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Relationships between the "Pompeii worms"

Hydrothennal vents Polychaeta

Bactcria association

Sources hydrothennales Polychètes

Associations bactériennes and their epibiotic bacteria

ABSTRACf

RÉSUMÉ

INTRODUCTION

Françoise GAILL a, Daniel DESBRUYERES b, Lucien LAUBIER C

a Centre de Biologie Cellulaire, Centre National de la Recherche Scientifique, École Pratique des Hautes Etudes, 67, rue Maurice Günsbourg, 94200 Ivry-sur-Seine, Fmnce. b Institut Français de Recherche pour l'Exploitation de la Mer, Centre de Brest, B.P. 337, 29273 Brest, France. C IFREMER, 66, avenue d'Iéna, 75116 Paris, France.

The morphological relationship betwccn the so-callcd "Pompeii worms" (Alvinella caudata and Alvinella pompejana) and their associatcd bacteria is describcd. Three main morphological modifications are observcd in Alvinella caudata: segmental division and the density and size of filarnentous bacteria incrcase in a gmdient manner from the anterior to the posterior part. Such a gradient does not occur in ALvinella pompejana, which is characterized by the presence of dorsal expansions associatcd with filamentous bacteria. The underlying cell epidermis aspects are qui te different as far as bacteria association types are concerned, espccially in the notopod part of Alvinella caudata. These results permit an overview of the possible functioning of the biological ensemble constitutcd by the worm, its tube and the associatcd bacteria.

Oceanol. Acta, 1988. Hydrothermalism, Biology and Ecology Symposium, Paris, 4-7 November, 1985, Proceedings, 147-154.

Relations entre les "vers de Pompéi" et leurs bactéries épibiontes

Les relations morphologiques entre les vers de Pompéi (Alvinella caudata et Alvinella pompejana) sont décrites. Trois types de modifications morphologiques sont observés chez Alvinella caudata : une division des segments ; la densité et la taille des bactéries filamenteuses qui augmentent selon un gradient antéro-postérieur. Un tel gradient n'existe pas chez Alvinella pompejana qui est caractérisée par la présence d'expansions dorsales associées à des bactéries filamenteuses. Les aspects de l'épiderme sous-jacent varient suivant le type d'association bactérienne présent, en particulier ceux de l'épiderme parapodial d'Alvinella caudata. Ces résultats nous conduisent à présenter une synthèse du fonctionnement du système biologique constitué par le ver, le tube et ses bactéries associées.

Oceanol. Acta, 1988. Actes du Colloque Hydrothcrmalisme, Biologie ct Écologie, Paris, 4-7 novembre 1985, 147-154.

The so-called "Pompei worms" (polychaetous annelids) live on active hydrothermal edifices on the East Pacific Rise at depths of about 2600 m. They secrete organic tubes (Vovelle, Gaill, 1986 ; Gaill, Hunt, 1986) on the peripheral surface of certain chimneys and zinc-sulphide

diffusers (Dcsbruyères et al., 1985) in zones of activ.e mixing of hOl, reducing, acidic, mctal-rich fluid and cold, well-oxygenated scawater. This mixing results in a sharp horizontal gradient, where the tcmperature changes within a few decimetres from 200° to 1.8°C. The Pompeii worms are confincd to the cooler part of the gmdient, betwcen approximately 20° and 60°C (Dcsbruyères et al., 1982),

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1

F. GAILL, D. DESBRUYÈRES, L. LAUBIER

this being the range of their temperature tolerance defined as accurately as is possible. A minor portion of the worm, the branchiae, is positioned at the opening of the tube bathing in ambient water, whereas the greater portion is enclosed within the tube these animaIs secrete. The Pompeü worms comprise two spccies: Alvinella pompe jan a and Alvinella caudata (Desbruyères, Laubier, 1986), primitively considered as two forms of the same species (Desbruyères, Laubier, 1980). Several polychaetous species inhabit the dcep sea hydrothermal chimneys, but the Pompeii worms are the most spectacular with respect to the density of the bacteria covering their dorsal part (Gaill et al., 1984 a). These bacteria have bcen described (Gaill et al., 1987; Desbruyères et al., 1985), but the biological significance of the bacteria/worm association remains unclear. The Pompeii worms are devoid of the endosymbiotic associations (Storch, Gaill, 1986) described and studied in vestimentiferans (Cavanaugh et al., 1981; Bosch, Grassé, 1984 a, b; Felbeck, 1981) and molluscs (Cavanaugh, 1983; Fiala-Medioni, 1984). They present only epibiotic bacteria. The purpose of this study was to determine more precisely the morphological relationship betwecn the bacteria and the worm epidermis and to inv~stigate whether the underlying cell epidermis is modified by the presence of bacteria associations and, - if so - in what manner. Answers to these questions would provide some indications conceming the local environ ment of the animal and its epidennal reactions, as well as information about what may be called the process of exosymbyosis.

