On the Origin, Growth, and Arrangement of Sponge-Spicules: a Study in Symbiosis

On the Origin, Growth, and arrangement of Sponge-spicules: A study in Symbiosis.

By Arthur Dendy, D.Sc, F.R.S., Late Professor of Zoology in the University of London and Fellow of King's College.

With Plates 1, 2, and 3, and 16 Text-figures.

CONTENTS. PAGE 1. INTRODUCTORY REMARKS ...... 2 2. SPEGIAL OBSERVATIONS ...... 0 (d) The Stephanotyles of Sceptrospongia coronata . 0 (b) The suppressed Triaenes of Stelletta haeckeli . ft (c) The Formation of Arnphitriaenes by incomplete Twinning 12 {d) The Development of adventitious Rays . . . .IS (e) The Formation of nodal Whorls in Discorhabds . . 18 (/) A revised Account of the Development of the Anisodisco- rhabd in Latrunculia bocagei .... 27 3. GENERAL CONCLUSIONS AS TO THE ORIGIN AND GROWTH OF SILICEOUS SPONGE-SPICULES . .... 36 (a) The primary co-operating Agents . . . . . 3(> (6) The Influence of the primary Protorhabd upon the Form of the Spicule 3!> (c) The Influence of the Mode of Division of the original Scleroplastid ...... 41 (a) The Formation of radiate Spicules . . .41 (|S) Variation in the number of primary Rays . . 43 (y) The Formation of Dragmata and Rosettes . . 44 (d) The Influence of adventitious Growing Points upon the Form of the Spicule 40 (a) Adventitious Growing Points associated with Protorhabds 40 (|3) Adventitious Growing Points without Protorhabd Formation ...... 47 NO. 277 B 2 ARTHUR DENDY

PAGE (e) Mechanical Influences affecting the Form of the Spicule . 49 (n) The Conversion of dermal Spicules into Plates . 49 (j9) The Mechanical Influence of the so-called Mother- cell 50 (y) Other Mechanical Influences . . . . 51 4. COMPARISON OF THE DEVELOPMENT OF CALCAREOUS SPONGE- SPICULES 52

5. COMPARISON or THE SPONGIN SPICULES OF DARWINELLA . . 53 6. CONSIDERATIONS AS TO THE NATURE OF THE SCLEROPLASTIDS . 54 (a) Their microscopical Characters, Mode of Growth, and Multi- plication ...... 54 (6) Their Specificity ...... 5.r» (c) The Occurrence of trimorphic Chelae and its Significance . 50 (d) The Want of Correlation between the Spicules and the Sponge 57 (e) The Dropping out of Spicule Categories in Sponge Phylo- geny ...... 58 (/) The sporadic Distribution of certain Spicule Types . . 60 (g) The Scleroplastids as symbiotic Organisms (Sclerocoeci) . 01 7. THE RELATIONS BETWEEN THE SOLEROCOCCI AND THE SPONGE . 02 8. THE TAXONOMIC VALUE OF SFONGE-SPICULES . . .07 9. LIST OF LITERATURE REFERRED TO 09 10. DESCRIPTION OF PLATES ...... 72

1. INTEODUCTOHY KEMAHKS. THERE are perhaps no structures met with in the animal kingdom of which the origin and significance are more difficult to explain than the spicules that form the skeleton of calcareous and siliceous sponges. The larger and usually less complex forms, technically known as megascleres, we are apt to accept as necessary constituents of the skeleton specially adapted to their position in the sponge, without stopping to reflect that there are many sponges which manage to get on very well without any spicules at all, sometimes, indeed, without skeleton of any kind. The smaller ones, or microscleres, met with amongst the siliceous sponges, are much more enig- matical, for they usually occur scattered promiscuously throughout the soft tissues, while they exhibit the most surprising variety SPONGE-SPICULES 3 of exquisitely symmetrical forms, the meaning of which it is impossible to interpret in terms of the physiological requirements of the sponge. At the same time these forms, as well as those of the megascleres, while exhibiting a remarkable degree of specific constancy, can be arranged in beautifully graduated series that leave little doubt in the mind of the trained observer that they owe their existence to some evolutionary process. The evolution of many of these series has been discussed in some detail in my memoir (1921 a) on ' The Tetraxonid Sponge- spicule : a Study in Evolution ', to which I would refer the reader who is unacquainted with the subject for descriptions and illustrations of many of the more remarkable types. I must, however, point out that the illustrations in that paper do very scant justice to the extraordinary beauty of the forms described. Much more artistic and at the same time more realistic representations of sponge-spicules are to be found in numerous monographs that deal systematically with this group of the animal kingdom, the best being perhaps those given by Schulze in his various memoirs on the Hexactinellida (1887 c, 1894 6, 1895, 1899 a, 1904, &c.) and by Lundbeck (1902, 1905, 1910) in his reports on the Porifera of the Danish Ingolf Expedition. I would refer the reader to the beautiful plates accompanying these memoirs, and to those illustrating my own reports on the sponges of the ' Sealark ' Expedition (1916 b, c ; 1921 b) and of the ' Terra Nova ' collection (1924 a) for some idea of the nature of the problem before us. Various attempts have been made by different investigators to trace the individual development of certain types of sponge- spicule. Perhaps the best known of these are Minchin's classical observations on the development of calcareous spicules (1898, 1908), to which I shall have occasion to refer later on. In the meantime I may venture to say that these observations seem to me to stop just at the most critical point. As regards siliceous spicules, the few observers who have paid much attention to the subject have usually been content to say that they originate within ' mother-cells ', and to give some details as to their growth stages. The so-called mother-cell has served B 2 4 ARTHUR DENDY as a convenient shelf on which to lay aside the problem of the origin of the spicule (compare Woodland, 1908). In 1917, in my Presidential Address to the Quekett Micro- scopical Club, I put forward certain views as to the origin and growth of siliceous spicules which formed the starting-point from which those enunciated in the present memoir have gradually developed. I pointed out ' that three factors may be concerned in the growth of a siliceous sponge-spicule : (1) the protorhabd, which is responsible for growth in length and serves as a foundation upon which silica is deposited ; (2) the formative cells, which are responsible for the arrangement of the silica ; and (3) the accessory silicoblasts, which supply the formative cells with the necessary material.' The name ' protorhabd ' was coined by me for the ' axial thread ', which had long been known to occupy the ' axial canal' of the spicule, regarded from the developmental point of view. The so-called formative cells were very problematical entities, which had been observed only in the case of L atrun- culia bocagei. In my memoir on the tetraxonid sponge- spicule (1921 a) I pointed out that although they might pro- visionally be spoken of as ' initial cells ' their real nature was somewhat doubtful, the condition of the material precluding exact cytological observations. According to the views to be elaborated in the following pages they are not to be regarded as themselves cells, or even as representing true cells, but as minute growing points, associated with the presence of still more minute granules, which seem to form the starting-points in spicule development. If we regard these granules merely from the point of view of their functioji as spicule initiators, we may call them ' scleroplastids ', on the analogy of chloro- plastids and other similar bodies in the vegetable kingdom, but if, as seems to me probable, they are really symbiotic organisms, then, from this point of vieAv, the name ' sclerococci' would seem to be preferable. Whatever we call them, they must be recognized as the foundations (usually elongated into protorhabds) around which silica is deposited by fchd silicoblasts of the sponge. These silicoblasts, temporarily or SPONGE-SPICULES 0 permanently enveloping the growing spicule, are the ' mpther- oells ' of earlier observers. The scleroplastids or sclerococci are so extremely minute, being of the same order of magnitude as bacteria, that it is difficult, if not impossible, to recognize them before a certain amount of silica has been deposited around them, for aa yet I know of no means by which they could be distinguished from

TEXT-FIG. 1.

r---

Young anisodiscorhabd of Latrunculia apicalis with projecting protorhabd (pr.). x 1,187.

ordinary bacteria and other granules that occur abundantly in the mesogloea. When they have elongated to form protorhabds, however, they are more readily detected, though even then it is doubtful if they have ever been seen absolutely naked, without any silica deposit. Apparently they are repeatedly enveloped (more or less completely) by the phagocytic silicoblasts, which usually, after depositing their silica, again leave them lying free in the mesogloea. The actual existence of the protorhabd as an independently growing entity is clearly demonstrated in Text-fig. 1, which represents a young anisodiscorhabd of Latruncnlia a pi- 6 ARTHUK DENDY calis, with the protorhabd projecting freely from the axial canal at the apex of the spicule. The protorhabd stains freely with such reagents as borax carmine and brazilin, and may then often be detected as a very slender, darkly stained thread, lying in the axial canal of the young spicule. The evidence that it is derived from a scleroplastid, and that the latter is a quasi- independent organism, capable of moving from place to place, multiplying by fission, and perhaps sometimes undergoing sexual union, will be set forth in the following pages. Much of this evidence is now published for the first time, but, in order to make the story as complete as possible, I have ventured to recapitulate certain facts, and to reproduce certain illustrations, already published in the papers referred to above.

2. SPECIAL OBSERVATIONS. (n) The Stephanotyles of Sceptrospongia coronata. In the genus Raphidotheca, amongst the Esperellinae, a very curious modification of the ordinary tylostylote megasclere, such as we find in Mycale and Esperella, gives rise to the exotyle. This is a tylostyle swollen out into a knob at the apex as well as at the base. Indeed the apical enlargement is much the more conspicuous of the two and may be ornamented with papillae or even take the form of a hollow cup (in Topsent's Gomphostegia (1896 c), a synonym of Baphidotheca). These exotyles are surface spicules, formed by modification of the outer ends of individual spicules in a more or less dense ' pile ' of radially arranged tylostyles or subtylostyles. If we examine carefully the exotyles of Eaphidotheca marshall-halli, the type of the genus, we shall find that the axial thread ends in the middle of the large apical knob, without any sign of branching, being surrounded by uninterrupted concentric laminae of silica (fig. 9, PI. 1). In the course of my examination of some deep-water sponges from the Indian Ocean, belonging to the Calcutta Museum, I have come across two specimens of a remarkable sponge closely SPONGE-SPICULBS 7 related to Raphidotheca, which I propose to describe in my report on those sponges under the name Sceptrospongia ooronata, there being, in my opinion, sufficient difference to merit generic as well as specific recognition. In this species the exotyles are replaced by very peculiar spicules which I propose to call ' stephanotyles ', occurring in the surface pile exactly as do the exotyles in Raphidotheca. In one specimen they are extraordinarily abundant, and though they may be regarded, in a sense, as abnormal forms of the tylostyle (compare figs. 1 and 2, PI. 1), yet we must recognize them as normal and constant features of the spiculation. They differ widely from the exotyles of Raphidotheca in the very peculiar behaviour of the axial thread (protorhabd), which results in the formation of a very remarkable crown of spines around the apical knob, each spine containing a branch of the axial thread. In uneroded spicules the axial thread is, as usual, extremely slender and difficult to follow, but when erosion has taken place, so that the axial thread is represented by relatively wide axial canal, this latter may be very conspicuous. Various degrees of erosion are represented in the illustrations. The manner in which the fully developed stephanotyle (fig. 8, PI. 1) has been, so to speak, arrived at, is clearly indicated by the series of variations met with in the number of spines, from only two, to as many as forty or fifty (figs. 4-8, PI. 1). The normal tylostyle, the apex of which is shown in fig. 3, PI. 1, forms the commencement of the series. In fig. 4, PI. 1, the axial thread has simply bifurcated and one branch runs out into each of two small spines. At the same time the apex of the spicule as a whole becomes enlarged and acuminate. Fig. 5, PL 1, represents a case in which the axial thread has divided into three branches, and there are probably three small spines. In fig. 6, PL 1, there are six branches of the axial thread and six spines, and now the crown-like character is becoming evident, the end of the spicule as a whole having enlarged to form an apical knob. The branches of the axial thread now at first radiate outwards from their point of origin at right angles to the long axis of the spicule ; they then turn 0 ARTHUR DENDY forwards and run, almost parallel with one another, into the spines. Fig. 7, PI. 1, shows a well-developed crown of about a dozen spines, each with its branch of the axial thread. Fig. 8, PI. 1, shows the highest stage of development of this very curious type of spicule ; there are in this case some forty or fifty spines, each with its axial thread (or canal) and all united together, except at their apices, to form a kind of fringe around the apical knob, which has now become fairly large. That the branch axial canals in the more or less eroded spieules really represent axial threads is shown by the fact that in some cases, after the spicules have been roasted on a hot plate, the charred and brown remnants of the thread can actually be seen. Occasionally the erosion extends in between the silica laminae from the branch axial canals for a short distance, giving rise to the appearance of short backward pro- longations of the canals, but these are of no importance. The reason why I have brought this case forward so promi- nently is that it seems to me to demonstrate, hardly less con- clusively than the case represented in Text-fig. 1, the character of the axial thread as a living and growing structure. Why it behaves in this very exceptional and extraordinary manner it is impossible to say. It may be an expression of the conflict between the protorhabd or axial thread, endeavouring to continue its growth, and the silicoblasts, endeavouring, so to speak, to seal up its end in silica. When these two activities are normally balanced we get a normal tylostyle with tapering apex and unbranched axial thread (figs. 1, 3, PI. 1). When the silicoblasts gain a complete mastery they form an apical knob in which the axial thread remains unbranched, as in the oxotyle of Eaphidotheca mar shall- h a 11 i (fig. 9, PI. 1). The branching of the protorhabd, resulting in the formation of the crown of spines, may represent a final effort to avoid being sealed up. All this, however, is mere speculation, the important thing is the demonstration of the activity of the protorhabd, to whatever course that activity may be due. I know no other case in which this phenomenon of terminal SPONGE-SPICULES 9 branching of the protorhabd is manifested to anything like the same extent, but it is worth noting that the same kind of thing occurs in the subtylostyles of Paresperella bidentata (Dendy, 1905), though here the process seems to be limited to the formation of two spines. In D e n d r o p s i s b i d e n t i f e r a (Eidley and Dendy, 1887), again, something very similar occurs at the base of certain of the styli, but again limited, at any rate usually, to the formation of two spines.

