Pompeii Worms" Polychètes and Their Epibiotic Bacteria Associations Bactériennes

Home , Alvinella pompejana, Alvinellidae

__------O-C-E-A-N-O-L-O-G-IC-A-A-C-T-A-1-98_8_-_N_O~S_P.~~~--

Hydrothennal vents Polychaeta Relationships between Bactcria association Sources hydrothennales the "Pompeii worms" Polychètes and their epibiotic bacteria Associations bactériennes

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.

ABSTRACf 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.

RÉSUMÉ 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.

INTRODUCTION diffusers (Dcsbruyères et al., 1985) in zones of activ.e mixing of hOl, reducing, acidic, mctal-rich fluid and cold, The so-called "Pompei worms" (polychaetous annelids) well-oxygenated scawater. This mixing results in a sharp live on active hydrothermal edifices on the East Pacific horizontal gradient, where the tcmperature changes within Rise at depths of about 2600 m. They secrete organic a few decimetres from 200° to 1.8°C. The Pompeii worms tubes (Vovelle, Gaill, 1986 ; Gaill, Hunt, 1986) on the are confincd to the cooler part of the gmdient, betwcen peripheral surface of certain chimneys and zinc-sulphide approximately 20° and 60°C (Dcsbruyères et al., 1982), ------147 F. GAILL, D. DESBRUYÈRES, L. LAUBIER