MATERIAL AND METHODS

Alvinella pompe jan a spccimens were collected at 200 50'N and 109°W with the research submersible Alvin (April­May 1979) and at 12°48'N and 103°56'W with the submersible Cyana (March 1982) at 2600 m depth. Ultrastructural studics were carried out on pieces of worm fixed with 0,4 M cacodylate-buffered glutaraldehyde (3% final concentration) at pH 7,2 and then posl-fixed with osmium tetroxyde (1 % final concentration), and embedded in Durcupan and Spurr resins. Thin sections were stained with aqueous uranyl acetate and lead citrate, and examined using a Philips EM 201 T E M (Centre de Biologie Cellulaire, CNRS, Ivry-sur-Seine, France). Scanning electron microscope observations were made on fixed samples dehydratcd with ethanol, critical point dried and sputter-coated with gold mctal. The samples wcre examined using a Cambridge Sloo SEM (IFREMER, Centre de Brest, France).

RESULTS

Whereas the cephalic and branchial regions of the two morphological forros within the species Alvinella are identical; the posterior sections of the body differ in their external anatomy, if not in thcir organization. In the caudata species, there is a gross dccrease in body diameter from the 49th to the 54th segment. The notopodia are elongated in this rcgion and bear four or five

bifurcatc digitations which are regular in form and arrangement In the antcrior region, dorsal segmentary giandular bourrelets are undivided (Fig. 1), whereas posteriorly they divide into an increasing number of subsections (Fig. 1 and 2 c, d). Concurrently the non-glandular inter­segmentary spaces increasc to a maximum size in the median region where the glandular bourrelets are not found. In the caudal region, the intcrsegmentary spaces are less dilated than in the median region, but are nevertheless more important than in the anterior region. In these spaces, bacilli, cocci and filamentous bacteria are associated with cuticular secretions, forming "cluster-like" structures (Fig. -l, 2 c, d). These associations are found along the cntire length of the animal (Fig. 1). However, the density and dimensions of filamentous bactcria increase posteriorly (Fig. 1, 2 d). Large filamentous bacteria, preferentially inserted in the apical portion of posterior notopods, form a veritable belt, easily visible to the naked eye (Fig. 2 a). The epidermis of the animal consists of an epithelium limited distally by a cuticle (Fig. 3 b) composed of a network of collagen fibres and bordered by an electron­dense epicuticle (Fig. 3 a). The epidermal cells issue

Figure 1

Parapodia transformation

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Increase of dorsal glandular torus parceiling

Increase of filamentous bacteria density

Diagram comparing morph%gica/ modifications in Alvinella cau­data and the associated bacteria densily. Schéma comparanl les modificaùons morphologiques d'A/vine//a caudata el la densité des bactéries associées.

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"POMPE" WORMS" AND THEIR EPIBIOTIC BACTERIA -----------------------------------------------------------------------------------------------------

Figure 2 A) posterior part of Alvinella caudata: the notopods present digitations (1, 2, 3). Filamentous bacteria are white. d, digitation, f, filamentous bacteria; p, notopod; s, setae; white arrow, minerai particles. B) Aspect of the dilated base of filamentous bacteria from the notopods. This base is covered by a portion of cuJicle which recovers microvillies. e, base of filamentous bacteria; f, ftlamentous bacteria; m, microvilli. C) Cluster-like associations in the inter­segmentary space (see 3 in fig. 2). a, inter­segmentary space; s, subsection of segments. D) Cluster-like associations in the caudal part where filamentous bacteria are numerous. f, ftlamentous bacteria; s, subsection of segments. A) Partie postérieure d'Alvinella caudata : les parapodes présentent des digitations (l, 2, 3). Les bactéries filamenteuses sont blanches. d, di­gitation; f, filaments bactériens; p, parapode; s, soie ; flèche blanche, particules minérales. B) Aspect dilaté de la base des filaments bactériens des parapodes. Cette base est recou­verte d'une portion de cuticule recouvrant les microvillosités ; e, base des filaments bacté­riens ; f, bactéries filamenteuses ; m, microvil­losités; C) Associations en bouquet de l'espace intersegmentaire (voir 3 sur la fig. 2). a, espace intersegmentaire ; s, partie de segment segmen­tée. D) Associations en bouquet de la partie postérieure où les bactéries filamenteuses sont nombreuses. f, bactéries filamenteuses ; s, partie de segment segmentée.

