The European flat oyster Ostrea edulis, the European native species presents a major ecological, economic, and cultural importance. O. edulis populations have experienced a severe decline due to overfishing, habitat destruction, parasitic diseases, and invasive species. This decline led to its classification as a threatened species under the OSPAR Convention. Despite this status, O. edulis plays a crucial role in coastal ecosystems by forming oyster reefs that enhance biodiversity, improve water filtration, and contribute to carbon storage. O. edulis is a protandrous sequential hermaphrodite, alternating between male and female phases in response to environmental conditions, particularly temperature and food availability. Populations are generally characterized by a male-biased sex ratio, which reduces reproductive success. Sex identification remains challenging due to the lack of reliable non-destructive methods. To improve sustainable seed production and support ecological restoration, a deeper understanding of the molecular mechanisms governing gametogenesis is required. This study combines histological and transcriptomic analyses of simultaneously hermaphroditic gonadal tissues of O. edulis to characterize gamete maturation stages and identify key genes and biological processes involved in the reproductive cycle. The identification of molecular markers aims to enhance reproductive management for aquaculture and population restoration.

For the first time, two Dynamic Energy Budget (DEB) models were developed for a chemosymbiotic deep-sea vesicomyid clam. A classical DEB model was applied and then an innovative DEB model was developed (named “farming”). The models were parameterized using data on host and symbionts, including original unpublished data. In the farming model the digestion of the sulfur-oxidizing bacterial symbionts for host nutrition was explicitly modeled. Unexpected results were obtained regarding the dynamics of host and symbionts with this model: the host appears to forgo a maximal ingestion for a lower and stable ingestion, revealing a new kind of homeostasis. Moreover, when the clam is adult, most of the oxygen consumed by the chemosynthetic symbiosis was predicted to be by the symbionts. A high host energy maintenance flux was predicted and consistent with the likely high energy demand of host ion homeostasis mechanisms to cope with symbiont protons and sulfates release.

Bivalves of the family Lucinidae thrive in sulfidic sediments thanks to their chemoautotrophic bacterial symbionts. However, how different Lucinidae species respond to sulfide deprivation and associated symbiont loss remains poorly understood. Here, we investigated the responses of Lucinoma borealis and Loripes orbiculatus, two species that co-occur in temperate seagrass beds, exposed to prolonged sulfide starvation. Using metabolomics, ultrastructural TEM analyses and 16S rRNA-based metabarcoding, we monitored and compared responses in gills and visceral mass over a 4-month period. Both host species as well as their symbionts survived sulfide-free conditions. Hosts tissues displayed limited impact on ultrastructure and metabolites. Despite decrease in numbers and activity level, symbionts remained present throughout the experiment and no evidence for bacteremia or infection was detected. Our results also revealed differences, in particular in host apoptosis response, suggesting species-specific stress strategies. Altogether, both holobionts can survive extended low-sulfide periods without critical damage and without completely losing their symbionts. These could be adaptations to the extended low-sulfide periods that are associated with low primary production and the cold season in seagrass beds. Adaptations could involve a switch in the symbionts' physiological state to preserve a dormant symbiotic population. These findings highlight the importance of stress tolerance mechanisms in coastal Lucinidae, and raise questions about the nature of host–symbiont dependency in these periods.

The polychaete Capitella is a typical member of the ‘thiobiome’, and is commonly used as an eutrophication indicator species in environmental assessment studies. To deal with a sulfide-rich and poisonous surrounding, cells in close contact with the environment, and thus able to play a major role in detoxication and survival, are circulating cells. This work aimed to morpho-functionally describe the circulating coelomic cells of Capitella from the English Channel inhabiting the sulfide-rich mud in Roscoff Harbor. In general, worms have three types of circulating cells, granulocytes involved in bacterial clearance and defense against microorganisms, eleocytes with an essentially trophic role and elimination of cellular waste, and erythrocytes which play a role in detoxification and respiration via their intracellular hemoglobin. By combining diverse microscopic and cellular approaches, we provide evidence that Capitella does not possess granulocytes and eleocytes, but rather a single abundant rounded cell type with the morphological characteristics of erythrocytes i.e. small size and production of intracellular hemoglobin. Surprisingly, our data show that in addition to their respiratory function, these red cells could exert phagocytic activities, and produce an antimicrobial peptide. This latter immune role is usually supported by granulocytes. Our data highlight that the erythrocytes of Capitella from the English Channel differ in morphology and bear more functions than the erythrocytes of other annelids. The simplicity of this multi-task (or polyvalent) single-cell type makes Capitella an interesting model for studies of the impact of the environment on the immunity of this bioindicator species.