(b) The suppressed Triaenes of Stelletta h a e c k e 1 i. In my report on the sponges collected by Professor Herdman in Ceylon (1905) I described very briefly and with very inadequate illustrations a number of abnormal spicules evidently derived from ordinary triaenes by suppression of the developing rays at various stages. I have since found very numerous and very beautiful examples of the same phenomenon in another specimen of this species obtained by Mr. Hornell at Okha- mandal,1 and it is upon this latter material that the following observations are based. The normal megascleres of this sponge are shown in figs. 20-2, PI. 1. They are anatriaenes (fig. 20), orthotriaenes (fig. 22), and oxea (fig. 21). The abnormal spicules represented in figs. 10-16, PI. 1, are mainly, if not exclusively, more or less completely suppressed anatriaenes, though it is impossible to say with certainty, in extreme cases of reduction, that we may not be concerned with some other form of megasclere. A similar suppression of the cladi has, indeed, been often observed in the case of the orthotriaenes but, curiously enough, the shaft of the orthotriaene appears very seldom to undergo suppression. The normal form of the anatriaene, which is still by far the most abundant, has nothing at all peculiar about it (fig. 20, PL 1). In the suppressed forms, which occur scattered in the

1 This specimen is mentioned in my ' Report on the Okhamandal Sponges (Dendy, 1916 a) as P.N. iv. 3, and the specimen is so labelled. 10 ARTHUR DENDY interior of the sponge, the shaft seems always to undergo reduction. In association with the reduced shaft we may have a perfect cladome (fig. 10, PI. 1), or one, two, or all three of the cladi may be suppressed (figs. 11-15, PI. 1). The suppression consists in the rounding off and shortening of the ray concerned, without diminution in thickness. In all cases that I have observed the axial canals (representing the protorhabds) remain distinct, and it is significant that they do not extend to the surface of the knob that represents the suppressed ray, as they do to the apex of a perfect ray, but are separated from it by a great thickness of laminated silica. This shows clearly that as soon as the protorhabds stopped elongating, the silica spread over its apex, or possibly the spreading of the silica over its apex was the cause of the cessation of the growth of the protorhabd. It looks almost like a race between protorhabd growth and silica deposition. Only if the two are exactly proportioned will the ray attain its full length and characteristic tapering form, with protorhabd extending to the very apex. When all four rays have been reduced to low, broadly rounded protuberances (figs. 13-15, PI. 1) the axial canals in the middle of the spicule still show a perfect tetract arrangement, with the shaft longer than the three equal cladi. This arrangement is still seen in fig. 14, PI. 1, where, externally, the cladi are no longer distinguishable. In fig. 15, PI. 1, however, there is very little difference in length between shaft and cladi and the axial canals have retained almost the form of the primitive tetract. Finally (fig. 16, PI. 1), we reach the last stage of suppression, in which the entire spicule takes the form of a perfect sphere of concentrically laminated silica, with a minute granule, about 0-0015 mm. in diameter (or a group of granules), in the centre, representing a scleroplastid or sclerococcus (or a group of such), and no axial canals at all. Such 'silica pearls', as Schulze (1893&) has termed them, are by no means uncommon in siliceous sponges. Schulze observed them in certain hexactinellids (e.g. Pheronema giganteum) and was much puzzled by them. Sollas (1888a) SPONGE-SPICULBS 11 regarded them as microscleres and termed them globules or spherules. In Gamin us sphaeroconia he considers them as normal constituents of the spiculation. Further examples are described by various writers; e.g. by Weltner (1901) in a fresh-water sponge, by Kirkpatrick (1908 c) in Cinachyra bar bat a (where they were also seen by Sollas), and particularly by Lebwohl (1914 a) in Tetilla, Characella, Sphinctrella, Pachastrella, and Yodomia ; but none of these observers seem to have found a well-graduated and regular series of reduction forms such as occurs in Stelletta haeckeli, nor to have suspected the nature of the central granule. It is obvious that the forms above described, in the case, of Stelletta haeckeli, represent progressive stages in the suppression of the rays of the tetract (triaene). The growth of the protorhabd has been stopped in each case by the over- whelming deposition of silica or by some other agency, and we thus have preserved to us a series of stages in the early development of the protorhabds (represented by the axial threads or canals) which, on account of their minute size and transient character, it would be extremely difficult, if not impossible, to find in the case of a normally developing spicule. Eeading the series in the natural order we should start, of course, with the scleroplastid (preserved as a sort of fossil in the centre of the siliceous pearl), from which (under normal conditions) the four rays of the tetract gradually develop. It seems to me that the evidence for the existence of an initial scleroplastid is conclusive, but how the scleroplastid comes to give rise to four elongating protorhabds, arranged as the axes of a tetrahedron, is not obvious. The observations to be described in the following sections, however, seem to me to indicate that the original scleroplastid divides into four, arranged in the form of a pyramid, and that each of these elongates outwards to form the protorhabd of one ray. The evidence for this is certainly indirect, but to my mind it is very convincing, for no other explanation of the observed facts seems to be forthcoming. 12 ARTHUR BENDY

(c) The Formation of Amphitriaenes by . incomplete Twinning. The amphitriaene, consisting of a shaft with three cladi at each end, is a type of spicule very rarely met with, and it may perhaps be doubted whether it is ever a normal constituent of the sponge skeleton. I have found four examples of this type in the Okhamandal specimen of Stelletta haeckeli referred to above. In all four the shaft is very short and some of the cladi are suppressed. All are evidently derived from anatriaenes and the cladi of the two ends point in opposite directions, those of one end towards those of the other. In the case represented in fig. 19, PI. 1, there are two fully developed cladi at one end and only one at the other, and the cladi are longer than the shaft. Fig. 18, PI. 1, represents a specimen with three cladi at one end and one at the other, and the shaft is rather longer than the cladi; while in fig. 17, PI. 1, is shown another specimen with a very short shaft bearing three cladi at one end and two at the other. Such forms might be regarded as being derived from a normal triaene by branching of an abbreviated shaft at each end, did we not know with a fair amount of certainty that the shaft and clacli of the normal triaene arise simultaneously and grow from a common centre. They can, however, be readily explained, on what we may call the seleroplastid hypothesis, as examples of incomplete twinning, in which the twins never separate. (We shall have to consider an example of complete twinning later on, in the case of Latrunculia bocagei.) Suppose that a seleroplastid (Text-fig. 2, A) divides into two (B) and then again into four (C), which arrange themselves in a pyramid. One of the four elongates to form the protorhabd of the shaft, but the other three divide again to form two groups of three (D), one group being associated with each end of the shaft and giving rise to the protorhabds of the cladi. Thip process would give rise to a perfect amphitriaene with all the six cladi developed, but if some of the daughter scleroplastids SPONGK-SPICULES 13 failed to elongate into protorhabds we should get forms with suppressed cladi, such as those actually met with. We have here an explanation of the formation of amphitriaenes which seems to fit in completely with the observed facts and which also harmonizes very well with the facts to be described in the following section.

(d) The Development of adventitious Rays. I shall now endeavour to prove that the protorhabd of each ray of the tetract spicule is formed by elongation of a separate scleroplastid by demonstrating that these protorhabds may

TES,T-HG. 2.

Diagram of hypothetical divisions of the scleroplastid in the formation of an amphitriaene. sometimes become dislocated instead of developing in their normal positions or in their normal associations with one another, and may even associate themselves in abnormal positions with some other protorhabd to which they do not properly belong, so as to give rise to spicules with adventitious rays. Instances of this kind are to be found amongst the megascleres of Stelletta haeckeli. Take, for example, what we may perhaps call the reversed anatriaene represented in fig. 23, PI. 1. Here the shaft of the spicule appears to be normally developed, greatly elongated and tapering gradually to a fine point at the apex. The three cladi, however, while retaining their normal shape, are separated from the base of the shaft and also reversed in position, so that they curve towards the base instead of towards the apex. Another possible explanation of the abnormal position of the 14 ARTHUR DENDY cladi in this case may be that the protorhabd of the shaft has elongated in both directions instead of in one only, and that the basal elongation has been abruptly stopped. Triaenes with the shaft elongated in two opposite directions are well known, for example, amongst the protriaenes of certain species of the genus Thenea, and are usually spoken of as mesotriaenes. This explanation, however, will not account for the reversal of the cladi, which seems to point to a re-orientation of scleroplastids. Take next the anatriaene represented in fig. 24, PL 1. Here we have a normally developed cladome and a normally developed shaft, but a short way below the normal cladome there is an attempt at a second, with only one fully developed cladus, and that in a reversed position. The two other adventitious cladi are represented merely by slight knobs on the shaft. In this case we meet with a feature which is very generally associated with the development of adventitious cladi, the axial canals of the latter, as they approach that of the shaft, becoming broken up into a series of minute dot-like cavities, usually irregular. This appearance seems to be restricted to adventitious cladi, and is not met with in the abnormal amphitriaenes described above (figs. 17-19, PL 1). The development of adventitious cladi is not confined to the anatriaenes. Fig. 25, PL 1, represents the cladome and part of the shaft of an orthotriaene in which two of the cladi have been reduced to mere knobs. The third is normally developed except that it has attached to it an adventitious cladus, in the position shown. This appears to be a favourite position for the development of such an adventitious ray, and sometimes two make their appearance there, side by side. In the case figured the axial canal of the adventitious ray appears to be completely disconnected from that of the normal cladus, indicating that the scleroplastid settled upon the normal protorhabd after the latter had received its first thin coating of silica. In other cases observed the axial canal of the adventitious ray is joined to that of the normal cladus by a row of dot-like cavities, as described above. This latter feature is very SPONGE-SPICULES 15 well shown in fig. 26, PI. 1, representing an adventitious cladus attached near the base of an abnormal spicule which may be a reduced orthotriaene. We come now to cases in which adventitious cladi have developed upon spicules that are not triaenes and normally have no cladi at all. Large oxea (fig. 21, PI. 1), i. e. linear diact. megascleres pointed at each end, form, as we have said above, a normal constituent of the spiculation in the genus Stelletta. Sometimes these are reduced to the stylote condition by the more or less complete suppression of one of the two opposite rays of the diact. Such stylote spicules with adventitious, hook-shaped cladi, have been figured by Thiele (1900) in the case of Stellettn ternatensis. He interpreted them simply as abnormal anatriaenes, but I find that in similar cases observed by myself in Stelletta haeckeli the cladi may be developed on the rounded inner end of the radially orientated spicule, while in the triaenes, of course, the cladomes are always at the outer ends. The spicules in question are therefore to be regarded as reduced oxea. This view is strongly supported by a case which I have observed in which two adventitious cladi are attached at just about the middle of a large oxeote spicule in which both rays are normally developed. The middle portion of this spicule is represented in fig. 27, PI. 2. It will be seen that the two cladi point in opposite directions, and that one of them is somewhat hook-shaped, as though it, were the cladus of an anatriaene. This anatriaene-like character is much better shown in the cases figured by Thiele. It is also fairly well shown in the very irregular case represented in fig. 29, PI. 2, but it is impossible in this instance to be certain of the nature of the shaft, whether it is a reduced oxeote or otherwise. A point of considerable interest remains to be emphasized before we leave the question of adventitious cladi. Sometimes these cladi appear singly, as though a single, definitive cladus- forming scleroplastid had settled upon the shaft; at other times, though only one cladus may be fully developed, there are distinct traces of two others, as though a scleroplastid had 16 ARTHUR DBNDY settled on the shaft before dividing into the definitive cladal scleroplastids. Both these cases have been met with in the examples described above, but fig. 28, PI. 2, represents a portion of a spicule, probably an oxeote with one ray suppressed, in which both occur together, each in a very typical manner. In the lower part of the figure, near the base of the spicule, is shown a single fully developed cladus, but two others are represented by their short axial canals, imbedded in the shaft, and all three axial canals meet in a point that lies on the axial canal of the shaft. The suppressed cladi are further indicated by slight protuberances on the surface of the shaft and by the curvature of the concentric layers of silica. Note also, again, the dotted character of the axial canals of the adventitious cladi. In the upper part of the same figure is seen a single adventitious cladus, the axial canal of which, for the most part broken up into dot-like cavities, altogether fails to meet the axial canal of the shaft. In all the cases described above the adventitious rays appear to be the cladi of triaenes, as indicated by their size and form, and in some cases by their arrangement in threes. We also meet with cases, however, in which the adventitious ray is evidently not a cladus, but a shaft. Such an instance is represented in fig. 29 (lower part), PI. 2, in which an adventitious shaft is shown coming off at a sharp angle from the proper shaft. That it cannot be regarded merely as a branch of the latter is clearly indicated by the fact that the two axial canals cross one another and the adventitious shaft is continued as a very slight protuberance on the opposite side of the main shaft. Higher up in the same figure is shown the remains of an adventitious shaft, broken off short on both sides, that completely crosses the main shaft. At the lower end of the main shaft, near the apex of the same spicule, is seen a still more remarkable feature. For a short distance a secondary or accessory axial canal (s.c.) lies parallel and completely confluent with the normal axial canal, curving away from the latter, however, at its two ends. A corresponding thickening of the silica is seen on the surface of SPONGE-SPICULES 17

the spicule (figs. 29 and 29 a, PL 2). This can only mean that an adventitious protorhabd attached itself lengthwise to the normal protorhabd before the siliceous layers began to be deposited. The extent to which abnormal megascleres with adventitious rays are developed varies greatly in different species. Usually they are very few and far between, but they are fairly abundant in some specimens of Stelletta haeckeli, though they may require a good deal of looking for. The climax is reached, however, in a sponge described by Oscar Schmidt (1868 a) under the very appropriate name Stelletta pathologica, though it is perhaps doubtful whether it belongs to the genus Stelletta at all. It may be a Pachastrella. In this sponge all, or nearly all, the megascleres are abnormal and appear to be formed of rays that have come together like the members of Empedoclean monsters, Avithout any regard to the propriety of their union. . The example which I figure (fig. 81, PL 2), from a prepara- tion of the type specimen in the British Museum (Natural History), was selected because it shows, in addition to adventitious rays in all sorts of positions, a group of secondary axial canals (s.c), four in number, embedded in the silica of the main shaft. These must indicate adventitious protorhabds which associated themselves with the principal protorhabd, but it is curious that there is no corresponding thickening of the silica deposits, as in the case represented in figs. 30 and 30 a, PL 2. A very similar case of the formation of what he terms ' secondary canals ' has been beautifully figured by Oscar Schmidt (1868 a, Taf. Ill, fig. 2, A) in his CallitesLacazii. Enough evidence has perhaps been brought forward now to justify us in concluding that the rays of the tetract spicule, although normally developing in such close association with one another as to grow out from a common centre, are yet capable, under certain circumstances, of separating from their original associations and making new ones. But to say that the rays themselves do this is of course merely a figure of speech. The ray itself is an inert mass of silica, incapable of NO. 277 c 18 ARTHUR DENDY any activity. We must look to something that underlies the ray, as it were ; something from which the ray develops and which, before being fossilized in silica, behaves like a living organism. We must look, in short, to a protorhabd, and beyond that to a scleroplastid, capable of multiplying by division and of moving from place to place. Moreover, we can hardly avoid concluding that different scleroplasts have different potentialities of development, that one may develop into a cladus and another into a shaft, and that whether shaft or cladus is formed is not a mere question of position.