10wards cellular extensions of microvil1i which traverse the cuticle perpendicularly (Fig. 3 e). These microvilli ramify 10wards the epicuticle and their more distal portions form epicuticular projections or ellipsoidal bodies on the exterior of the epicuticle (Fig. 3 a). Also, SEM observations of the surface integument reveal a granular appearance associated with the presence of epicuticular projections (Fig. 4 d). The epithelium in the dorsal intersegmentary zones consists of a layer of cells overlying a basal lamina. This tissue is traversed by abundant blood vessels and is proximal to numerous nerve terminations. The dorsal epithelium comprises unspecialized cells and mucous cells which are much less numerous in the dorsal glandular flanges than in the ventral epithelium; they are absent in the intersegmentary zone where unspecialized epidermal cells occur. These cells are characterized by the presence of Golgi apparatus, an abundance of granular endoplasmic reticulum and the presence of numerous mitochondria. Inclusions are present in large numbers in the cytoplasm: vesicles of all sizes near the cellular apex, lysosomes of various sizes, and spherulites, small vesicles containing hererogeneous inaterial (Fig. 3 b). The course of the plasma membrane in the lateral cellular portions is very

sinuous. Electron-dense accumulations, similar 10 glycogen "rosettes", are visible near the membranous folds and arounds electron-clear areas (Fig. 4 e). Isolated bacteria (Gaill et al., 1984 a) are present on the epidermal surface, sorne of which are inserted at depth in the cuticle (Fig. 4 d). The cluster-like associations are very numerous in the intersegmentary zones. Their presence around the cuticular pinnules does not modify the general aspect of the underlying cells in the anterior region of the animal (Fig. 3 d). In contrast, in the caudal region where bacterial filaments are more numerous, the epidermal cells contain a large concentration of glycogen vesicles (Fig. 3 e). Mitochondria are also more numerous but Golgi apparatus, endoplasmic reticular networks and lysosomes, signifiers of exchange with the exterior, diminish in importance. Cellular activity seems rather directed 10wards storage.

The same type of cellular modifications was observed in the epidermis of the parapodial digitations of the caudal region. However, cytoplasmic glycogen is quantitatively less significant there, whereas "spherocrystals" abound, and the cytoplasm is dotted with electron-clear zones

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F. GAILL, D. DESBRUYÈRES, L. LAUBIER ~ ---------------------------------------------------------------1

Figure 3 k A) Subcuticular bacteria in Alvinella camlala. b, r bacteria; e, epicuticle; f, collagenous fibril; p, , epicuticular projections. , B) General aspect of the cell epùJermis of ~ Alvinella caudata. c, cuticle; p, epùJermis. t C) The cell epidermis of the posterior notopods f (Alvinella caudata). b, filamen/ous bacteria; m, le microvilli; s, spherocrystal. D) The base of a cluster-/ike assocation f

(Alvinella caudata). c, cuticle; f, collagenous J fibre; l, lysosome; q, glycogen particles; v, " vacuole. E) Aspect of the dorsal part of the Pompeii t worm epidermis. c, cuticle; g, glycogen f particles; m, microvilli; n, mitochondria; p, epi- t cuticular projections. , A) Bactéries sous-cuticulaires chez Alvinella cau- ~

dota. b, bactéries ; e, epicuticule ; f, fibres de ~ collagène; p, projections épicuticulaires. r B) Aspect général des cellules épidenniques d' AI- ~ vinella caudata. c; cuticule; p, épidenne. 1 C) Les cellules épidenniques des parapodes postérieurs (Alvinella caudata). b, bactéries ma- i menteuses ; m, microvillosités ; s, sphéro- i cristal. t D) Bases des associations en bouquet f (Alvinella caudata). c, cuticule ; f, fibres de colla- t gène; l, lysosome; q, glycogène; v, vacuole. r E) Aspect des cellules épidenniques de la panie t dorsale du ver de Pompéi. c, cuticule ; g, glyco-gène ; m, microvillosités ; n, mitochondries ; r. p, projections épicuticulaires. t