this being the range of their temperature tolerance defined bifurcatc digitations which are regular in form and as accurately as is possible. A minor portion of the worm, arrangement the branchiae, is positioned at the opening of the tube In the antcrior region, dorsal segmentary giandular bathing in ambient water, whereas the greater portion is bourrelets are undivided (Fig. 1), whereas posteriorly they enclosed within the tube these animaIs secrete. The divide into an increasing number of subsections (Fig. 1 Pompeü worms comprise two spccies: Alvinella and 2 c, d). Concurrently the non-glandular inter pompejan a and Alvinella caudata (Desbruyères, Laubier, segmentary spaces increasc to a maximum size in the 1986), primitively considered as two forms of the same median region where the glandular bourrelets are not species (Desbruyères, Laubier, 1980). found. In the caudal region, the intcrsegmentary spaces are Several polychaetous species inhabit the dcep sea less dilated than in the median region, but are nevertheless hydrothermal chimneys, but the Pompeii worms are the more important than in the anterior region. In these most spectacular with respect to the density of the bacteria spaces, bacilli, cocci and filamentous bacteria are covering their dorsal part (Gaill et al., 1984 a). These associated with cuticular secretions, forming "cluster-like" bacteria have bcen described (Gaill et al., 1987; structures (Fig. -l, 2 c, d). These associations are found 1 Desbruyères et al., 1985), but the biological significance along the cntire length of the animal (Fig. 1). However, t of the bacteria/worm association remains unclear. The the density and dimensions of filamentous bactcria increase 1 Pompeii worms are devoid of the endosymbiotic posteriorly (Fig. 1, 2 d). Large filamentous bacteria, , associations (Storch, Gaill, 1986) described and studied in preferentially inserted in the apical portion of posterior 1 vestimentiferans (Cavanaugh et al., 1981; Bosch, Grassé, notopods, form a veritable belt, easily visible to the naked 1 1984 b; Felbeck, 1981) and molluscs (Cavanaugh, eye (Fig. 2 a). a, 1 1983; Fiala-Medioni, 1984). They present only epibiotic The epidermis of the animal consists of an epithelium 1 bacteria. limited distally by a cuticle (Fig. 3 b) composed of a The purpose of this study was to determine more precisely network of collagen fibres and bordered by an electron 1 the morphological relationship betwecn the bacteria and dense epicuticle (Fig. 3 a). The epidermal cells issue ~ the worm epidermis and to inv~stigate whether the ! underlying cell epidermis is modified by the presence of Parapodia transformation 1 bacteria associations and, - if so - in what manner. 1 Answers to these questions would provide some co , indications conceming the local environ ment of the animal _\,?-.tf.Jlk\ , and its epidennal reactions, as well as information about ~~~" ! what may be called the process of exosymbyosis. 1 \ 1 MATERIAL AND METHODS 1 f Alvinella pompejan a spccimens were collected at 200 50'N and 109°W with the research submersible Alvin (April May 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 Increase of dorsal glandular torus parceiling 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 Increase of filamentous bacteria density morphological forros within the species Alvinella are ~ identical; the posterior sections of the body differ in their Figure 1 external anatomy, if not in thcir organization. Diagram comparing morph%gica/ modifications in Alvinella cau r In the caudata species, there is a gross dccrease in body data and the associated bacteria densily. ~ diameter from the 49th to the 54th segment. The Schéma comparanl les modificaùons morphologiques d'A/vine//a f notopodia are elongated in this rcgion and bear four or five caudata el la densité des bactéries associées. ~ ~ ------" - 148 ~ 1 "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 inter segmentary space (see 3 in fig. 2). a, inter segmentary 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, di gitation; 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 recou verte d'une portion de cuticule recouvrant les microvillosités ; e, base des filaments bacté riens ; f, bactéries filamenteuses ; m, microvil losités; C) Associations en bouquet de l'espace intersegmentaire (voir 3 sur la fig. 2). a, espace intersegmentaire ; s, partie de segment segmen tée. 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 sinuous. Electron-dense accumulations, similar 10 the cuticle perpendicularly (Fig. 3 e). These microvilli glycogen "rosettes", are visible near the membranous folds ramify 10wards the epicuticle and their more distal portions and arounds electron-clear areas (Fig. 4 e). Isolated bacteria form epicuticular projections or ellipsoidal bodies on the (Gaill et al., 1984 a) are present on the epidermal surface, exterior of the epicuticle (Fig. 3 a). Also, SEM sorne of which are inserted at depth in the cuticle observations of the surface integument reveal a granular (Fig. 4 d). appearance associated with the presence of epicuticular The cluster-like associations are very numerous in the projections (Fig. 4 d). intersegmentary zones. Their presence around the cuticular The epithelium in the dorsal intersegmentary zones pinnules does not modify the general aspect of the consists of a layer of cells overlying a basal lamina. This underlying cells in the anterior region of the animal (Fig. tissue is traversed by abundant blood vessels and is 3 d). In contrast, in the caudal region where bacterial proximal to numerous nerve terminations. The dorsal filaments are more numerous, the epidermal cells contain a epithelium comprises unspecialized cells and mucous cells large concentration of glycogen vesicles (Fig. 3 e). which are much less numerous in the dorsal glandular Mitochondria are also more numerous but Golgi flanges than in the ventral epithelium; they are absent in apparatus, endoplasmic reticular networks and lysosomes, the intersegmentary zone where unspecialized epidermal signifiers of exchange with the exterior, diminish in cells occur. These cells are characterized by the presence of importance. Cellular activity seems rather directed 10wards Golgi apparatus, an abundance of granular endoplasmic storage. reticulum and the presence of numerous mitochondria. Inclusions are present in large numbers in the cytoplasm: The same type of cellular modifications was observed in vesicles of all sizes near the cellular apex, lysosomes of the epidermis of the parapodial digitations of the caudal various sizes, and spherulites, small vesicles containing region. However, cytoplasmic glycogen is quantitatively hererogeneous inaterial (Fig. 3 b). The course of the less significant there, whereas "spherocrystals" abound, plasma membrane in the lateral cellular portions is very and the cytoplasm is dotted with electron-clear zones ------149 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 pompejan a activity is revea1ed in numerous vesic1es, well-developed t ____d_ifti_e_rs_pro __ fo_u_n_dl_y_f_ro_m_th_a_t _Of_A_l_v_ine_l_la_ca_uda__ ta_._Th_e_bod_y ___ G_o_lgt_·_a_pp_ara _t_u_s _an_d_th_e_p_r_e_se_n_ce_o_f_ly_s_os_o_m_e_s_(F_ig_. _4_c_)_. __ t 150 "POMPEII WORMS" AND THEIR EPIBIOTIC BACTERIA