(e) The Formation of nodal Whorls in Discorhabds. In the discorhabd type of spicule, met with in the tetraxonid sub-family Spirastrellinae, we find a straight or curved shaft with whorls of outgrowths developed upon it in definite positions. In certain cases there is very good reason for believing that the position of these whorls is nodal, that is to say that, supposing the shaft to have been a vibrating rod at the time when the whorls were first laid down, these were formed in those situations where the rod exhibited least movement. The main point to be established here is that these whorls must have been initiated by scleroplastids which settled on the shaft of the developing spicule. Let us consider very briefly the principal types of discorhabd. The spicule is termed an isodiscorhabd when the two ends are symmetrically developed on either side of the median transverse plane. It is an anisodiscorhabd when the two ends are developed asymmetrically. It is also sometimes convenient to classify these spicules according to the original form of the shaft, before it has been modified by secondary deposition of silica ; thus we have the oxydiscorhabd in Avhich the shaft is oxeote or pointed at both ends, and the sigmodiscorhabd in which the two ends of the original shaft are sharply recurved, like the ends of an ordinary sigmoid microsclere. SPONGE-SPICULES 19 In the oxydiscorhabd of Barbozia primitiva (Dendy, 1921 b), which is also an isodiscorhabd, there are two whorls of sharp spines, each placed about half-way between the middle of the shaft and one of its sharply pointed extremities (Text-fig. 3, C). Here again we derive valuable evidence from an abnormal spicule. The megascleres in this sponge are also oxeote, but normally quite smooth, without any trace of whorls or spines. Text-fig. 3, D, however, represents a megasclere with two very irregular whorls of blunt protuberances developed in situations corresponding to those of the whorls of spines in the normal oxydiscorhabd, presumably nodal. I do not see how the peculiar abnormal condition of this spicule is to be explained except on the hypothesis that one or more scleroplastids have settled upon the developing shaft and multiplied to form two nodal groups. The irregularity of these groups as compared with the well-defined whorls of the normal oxydiscorhabd is probably to be connected with the fact that they have developed on a megasclere instead of on a microsclere, so that the shaft with which they are associated is several times larger than it would normally be. Take next the oxydiscorhabd of Didiscus placo- spongioides (Dendy, 1921 6), which formed the subject of a special memoir by Professor J. W. Nicholson and myself (Dendy and Nicholson, 1917). This is an anisodiscorhabd. with only two whorls, one about the middle and the other on one side of it (Text-fig. 3, A). As the result of a series of very elaborate mathematical calculations Professor Nicholson was able to show that the position of these two whorls upon the shaft, as determined at the critical point of development (Text-fig. 3, B), corresponds very remarkably with the theoretical position of two of the principal nodes. There must, of course, be another node on the other side of the central whorl, but no whorl is developed in this position, a fact which can only be explained by the absence of some co-operating factor—evidently the scleroplastids, the divisions of which are presumably only sufficient to form two whorls. We come now to the anisodiscorhabds of the genus Latrun- 02 20 ARTHUR DBNDY culia, which we may consider very briefly here, as I have already dealt with them on more than one occasion (1917, 1921 a), and propose to give a somewhat detailed account of their development in the next section of this paper.

TEXT-ITG. 3.

A, oxydisoorhabd of Didiscus plaoospongioides (fully- developed), x 1,065. B, ditto, at critical point of development, the whorls just appearing, x 1,065. C, oxydisoorhabd of Bar- bozia primitiva. x 1,095. D, abnormal megasclere of Barbozia primitiva, with two irregular whorls of outgrowths. X730.

In Latrunculia bocagei the fully developed anisodiscorhabd (fig. 32, PI. 2) consists of a straight shaft with a spiny ' manubrium ' (mn.) at one end (the base) and a crown of capitate spines at the other (the apex). Situate near the middle of the shaft is the ' median whorl' (m.io.), which is tripartite, consisting of three horizontally flattened outgrowths, SPONGB-SPICULBS 21 or lobes, with crenate margins. Above the median whorl comes the ' subsidiary whorl' (SM.), closely resembling the median whorl, but with the edges of the three lobes upturned. Above the subsidiary whorl, and immediately beneath the crown of spines, comes the ' apical whorl' (a.w.), which has a crenate margin but is not divided into lobes. It is formed from the ' apical knob ' of the young spicule, in which all these parts are more easily recognized (compare figs. 46, 47, PI. 3). In Latrunculia apicalis the amsodiscorhabd (fig. 33, PI. 2) exhibits the same structure except that the crown of short, knobbed spines is replaced by a single long, terminal spine, with a few small, lateral spines on its base. This striking difference is due to the fact that the protorhabd continues its growth in this species after the rudiments of the whorls and knobs have been laid down (Text-fig. 1), while in Latrun- culia bocagei it is stopped abruptly with the formation of the apical knob of silica (fig. 47, PI. 3). In Latrunculia bocagei the slender shaft of the young spicule very early becomes weighted at one end owing to the development of the apical knob (figs. 38-40, PI. 2). Con- siderably later another knob, the rudiment of the manubrium, makes its appearance at the opposite end (figs. 41-5, PL 2). The two tripartite whorls begin to develop before there is any trace of the manubrium, and it is highly significant that they are both displaced towards the weighted end, as shown in figs. 39-45, PL 2, and figs. 46-7, PL 3, exactly as would be expected in the ease of the nodes of a vibrating rod. In Latrunculia apicalis there is a curious and significant difference in the position of the whorls. The apical knob and the manubrium appear simultaneously (or nearly so) and, the shaft being approximately equally weighted at both ends, the median whorl appears approximately half-way between them. The position of the subsidiary whorl requires further investigation. I owe to my friend Mr. Maurice Burton the opportunity of studying quite recently the development of the anisodiscorhabd in a third species of Latrunculia, as yet unidentified, 22 ARTHUR DBNDY collected by Dr. Gilchrist in the neighbourhood of the Cape of Good Hope. This species is very closely related to Latrun- culia bocagei. The anisodiscorhabds are almost identical in structure, but the spines of the apical crown are not capitate (this is a trifling difference). There is, however, a difference in the position of the whorls, which are displaced, not towards the apex but slightly towards the base of the spicule (figs. 55-7, PI. 3). This fact is fully accounted for when we come to study the development of the spicule, for we find that, instead of the apical knob appearing before the manubrium, as in L a t r u n - culia bocagei, the latter is slightly ahead of the former in its development (figs. 55, 56, PI. 3). Thus the nodes are pulled towards the base instead of towards the apex of the spicule. I do not see how the very definite and constant relation between the order of appearance of the terminal knobs and the position of the tripartite whorls can be explained except on the' vibratory theory '. In Didiscus, where no knobs are developed, the whorls are not displaced towards either end, but, owing to the tapering form of the shaft, the subsidiary whorl is pulled in towards the middle.1 I endeavoured to show long ago (1917, 1921 a) that the formation of the median and subsidiary tripartite whorls of siliceous outgrowths in the anisodiscorhabd Latrunculia was to be attributed to the presence of two groups each of three so-called ' formative ' or ' initial cells ', which settled upon two of tho principal nodes, a view Avhich received further support from the fact that the subsidiary whorl, though usually appearing on the apical side of the median whorl, occasionally appears on the other side (sometimes it is wanting altogether). The absence of a whorl of outgrowth on the third node was supposed to be due to the fact that only two whorls of ' initial 1 I am indebted to my friend Professor Dale, of King's College, and to my colleagues in the physical laboratory, especially Dr. Flint, for mathematical and experimental investigations of the positions of the nodes in a vibrating rod weighted at one end. Their results support, in a striking manner, my views as to the nodal positions of the tripartite whorls in Latrunculia. I hope that it may be possible to publish more exact observations on these positions in a subsequent paper. SPONGE-SPICULES •23 cells ' were produced, so that one node had to go without. Substituting the conception of scleroplastids for that of initial cells I still adhere to this view, which has received unexpected and startling confirmation from the study of the phenomenon of twinning recently observed by me in Latrunculia bocagei. In the course of a prolonged re-examination of my preparations of this sponge I came across several instances in which the half-grown anisodiscorhabds occur in pairs, as shown in figs. 34 and 35, PI. 2. In such a pair the twin spicules, both in exactly the same state of development, lie close together, side

TEXT-FIG.

Diagrams of hypothetical divisions of the scleroplastid in the formation of (A) a normal and (B) a twin anisodiscorhabd in Latrunculia bocagei. by side, with their long axes approximately parallel, and oriented in the same sense. They are very much smaller than the normal, single spicule at a corresponding stage of development (compare fig. 46, PI. 3) and, most remarkable of all, there is only one nodal whorl in each spicule. It would appear that each pair must have developed from a single scleroplastid with limited powers of division, so that instead of giving rise to a single spicule with two whorls, it has given rise to two spicules each with only one whorl. It is not possible to determine the number of lobes in each whorl, as they are very ill-defined, but, assuming it to be three, as it is in the normally formed whorls, then the manner in which the divisions of the original scleroplastid may be supposed to have taken place in the case of the twin spicules is indicated in the accompanying diagram (Text-fig. 4, B), in which the 24 ARTHUR DENDY protorhabds are represented by the vertical straight lines and the scleroplastids by the dots. Text-fig. 4, A, represents similarly the divisions which may be supposed to take place in the case of the normal anisodiscorhabd. This case appears to me to afford almost conclusive evidence of the real existence of mobile and dividing scleroplastids, even though these cannot actually be seen. It also suggests very strongly that the protorhabds and the scleroplastids that give rise to the whorls have a common origin.

TEXT-FIG. 5.

Abnormal anisodisoorhabd of Latrunculia apicalis, with adventitious apical prolongation, x 700. A curiously abnormal form of the anisodiscorhabd of Latrunculia apicalis, already described by me in 1917, with a figure which I venture to reproduce here (Text-fig. 5), shows an imperfection in the subsidiary whorl on one side, due apparently to the absence of one of the three lobes, while on the same side a long, tapering branch is given off obliquely from just above the apical whorl, resembling the apical prolongation. May not this be explained by supposing that the scleroplastid that should have formed the missing lobe of the subsidiary whorl has settled down in the wrong place and given rise, under the influence of its new surroundings, to an adven- SPONGB-SPICULBS 25 titious apical prolongation ? The appearances are certainly very suggestive, but in the absence of further evidence I do not wish to press the point, and it would be difficult to reconcile such an explanation with what has been said above as to the specificity of the scleroplastids in the case of Stelletta haeckeli. Another and very beautiful indication that the whorls of the discorhabd originate extraneously from groups of separate scleroplastids and are not mere outgrowths from the shaft of

TEXT-FIG. 6.

f Sigmosceptrella fibrosa. A, sigmoid crepis of isodiscorhabd. x 1,750. B, later stage of same, showing commencement of tripartite whorls and apical tufts of outgrowths. X 1,750. C, one of the tripartite whorls of older spicule. X 1,750. the spicule, is afforded by the development of the sigmo- discorhabds of the genus Sigmosceptrella. These spicules are isodiscorhabds developed around a sigmoid crepis -1 with strongly recurved extremities (Text-fig. 6, A). The fully developed spicule shows no indication of its sigmoid origin

1 This term was first employed by Sollas (1888 a) for the original spicule forming the foundation upon which a desma (monocrepid or tetracrepid) is built up. It may be extended to all similar foundation spicules that differ conspicuously in form from the completed spicule. 26 ARTHUR DBNDY but consists of a stout, straight shaft with a bunch of spines at each end and two whorls of spinose outgrowths arranged symmetrically (Text-fig. 7, B, C). In Sigmosceptrella fibrosa each whorl consists of three outgrowths or lobes each dividing typically into two spines (Text-fig. 6, C). In Sigmosceptrella quadrilobata, on the other hand, each whorl consists of four similar lobes (Text-fig. 7, D.) I

TBXT-FIO. 7.

Sigmosceptrella quadrilobata. A, sigmoid crepis of isodiscorhabd, showing commencement of quadripartite whorls of outgrowths, x 1,750. B, C, fully developed, or nearly developed, isodiscorhabds (side views), x 770. D, ditto (end view), showing apical tuft of spines and one of the quadripartite whorls, x 770. should not like to say, however, that the number of spines into which each lobe divides is in either case constant. The terminal spines and the whorled outgrowths or lobes alike originate as secondary growing points, differing from the rest of the spicule (but agreeing with the protorhabd) in their more darkly staining character. We may confine our attention to those that form the whorls. In Sigmosceptrella fibrosa each whorl is developed from three growing points, one appearing at the end of the recurved portion of the crepis and two making their appearance on the main shaft nearly opposite to the former (Text-fig. 6, B). All three seem to appear simultaneously. In Sigmosceptrella quadrilobata two growing points appear at the end of the recurved hook and SPONGE-SPICULBS 27 two on the main shaft, making a whorl of four (Text-fig. 7, A). In both species the further deposition of silica envelopes the main shaft and its recurved hooks in a common envelope, so that the two whorls of outgrowths ultimately seem to arise from a short, thick, straight shaft (Text-fig. 7, B, C) and the sigmoid character is completely concealed. It appears sufficiently clear that the position of the whorls in the sigmodiscorhabd is determined with regard to the crepis as a whole, and that they cannot be regarded merely as outgrowths of its curved shaft. Their formation ignores, so to speak, the curvature of the shaft, and this can only mean that the growing points are of extraneous origin. I think there are here good grounds for concluding that each whorl owes its existence to a group of scleroplastids, and that the number of secondary growing points in each depends upon the number of divisions undergone by a parent scleroplastid.