~ 1 1 1 f r t i f 1 t

(Fig. 3 c). The exchange surface area of the epidermal cells is thick and regularly attenuated posteriorly. The notopodia 1 and cytoplasm is diminished; microvilli are shorter and are all similar along the length of the body, with 1 non-branching (Fig. 3 c). exception of the two modified anterior segments. The Filamentous bacteria are inserted at depth in the cutic1e of segmentary glandular bourrelets are evenly subdivided 1 the parapodial digitations, when they are present (Fig. 2 along the length of the body, except for sorne post- 1 b). Their dilated discoid bases are covered by a relatively branchial segments. The size of the intersegmentary spaces 1 smOOth portion of cutic1e, as observed by SEM; c1usters increases only slightly from anterior to posterior; c1uster- ! of small-sized spherical elements occasionally surround the like associations occur there but in a lesser density than in f insertions (Fig. 2 b). When the load of filamentous Alvinella caudata. Epidermal formations (Fig. 4), unique bacteria becomes too great, the cuticle breaks away in in annelid polychaetes, occupy the intersegmentary spaces t small flakes until it disappears entirely from sorne areas; on the dorsal side, between the two rows of parapodia and ~ this is apparent in areas still containing impressions of positioned ventral or dorsal to the notopodia themselves. l, bacterial insertions on the microvilli. In areas devoid of Expansions, attaining or surpassing 10 mm in length, 1 cutic1e, the proximal extremity of the filamentous bacteria issue from epidermal "cupuliform" structures (Fig. 4 a) F is attached to the same microvilli upon which it rests, at arranged in a single or two altemating rows. ~ the same time, remaining separated by a network of very The epidermal "cupuliform" structures are composed of fine fibres (Fig. 3 c). 1EM studies show that shedding of glandular cells (Fig. 4 a). They are devoid of cutic1e, and r the cutic1e of the parapodial digitations is progressive; it c1ustered microvilli are present at their apex. At the distal t,',

gradually undergoes a reduction in thickness, the number extremity of these microvilli, spherical elements, with the 1

of collagen fibres decreases and the epicutic1e, whose appearance of mucous secretions (Fig. 4 b), are emitted ~ surface appears smooth under the SEM, separates at the and fuse together to form a reticulated mass of a fibrillar ~ level of the bacterial insertions. appearance at the periphery (Fig. 4 e). Cellular secretory ~ The general body form of the species Alvinella pompe jan a activity is revea1ed in numerous vesic1es, well-developed t

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150

--At the base of the expansion, the surface was observed by SEM to be smooth and consisting, at least exteriorly, of a "anhyste" substance. Further away from the insertion, most of this surface coating is broken up, revealing a shaggy "hirsute" structure composed of more or less entangled bacterial filaments. A transverse section of the expansion observed by SEM (Fig. 4 b) showed that the secretion was distributed in pseudo-concentric layers. Sheathed fIlamentous bacteria of a slender diameter (0.3 J.Illl) colonize the peripheral internaI portions of the expansion (Fig. 4 e), whereas larger-sized fIlamentous bacteria (cells 10 ~ x 1.0 ~) seem to develop around the distal regions. Bacterial colonization of the expansion is never central and the bacteria multiply in the polysaccharide matrix of the expansion (Fig. 4 e). These expansions are general in AlvineUa pompejana, the same in small individuals, but they may also be seen in lesser quantities in AlvineUa caudata within the zone of reduction of the body diameter (segments 50-65). They remain in this case limited to segmentary zones latero-dorsal to sorne segments and at the apex of the terminal digitation of the notopodia. Nevertheless, the frequency of these structures in Alvinella caudata is very reduced.

Figure 4 A) Dorsal expansion of Alvinella pompejana. c, cU/icle; g, glandular cells; n, matrix of the dorsal expansion; p, columnar cells; m, muscles. B) Cross·section of the dorsal expansion (SEM). White arrows, filamentous bacteria; C) Glandular cells which secrete the dorsal expansion (TEM). e, glandular cells; l, lyso-somes; s, secretions; v, microvilli; x, mitochon-dria. D) Insertion of bacteria insük the cU/icle (Alvinel-Ia caudata) (SEM). E) Aspect of the matrix of the dorsal expansion (TEM). n,fibrillar matrix; b, bacteria. A) Expansion dorsale d' Alvinella pompejana. c, cuticule; g, cellule glandulaire; n, matrice des ex­pansions dorsales ; p, cellule de soutien ; m, muscles. B) Coupe transversale d'une expansion dorsale (MEB). Flèche blanche, bactéries fIlamenteuses. C) Cellules sécrétrices de l'expansion dorsale (MET). e, cellule sécrétrice ; l, lysosome ; s, sécré-tions ; v, microvillosités ; x, mitochondries. D) Insertion bactérienne dans la cuticule (Alvinel-la caudata) (MEB). E) Aspect de la matrice de l'expansion dorsale (ME1). n, matrice fibrillaire ; b, bactéries.