At the base of the expansion, the surface was observed by DISCUSSION AND CONCLUSIONS --SEM to be smooth and consisting, at least exteriorly, of a "anhyste" substance. Further away from the insertion, With the exception of particular cases at deep-sea and most of this surface coating is broken up, revealing a coastal hydrothermal vents, bacterial epibioses in the shaggy "hirsute" structure composed of more or less marine environment have been scantily described. entangled bacterial filaments. A transverse section of the Cases of epibiosis are, evidently, more widespread in expansion observed by SEM (Fig. 4 b) showed that the hydrothermal communities, where practically aIl available secretion was distributed in pseudo-concentric layers. surfaces are heavily colonized by bacteria Jannasch and Sheathed fIlamentous bacteria of a slender diameter (0.3 Wirsen (1979) described major populations of fIlamentous J.Illl) colonize the peripheral internaI portions of the bacteria of the periostracum of Calyptogena magnifica and expansion (Fig. 4 e), whereas larger-sized fIlamentous thick covering of fme filaments attributed to the genera bacteria (cells 10 ~ x 1.0 ~) seem to develop around Hyphomonas or Hyphomicrobium on the shell of the the distal regions. Bacterial colonization of the expansion giant mussel of the Galàpagos vents. Associated with is never central and the bacteria multiply in the these fIlaments are curious trichomes, tentatively polysaccharide matrix of the expansion (Fig. 4 e). These attributed to Calothrix, a chemosynthetic cyanobacteria expansions are general in AlvineUa pompejana, the same (Jannasch, Wirsen, 1979). The black abalone, Haliotis in small individuals, but they may also be seen in lesser cracherodii, of the White Point infra-littoral hydrothennal quantities in AlvineUa caudata within the zone of reduction vents, carries on its shell a thick covering of fllamentous of the body diameter (segments 50-65). They remain in bacteria of the genus Thiothrix (Stein, 1984). A number this case limited to segmentary zones latero-dorsal to sorne of similar cases have also been observed in the patellifonn segments and at the apex of the terminal digitation of the gastropods that colonize the tubes of Riftia pachyptila. notopodia. Nevertheless, the frequency of these structures .The unavoidable sUSpicion must be that these epibioses in Alvinella caudata is very reduced. are functionally advantageous, to the point of an actual

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 ex pansions 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.