(/) A revised Account of the Development of the Anisodiscorhabd in Latrunculia bocagei. It is, of course, of first-class importance for the establishment of our thesis that we should have detailed an accurate know- ledge concerning the processes of development which the sponge-spicule actually undergoes. Unfortunately very little information of this kind is at present forthcoming. The theory that the spicule arises and develops in a so-called mother-cell seems to have exercised a fatal influence upon progress in this direction, and very few observers have succeeded in going beyond the demonstration of a nucleus and a cell-membrane in close association with the young spicule. The problem is really a very difficult one. It is very seldom that material of a suitable nature is forthcoming, and then it is usually in a state of preservation by no means satisfactory for cytological study. For the investigation of all the various factors that may take part in the process some complex type of spicule is needed, and it was by the study of the discorhabd that the existence of scleroplastids was first suggested. I have already on two separate occasions (1917, 1921 a), 28 ARTHUR DENDY given some account of the development of the anisodiscorhabd of Latrunculia bocagei, but only ordinary spirit-preserved material has so far been available, which could not be expected to yield completely satisfactory results. These results, however, have proved sufficiently interesting to justify me in making once more a thorough examination of my preparations, as a consequence of which I am able to correct some erroneous interpretations and to extend my observations in certain directions. Moreover, these observations have been to a large extent confirmed by the study of the South African species of Latrunculia already referred to. In the earliest stage observed (fig. 36, PI. 2) we have a long, slender rod, darkly stained (by brazilin) and highly refringent. This is evidently a protorhabd already silicified. It is already much thicker than the naked protorhabd would be, as will be obvious on comparison with figs. 39-44, PL 2, in which the original protorhabd is clearly recognizable as a very slender, darkly staining ' axial thread '. The silicified protorhabd represented in fig. 36, PI. 2, is enclosed in an irregular granular mass containing what looks like a badly fixed nucleus. This may very well represent a nucleated silicoblast. As yet the young spicule shows no indication either of the terminal enlargements or of the whorled outgrowths. The next stage is perhaps represented by fig. 37, PI. 2, in which the protorhabd, though less highly silicified, is somewhat longer and is surrounded near the middle of its length by a whorl of three elongated, darkly staining bodies (of which two alone are visible). It is to these bodies that I at first (1917) gave the name of ' formative cells ', which I afterwards (1921 a) changed to ' initial cells '. I have now satisfied myself that they are not cells at all, though exactly what they are it is extremely difficult to say, as I have never been able to repeat my original observation of this stage. The facts that they form only a single whorl and that they lie so near the middle of the shaft seem to indicate that they are scleroplastids rather than growing points, and their elongated form may be due to their approaching division into two whorls, but their SP0NGE-SP1UULBS 29 size and shape are both against this view. They certainly look more like growing points, but they do not appear to be attached to the shaft, nor does it seem likely, from the consideration of numerous other cases, that they should be attached before the development of the apical knob, unless, indeed, we are dealing with a quite abnormal spicule. No silicoblast was observed in this case, but later stages appear sometimes with and sometimes without a silicoblast, and there can be little doubt that the association of the latter with the developing spicule is a temporary one, repeated at intervals until the full comple- ment of silica has been secreted. A somewhat more intelligible stage is represented in fig. 38, PI. 2, which shows the young spicule again enveloped in an irregular silicoblast, this time with a large and very distinct nucleus of characteristic appearance—oval and coarsely granular, without the single large and well-defined nucleolus usually so conspicuous in the amoebocytes of the mesogloea (cf. fig. 41, am., PI. 2). The silicified shaft of the spicule has now enlarged at one (the apical) end so as to form a small knob. Embedded in the cytoplasm of the silicoblast, near the middle of the shaft, are visible two small, darkly staining bodies corresponding closely in position with the bodies observed at the preceding stage, but rather farther from the shaft. No doubt there are really three of these, the third being concealed by the shaft. Each lies at the apex of a V-shaped disturbance in the cytoplasm of the silicoblast, pointing downwards, as if it had recently changed its position, and we are again tempted to conclude that we have before us the actual scleroplastids, which have not yet divided for the last time and taken up their definitive positions on the shaft as two nodal whorls. They seem too large, however, to be merely scleroplastids, and the observation of further cases will be necessary before their exact nature can be established. That they have something to do with the formation of the whorls can hardly be doubted. A more advanced stage is represented in fig. 39, PI. 2, which I give because it shows two silicoblasts associated with the young spicule, neither of which has yet enveloped it, the 30 ARTHUR DENDY granular substance which partly surrounds the upper portion being probably rnesogloeal. It is obvious that there are in this case already two whorls of secondary or adventitious1 growing points, situate in the characteristic nodal positions, pulled towards the apical knob. Only one growing point, however, is distinctly seen, that on the right side of the median whorl, and the subsidiary whorl is hardly visible at all. We may pass on at once to fig. 40, PI. 2, which shows a young spicule in nearly the same stage of development, but without any silicoblast. Both whorls of growing points are now well developed. The upper (subsidiary) whorl is, however, rather less advanced than the lower (median) whorl. We may suppose that each of the three scleroplastids of the original single whorl has divided into two, forming altogether two whorls which have separated from one another and taken up nodal positions. Owing to the position in which the spicule is lying, only two of the three growing points in each whorl are visible. Each growing point appears as a slender crescent, staining fairly darkly with borax carmine or brazilin and forming a kind of thickening on the edge of a very thin and perfectly transparent plate of silica. This plate of silica is placed longitudinally and radially between the shaft, and the growing point and the corresponding plates of the two whorls are continuous with one another. These thin, radial plates of silica, forming longitudinal flanges on the shaft, are broadest at the growing points, each of which forms a protuberance standing out at some distance from the shaft. That they appear to stand out further from the shaft on one side than on the other is due to the effect of foreshortening on one side owing to the position of the spicule. At one end each plate no doubt becomes connected with the apical knob, though this can only be seen on the left side. Towards the other end of the spicule they gradually narrow down until their free edges meet the shaft. It is to be noted that the texture of the growing points is not perfectly uniform. Usually a single minute granule, of darker appear-

1 So-called to distinguish them from the primary growing points found by the ends of the original protorhabd, now sealed up in silica. SPONGE-SPICULES 31 ance, is recognizable. I propose to call these the ' growing- point granules ' ; I shall have more to say about them presently. One more feature shown in fig. 40, PI. 2, demands our notice, and that is the appearance of a peculiar curvature or kink in the shaft at the level of each whorl of growing points. The significance of this also will be discussed later on. The next stage (fig. 41, PI. 2) differs from the preceding only in the commencing development of the manubrium, which appears as an elongated knob or thickening at the basal extremity of the shaft. Now that we have indicated the manner in which all the principal parts of the spicule first make their appearance, and the relations which they bear to one another, we may save time and space by continuing with a concise account of the further history of each of these parts, instead of describing in detail all the later stages passed through by the spicule as a whole. The principal parts now to be recognized are (1) the shaft, containing the protorhabd ; (2) the median tripartite whorl, represented by three growing points ; (3) the subsidiary whorl, also represented by three growing points ; (4) the apical knob, from which the apical whorl and crown of capitate spines will be developed ; and (5) the basal knob or manubrium. A refer- ence to fig. 32, PI. 2, representing the fully developed spicule, will be useful at this stage of our inquiry. The shaft is at first a very slender rod, staining lightly with borax carmine or brazilin and containing the much more slender and more darkly staining protorhabd, which extends into the basal and apical knob at either end (figs. 39-44, PI. 2, and fig. 47, PI. 3). (In Latrunculia apicalis, as already pointed out, the growth of the protorhabd is not stopped with the formation of the apical knob, but extends far beyond it, so that the shaft has a long apical prolongation (Text-fig. 1, and fig. 33, PI. 2).) The shaft is connected with the growing points of the tripartite whorls by the three longitudinal, radial plates of silica, which project from it like flanges or ribs. These at first are very thin and transparent, though 32 ARTHUE DBNDY sometimes they have a finely granular appearance, due apparently to adhering protoplasm or mesogloeal substance (fig. 41, PI. 2). They very soon extend up to the apical knob (figs. 40-5, PI. 2), later on to the basal knob as well (fig. 45, PI. 2). The apparent width of these radial plates of silica varies with the point of view, and the foreshortening on the one side or the other frequently gives the young spicule a very asymmetrical appearance (compare figs. 40-5, PL 2, and fig. 46, PI. 3). The shaft also frequently appears to be asymmetrically inserted into the terminal knobs (figs. 41-5, PL 2), but this again is probably due merely to the point of view. The shaft as a whole continues to thicken and the grooves between the longitudinal ribs become filled with silica, so that presently it appears circular, or nearly so, in transverse section (figs. 52, 54, PL 3), becoming triangular as it approaches a tripartite whorl (fig. 53, PL 3). A very common feature of the shaft at an early stage of its development, while it is still very slender, is the appearance of a more or less sharp bend or kink just at the level of each tripartite whorl (figs. 40-4, PL 2). This kink must, of course, involve the protorhabd, and yet we find at later stages that the shaft and its axial thread are both perfectly straight (fig. 47, PL 3). If the kinks are normal we must assume that at the time when they are formed the silica is still soft and flexible, and that they straighten out again with the further growth of the spicule. In this connexion we may mention another feature that is not infrequently observed. It will be seen on examination of figs. 40, PL 2, and figs. 55 and 56, PL 3, that certain radial markings occur on one of the thin longitudinal plate of silica, passing outwards at right angles from the shaft towards the surface of one of the growing points of the median whorl. I do not attribute any special importance to these appearances. They may possibly indicate the commencement of thickening of the radial plate, or they may be merely creases due to the stresses set up by the growth of the whorl. They may be connected with the kinking of the shaft, and both radial SPONGE-SPICULBS . 3B markings and kinks may be artifacts due to shrinkage, which would explain why the kinks are not visible in the axial threads of older spicules that have hardened before being subjected to dehydration. The two tripartite whorls may be considered together, for the development, so far as my observations go, appears to be identical in the two cases. The three growing points of each appear as protuberant thickenings on the edges of the radial plates of silica (figs. 39-45, PI. 2, and fig. 50, PI. 3). In stained preparations these thickenings appear darker in colour than the radial plates themselves, but this may be due partly to the greater thickness of silica. It is, however, a very common, probably universal, feature of such secondary growing points. Very often a minute, dark-looking granule is found in or on the growing point. Such granules are shown in many of the figures. Fig. 48, PI. 3, shows the lower part of a broken shaft at a very early stage of development, with an associated silicoblast. The position in which the two tripartite whorls will develop, as indicated by two kinks in the slender shaft, is evidently abnormal.1 The feature to which I wish to call attention, however, is the presence, nearly opposite to the upper kink, on the right-hand side, of a very distinct and quite detached granule (gr.), whose staining and refractive properties appear to be identical with those of the shaft itself, which is already to some extent silicified. A well-marked growing point, with a much less well-defined granule, is seen on the left- hand side nearly opposite the lower kink in the shaft. Fig. 49, PI. 3, shows a very young whorl, with a growing point on the left that appears still to be quite detached from the shaft, and contains a very distinct, more darkly stained granule at its apex. Is it possible that these growing-point granules actually represent the hypothetical scleroplastids and that some mutual attraction exists between them and the protorhabd ? May this be the true explanation of the kinking of the shaft ? We must await further evidence before these questions can be answered.

1 It may be one of the cases in which the subsidiary whorl lies towards the basal instead of towards the apical end of the spicule. NO. 277 D 34 , ARTHUR DBND"y Fig. 50, PI. 3, taken from a broken spicule, shows a very beautiful example of the median tripartite whorl at a very early stage of its development. The shape of the growing points is clearly seen, and the shading is intended to represent the staining intensity. The slender protorhabd is seen in the middle of the shaft. It must not be supposed that there is absolutely no variation in the number of these growing points. Occasionally there is an abnormality in this respect, and fig. 51, PI. 3, shows a whorl with an extra growing point at a different level from the other three. Before passing on to consider the later stages in the development of the tripartite whorls, I wish to correct an erroneous interpretation that I formerly (1917, 1921 a) placed upon the appearances exhibited by the earlier stages. This misinter- pretation arose from my not recognizing the existence of the thin longitudinal plates of silica by which the growing points are connected with the shaft. The free edges of these plates appeared to me as a thin membrane or sheath formed by the supposed ' formative ' or ' initial cells ' and enveloping the growing spicule. The projecting growing points on the edges of the plates I regarded as pockets bulging out from the sheath, and I spoke of them as ' silica pockets ' into which silica was supposed to be secreted by the silicoblasts. Some of the figures which I published in my two previous papers certainly seem to afford strong justification for such an interpretation, but I arn convinced now that it is erroneous and that the true facts are as stated above. Eeturning to the history of the tripartite whorls, it is to be observed next that the growing points begin to flatten and spread out at right angles to the shaft of the spicule, which by this time has become greatly thickened. This is well shown in fig. 46, PI. 3, in the fr\vo growing points on the right. It is still better shown in fig. 52, PI. 3, taken from a broken-off fragment of a young spicule, so that one cannot say which of the two whorls is represented. Each growing point has now developed into a broad, flat lobe or plate, forming a segment of a disc, but rather widely separated by a deep notch from its SPONGE-SPICULES 35 neighbour on either side. Each lobe is a wedge-shaped plate of silica, thin at its outer edge and thickening very considerably towards its junction with the shaft. At first the outer edge or margin of each lobe is perfectly smooth and even (fig. 52, PL 3). Later on it becomes crenate, or scalloped, with about half a dozen projecting points (fig. 53, PI. 3), which stain lightly at their apices (the other parts being unstained or nearly so) and look like another set of secondary, or perhaps we should say tertiary, growing points. The whorl, though perhaps not yet of full size, has now attained the condition characteristic of the adult spicule. We have to deal next with the later development of the apical knob, from which arise (1) the apical whorl and (2) the apical crown of capitate spines. Whether or not any selero- blasts are concerned in the development of the apical knob I have been unable to determine. I have once seen a couple of minute granules associated with it (fig. 42, PI. 2), but it is quite impossible to say what these really are. The knob, which may be at first sight elongated (figs. 38-40, PI. 2), very soon assumes an almost spherical form (figs. 41-4, PI. 2). As it increases in size it becomes slightly flattened in a plane at right angles to the shaft, and a very slightly prominent, equatorial or sub-equatorial rim makes its appearance. This is best shown in fig. 46, PI. 3. This rim grows out into an annular plate of silica, forming the apical whorl. The apical whorl is not divided into lobes like the tripartite whorls, but its edge becomes scalloped by the development of marginal growing points much as in the case of the latter. There is some indication that these growing points bifurcate (fig. 54, PI. 3). A short, rib-like thickening extends radially into each of them, and the growing point itself stains distinctly as in the cases already described. I have seen no granules in association with these growing points, but I am strongly disposed to attribute their development to the presence of scleroplastids that settle down on the margin of the undivided whorl, and to extend the same explanation to the marginal growing points of the two tripartite whorls. D2 36 ARTHUR DENDY The edges of the apical whorl turn upwards around the upper half of the apical knob, which may be compared to a teacup turned upside down in a saucer. From the strongly convex upper surface of the apical knob a number of short, capitate spines grow out to complete this part of the spicule (fig. 32, PL 2). These also are formed from secondary growing points, with which, presumably, scleroplastids are associated. The basal knob, which develops into the manubrium at the other end of the shaft, appears, as we have already said, somewhat later than the apical knob and is from the first more elongated in form (figs. 41-5, PI. 2). It gradually grows larger and presently assumes the characteristic form of a stonemason's mallet, as shown in fig. 46, PI. 3. Presently secondary growing points appear upon its surface, which develop into short, sharp spines. These tend to be arranged in two whorls (fig. 32, PL 2) not nearly so well defined, however, as the whorls already described. In the above account of the development of the anisodiscorhabd of Latrunculia I have not dwelt upon the dislocated nodal positions of the median and subsidiary whorls, because this has been sufficiently dealt with in the preceding section of this paper, but I would again remind the reader that the peculiar disposition of these tripartite whorls, and especially the suppression of one of the two in cases of twin-formation, oblige us to postulate the existence of individual mobile scleroplastids as initiators of the three original growing points in each, and it seems only logical to extend this conception to all the other secondary (or tertiary) growing points that arise in the course of development.