"POMPEII WORMS" AND THEIR EPIBIOTIC BACTERIA

DISCUSSION AND CONCLUSIONS

With the exception of particular cases at deep-sea and coastal hydrothermal vents, bacterial epibioses in the marine environment have been scantily described. Cases of epibiosis are, evidently, more widespread in hydrothermal communities, where practically aIl available surfaces are heavily colonized by bacteria Jannasch and Wirsen (1979) described major populations of fIlamentous bacteria of the periostracum of Calyptogena magnifica and thick covering of fme filaments attributed to the genera Hyphomonas or Hyphomicrobium on the shell of the giant mussel of the Galàpagos vents. Associated with these fIlaments are curious trichomes, tentatively attributed to Calothrix, a chemosynthetic cyanobacteria (Jannasch, Wirsen, 1979). The black abalone, Haliotis cracherodii, of the White Point infra-littoral hydrothennal vents, carries on its shell a thick covering of fllamentous bacteria of the genus Thiothrix (Stein, 1984). A number of similar cases have also been observed in the patellifonn gastropods that colonize the tubes of Riftia pachyptila. . The unavoidable sUSpicion must be that these epibioses are functionally advantageous, to the point of an actual

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t=. GAILL, D. DESBRUYËRES, L. LAUBIER

symbiosis in the mutualistic sense of the term. Very different, without doubt, is the case of bacterial cpibiosis present on the branchial filaments of limpets from vents of the Juan de Fuca Ridge. Thcse bacterial filaments are frequenüy endocytozed by the epithelial cells and then lysed in cytoplasmic organelles similar to lysosomes (De Burgh, Singly, 1984). The importance of this process in the nutrition of the animal is again, however, difficult to estimate. In the polychaetes of hydrothermal vents of the East Pacific Rise, several cases of bacterial epibiosis were observed in our samplcs collectcd at l3°N. In the polynoids of the genus Lepidonotopodium, bacteria are frequenüy present on the brisües of the notopodia and on the elytra (pettibone, 1984). In llesiolyra bergi, which colonizes the tubes secreted by Alvinella, the notopodial brisües serve as a substrate for an abundant covering of filamentous bacteria. The digestive tract of the Serpulidae shows numerous epibioses whose functional significance is againenigmatic (Desbruyères et al., 1985). However, none of these epibioses possess the characteristics of those of Alvinella; the exceptional quantitative significance, diversity of bacterial epibiotic bases, and consistency of bacterial colonization in aIl individuals observed, irrespective of their size and segment numbers. Exarnination of another Alvinellid, Paralvinella grasslei (Desbruyères, Laubier, 1982), which lives in the same environment as Alvinella, reveals a very differcnt aspect of this epibiosis. Although its integument bas a very similar structure 10 that of Alvinella (Lepescheux, in press), intersegmentary epibiosis is extrcmely reduced and. bacterial epibiosis on the parapodia is non-existent.

Sorne epicuticular hacteria are observed in the two Alvinella species (Fig. 3 a; Gaill et al., 1987) but they are less numerous than epibiotic ones. This kind of association has been mentioned in sorne gutless annelids (Giere, 1981; Richards et al., 1982), and it scems that it is the second case where such an occurrence is mentioned in enteric polychaetes (Hausmann, 1982), the polynoids elytra being apart. SEM and MET observations show tbat the bacteria distribution is not a random one. It is more accurate in the caudata species where three main aspects were pointed out segmental division, and the density and size of filarnentous bacteria increase from the anterior to the posterior part in a gradient manner. Such a gradient does not occur in the pompe jan a species, which is characterizcd by the presence of dorsal expansions associated with filarnentous bacteria. The underlying cell epidermis aspects arc quite different with respect to the bacteria association types (which are always extracellular), each type having a single morpho­logical relationship with the cclI surface: • One such type is directly inserted on the cell epidermis close to the ceU microvilli. This is the case of the filarnentous bacteria associatcd with Alvinella caudata parapods. These epidermal parts lack cuticle, and the activity of the supportive cells as weIl as the exchange surfaces are reduccd, which suggests a stomge rather than an exchange activity, even if the bacteria arc direcüy in contact with the cell microvilli. It is not known whether the cuticle disappearance is a consequence of bacterial activity or of cclI activity relatcd to the worm physiology.