------151 . ····1

t=. GAILL, D. DESBRUYËRES, L. LAUBIER

symbiosis in the mutualistic sense of the term. Very • Cluster-like assoclauons arc charactcrizcd by their different, without doubt, is the case of bacterial cpibiosis morphological relationship with the cuticle fibre and the present on the branchial filaments of limpets from vents surrounding mineraI storage which does not exist in the of the Juan de Fuca Ridge. Thcse bacterial filaments are other types of bacteria association (Gaill et al., 1984 b). frequenüy endocytozed by the epithelial cells and then The undcrlying cclI epidermis does not differ from the lysed in cytoplasmic organelles similar to lysosomes (De other supportive ceUs of the worm. A fcw of these bacteria Burgh, Singly, 1984). The importance of this process in can he in contact with the cclI epidermis by the way of the the nutrition of the animal is again, however, difficult to epicuticular projections. It seems tbat the collagenous estimate. fibres act as a physical substrate which permits the In the polychaetes of hydrothermal vents of the East bacteria to exist in a pcculiar microenvironment where Pacific Rise, several cases of bacterial epibiosis were numerous worm secretions are available. observed in our samplcs collectcd at l3°N. In the • The third type of association occurs only in the polynoids of the genus Lepidonotopodium, bacteria are pompejana species, and is in direct relationship with the frequenüy present on the brisües of the notopodia and on cell epidermis. The underlying cells differ markedly from the elytra (pettibone, 1984). In llesiolyra bergi, which the supportive ceUs in their secretory activity. Bactcria are colonizes the tubes secreted by Alvinella, the notopodial inserted in the secretions of the worm without contact brisües serve as a substrate for an abundant covering of with the cell epidermis. filamentous bacteria. The digestive tract of the Serpulidae A single observation suggesLS that the attachment of shows numerous epibioses whose functional significance bacteria on the cuticle of Alvinella is not a passive is againenigmatic (Desbruyères et al., 1985). However, phenomenon comparable to the attachment of bacteria on none of these epibioses possess the characteristics of those an inert submerged surface. The preferential attachment of of Alvinella; the exceptional quantitative significance, bacteria in the intcrsegmentary space on unusual epidermal diversity of bacterial epibiotic bases, and consistency of secretions leads us to hypothesize an actual mUlualistic bacterial colonization in aIl individuals observed, association between the worm and the bacterial epibionts. irrespective of their size and segment numbers. In our study, two different reactions of the epidermisof the Exarnination of another Alvinellid, Paralvinella grasslei Pompeii worms have been established. In the case of the (Desbruyères, Laubier, 1982), which lives in the same dorsal intersegmentary spaces, particularly in the posterior environment as Alvinella, reveals a very differcnt aspect of region, the epidermal ceUs play a role in the storage of this epibiosis. Although its integument bas a very similar organic and mineral compounds. Experiments with in situ structure 10 that of Alvinella (Lepescheux, in press), labelling have shown that absorption of low molecular intersegmentary epibiosis is extrcmely reduced and. weight organic compounds takes place in that zone bacterial epibiosis on the parapodia is non-existent. (Alayse-Danet et al., 1985; 1986). In contrast, at the level of the parapodial digitations, the cellular exchange surfaces Sorne epicuticular hacteria are observed in the two are rcduced. In that rcgion, Laubier et al. (1983) found Alvinella species (Fig. 3 a; Gaill et al., 1987) but they are evidence of significant concentrations of sulphur and Gaill less numerous than epibiotic ones. This kind of et al. (1984 b) have locatcd, in the lysosomes and association has been mentioned in sorne gutless annelids spherocrystals, high concentraûons of seveml mineml (Giere, 1981; Richards et al., 1982), and it scems that it is elements: S, As, Zn, Fe, Cu, P and Al. the second case where such an occurrence is mentioned in Results of in situ labelling experiments (Alayse-Danet et enteric polychaetes (Hausmann, 1982), the polynoids al., 1986) show a weak absorption of dissolved organic elytra being apart. compounds by the epidermis of the parapodial digitations. SEM and MET observations show tbat the bacteria Cellular characteristics also show that the uptake is distribution is not a random one. It is more accurate in the rcduced where the epidermis stores waste products and caudata species where three main aspects were pointed out mineraI clements. The nature of these stored materials segmental division, and the density and size of filarnentous indicates that they have bcen translocated from the internal bacteria increase from the anterior to the posterior part in a environment of the animal (Gaill et al., 1984 b), and that gradient manner. Such a gradient does not occur in the the parapodial digitations and their epibioûc bacteria may pompejan a species, which is characterizcd by the presence be involved in detoxification processes of the animal of dorsal expansions associated with filarnentous bacteria. (Cosson et al., 1986). Baross and Deming (1985) The underlying cell epidermis aspects arc quite different incubatcd a specimen of Alvinella caudata at in situ with respect to the bacteria association types (which are temperature and pressure conditions in hydrothermal fluid: always extracellular), each type having a single morpho microbial sulphate reduction was evident in an intense logical relationship with the cclI surface: precipitation of sulphur, indicating that the filarnentous • One such type is directly inserted on the cell epidermis bacteria Lake part in the metabolism of sulphur close to the ceU microvilli. This is the case of the compounds. filarnentous bacteria associatcd with Alvinella caudata The functional significance of the epibioûc bacteria of the parapods. These epidermal parts lack cuticle, and the dorsal expansions in Alvinella pompejan a is even more activity of the supportive cells as weIl as the exchange enigmatic. The extent of this association is unique in surfaces are reduccd, which suggests a stomge rather than marine invertebrates. These filamentous bacteria are an exchange activity, even if the bacteria arc direcüy in extracellular but seem LO divide actively in a contact with the cell microvilli. It is not known whether polysaccharide substance secreted by the animal. The the cuticle disappearance is a consequence of bacterial incorporation of 3H-Thymidine by the bacteria at the base activity or of cclI activity relatcd to the worm physiology. of the expansion in in situ experiments confirms this