3. GENERAL CONCLUSIONS AS TO THE ORIGIN AND GROWTH OF SILICEOUS SPONGB-SPICULES. (a) The primary co-operating Agents. It is evident from the observations described in the preceding pages that the siliceous sponge-spicule owes its existence to the co-operation of two primary agents, the sclerococcus (in SPONGB-SPICULES 37 the role of scleroplastid) and the silicoblast. The relation between these two appears to be brought about in the first instance by phagocytosis, the silicoblast enveloping the sclerococcus, usually after the latter has elongated into a protorhabd, and depositing silica around it. As the young spicule is sometimes seen without a silicoblast and sometimes with one, we must suppose that its envelopment by the latter is a temporary performance repeated at intervals (presumably by different silicoblasts), and this may well account for the laminated character of the siliceous deposit. Phagocytosis is probably of extremely common occurrence in sponges. Indeed, it was in sponges that some of the classical instances of this phenomenon were observed by Metschnikoff (1892), and some years ago (1914 b) I myself had occasion to lay stress upon some remarkable cases in Grantia com- press a. There can be little doubt that it was this phagocytic envelopment of the spicule that gave rise to the idea of ' mother- cells '• in which the young spicules were supposed actually to arise. Moreover, it is not necessary to suppose that the phagocyte is always a silicoblast, it seems highly probable that the fully formed spicule is often enveloped phagocytically by an ordinary wandering cell and thus carried from place to place. We shall return to this point later on. Examples of silicoblasts associated with the developing spicule have already been given in Section 2 of this paper (figs. 36, 38, 39, 42, PI. 2). In further illustration of the point under discussion I may now briefly call attention to certain typical cases in which the phagocyte appears as a so-called mother-cell. Pig. 58, PI. 3, represents a trichodragma, or bundle of fine, hair-like spicules, enveloped in a phagocyte whose nucleus forms a conspicuous projection on one side, while the cytoplasm is stretched thinly over the surface of the bundle. Pig. 59, PI. 3, represents a young chela of Cladorhiza inversa, with a silicoblast apparently attached to one side but probably really enveloping the spicule. Pig. 60, PL 3, shows a fully developed chela of the same kind, with its ' mother-cell', probably an exhausted silicoblast, with a 38 ARTHUR DENDY conspicuous nucleus lying in the curve of the spicule. This should be compared with fig. 47, PI. 3, representing a young anisodiscorhabd, also enclosed in a ' mother-cell' which is probably merely an exhausted silicoblast. The colloidal silica, when first deposited on the spicule, is doubtless in a liquid condition, but presently becomes hard and brittle. I have already (1916 e) pointed out that in certain cases (e.g. Collosclerophora) the silicoblasts may discharge their silica into the surrounding mesogloea without associating themselves with scleroplastids or protorhabds, and that then the silica remains soft and jelly-like, unless artificially dehydrated. It seems unnecessary to spend much time in comparing the views enunciated above with the generally accepted ideas as to the origin and development of the siliceous sponge spicule. Most of these ideas are dominated by the conception of the ' mother-cell', inside which the spicule is supposed to be secreted, and sometimes actually to complete its development, though it is admitted that in very large spicules there must be the co-operation of secondary silicoblasts. The axial thread is supposed to be the first part secreted. It is composed of a protein substance (' spiculin '). It is then surrounded by alternate layers of silica and spiculin, the latter much thinner than the former. Schulze (1904) has given a very detailed account of the structure of the gigantic spicules in the hexactinellid genus Monoraphis. He believes the axial thread and the layers of spiculin to be of the same nature and actually in connexion with one another at the ends of the growing spicule, as though the thin spiculin layers were actually outgrowths of the axial thread. However this may be in Monoraphis it is obvious that such rv view cannot be applicable to the case of siliceous pearls, where the spicule is spherical and the lamina- tion concentric. The probability seems to be that there is much more in common between the spiculin layers and the silica layers than between the former and the axial thread. Even the silica layers (siphons) contain a certain amount of organic matter, as indicated by their turning brown on heating, SPONGE-SPICULES 3& though to a less extent than the spiculin layers. The latter, on the other hand, probably contain a certain amount of silica, so that the difference between the successive laminae seems to consist merely in the relative proportions of silica, water, and organic matter. The case seems to be closely com- parable to that of the starch grain, so far as the parts outside the axial thread are concerned. It is extremely difficult to believe that a non-crystalline body of such complex structure as a siliceous sponge-spicule could originate, and in some cases attain its full development, as an intracellular secretion. There must be a definite foundation upon which the secretion can be laid down, and that foundation is the living protorhabd, or, in the case of silica pearls, the scleroplastid before it has elongated to form a protorhabd.

(b) The Influence of the primary Protorhabd upon the ]?orni of the Spicule. The protorhabd forms the long axis of the spicule, and the extent to which it elongates determines very largely the form of the spicule. This is very well illustrated by the reduced triaenes of Stelletta haeckeli described in Section 2 (figs. 10-16, PI. 1). These, however, are abnormalities. The varying extent to which growth in length takes place may, however, give rise to constant specific differences. In the anisodiscorhabd of Latrunculia apicalis, for example, as I have already pointed out, the growth of the protorhabd is continued even after the apical knob of silica has been formed and gives rise to an apical prolongation. This apical prolongation is entirely absent in Latrunculia bocagei, where the growing end of the protorhabd becomes sealed up in the apical knob (compare Text-fig. 1, and figs. 32, 33, PI. 2). If, then, the rate of deposition of the silica and the rate of growth of the protorhabd are accurately adjusted to one another the spicule will gradually taper off to a point, the apex coinciding with the end of the protorhabd (or nearly so). If the silica deposition overtakes the protorhabd growth the spicule may end bluntly, or even in a knob (fig. 9, PI. 1). It 40 ARTHUR DBNDY is very rarely that the protorhabd grows so rapidly as to project for a time beyond the silica altogether, as shown in Text-fig. 1. Usually the protorhabd remains unbranched, so that if the .spicule itself exhibits branching at all it is due to the association of several protorhabds or to the formation of secondary growing points without protorhabd formation. Cases of actual bifurcation of the protorhabd are, however, by no means uncommon. The best known, perhaps, is that afforded by the cladi of the dichotriaene in many species of Thenea, Stel- letta, and Geodia (Text-fig. 8). The branching of the protorhabd appears to be always apical, and this must necessarily

TEXT-FIG. 8.

Dichotriaene of Ecionemia laviniensis (end view), showing oladome with axial canals, x 120.

be the case when it is only the apex of the growing protorhabd that remains free from a coating of silica. If lateral branches appear on the spicule in such cases, they must be adventitious and of extraneous origin. We have discussed abnormal examples of this kind in Section 2, and a normal example, in which the branches are represented by short spines, will be described later on (Text-fig. 12). A very unusual type of apical branching of the protorhabd, giving rise to a crown of spines, has already been described in the case of Sceptrospongia coronata (figs. 4-8, PI. 1). Though the protorhabd usually grows in a straight line this is by no means always the case. In the spiral spicules of the Spirastrellinae, and especially in the Spinispirae of Trachy- SPONGB-SPICULBS 41 cladus (Text-fig. 9), we must assume that it grows spirally and thus determines the spiral form of the spicule as a whole. There is no reason to suppose that this spiral form is due to any outside agency. A spiral axis may, however, be formed sympodially, as in the dichotriacts of Thenea (Dendy, 1924 b).

(c) The Influence of the Mode of Division of the original Scleroplastid. (a) The Formation of radiate Spicules. We have already had occasion to notice that the tetract spicule may be regarded as owing its characteristic symmetry

TEXT-FIG. 9.

Spinispirac of Traohycladus stylifer. x900. to the association of four protorhabds derived from a scleroplastid that divides into four daughter scleroplastids, these latter arranging themselves in the form of a pyramid and then elongating centrifugally to form the rays of the spicule, while their inner ends all join together at the centre. In my memoir on ' The Tetraxonid Sponge-spicule: a Study in Evolution ' (1921 a) I endeavoured to show how all the endlessly varied spicules of the Tetraxonida may be derived from this fundamental type. In the hexactinellid sponges the fundamental symmetry is quite different, being based upon three axes which cross one another at right angles, forming six rays meeting in a common centre. It is difficult to decide in this case whether the original scleroplastid divides into six, which elongate in one direction 42 ARTHUR DBNDY only, or into three which elongate each in two opposite directions. On the analogy of the tetract spicule the first alternative seems more probable, and the absence of any indication of crossing perhaps supports this view. That the protorhabds of a radiate spicule sometimes do elongate each in two opposite directions seems to be established, however, by a very curious type of spicule found in the siliceous deposits at Oamaru, in New Zealand, and referred by Hinde and Holmes (1892) to the genus Dactylocalycites. It has, however, nothing to do with that genus, which has tetract spicules (phyllo- or discotriaenes). Indeed, it exhibits a quite unique type of symmetry. I have several examples of this spicule in my possession, and I find that its peculiar features are so constant that they must unquestionably be regarded as normal. It consists of a thin, flat plate of silica of elliptical shape and nearly smooth outline (fig. 61, PL 3). About ten axial canals are present, all lying in the plane of the ellipse. These cross one another at various angles near the centre of the spicule, but the points of intersection are not all coincident. The pair of canals which lie next to the short axis of the ellipse have their points of intersection a little to one side of those of the other canals (which are very nearly coincident with one another). These two crossed canals form an X, the limbs of which, in conjunction with the margin of the ellipse, define four triangular spaces, two small, which are bisected by the short axis of the ellipse, and two large, bisected by the long axis. The larger triangles are each further subdivided by the remaining axial canals into some eight or nine narrow, wedge-shaped segments. The smaller triangles are not subdivided, but each forms a single wedge-shaped segment much larger than any of the remainder. Near the margin of the spicule is a series of larger and smaller perforations, the larger ones radially elongated. The form and arrangement of these is sufficiently well shown in the figure. Sometimes an axial canal, complete on one side, is only partially developed on the other. Such a case is seen on the right side of the figure. Evidently the protorhabd, which SPONGE-SPICULES 43 must have occupied the axial canal at first, has stopped growing prematurely. Silica must have been deposited around the protorhabds in such a way that all the rays of the spicule have become fused into a thin plate. The sub-marginal perforations may be regarded as remnants of the original intervals between the rays. There is no trace of a shaft projecting at right angles from the middle of the plate, as in the spicules of Dactylocalycites. There is, of course, no actual proof that the protorhabds in this remarkable spicule have all been formed by the division of an original scleroplastid, but it seems far more likely that this is the case than that they should have come together secondarily and arranged themselves in a pattern of such remarkable and constant symmetry, which seems to be the only possible alternative.

(j8) Variation in the number of primary Kays. In each of the great groups of spicule-bearing sponges the typical number of primary rays is highly characteristic, viz. three in the Calcarea, four in the Tetraxonida, and six in the Hexactinellida. Nevertheless, we frequently meet with cases of rneristic variation in either a plus or a minus direction. In the Tetraxonida, for example, increase in the number of rays gives rise to the true asters, which often have a dozen or more rays springing from a common centre ; sometimes, as in the sterrasters of Geodia, far more. In many cases axial canals have been observed in these rays and there can be little doubt that the rays are formed by deposition of silica around primary protorhabds growing out from a common centre. The increase in the number of rays is readily explained on the assumption of an original scleroplastid dividing many tirues instead of twice only. All stages between the primitive tetract and the many-rayed aster on the one hand, and the simple linear spicule on the other, are actually met with in certain species. The reduction in the number of rays is often due to suppression of rays which have already commenced to elongate, as in 44 ARTHUR DENDY the case of the suppressed triaenes of Stelletta haeckeli described in Section 2. There are innumerable cases, on the other hand, in which there is no indication of the suppressed rays, as, for example, in the common oxeote type of spicule, which may be regarded as composed of two rays coming off from the centre in opposite directions, though often forming a slight angle with one another. This form is explained at once on the supposition that the original scleroplastid has divided once only instead of twice. The single-rayed stylote spicule, again, may be regarded as initiated by a scleroplastid that has elongated in one direction only without dividing at all. In the hexactinellids, curiously enough, there seems to be no meristic variation in a plus direction, the so-called hexasters being formed by branching of the six primary rays. We frequently get suppression of rays, however, in this group.

(y) The Formation of Dragmata and Bosettes. Closely akin to the formation of asters is the development of dragmata and rosettes of microscleres amongst the Tetra- xonida. The dragmata are compact bundles of linear spicules lying side by side parallel with one another. Such bundles are formed by microxea (trichodragmata, Text-fig. 10, A), by toxa (toxodragmata), and by sigmata (sigmodragmata, Text- fig. 10, B). The trichodragmata in particular are very widely distributed amongst the tetraxonida and of curiously sporadic occurrence. The rosettes (Text-fig. 11), so far as I am aware, are always formed of anisochelae, radially arranged in a ball, with their small ends meeting in the centre. They are frequently met with in certain Esperelline genera. Whether originating in dragmata or in rosettes the spicule in question probably always separate from one another sooner or later. Each is independent and complete in itself, and, moreover, toxa and sigmata at least more often originate separately. The dragmata and rosettes are to be explained, like the asters, as due to multiple fission of a scleroplastid whose SPONGB-SPICULES 45 products remain close together and give rise to spicules before separating. They do not unite together to form a single spicule as in the case of the aster. They differ from the twin spicules of Latrunculia in that the form of the spicules appears to be

TEXT-HG. 10.

Biemna no vae-zealandiae. A, trichodragma. x 325. B, sigmodragma. X 325.

TEXT-ITO. 11.

Pseudoesperia carteri, rosette of anisochelae. X280. unaffected by the association ; unless, indeed, the unequal- ended character of the anisochelae be attributable to their origin in rosettes in which the inner ends of the spicules have no room to develop properly. The reader should also compare the incomplete twinning that gives rise to the amphitriaene, as described in Section 2 (figs. 17-19, PI. 1). 46 ARTHUR DENDY

(d) The Influence of adventitious Growing Points upon the Form of the Spicule. (a) Adventitious Growing Points associated with Protorhabds. A great many siliceous spicules, both mega- and microscleres, are ornamented with larger or smaller spines arranged in a great variety of ways ; such are the well-known acanthoxea, acanthostyli, acanthostrongyla, and so forth. In certain cases it has been demonstrated that these spines contain axial canals

TEXT-FIG. 12.

Acanthostrongyle of Lithoplocamia lithistoid.es, showing primary and secondary axial canals, x 300. and their growth must therefore have been based upon that of a protorhabd. In the acanthostrongyla of Lithoplo- camia lithistoides (Dendy, 1921 6) this is particularly obvious, though only in the more or less eroded spicule (Text- fig. 12). It will be seen from the figure that the main axial canal of the spicule pursues an uninterrupted course from end to end, and that numerous secondary canals run outwards into the projecting spines and actually reach their apices. These secondary canals, however, do not quite meet the main canal, which seems to show that the scleroplastids, one for each spine, settled on the main protorhabd and began to grow out SPONGE-SPICULES 47 into adventitious protorhabds after the main protorhabd had received its first thin coating of silica. Sollas showed, many years ago (1879 a), that axial canals are also present in the spines of the remarkable pseudopolyact spicules of Trikentrion muricatum (Plectronella papillosa). We cannot say, however, that all spines are based upon protorhabds. In the great majority of cases axial canals have not been demonstrated, and it is probable that the mode of formation of the spines is more in accordance with what is described under the next heading.

(j8) Adventitious Growing Points without Protorhabd Formation. I have dealt at some length with this type of secondary growing point in my account of the development of the anisodiscorhabd of Latrunculia. In the case of those growing points that are responsible for the initiation of nodal whorls of outgrowths, the evidence for the association of scleroplastids with the growing points, though almost entirely indirect, is, as we have already seen, very strong. Apparently, wherever a scleroplastid settles it forms a centre of activity for the deposition of silica, but in the cases now under discussion it does not elongate into a protorhabd and is usually quite indistinguish- able. We may assume that its growth is prevented by the fact that it is immediately sealed up in silica, as in the final stage of reduction of the triaene in Stelletta haeckeli (fig. 16, PI. 1), or in the apical knob of the anisodiscorhabd of Latrun- culia bocagei (fig. 47, PI. 3) as contrasted with that of Latrunculia apicalis (Text-fig. 1) ; but it is difficult to understand why it does not remain visible as an imbedded granule, as in the case of the ' silica pearl'. We must remember, however, that even the protorhabd, or axial thread, is not always recognizable without special treatment. Sometimes these secondary or adventitious growing points are symmetrically arranged and the spicule then assumes a very definite and characteristic form. Thus the development of the teeth or flukes on the chelae (and ancorae) of the Desmaci- 48 ARTHUR DENDY donidae (Text-figs. 11, 15) appears to be due to the establishment of secondary growing points. The most usual number of such outgrowths is three at each end, but it is subject to variation, sometimes even in the same sponge, but more often from species to species. This meristic variation can be explained, as in the case of the rays of the aster, by variation in the number of divisions of a scleroplastid. I see no other way of accounting for it. In other cases the entire spicule acquires an irregular, branching character, as in the remarkable desmas of the

TEXT-ITO. 13.