152

• Cluster-like assoclauons arc charactcrizcd by their morphological relationship with the cuticle fibre and the surrounding mineraI storage which does not exist in the other types of bacteria association (Gaill et al., 1984 b). The undcrlying cclI epidermis does not differ from the other supportive ceUs of the worm. A fcw of these bacteria can he in contact with the cclI epidermis by the way of the epicuticular projections. It seems tbat the collagenous fibres act as a physical substrate which permits the bacteria to exist in a pcculiar microenvironment where numerous worm secretions are available. • The third type of association occurs only in the pompejana species, and is in direct relationship with the cell epidermis. The underlying cells differ markedly from the supportive ceUs in their secretory activity. Bactcria are inserted in the secretions of the worm without contact with the cell epidermis. A single observation suggesLS that the attachment of bacteria on the cuticle of Alvinella is not a passive phenomenon comparable to the attachment of bacteria on an inert submerged surface. The preferential attachment of bacteria in the intcrsegmentary space on unusual epidermal secretions leads us to hypothesize an actual mUlualistic association between the worm and the bacterial epibionts. In our study, two different reactions of the epidermisof the Pompeii worms have been established. In the case of the dorsal intersegmentary spaces, particularly in the posterior region, the epidermal ceUs play a role in the storage of organic and mineral compounds. Experiments with in situ labelling have shown that absorption of low molecular weight organic compounds takes place in that zone (Alayse-Danet et al., 1985; 1986). In contrast, at the level of the parapodial digitations, the cellular exchange surfaces are rcduced. In that rcgion, Laubier et al. (1983) found evidence of significant concentrations of sulphur and Gaill et al. (1984 b) have locatcd, in the lysosomes and spherocrystals, high concentraûons of seveml mineml elements: S, As, Zn, Fe, Cu, P and Al. Results of in situ labelling experiments (Alayse-Danet et al., 1986) show a weak absorption of dissolved organic compounds by the epidermis of the parapodial digitations. Cellular characteristics also show that the uptake is rcduced where the epidermis stores waste products and mineraI clements. The nature of these stored materials indicates that they have bcen translocated from the internal environment of the animal (Gaill et al., 1984 b), and that the parapodial digitations and their epibioûc bacteria may be involved in detoxification processes of the animal (Cosson et al., 1986). Baross and Deming (1985) incubatcd a specimen of Alvinella caudata at in situ temperature and pressure conditions in hydrothermal fluid: microbial sulphate reduction was evident in an intense precipitation of sulphur, indicating that the filarnentous bacteria Lake part in the metabolism of sulphur compounds. The functional significance of the epibioûc bacteria of the dorsal expansions in Alvinella pompe jan a is even more enigmatic. The extent of this association is unique in marine invertebrates. These filamentous bacteria are extracellular but seem LO divide actively in a polysaccharide substance secreted by the animal. The incorporation of 3H-Thymidine by the bacteria at the base of the expansion in in situ experiments confirms this

. ····1

--hypothesis. (Alayse-Danet et al., 1986). Electron microscoPIC observations, as weIl as the absence of incorporation of 14C-bicarbonate lead us 10 surmise a heterotrophic or mixotrophic metaboIism coupled to the sulphur cycle, as in bacteria of the genera Thiothrix or Beggiatoa. The observations described here, 10gether with the experimental data (Alayse-Danet et al., 1985; 1986) do not adequately explain the functional relationship between the epibiotic bacteria and the Pompeü worm. Nevertheless, the work of Tuttle et al. (1983) as weIl as the results of cultures of epibiotic bacteria (Gaill et al., 1986) on various media (Baross media, Strohl media and the 2216E media of Oppenheimer and ZoBeIl) show that the metabolism of this apparent bacterial "community" is very diverse, ranging from strict chemoautotrophy to heterotrophy. Let us now propose an overview of the functioning of the biological "ensemble" constituted by the worm and its tube, as well as the tube internaI environment, and the associated bacteria. Through the tube, intemally lined by