152 "POMPE Il WORMS" AND THEIR EPIBIOTIC BACTERIA --hypothesis. (Alayse-Danet et al., 1986). Electron fIlamentous bacteria (Gaill, Hunt, 1986) passes a hot (20°- microscoPIC observations, as weIl as the absence of 60°), acidic fluid with Iittle dissolved oxygen and a high incorporation of 14C-bicarbonate lead us 10 surmise a concentration of reducing gases and metal species. This heterotrophic or mixotrophic metaboIism coupled to the tube is an environment favourable to the multiplication of sulphur cycle, as in bacteria of the genera Thiothrix or chemosynthetic bacteria which provide food for the worm; food in particulate form (Desbruyères et al., 1983; Baross, Beggiatoa. The observations described here, 10gether with the Deming, 1985) collected by the protractile buccal tentacles, in the form of dissolved organic material experimental data (Alayse-Danet et al., 1985; 1986) do not absorbed through the body wall in the dorsal adequately explain the functional relationship between the intersegmentary zones and by the branchiae. However, this epibiotic bacteria and the Pompeü worm. Nevertheless, exceptional environment, rich in organic matter, is also the work of Tuttle et al. (1983) as weIl as the results of 10xic because of its metal content, its low oxygen fugacity cultures of epibiotic bacteria (Gaill et al., 1986) on and acidic pH. The Pompeii worms have blood pigments various media (Baross media, Strohl media and the 2216E well adapted to life in such an environment (Terwilliger, media of Oppenheimer and ZoBeIl) show that the Terwilliger, 1984). The ensemble of fùamentous chemo metabolism of this apparent bacterial "community" is very organotrophic genera Sphaerotilus, Herpetosiphon . or diverse, ranging from strict chemoautotrophy to Streptothrix may actively participate in the fluid heterotrophy. detoxification. Thus, this appears 10 be a complex Let us now propose an overview of the functioning of the symbiotic association in the mutualistic sense, although biological "ensemble" constituted by the worm and its well opened 10 the exterior. The worm provides the carbon tube, as well as the tube internaI environment, and the source for the bacteria in the form of respired CÛ2 and associated bacteria. Through the tube, intemally lined by organic material, the bacteria provide particulate and dissolved organic matter for the worm and, in the case of . the chemo-organotrophs, participate in detoxification.