Tetracrepid desma of Discodermia emarginata. x 153. A, very early stage (orepis). B, older stage. C, end of branch of fully grown spicule. Lithistidae. It has long been known that these spicules are of two principal types, which Sollas has termed tetracrepid and monocrepid respectively. In the former case the crepis, or primary spicule, is four-rayed, like an ordinary tetract. In the latter it is diact or monact, with only a single axis. We may confine our attention to the former. The tetracrepid desma (Text-fig. 13) frequently shows four straight axial canals radiating from a common centre, indicating the presence of four protorhabds, arranged in the usual tvay, as the foundation around which silica is deposited. At first the young spicule is strictly tetract (Text-fig. 13, A), but presently secondary growing points are established which cause the rays to branch SPONGE-SFICULES 49 out very irregularly in all directions (B). The axial canals seem never to be continued into these branches, which may become very numerous and form interlocking processes (C) whereby the desmas may be all connected together into a continuous skeleton. . It cannot as yet be demonstrated that the secondary branches of the desma are initiated by scleroplastids, but it seems not unlikely. There must • be something to account for the fact that the spicule departs so widely from the type defined by its protorhabds and we can hardly attribute the branching solely to the behaviour of the silicoblasts.

(e) Mechanical Influences affecting the Form of the Spicule. There can be no doubt that the form of the spicule is often determined to a large extent by purely mechanical causes, usually dependent upon the situation in which it develops. We must content ourselves with a few illustrations. (a) The Conversion of dermal Spicules into Plates. In the Lithistid genus Discodermia there is a surface layer of thin siliceous plates, with irregularly rounded outlines and usually a short spike projecting inwards from the middle of each (Text-fig. 14). Development shows that this spicule is a modified tetract (a discotriaene) in which the three cladi have flattened out and fused together while the shaft is represented by the spike (which in some cases is completely suppressed, leaving a mere disc). These spicules develop in situ and are not, as in the case of the dermal spicules (anisodiscorhabds) of Latrunculia, transported, fully formed, from the interior of the sponge to the surface. The flattening and fusion of the cladi is probably due to pressure against the overlying dermal membrane. One little feature of this type of spicule deserves special notice. The edge of the disc is frequently notched in one or more places. The meaning of these notches is quite unintel- ligible so long as the spicule is considered merely as an isolated NO. 277 E 50 ARTHUR DENDY structure, and I am not aware that it has hitherto received any explanation. But when the spicules are examined in their natural relations it is' at once seen that the discs overlap one another, like scales, and that the notch in one disc is due to the presence of the shaft of another passing inwards and interrupting the growth. There can be no doubt about the purely mechanical explanation of the notch.

TEXT-FIG. 14.

Discotriaenes of Diseodermia tuberosa. A, disc of fully developed spicule. x 73. B, C, ditto, showing marginal notches. X 73. D, tetract crepis. X120.

(|S) The Mechanical Influence of the so-called Mother-cell. We have already seen that the developing spicule may be completely enveloped, for a longer or shorter period, by a phagocyte, which may be silicoblast. In some cases this envelopment appears to be merely temporary ; in others it may possibly persist throughout the entire development of the spicule. Sometimes the cell-membrane forms a thin envelope which apparently exercises a restraint upon the growing spicule, manifested more especially in curvature of the long axis, as in toxa, sigmata, and chelae (figs. 59, 60, PI. 3). This factor was discussed long ago by Sollas (1888 a), in his report on the Tetractinellida of the ' Challenger ' Expedition, but it seems to me that the conception of scleroplastids is required SPONGE-SPICULES 51 to account for the first appearance of the teeth or flukes. Their spreading out into concave films may well be explained by pressure against the restraining envelope. A very interesting case is that of the sterraster of Geodia. This spicule has an immense number of very slender rays, diverging from a common centre. The rays are at first separate, but presently the interspaces between them get filled up with silica, and a peculiar ornamentation develops at the surface of the solid mass thus formed. In one place this is interrupted by a depression known as the hilum, in which the nucleus of the silicoblast lies during the later part of the development. Obviously the presence of the nucleus serves as a mechanical hindrance to the deposition of silica.

(y) Other Mechanical Influences. With the mechanical forces referred to in the last two sub- sections, which may all be resolved into pressure or restraint upon some part or another of the growing spicule, may be contrasted those forces that result in the appearance of whorls of outgrowths upon nodal points. The formation of such whorls has already been discussed in previous papers (Bendy, 1917, 1921 a ; Dendy and Nicholson, 1917) and dealt with in summary fashion in earlier parts of the present communication. so that it is unnecessary to say much about it here. The vibratory theory, propounded by myself with the mathematical support of Professor Nicholson, assumes that the protorhabd, perhaps already with a thin coating of silica, is in a state of active vibration, and that the adventitious scleroblasts settle upon the nodes because these are the positions in which there is least movement. The position of the whorls in the discorhabds of Didiscus and Latrimculia affords striking confirmation of this view. In Didiscus, where alone it has been possible to make measurements of sufficient accuracy for mathematical treatment, the observed positions agree in a striking manner with those derived from Professor Nicholson's elaborate calculations. In Latrunculia bocagei there is a displacement of the whorls such as might be expected from the fact that the E2 52 ARTHUR DENDY young spicule is weigh'ted at one end by the early development of the apical knob, while there is no such displacement in the case of Latrunculia apicalis, where the apical and basal knobs develop simultaneously and the shaft is therefore weighted at both ends symmetrically. In a third species, in which the knob at the basal end of the spicule develops first, the displacement of the whorls is in that direction. I have suggested (1917) that the supposed vibration of the protorhabd may be caused by the shaking that must result from the passage of streams of water through the canal system of the sponge. Another possibility presents itself in view of the symbiotic theory of spicule origin. If the protorhabd is a living organism akin to a bacillus, it may sometimes have a vibratory movement of its own. Another mechanical factor that ought certainly to be taken into account is that of surface tension. If surface tension can bring about the arrangement of the drops of water along a telegraph wire, or of the microscopic beads on a spider's web, it is certainly worth while considering whether it may not also play a part in the distribution of secondary growing points on the axis of the spicule or on the margin of a whorl. This is a problem that remains to be investigated.

4. COMPARISON OF THE DEVELOPMENT OF CALCAREOUS SPONGE-SPICULES. It is obvious that the views set forth in this paper as to the origin and development of siliceous sponge-spicules differ fundamentally from the conclusions arrived at by Minchin as the result of his elaborate study of spicule-formation in the Calcarea. It will be remembered that, according to Minchin (1898, 1908), in the formation of a triradiate calcareous spicule, or spicule system,, three cells (actinoblasts) derived from the 1 dermal epithelium' come together in the mesogloea and arrange themselves in a trefoil. The nucleus of each divides into two and a minute spicule-ray appears between the sister nuclei of each pair. Then each actinoblast completes its 9PONGE-9PICULES 53 division, and one of each pair of ' formative cells ' thus produced migrates to the end of the growing ray and leaves the spicule, while the other remains for a time at the base of the spicule ; but when this is sufficiently thickened by the deposition of calcite, the latter also migrates to the end of the ray, to which it may remain attached for some time. The three rays of the spicule are thus at first separate from one another but soon become united in a common centre from which they radiate. In the quadriradiate spicule the fourth ray is said to be derived from a distinct source, viz. a gastral actinoblast, derived from a porocyte. The other types of spicule need not concern us. There are two features about this account that render it very difficult to accept. In the first place it altogether fails to explain the existence of what Minchin himself terms the ' organic axial thread ', a well-known feature of the calcareous spicule, and evidently of the same nature as the protorhabd of the siliceous spicule. In the second place it seems highly improbable that the actinoblasts (or calcoblasts as I prefer to call them) should come together in threes in such a definite manner unless there were something to attract them. I assume, therefore, that each triradiate spicule is really initiated by a group of three very minute scleroplastids, probably formed by division of a single one, and that these are enveloped by the calcoblasts either before or after they have begun to elongate. In the formation of the quadriradiate it seems most probable that the fourth ray is initiated by an adventitious scleroplastid. It appears to me that this view is quite as con- sistent with Minchin's actual observations as that adopted by him, and it completely harmonizes with my suggestions as to the origin and growth of the siliceous spicule.

5. COMPARISON OF THE SPONGIN SPICULES OF DARWINELLA. In the genus Darwinella alone amongst the Euceratosa (or true horny sponges) have spicules been observed, and these are very different from those of either the siliceous or cal- 54 ARTHUR DENDY careous sponges, being composed of spongin, a proteid substance closely related in chemical composition to silk. The spicules consist of three or four slender, tapering rays diverging from a common centre, and they are quite detached from the ordinary spongin skeleton, which coexists with them. No axial thread or canal has been observed, though I have examined them carefully, by different staining methods, from this point of view. Nevertheless it seems almost certain that the spongin must be deposited around some axis, and the fact that this has not yot been shown to exist may perhaps be explained by its close agreement in chemical and physical properties with the enveloping spongin. The spongin appears to be secreted by a, surrounding sheath of spongoblasts as in the case of the ordinary fibres of the horny skeleton. There is no evidence at all of origin within, or envelopment by a single ' mother- cell '.

6. CONSIDERATIONS AS TO THE NATURE OF THE SCLEROPLASTIDS. (a) Their microscopical Characters, Mode of Growth, and Multiplication. In the almost complete absence of direct observation, very little can be said as to the microscopic characters of the scleroplastids. They have probably been seen by many observers, but not recognized, amongst the countless granules (including bacteria) with which the mesogloea of the sponge is usually filled. It is only in cases where they have been, so to speak, picked out by silicoblasts to form the centre of ' silica pearls ' that they have so far been recognized at all. We learn from such cases that the scleroplastid is a minute granule about the size of a micrococcus. After it has elongated to form a protorhabd it is more readily distinguishable, and we can often see this protorhabd, as the axial thread, lying in the axial canal of the spicule, where it can be stained fairly darkly by such dyes as borax, carmine, and brazilin. The protorhabd is always, so far as my experience goes, very slender, and never seems to SPONGE-SPICULES 55 increase in diameter beyond that of the scleroplastid, though the axail canal in eroded spicules often becomes relatively wide. It seems to be generally admitted that it is composed of a proteicl substance, the so-called ' spiculin '. The growth in length of the protorhabd is clearly shown in the development of the anisodiscorhabd of Latruncnlia apicalis (Text-fig. 1). This growth appears to be strictly limited for each type of spicule, but it may bo very great. In the huge transfixing spicule of Monoraphis, which may be two or three feet long and as thick as a lead pencil, the protorhabd has a corresponding length, and it cannot be very much shorter in the spicules that form the ' glass-rope ' of Hyalonema. The existence of such huge spicules is hardly reconcilable with the view that the spicule is the intracellular product of a single mother-cell, but is readily explained by the hypothesis of an elongating protorhabd attacked by relays of silicoblasts. In spite of my previous conception of ' initial cells ' as agents in spicule-formation, there is no evidence that the scleroplastids are cells in the proper sense of the term. They tire certainly not nucleated masses of protoplasm, but seem rather to resemble bacteria, with which I believe them to have much in common. The fact that they multiply by fission appears to me to be demonstrated by the evidence brought forward in Section 2 of this paper, and it also seems clear that fundamental differences in the mode of division and in the arrangement of the products are responsible for the differences between the chief types of spicule. Abnormalities in these processes result in the formation of abnormal spicules, such as complete and incomplete twins and spicules with adventitious rays.

(b) Their Specificity. It is evident that the form of the spicule is determined primarily by the scleroplastids, and it must also be evident that, when we come to details of structure, it is not merely a question of the mode of division and arrangement of the products. We have already seen strong reason for believing 5G ARTHUR DENDY that the scleroplastids have, in some degree, what, for want of a better term, we may call a ' specificity ' of their own ; that, in the triaene, for example, one is predestined to form the shaft and the others the cladi, and that whether shaft or cladi will be formed is not determined merely by position. The form of the spicule, within the limits of the species, especially in the case of microscleres, is an extraordinarily constant feature. Take, for example, the remarkable chelae and ancorae of the Desmacidonidae (Text-figs. 11, 15). There may be many thousands of each kind scattered promiscuously throughout the mesogloea of the sponge, and no substantial variation may be discernible amongst them. Each kind has well-defined specific character which it rarely, if ever, fails to show, though of course there is a certain amount of variation as regards size and (in a few cases) as regards the number of the teeth or flukes. This specificity, together with the fact that it is handed on by heredity from generation to generation of scleroplastids, is very beautifully demonstrated in the curious case of Cina- chyra vaecinata (Dendy, 1921 b), in which the shafts of all the triaenes (both protriaenes and anatriaenes) end in elongated knobs, while the cladi are pointed in the usual manner. It might, of course, be argued that this very unusual condition is due to some peculiarity of the silicoblasts, but, if so, why does it occur only on the shafts ? (c) The Occurrence of trimorphic Chelae and its Significance. We come now to a very remarkable and suggestive phenomenon. In many species of Mycale and Esperiopsis three distinct categories of chelae are met with, all in large numbers. Sometimes the differences depend principally upon size, in other cases there is a great difference in shape also. Take, for example, Mycale novae-zealandiae (Dendy, 1924a). Here the difference in size between the three types of anisochela is very marked. The difference in shape is hardly less obvious, and is seen to depend primarily upon the angle that SPONGE-SPICULES 57 the median tooth or fluke at the large end of the spicule makes with the shaft, this angle being least in the smallest spicule and greatest in the largest, so that one of the three categories is intermediate both as regards size and shape between the other two. These differences are clearly shown in Text-fig. 15. Were it not for the very frequent occurrence of this phenomenon of trimorphism I should be inclined to pay but little attention to it. Under the circumstances, however, it seems worth while to suggest that it may be due to hybridization ; the

TEXT-TIG. 15.

Trimorphic anisochelae of Mycale novae-zealandiae, front and side views, x 485. three forms representing respectively the two parents and the hybrid. Such an interpretation implies the power of conjuga- tion on the part of the scleroplastids, and, if it could be established, would afford strong evidence that the latter are quite distinct organisms from the sponge.

(cZ) The Want of Correlation between the Spicules and the Sponge. The facts that sponge-spicules occur in such an immense variety of forms and that it is possible, as I have endeavoured to show in my memoir on the Tetraxonid sponge-spicule (1921 a), to arrange these forms in what appear to be well- defined evolutionary series, also seem to me to indicate that the scleroplastids are separate organisms; for it is quite 58 ARTHUR DBNDY impossible, in the vast majority of cases, to correlate these series with the evolutionary stages of the canal system and other parts of the sponge. That the spicules actually have undergone a course of evolution, suggesting that of independent organisms rather than that of mere organs or parts, is indicated not only by the seriation of the almost endlessly varied forms, but also by the fact that it is sometimes possible to trace in the actual development of the individual spicule a recapitulation of what must be ancestral stages. The best instance of this kind known to me is that of the remarkable pseudopolyact spicules of C yam on vickersii, which I have discussed in the memoir referred to (1921 b). Except in a few cases, which are readily explained, the spicules appear to vary quite independently of the sponge itself, and the smaller ones among them, in spite of their very definite and frequently complex forms, seem to have no physiological significance for the sponge whatever. It is true that in many cases the sponge utilizes the larger spicules (megascleres), and sometimes even the smaller ones, in a singularly appropriate manner, as we shall see presently, but the latter (microscleres) are usually found scattered anyhow in the mesogloea, without orientation and without arrangement, and many of them, apart from the impossibility of correlating their very remarkable and specific forms with any physiological requirements on the part of the sponge, are too minute to be of any con- ceivable use.