REFERENCES

Alayse·Danet A.M., GaiU F., Desbruyères D., 1985. Pre1iminaIY studies on the relationship between the "Pompeii wonn", Alvinella pompajana (polychaeta: ampharetidae), and its epibiotic bacteria, in: Proceedings of XIXXth ElUopean Marine Biology Symposium, edited by P.E. Gibbs, Cambridge University Press, 167-172. Alayse·Danet A.M., GaiIJ F., Desbruyères D., 1986. In silu experiments on the Pompeii wonns, Mar. &01., 7, 3, 233-240. Baross A., Deming J., 1985. The role of bacteria in the ecology of black smoker environment, in: The hydrothermaI vents of the Eastern Pacifie: an overview, edited by M. Jones, Bull. Biol. Soc. Wash., 6, 355-372. Bosch C., Grassé P.P., 1984 a. Cycle partiel des bactéries cbimioautotrophes symbiotiques et leurs rapports avec les bactériocytes chez Riftia pachyptila Jones (pogonophore vestimentifère). 1: Le trophosome et les bactériocytes, CR. Acad. Sei. Paris, 299, 9, 371-376. Bosch C., Grassé P.P., 1984 b. Cycle partiel des bactéries cbimioautotrophes symbiotiques et leurs rapports avec les bactériocytes chez Riftia pachyptila Jones (pogonophore vestimentifères. Il: L'évolution des bactéries symbiotiques et des bactériocytes, CR. Acad. Sci. Paris, 299, 10,413-419. Cavanaugh C.M., 1983. Symbiotic chemoautotrophic bacteria in marine invertebrates from sulphide-rich habitats, NatlUe, 302, 58-61. Cavanaugh C.M., Gardiner S.L., Jones M.L., Jannasch H.W., Waterbury J.B., 1981. Prokaryotic ce1ls in the hydrothennal vent tube . wonn Riftia pachyptila Jones: possible chemoautotrophic symbionts, Science, 213, 340-341. Cosson-Mannevy M.A., Cosson R., Galll F., 1986. Mise en évidence de protéines de type métallothionéine chez deux invertébrés des sources hydrothennales, Riftia pachyptila et Alvinella pompejana, CR. Acad., Sci. Paris, 302, 9,347-351. De Burgh M.E., Singly C.L., 1984. Bacterial colonization and endocytosis on the gill of a new limpet species from a hydrothennal vent, Mar. Biol., 84, 1-6. Desbruyères D., Laubler L., 1980. Alvinella pompejana gen. sp. nov., Ampharetidae abberrant des sources hydrothennales de la ride Est-Pacifique, Oceanol. Acta, 3, 3, 267-274. Desbruyères D., Laubier L., 1982. Paralvinella grasslei new genus, new species of Alvinellidae (polychaeta: Ampharetidae) from the Ga1àpagos Rift geothennal vents, Proc. Biol. Soc. Wash., 95, 3,484-494. Desbruyères D., Laubier L., 1986. Les Alvinellidae, une

"POMPE Il WORMS" AND THEIR EPIBIOTIC BACTERIA

fIlamentous bacteria (Gaill, Hunt, 1986) passes a hot (20°-60°), acidic fluid with Iittle dissolved oxygen and a high concentration of reducing gases and metal species. This tube is an environment favourable to the multiplication of chemosynthetic bacteria which provide food for the worm; food in particulate form (Desbruyères et al., 1983; Baross, Deming, 1985) collected by the protractile buccal tentacles, in the form of dissolved organic material absorbed through the body wall in the dorsal intersegmentary zones and by the branchiae. However, this exceptional environment, rich in organic matter, is also 10xic because of its metal content, its low oxygen fugacity and acidic pH. The Pompeii worms have blood pigments well adapted to life in such an environment (Terwilliger, Terwilliger, 1984). The ensemble of fùamentous chemo­organotrophic genera Sphaerotilus, Herpetosiphon . or Streptothrix may actively participate in the fluid detoxification. Thus, this appears 10 be a complex symbiotic association in the mutualistic sense, although well opened 10 the exterior. The worm provides the carbon source for the bacteria in the form of respired CÛ2 and organic material, the bacteria provide particulate and dissolved organic matter for the worm and, in the case of

. the chemo-organotrophs, participate in detoxification.