REFERENCES

Alayse·Danet A.M., GaiU F., Desbruyères D., 1985. fami1le nouvelle d'annélides polychètes inféodées aux sources Pre1iminaIY studies on the relationship between the "Pompeii hydrothennaIes sous-marines : systématique, biologie et écologie, wonn", Alvinella pompajana (polychaeta: ampharetidae), and its Cano J. Zool., 64, 10,2227-2245. epibiotic bacteria, in: Proceedings of XIXXth ElUopean Marine Desbruyères D., Crassous P., Grassle J.P., Biology Symposium, edited by P.E. Gibbs, Cambridge University Khripounotf A., Reyss D., Rio M., Van Praët M., 1982. Press, 167-172. Données écologiques sur un nouveau site d'hydrothennaIisme actif Alayse·Danet A.M., GaiIJ F., Desbruyères D., 1986. In de la ride du Pacifique oriental, CR. Acad. Sei. Paris, 295, 489- silu experiments on the Pompeii wonns, Mar. &01., 7, 3, 233-240. 494. Baross A., Deming J., 1985. The role of bacteria in the Desbruyères D., Gaill F., Prieur D., Rau G.H., 1983. ecology of black smoker environment, in: The hydrothermaI vents Unusual nutrition of the "Pompeii wonn" Alvinella pompejana of the Eastern Pacifie: an overview, edited by M. Jones, Bull. Biol. (polychaetous annelid) from a hydrothennal vent environment: Soc. Wash., 6, 355-372. SEM, TEM, 13C and ISC evidence, Mar. Biol., 75, 201-205. Bosch C., Grassé P.P., 1984 a. Cycle partiel des bactéries Desbruyères D., Galll F., Laubler L., Fouquet Y., 1985. cbimioautotrophes symbiotiques et leurs rapports avec les Polychaetous anne1ids from hydrothermal vent ecosystems: an bactériocytes chez Riftia pachyptila Jones (pogonophore ecological overview, in: The hydrothennal vents of the Eastern vestimentifère). 1: Le trophosome et les bactériocytes, CR. Acad. Pacific: an overview, edited by M. Jones, Bull. Biol. Soc. Wash., Sei. Paris, 299, 9, 371-376. 6, 103-116. Bosch C., Grassé P.P., 1984 b. Cycle partiel des bactéries Felbeck H., 1981. Chemoautotrophic potential of the cbimioautotrophes symbiotiques et leurs rapports avec les hydrothennal vent mbe wonn Riftia pachyptila Jones bactériocytes chez Riftia pachyptila Jones (pogonophore (Vestimentifera), Science, 213, 336-338. vestimentifères. Il: L'évolution des bactéries symbiotiques et des Flala-Medionl A., 1984. Mise en évidence par microscopie bactériocytes, CR. Acad. Sci. Paris, 299, 10,413-419. électronique à transnusslOn de l'abondance de bactéries Cavanaugh C.M., 1983. Symbiotic chemoautotrophic bacteria symbiotiques dans la branchie de mollusques bivalves des sources in marine invertebrates from sulphide-rich habitats, NatlUe, 302, hydrothennales, CR. Acad. Sci. Paris, 298, 487-492. 58-61. Galll F., Hunt S., 1986. Tubes of deep sea hydrothennal vent Cavanaugh C.M., Gardiner S.L., Jones M.L., Jannasch wonns Riftia pachyptila (Vestimentifera) and Alvinella pompejana H.W., Waterbury J.B., 1981. Prokaryotic ce1ls in the (Anne1ida), Mar. &01. Progr. Ser., 34, 267-274. hydrothennal vent tube . wonn Riftia pachyptila Jones: possible Galll F., Desbruyères D. Prieur D. Gourret J.P., 1984 a. chemoautotrophic symbionts, Science, 213, 340-341. Mise en évidence par la microscopie électronique des communautés Cosson-Mannevy M.A., Cosson R., Galll F., 1986. Mise bactériennes épibiontes des "vers de Pompéi" (Alvinella en évidence de protéines de type métallothionéine chez deux pompejana), CR. Acad. Sci. Paris, 298, 553-558. invertébrés des sources hydrothennales, Riftia pachyptila et Gaill F., Halpern S., Quintana C., Desbruyères D., Alvinella pompejana, CR. Acad., Sci. Paris, 302, 9,347-351. 