor with metameric gut- pouches. W. K. BEOOKS ON THE GENUS SALPA. 203 At present, however, the evidence all tends to show that the ancestors of the tunicates had no such structure, and that the presence of coelomic vesicles in pyrosoma is an adaptation to its peculiar mode of development. Since the critical review of the literature of the embryology of salpa will require all tJie space which remains, I must bring this theoretical discussion to an end, and must reserve, for publication at another time, the sections to which I have referred, on the pelagic origin of the veiled medusae, and on the morphological significance of metamerism and other vegetative duplications in animals. PART THREE. A CRITICAL DISCUSSION OF MY OWN OBSERVATIONS AND THOSE OF OTHER WRITERS, ON THE SEXUAL AND THE ASEXUAL DEVELOPMENT OF SALPA. CHAPTERS IX, X, XI, XII, XIII, AND XIV. CHAPTER IX. THE ORIGIN AND MATURATION OF THE EGG OF SALPA. SECTION 1. The Embryonic Germ Cells. Many names have been proposed by various authors for the mass of cells, Plate XXI, m, n, and Plate XXXIV, m, n, and Plate XLI, Fig. 7, n, which runs along the haemal region of the stolon, and gives rise to the reproductive organs of the chain-salpa; but a technical term does not seem necessary, and I shall call it the germinal mass. It makes its appearance very early in the life of the embryo, in the form of a sharply defined, compact, subspherical mass of cells, which at first differ very slightly from the other cells of the germ layers of the body of the embryo, although they are, as we shall see, quite different from the follicle cells, which are so numerous during the embryonic stages. In Salpa pinnata, the species in which I have studied it most thoroughly, it is on the middle line of the haemal side of the body at n in Plate XXXV, and between the placenta y" and the eleoblast k. It marks the point where the