fami1le nouvelle d'annélides polychètes inféodées aux sources hydrothennaIes sous-marines : systématique, biologie et écologie, Cano J. Zool., 64, 10,2227-2245. Desbruyères D., Crassous P., Grassle J.P., Khripounotf A., Reyss D., Rio M., Van Praët M., 1982. Données écologiques sur un nouveau site d'hydrothennaIisme actif de la ride du Pacifique oriental, CR. Acad. Sei. Paris, 295, 489-494. Desbruyères D., Gaill F., Prieur D., Rau G.H., 1983. Unusual nutrition of the "Pompeii wonn" Alvinella pompejana (polychaetous annelid) from a hydrothennal vent environment: SEM, TEM, 13C and ISC evidence, Mar. Biol., 75, 201-205. Desbruyères D., Galll F., Laubler L., Fouquet Y., 1985. Polychaetous anne1ids from hydrothermal vent ecosystems: an ecological overview, in: The hydrothennal vents of the Eastern Pacific: an overview, edited by M. Jones, Bull. Biol. Soc. Wash., 6, 103-116. Felbeck H., 1981. Chemoautotrophic potential of the hydrothennal vent mbe wonn Riftia pachyptila Jones (Vestimentifera), Science, 213, 336-338. Flala-Medionl A., 1984. Mise en évidence par microscopie électronique à transnusslOn de l'abondance de bactéries symbiotiques dans la branchie de mollusques bivalves des sources hydrothennales, CR. Acad. Sci. Paris, 298, 487-492. Galll F., Hunt S., 1986. Tubes of deep sea hydrothennal vent wonns Riftia pachyptila (Vestimentifera) and Alvinella pompejana (Anne1ida), Mar. &01. Progr. Ser., 34, 267-274. Galll F., Desbruyères D. Prieur D. Gourret J.P., 1984 a. Mise en évidence par la microscopie électronique des communautés bactériennes épibiontes des "vers de Pompéi" (Alvinella pompejana), CR. Acad. Sci. Paris, 298, 553-558. Gaill F., Halpern S., Quintana C., Desbruyères D., 1984 b. Présence intracellulaire d'arsenic et de zinc associés au soufre chez une Polychète des sources hydrothennales, CR. Acad. Sei. Paris, 298, 331-335. Gaill F. Desbruyères D., Prieur D.. Alayse-Danet A.M., 1986. Les relations d'une polychète avec ses bactéries épibiontes (Alvinella pompejana des sources hydrotherma1ès profondes), in: 2nd International Colloquiwn of Marine Bacteriology, GERBAM, IFREMER, CNRS, 401-406. Gaill F., Desbruyères D., Prieur D., 1987. Bacterial communities associated with "Pompeii wonns" from the East Pacific Rise hydrothennal vents: SEM, TEM observations, Microbiol. &01., 13, 129-134.

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Clerc O., 1981. The gutless marine oligochate Phallodri/us leukodermalus. Structural studies on an aberrant tubificid associated with bacteria, Mar. Ecol. Progr. Ser., S, 353-358. Hausmann K., 1982. Electronenmikroskopische untersuchungen an Anailides mucosa (Annelida Polychaeta). Cuticula and cilien, schleimzellen und schleimextrusion, Ile/go/. W~s. Meeresunlers., 3S,79-96. Jannasch H.W., Wirscn C,O., 1979. Chemosynthetic primary production at East-Pacifie sea floor spreading centers, Bioscience, 29, 592-598. Laubler L., Dcsbruyères D., Chassard-Bouchard C" 1983. Microanalytical evidence of sulfur accumulation in a polydtaete from deep-sea hydrothermal vents, Mar. Biol. Leu., 4, 113-116. Lepescheux L., in press. Biologica1 analogues of blue phases in marine worms, Tissue Ce//. Pettlbone M.H., 1984. A new-sca1e worm commensal with deep­sea mussels on the Galàpagos hydrolhcrmal vent (Polychaete: Polynoidae), Proc. Biol. Soc. Wash.,97, l, 226-239. Richards K.S., Fleming T,P., J;Imlcson B.C.M., 1982.

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An ultrastructural study of the distal epidermis and the occurrence of subcuticular bacteria in the gutJess tubificid Phallodrilus a/bidus (Oligochaeta, Annelida), Ausl. J. Zoo/., 30, 327-336. Stein J.L., 1984. Subtidal Gastropods consume sulfur-oxidizing bacteria: evidence from coastal hydrothermal vents, Science, 223, 696-698. Storch V., cam F., 1986. Ultrastrucwral observations on fccding appendages and gi11s of A/vinella pompejana (Anne1ida, Polychaeta), Ile/go/. W~s. Meeresunlers., 40, 309-319. Tcrwilliger A., Terwilliger M., 1984. Hemoglobin from the Pompeü worm, A/vine/la pompejana, an annelid from a deep-sea hot hydrothermal vent environment, Mar. Bio/. Leu., S, 191-201. Tuttle J.H., Wirscn C.O., Jannasch H.W., 1983. Microbial activities in the emiued hydrothermal waters of the Galapagos rift vents, Mar. Bio/., 73, 293-299. VoveUe J., cam F., .1986. Données morphologiques, histochimiques et nûcroanalytiques sur l'élaboration du tube organominéral d'A/vine//a pompejana, polychète des sources hydrothennales, et leurs implications phylogénétiques, Zool. Scr., IS,33-43.

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