1984 b. Présence intracellulaire d'arsenic et de zinc associés au De Burgh M.E., Singly C.L., 1984. Bacterial colonization soufre chez une Polychète des sources hydrothennales, CR. Acad. and endocytosis on the gill of a new limpet species from a Sei. Paris, 298, 331-335. hydrothennal vent, Mar. Biol., 84, 1-6. Gaill F. Desbruyères D., Prieur D.. Alayse-Danet Desbruyères D., Laubler L., 1980. Alvinella pompejana gen. A.M., 1986. Les relations d'une polychète avec ses bactéries sp. nov., Ampharetidae abberrant des sources hydrothennales de la épibiontes (Alvinella pompejana des sources hydrotherma1ès ride Est-Pacifique, Oceanol. Acta, 3, 3, 267-274. profondes), in: 2nd International Colloquiwn of Marine Desbruyères D., Laubier L., 1982. Paralvinella grasslei new Bacteriology, GERBAM, IFREMER, CNRS, 401-406. genus, new species of Alvinellidae (polychaeta: Ampharetidae) from Gaill F., Desbruyères D., Prieur D., 1987. Bacterial the Ga1àpagos Rift geothennal vents, Proc. Biol. Soc. Wash., 95, communities associated with "Pompeii wonns" from the East Pacific 3,484-494. Rise hydrothennal vents: SEM, TEM observations, Microbiol. Desbruyères D., Laubier L., 1986. Les Alvinellidae, une &01., 13, 129-134. ------153 1 F. GAILL, D. DESBRUYÈRES, L. LAUBIER 1 1 - Clerc O., 1981. The gutless marine oligochate Phallodri/us An ultrastructural study of the distal epidermis and the occurrence 1 leukodermalus. Structural studies on an aberrant tubificid associated of subcuticular bacteria in the gutJess tubificid Phallodrilus a/bidus 1 with bacteria, Mar. Ecol. Progr. Ser., S, 353-358. (Oligochaeta, Annelida), Ausl. J. Zoo/., 30, 327-336. Hausmann K., 1982. Electronenmikroskopische untersuchungen Stein J.L., 1984. Subtidal Gastropods consume sulfur-oxidizing 1 an Anailides mucosa (Annelida Polychaeta). Cuticula and cilien, bacteria: evidence from coastal hydrothermal vents, Science, 223, 1 schleimzellen und schleimextrusion, Ile/go/. W~s. Meeresunlers., 696-698. 3S,79-96. Storch V., cam F., 1986. Ultrastrucwral observations on 1 Jannasch H.W., Wirscn C,O., 1979. Chemosynthetic primary fccding appendages and gi11s of A/vinella pompejana (Anne1ida, production at East-Pacifie sea floor spreading centers, Bioscience, Polychaeta), Ile/go/. W~s. Meeresunlers., 40, 309-319. 29, 592-598. Tcrwilliger A., Terwilliger M., 1984. Hemoglobin from the Laubler L., Dcsbruyères D., Chassard-Bouchard C" Pompeü worm, A/vine/la pompejana, an annelid from a deep-sea hot 1983. Microanalytical evidence of sulfur accumulation in a hydrothermal vent environment, Mar. Bio/. Leu., S, 191-201. polydtaete from deep-sea hydrothermal vents, Mar. Biol. Leu., 4, Tuttle J.H., Wirscn C.O., Jannasch H.W., 1983. 113-116. Microbial activities in the emiued hydrothermal waters of the Lepescheux L., in press. Biologica1 analogues of blue phases in Galapagos rift vents, Mar. Bio/., 73, 293-299. marine worms, Tissue Ce//. VoveUe J., cam F., .1986. Données morphologiques, Pettlbone M.H., 1984. A new-sca1e worm commensal with deep histochimiques et nûcroanalytiques sur l'élaboration du tube sea mussels on the Galàpagos hydrolhcrmal vent (Polychaete: organominéral d'A/vine//a pompejana, polychète des sources Polynoidae), Proc. Biol. Soc. Wash.,97, l, 226-239. hydrothennales, et leurs implications phylogénétiques, Zool. Scr., Richards K.S., Fleming T,P., J;Imlcson B.C.M., 1982. IS,33-43.

154