(c) The Dropping out of Spieule Categories in Sponge Phylogeny. The classification of the tetraxonid sponges is often rendered extremely difficult by the fact that entire spicule categories frequently drop out in the course of evolution. Many species and genera have been based upon the absence of some type of spicule that is present in evidently closely related forms. Nay, this process has gone beyond the making of genera and SPONGE-SPICULES 59 species, and been extended, in at least one case, to the supposed family, for it is now generally recognized that Sollas's ' Epi- polasidae ' are nothing but a collection of Stellettid genera in which the triaenes have disappeared and which are severally related to quite distinct genera of normal stellettids. I havo gone into this question in my report on the Homosclerophora and Astrotetraxonida of the ' Sealark ' Expedition (1916 c), where I have shown that pairs of closely related species aro frequently met with, one with triaenes and the other without— one normal and the other ' lipotriaenose '. Similarly, amongst the Desmacidonidae, we frequently come across ' lipochelous ' species, in which the chelae, so characteristic of the family, are entirely wanting. Thus, the species of Biemna may be regarded as lipochelous Mycales, and the same relation exists between Aulospongus and Microciona, Crella, and Yvesia. The loss of spicules may extend to more than one category. The genus Chondrilla evidently consists of corticate stellettids from which all the megascleres have disappeared, leaving only asters, and the aster of Chondrilla sacciformis is so different from that of other Ohondrillas as to suggest the origin of that species from a distinct genus of Stellettidae. Finally, in Chondrosia, both microscleres and megascleres are entirely wanting, and we are left with a sponge altogether devoid of spicules. This circumstance has led certain spongologists to associate Chondrosia with such primitive forms as Oscarella and Halisarca, but the histology and canal system clearly demonstrate its true position near Chondrilla. This dropping out of entire spicule-categories in the course of evolution might, as I have already (1921 a) suggested, be explained by the loss of Mendelian factors from the germ-plasm of the sponge, but it seems much more likely that it is due to failure of infection by the appropriate scleroplastids. We have here a further indication that for each kind of spicule a particular kind of scleroplastid is required, and that the scleroplastids are distinct organisms. 60 ARTHUR DENDY

(/) The sporadic Distribution of certain Spicule Types. Particular lines of spicule evolution seem to be so intimately associated with corresponding lines of phylogenetic descent amongst the sponges that there is very rarely any transference of scleroplastids from one line to another. Most of the cases of apparently anomalous spicule associations are to be explained by convergent evolution of quite distinct types of spicule, as in the case of the numerous forms of pseudaster. This is discussed in my memoir on the tetraxonid sponge-spicule (1921 a). There are, however, a few cases which seem to go beyond the probabilities of mere convergence. The trichodragma, for example (Text-fig. 10, A), is extraordinarily widely and sporadically distributed, both amongst the Astro- and (Sigmatotetraxonida, and suggests cross-infection from one line of sponge descent to another. It occurs in some stellettids, in some tetillids, and in some desmacidonids, but may be absent in one species of a genus and present in another. It does not seem as if mere dropping out of the scleroplastids along particular lines of descent could explain this distribution, for apparently it has not only dropped out but come in again. As, however, the trichodragma is a very simple type, it may have arisen independently on more than one occasion. The very regular and apparently typical asters of Vibu- linus stuposns form another case that is very difficult to explain by convergent evolution. They may be modified acanthostyles, such as are known in other genera, but they may equally well be due to cross-infection. Lastly, we have the curious spongin spicules of Darwinella, in a group where there ought to be no spieules at all, which may quite possibly be formed by the deposition of spongin instead of silica around triradiate or quadriradiate protorhabds which have been accidentally introduced as scleroplastids from some other line of sponge-descent. SP0NGE-6PICULES 61.

{g) The Scleroplastids as symbiotic Organisms (Sclerococci). Taken altogether the evidence seems to me to point very strongly to the interpretation of the scleroplastids, or sclerococci as they may be called from this point of view, and of the protorhabds into which they so frequently develop, as symbiotic organisms, and this is the interpretation that, after very careful consideration, I feel justified in inviting my zoological colleagues to accept. The occurrence of symbiotic bacteria is by no means unknown in the animal kingdom, and it has even been suggested, though I believe by no means proved, that ' mitochondria are, in reality, bacterial organisms, symbiotically combined with the tissues of higher organisms '-1 If not actually bacteria I would suggest that the sclerococci are at least descended from that group. It is doubtful whether they are now capable of existing apart from the sponge. If they were we should expect a far more sporadic and irregular distribution of spirals types than actually exists. The view that they are related to the bacteria is supported by the very minute size of the sclerococcus as observed ' fossilized ' in the centre of the siliceous pearl, and by the form of some of the smaller and simpler microscleres. I would refer the reader more especially in this connexion to the minute spirulae (only about 0-012 mm. in length) which crowd the mesogloea of Trachycladus to such an extent as to give it a quite characteristic opacity (Text-fig. 9). It is possible that if this sponge were examined alive one might find some of these spirulae in an unsilicified condition and even exhibiting bacteroid movements. It is quite probable that unsilicified sclerococci and protorhabds have frequently been observed in different sponges and mistaken for ordinary bacteria. I have myself seen what I took to be elongated rod-shaped bacteria of rather large size in a living siliceous sponge from Plymouth, x I. E. Wallin (' Amer. Journ. Anat.', vol. xxx, no. 1. Quoted in' Science Progress \ October 1924). 62 ARTHUR DENDY and I have found almost if not quite naked protorhabds in another case, but this is a line of investigation as yet practi- cally untouched. The enormous length attained by the protorhabds (axial threads) in some large spieules, such as the transfixing spicule of Monoraphis, seems opposed to the view that they are actually bacteria, and suggests that they have acquired special powers of growth as a result of their association with the sponge. It is generally admitted that the ' axial thread ' consists of a proteid substance, and its power of growth is unquestionable (compare Text-fig. 1); why, then, should we not regard it as a protoplasmic organism ?

7. THE EELATIONS BETWEEN THE SCLEROCOCCI AND THE SPONGE. The relations between the sclerococci and the sponge in which they occur are obviously very intimate and very definite. We may suppose that they had their origin in that tendency which animals in general exhibit to isolate intrusive foreign bodies by the secretion around them of some kind of envelope. The analogy with pearl-formation is very close, for it has been shown that even in the pearl the centre is frequently formed by some minute parasite. In the case of the siliceous sponge, however, it seems to be only the sclerococci or protorhabds that excite the silicoblasts to pour out their secretion, though when this process has once been commenced it may be continued by other silicoblasts laying down silica upon the foundations already established. The same is doubtless true, mutatis mutandis, for calcareous sponges. In neither of these groups do we ever find silica or calcite deposited around ordinary foreign bodies, such as grains of sand. In the horny sponges, on the other hand, we frequently find sand-grains and foreign spieules enveloped in spongin, and in the pseudocerotose sponges the proper siliceous spieules of the sponge are regularly united together by spongin into a coherent skeleton. The fact that silicoblasts (or calcoblasts) always seek out SPONGE-SPICULES t)3 scleroplastids and protorhabds, or already partially formed spicules, for the scene of their activities, points to the existence of a chemotactic attraction between the two, and a similar attraction may also be responsible for the manner in which adventitious scleroplastids settle upon partially formed spieules and start secondary growing points. If the scleroplastid is at once completely enveloped by silica no protorhabd will be formed. This is the case with the silica pearls and apparently with most secondary growing points. ' More usually the scleroplastid elongates into a protorhabd, the growth of which is so accurately adjusted to the rate of silica-deposition that a long, tapering ray results. Another important question that has to be answered is that of the manner in which the individual sponge first became infected with its sclerococci. It seems to me obvious that not all the sclerococci are destroyed by envelopment in silica, some of each kind must normally remain over to be transferred to the next generation, and this transference is probably effected by egg-infection.1 Herein we find our justification for regarding the association of the sclerocoeci and the sponge as one of symbiosis rather than of parasitism. The sclerococci on the whole benefit by finding food and shelter, though many of them are destroyed. The sponge obviously benefits by finding materials for the formation of a very efficient skeleton. r In free-swimming embryos of siliceous sponges the different spicule categories are sometimes already present and arranged in a very definite manner. The best example of this known to me is the embryo of Esperia lorenzii described and figured by Maas (1892 c), in which the megascleres occur in a compact bundle in the middle, surrounded by the microsclercs in more or less regular order. These altogether form only a minute fraction of the number of spicules present in the fully grown sponge, so that it is obvious that some of the sclorococci

1 Compare the observations of Buchner upon the transmission of luminous bacteria in sepia (' Thier und Pflanze in intrazellularer Nym- biose', by Professor P. Buchner. Reviewed by Professor Gamble in 1 Nature ', April 29 and May 6, 1922). 64 ARTHUR DENDY must escape immediate attack by silicoblasts and multiply very greatly by fission. The phagocytic character of the sponge ovum in its amoeboid phase (compare Dendy, 1914 b) is probably at least partly responsible for its infection by sclerococci. We may suppose that it simply engulfs them along with other food-particles, but fails to digest them. It may be argued against the views here put forward as to the nature and origin of sponge-spicules that these spicules are frequently adapted in form and arrangement to the requirements of the sponge in building up its skeleton. It is true that in certain situations the spicules exhibit an appropriate form. This, however, is sometimes due to purely mechanical causes, as in the case of the plate-like dermal spicules of Discodermia (Text-fig. 14), which develop in situ. More often the apparent adaptation seems to be due to the fact that the sponge is able to make use of certain forms of spicule that happen to have arisen in the course of evolution and disposes of them in the most appropriate manner. Evans (1899 b) long ago described how the amphidiscs of Ephydatia are developed in the mesogloea and carried by amoebocytes to their position in the wall of the granule, to which they seem so remarkably adapted. In typical species of Latrunculia, again, the curious anisodiscorhabds (figs. 32, 33, PI. 2) appear wonderfully adapted to form a superficial armour. Each has a manubrium by which it is attached to the surface of the sponge and one or more projecting spines, and they are arranged with the utmost regu- larity, closely together side by side in an uninterrupted layer. The study of the development of these spicules, however (see Section 2 (/)), entirely precludes the idea that their remarkable and complex form is due in any way to the position which they are destined to occupy, for they attain their full development in the mesogloea of the sponge, at some distance beneath the surface, to which they must be transported in some way by the sponge, presumably by the activity of amoebocytes. If the sponge finds itself in possession of simple linear spicules it usually arranges them in bundles or ' fibres ' to form a reticu- SPONGB-SPICULES 65 late skeleton, as in the Chalininae and many other Sigmato- tetraxonida. A very curious illustration of this principle is found in some of the so-called Pharetronidae amongst the Calcarea. In this family a not uncommon type of spicule is the ' tuning-fork ', in which two rays of a triradiate are turned forwards and lie close together, parallel with one another

TEXT-FIG. 16.

A, reduced triradiate of Kebira uteoides. X 140. B, reduced triradiate of Grantiopsis cylindrica. x 140. C, tuning- fork spicule of Lelapia antiqua. x 140.

(Text-fig. 16, C). The spicule thus becomes suitable for forming compact bundles or ' fibres ', and in the genus Lelapia is utilized accordingly by the sponge. In the related genus Kebira the bundled spicules are reduced very nearly to simple linear forms by the almost complete suppression of two of the rays (Text-fig. 16, A). That the modification of these spicules may have come before, and not as a consequence of, their association in bundles, is indicated very clearly by the case of Grantiopsis cylindrica, in which the triradiates of the KO. 277 F 66 ARTHUR DBNDY tubar skeleton, still occupying their primitive positions in the walls of the radial chambers, have already been reduced to a simple linear form by the almost complete suppression of two of the rays (Text-fig. 16, B). It would be easy to give other examples to show that the sponge selects such spicules as it can make use of and disposes of them in the most appropriate manner in the formation of its skeleton. Those that it is unable to find a use for, including the great majority of the microscleres, it either leaves scattered promiscuously through the mesogloea, or removes to the surface, whence they are probably sooner or later thrown off. It seems very wonderful that the sponge should have this selective power and this ability to make appropriate use of the materials with which it is, accidentally so to speak, provided. But something very similar occurs amongst the Protozoa. Foraminifera such as Haliphysema select broken sponge-spicules from amongst their surroundings for the purpose of building up their skeleton, other genera select different materials. Euglypha is said to secrete siliceous or chitinous plates of definite form in the interior of the cytoplasm, which it afterwards builds up into a regular skeleton on the surface. The only other cases known to me in which anything occurs that is at all similar to the manner in which I believe sponge- spicules to be formed are afforded by those very anomalous sponges Merlia and Astrosclera. In these genera there exists a solid calcareous skeleton of a very peculiar type, in addition to the ordinary skeleton of siliceous spicules, which is fully developed. According to Kirkpatrick (1912 6, e) the calcareous skeleton is composed of spherules of carbonate of lime, each deposited around a minute symbiotic alga (or ' monad ') as its centre. The parallel is very close, but the spherules of calcium carbonate are not spicules and they are not composed of the material ordinarily used by the sponge in spicule-formation ; nor does the symbiotic organism appear to be the same. The whole process is exceptional and anomalous, and although I have been aware of my friend Mr. Kirkpatrick's very interesting observations since they were first published, SPONGE-SPICULES 07 I have not been consciously influenced by them in arriving at my own conclusions with regard to spicule-formation. I believe, however, that they afford considerable supp*ort, of a collateral character, to my views. Other cases of a similar nature will probably be found to exist, and I would suggest that the skeleton of the Eadiolarians and of the Alcyonarians is well worth investigating from this point of view. The perfection of skeleton formation in sponges, so far at least as the production of a hard protective and supporting structure is concerned, is met with in those forms in which the spicules, whether siliceous or calcareous, are all united together into a stony framework by the secretion of silica or calcite around them collectively. I will give only one illustration, which I have recently come across in the course of my investigations of certain deep-water sponges from the Indian Ocean belonging to the Calcutta Museum. In a hexactinellid sponge, to which I have given the manu- script name Hexactinella minor, the skeleton at first consists of regular hexacts with slender rays, irregularly scattered through the soft tissues. After these have been fully formed a wave of secondary silica deposition passes over them, spreading from spicule to spicule wherever the rays happen to come into contact, and uniting them all into an irregular network in which it is hard to detect the original hexacts (fig. 62, PI. 3). We must assume that this secondary deposit is secreted by silicoblasts that wander over the surfaces of the spicules and pass from one to another as opportunity offers.

8. THE TAXONOMIC VALUE OF SPONGE-SPICULES. We can hardly take leave of our subject without referring to the possible bearings of our conclusions upon the classification of the Porifera. It might be argued that if the spicules are merely fossilized sclerococci and protorhabds, so to speak, their form being determined rather by these organisms themselves than by the sponge, we are not justified any longer in attributing to them prime importance as indices of the genetic F2 68 ARTHUR DENDY affinities of the sponges in which they are found. They might still be used for the purposes of a purely empirical and formal classification, but though such a classification might express the phylogenetic relationships of the sclerococci it could hardly be expected to express those of the sponges. Such an argu- ment, however, would, in my opinion, be entirely superficial, for if, as I maintain, the sclerocoeei are, with very rare exceptions, transmitted from generation to generation of sponges solely by egg-infection, their phylogeny must run in parallel lines with that of the sponges themselves, so that the solution of the one problem carries with it the solution of the other. Were it not for the existence of the spicules it would be extremely difficult to classify the Porifera at all, as we see in the many cases in which characteristic spicule categories have dropped out at some period or other of the ancestral history. The vast majority of the so-called ' species ' of sponges are, in fact, based upon small differences in spiculation, and without the spicules such ' species ' could have no existence. Are we, then, justified in speaking of them as ' species ' at all ? It depends upon how we define the term. If, as I believe, the term ' species ' is rightly applied to any distinguishable group of individuals closely resembling one another owing to com- munity of descent, and interbreeding to produce offspring like themselves but differing from the individuals of all other species, then there is no reason why we should not continue to use it in the case of the sponges and to define our species mainly in terms of spiculation. The only trouble is that if the symbiotic sclerococci were removed the specific differences would in most cases vanish with them, and this has probably happened in certain cases. The case is analogous to that of the lichens, amongst which botanists do not hesitate to recognize genera and species, although ' strictly speaking, both Fungi and Algae should be classified in their respective orders ; but the lichens exhibit among themselves such an agreement in their structure and mode of life, and have been so evolved as consortia that it is more convenient to treat them as a separate class '.1 The 1 Strasburger, ' A Text-book of Botany,' 3rd English edition. SPONGE-SPICULES 69 sponges, as a group, of course differ from the lichens (as regards their symbiotic character) in that there are many in which sclerococci, and consequently spicules, do not occur, and the absence of the spicules does not materially affect the essential sponge organization as expressed in the histology and canal system.

LIST OF LITERATURE REFERRED TO. Dendy, A. (1905).—" Report on the Sponges collected by Professor Eerdman, at Ceylon in 1902 ",' Rep. Pearl Oyster Fish. Gulf of Manaar. Roy. Soc.', vol. iii, 1905, pp. 56-246. (1914 6).—"Observations on the Gametogenesis of Grantia com- pressa", ' Quart. Journ. Micr. Sci.1, N.S., vol. lx, Part III, 1914 pp. 313-76. (1916 a).—" Report on the Non-Calcareous Sponges collected by Mr. James Hornell at Okhamandal in Kattiawar in 1905-6 ", ' Report to the Government of Baroda on the Marine Zoology of Okhamandal in Kattiawar ', Part II, pp. 93-146. London, Williams and Norgate, 1916. (1916 6).—" Report on the Hexactinellid Sponges (Triaxonida) collected by H.M.S. ' Sealark ' in the Indian Ocean ", ' Trans. Linn. Soc.', 2nd series, Zoology, vol. xvii, 1916, pp. 211-24. (1916 c).—" Report on the Homosclerophora and Astrotetraxonida collected by H.M.S. ' Sealark' in the Indian Ocean ", ibid., pp. 225-71. (1916 e).—" On the Occurrence of Gelatinous Spicules, and their Mode of Origin, in a New Genus of Siliceous Sponges ", ' Proc. Roy. Soc.', vol. lxxxix, Series B, pp. 315-22. (1917).—" The Chessman Spicule of the Genus Latrunculia : a Study in the Origin of Specific Characters ". Presidential Address, Quekett Microscopical Club. ' Journ. Quekett Microscopical Club ', vol. xiii, pp. 231-46. (1921 a).—" The Tetraxonid Sponge-spicule : a Study in Evolution ", ' Acta Zoologioa', vol. ii, pp. 95-152. (1921 b).—" Report on the Sigmatotetraxonida collected by H.M.S. ' Sealark' in the Indian Ocean", ' Trans. Linn. Soc.', 2nd series, Zoology, vol. xviii, Part I. (1924a).—" Porifera. Part I. Non-Antarctic Sponges". British Antarctic (' Terra Nova ') Expedition, 1910. ' Natural History Report, Zoology', vol. vi, no. 3, pp. 269-392. British Museum, London, 1924. (1924 6).—" On an Orthogenetic Series of Growth-Forms in certain Tetraxonid Sponge-spicules ", ' Proc. Roy. Soc.', Series B, vol. xcvii, pp. 243-50. 70 ARTHUR DBNDY Dendy, A., and Nicholson, J. W. (1917).—" On the Influence of Vibrations upon the Form of certain Sponge-spioules ", 'Proc. Roy. Soc.', Series B, vol. lxxxix, pp. 573-87. Evans, B. (1899 6).—" The Structure and Metamorphosis of the Larva of Spongillalacustris",' Quart. Journ. Micr. Sci.', N.S., vol. xlii,pp. 363-476. Hinde, G. J., and Holmes, W. M. (1892).—" On the Sponge Remains in the Lower Tertiary Strata near Oamaru, Otago, New Zealand", ' Journ. Linn. Soc.', vol. xxiv, no. 151, pp. 177-262. Kirkpatrick, R. (1908 c).—' Tetraxonida. National Antarctic Expedition Natural History ', vol. iv, 1908, pp. 1-56. (1912 6).—"Note on Merlia normani and the ' Monticuliporas' ", ' Proc. Roy. Soc.', Series B, vol. lxxxv, pp. 562-3. (1912 e).—" Note on Merlia ",' Nature ', June 6, vol. lxxxix, p. 353. Lebwohl, F. (1914 a).—" Japanische Tetraxonida. I. Sigmatophora. II. Astrophora metastrosa ",' Journ. of the College of Science Imperial University of Tokyo ', vol. xxxv, March 15, 1914. Lundbeck, W. (1902).—" Homorrhaphidae and Heterorrhaphidae", ' Danish Ingolf-Expedition', vol. vi, no. 1. (1905).-—" Desrnacidonidae (Pars.). Porifera, Part II ", ibid., no. 2. (1910).—" Desmacidonidae (Pars.). Porifera, Part III ", ibid., no. 3. Maas, O. (1892 c).—" Die Metamorphose von Esperia lorenzi O.S., nebst Beobachtungen an anderen Schwammlarven",' Mitt. Zool. Stat. Neap.', x, Heft 3, pp. 408-40. Metschnikoff, E. (1892).—" Lecons sur la pathologie compar6e de l'in- flammation, faites a l'lnstitut Pasteur en avril et mai 1891 ", Paris. Minchin, E. A. (1898).—" Materials for a Monograph of the Ascons. I. On the Origin and Growth of the triradiate and quadriradiate Spicules in the Family Clathrinidae ", ' Quart. Journ. Micr. Sci.', N.S., no. 160, vol. xl, Part IV, pp. 469-587. (1908).—" Materials for a Monograph of the Ascona. II. The Formation of Spicules in the Genus Leucosolenia with some Notes on the Histology of the Sponges ", ibid., vol. Hi, pp. 301-55. Ridley, S. O., and Dendy, A. (1887).—" Monaxonida. Report on the Scientific Results of the Voyage of H.M.S. ' Challenger' ", ' Zoology', vol. xx. Schmidt, O. (1868 a).—' Die Spongien der Kiiste von Algier, mit Nach- tragen zu den Spongien des Adriatischen Meeres ' (Drittes Supplement). Leipzig, 1868. Schulze, F. E. (1893 6).—" t)ber die Ableitung der Hexactinelliden-Nadeln von regularen Hexactine ", ' Sitzb. der Kon. preuss. Akad. der Wiss. zu Berlin', 1893, vol. xlvi, pp. 991-7- (1894 6).—" Hexactinelliden des Indischen Oceans. I. Theil. Die Hyalonematiden ". ' Abhand. der K. preuss. Akad. der Wiss. zu Berlin', 1894. SPONGE-SPICULES 71 Schulze, F. E. (1895).—" Hexactinelliden des Indischen Oceans. II. Theil. Die Hexasterophora ", ibid., 1895.. . (1887 c).—" Hexactinellida. Report on the Scientific Results of the Voyage of H.M.S. ' Challenger ' ", ' Zoology ', vol. xxi. London, 1887. r (1899 a).—' Amerikanische Hexactinelliden nach dem Materiale det Albatross-Expedition.' Herausgegeben mit Unterstiitzung d. Kgl. preuss. Akademie der Wiss. Jena. (1904).—" Hexactinellida ", ' Wiss. Ergebn. Deutsch. Tiefsee-Exped. Valdivia', Bd. 4, viii. Sollas, W. J. (1879 a),—'' On Plectronella papillosa, a new genus and species of Echinonematous Sponge ", ' Ann. Mag. Nat. Hist.', vol. iii, 1879, pp. 17-28. (1888a).—"Report on the Tetractinellida collected by H.M.S. 'Challenger' during the years 1873-6'% '"Challenger" Reports', xxv. Thiele, J. (1900).—" Kieselschwamme vonTernate ",' I. Abh. Senckenb. nat. Ges. Frankfurt a. M.', Bd. 25, pp. 17-80. Topsent, E. (1896 c).—" Campagnes du Yacht ' Princesse-Alice '. Sur deux curieuses Esperellines des Acores ", ' Bull. Soc. zool. France ', vol. xxi, no. 7, pp. 147-50. Weltner, W. (1901).—" Siisswasserspongien von Celebes." (Spongilliden- studien, IV.) ' Arch. Nat.', Jahrg. 67, Festschr. Ed. von Martens, Beiheft, pp. 187-204. Woodland, W. (1908).—" Studies in Spicule Formation. VIII. Some Observations on the Scleroblastic Development of the Hexactinellid and other Siliceous Sponge Spicules " ' Quart. Journ. Micr. Sci.', vol. Hi, pp. 139-57. 72 ARTHUR DENDY

DESCBIPTION OF PLATES 1, 2, AND 3. Illustrating Professor Dendy's paper ' On the Origin, Growth, and Arrangement of Sponge-spicules: a Study in Symbiosis '.

EXPLANATION OF LETTERING. a.k., apical knob ; a.m., amoebocyte in mesogloea ; a.m., apical whorl; e g.p., extra growing point; e.sp., edge of silica plate; gr., granule; g.p., secondary growing point; hex., primary hexacts ; hi., kink in shaft; mn., manubrium ; TO.*., membrane of silicoblast; m.w., median whorl; n.s., nucleus of silicoblast; pr., protorhabd ; s.c, secondary or accessory axial canals ; Si., silicoblast; s.w., subsidiary whorl.

PLATE 1. Figs. 1-8. Sceptrospongia coronata Dendy M.S. Fig. 1.—Tylostyle. xl90. Fig. 2.—Stephanotyle. X190. Fig. 3.—Apex of tylostyle. x 1,070. Figs. 4—8.—Apices of stephanotyles, representing various stages of evolution actually existing side by side, x 1,070. Fig. 9.— Raphidotheca marshall-halli, apex of exotyle. X650. Figs. 10-26. Stelletta haeckeli Sollas. Okhamandal specimen (R.N. iv. 3). Figs. 10-15.—Suppressed triaenes. x280. Fig. 16.—Silica pearl, with central granule (scleroplastid). x 280. Figs. 17-19. Abnormal amphitriaenes, produced by incomplete twinning, x 280. Kg. 20.—Normal anatriaene. X 65. Fig. 21.—Normal oxeote. x65. Fig. 22.—Normal orthotriaene. x 65. Fig. 23.—Reversed anatriaene. x280. Fig. 24.—Anatriaene with adventitious cladi. X280. Fig. 25.—Suppressed orthotriaene with adventitious cladus. x 230. Fig. 26.—Abnormal orthotriaene. x 170. PLATE 2. Figs. 27-9. Stelletta haeckeli Sollas. Ceylon Specimen (B.M. Coll. 82.8.2.5). Fig. 27.—Middle portion of large oxeote spicule, with adventitious cladi. x 280. Fig. 28.—Stylote (?) spicule, with adventitious cladi. x 280. SPONGB-SPICULES 73 Fig. 29.—Anatriaene with adventitious rays, x 280. Fig. 29 a.—End of shaft more highly magnified, showing secondary axial canal, x 650. Fig. 30.—Stelletta haeckeli Sollas. Okhamandal specimen (R.N. iv. 3). End of style (?) with adventitious cladi. x280. Fig. 31.—Stelletta pathologica Schmidt. Abnormal megasclere, from type specimen in B.M. Coll. X170. Fig. 32.—Latrunoulia bocagei Ridley and Dendy. 'Challenger' specimen. Anisodiscorhabd. x 840. Fig. 33.—Latrunculia apicalis Ridley and Dendy. ' Challenger' specimen. Anisodiscorhabd. x 840. Figs. 34-45. Latrunculia bocagei Ridley and Dendy. ' Challenger' Specimen. Figs. 34, 35.—Two pairs of twin anisodiscorhabds. X 1,900. Figs. 36-45.—Early stages in the development of the anisodiscorhabd. X 1,900.

PLATE 3. Figs. 46-54. Latrunculia bocagei Ridley and Dendy. ' Challenger' Specimen. Figs. 46-7.—Later stages in the development of the anisodiscorhabd. X 1,900. Fig. 48.—Basal portion of young anisodiscorhabd with abnormally placed whorls just commencing, x 1,900. Fig. 49.—Fragment of young anisodiscorhabd showing whorl of growing points, of which two only are visible. X 1,900. Fig. 50.—Fragment of young anisodiscorhabd showing median whorl of three growing points and protorhabd. x 1,900. Fig. 51.—Median whorl of young anisodiscorhabd with extra growing point, x 1,900. Fig. 52.—Tripartite whorl of young anisodiscorhabd in which the growing points have developed into three flat lobes, x 1,900. Fig. 53.—Later stage of tripartite whorl in which the edges of the lobes have become crenate. x 1,900. Fig. 54.—Apical whorl of not quite fully developed anisodiscorhabd. X 1,900. Figs. 55-7.—Stages in the development of the anisodiscorhabd in Latrunculia sp. (Gilchrist Coll., South Africa), x 1,900. Fig. 58.—Trichodragma of Esperella fusca Ridley and Dendy, surrounded by silicoblast. x 1,900. Fig. 59.—Young anisochela of Cladorhiza inversa Ridley and Dendy, surrounded by silicoblast. X 1,900. 74 ARTHUR DBNDY Fig. 60.—Fully developed anisochela of Cladorhiza in versa Ridley and Dendy, surrounded by silicoblast. x 1,900. Fig. 61.—Fossil sponge spicule from Oamaru, New Zealand, showing crossing of axial canals. X 280. Fig. 62.—Part of skeleton of Hexactinella minor Dendy M.S., showing how the primary hexacts are gradually united in a continuous ramework by secondary silica deposition, x 125. .nrtfluruenay i tMuyr-Sco. Voi. 70, MS. 9

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