High latitude seeps are dominated by Oligobrachia siboglinid worms. Since these worms are often the sole chemosymbiotrophic taxon present (they host chemosynthetic bacteria within the trophosome organ in their trunk region), a key question in the study of high latitude seep ecology has been whether they harbor methanotrophic symbionts. This debate has manifested due to the mismatch between stable carbon isotope signatures of the worms (lower than -50‰ and usually indicative of methanotrophic symbioses) and the lack of molecular or microscopic evidence for methanotrophic symbionts. Two hypotheses have circulated to explain this paradox: (1) the uptake of sediment carbon compounds with depleted δC13 values from the seep environment, and (2) a small, but significant and difficult to detect population of methanotrophic symbionts. We conducted 16S rRNA amplicon sequencing of the V3-V4 regions on two species of northern seep Oligobrachia (Oligobrachia webbi and Oligobrachia sp. CPL-clade), from four different high latitude sites, to investigate the latter hypothesis. We also visually checked the worms’ symbiotic bacteria within the symbiont-hosting organ, the trophosome, through transmission electron microscopy. The vast majority of the obtained reads corresponded to sulfide-oxidizers and only a very small proportion of the reads pertained to methane-oxidizers, which suggests a lack of methanotrophic symbionts. A number of sulfur oxidizing bacterial strains were recovered from the different worms, however, host individuals tended to possess a single strain, or sometimes two closely-related strains. However, strains did not correspond specifically with either of the two Oligobrachia species we investigated. Water depth could play a role in determining local sediment bacterial communities that were opportunistically taken up by the worms. Bacteria were abundant in non-trophosome (and thereby symbiont-free) tissue and are likely epibiotic or tube bacterial communities. The absence of methanotrophic bacterial sequences in the trophosome of Arctic and north Atlantic seep Oligobrachia likely indicates a lack of methanotrophic symbionts in these worms, which suggests that nutrition is sulfur-based. This is turn implies that sediment carbon uptake is responsible for the low δ13C values of these animals. Furthermore, endosymbiotic partners could be locally determined, and possibly only represent a fraction of all bacterial sequences obtained from tissues of these (and other) species of frenulates.

The European flat oyster (Ostrea edulis L.) is a native bivalve of the European coasts. Harvest of this species has declined during the last decades because of the appearance of two parasites that have led to the collapse of the stocks and the loss of the natural oyster beds. O. edulis has been the subject of numerous studies in population genetics and on the detection of the parasites Bonamia ostreae and Marteilia refringens. These studies investigated immune responses to these parasites at the molecular and cellular levels. Several genetic improvement programs have been initiated especially for parasite resistance. Within the framework of a European project (PERLE 2) that aims to produce genetic lines of O. edulis with hardiness traits (growth, survival, resistance) for the purpose of repopulating natural oyster beds in Brittany and reviving the culture of this species in the foreshore, obtaining a reference genome becomes essential as done recently in many bivalve species of aquaculture interest. Here, we present a chromosome-level genome assembly and annotation for the European flat oyster, generated by combining PacBio, Illumina, 10X linked, and Hi-C sequencing. The finished assembly is 887.2 Mb with a scaffold-N50 of 97.1 Mb scaffolded on the expected 10 pseudochromosomes. Annotation of the genome revealed the presence of 35,962 protein-coding genes. We analyzed in detail the transposable element (TE) diversity in the flat oyster genome, highlighted some specificities in tRNA and miRNA composition, and provided the first insight into the molecular response of O. edulis to M. refringens. This genome provides a reference for genomic studies on O. edulis to better understand its basic physiology and as a useful resource for genetic breeding in support of aquaculture and natural reef restoration.

Deep-sea mussels Bathymodiolus azoricus, from Azorean hydrothermal vents, house two types of symbionts in their fleshy gills: methane-oxidizing (MOX) and sulfide-oxidizing (SOX) Gamma-proteobacteria. As soon as the mussels are collected, their symbionts are deprived from their environmental nutrient flux, and cannot rely on their usual metabolism. Recent studies have shown that the gill cells undergo high rates of apoptosis, as well as regionalized cell proliferation. This study follows the fate of the symbionts and of the hosting bacteriocytes at the ultrastructural level, during an extended starvation period. Just upon collection, we evidenced an apico-basal journey of the symbionts in the bacteriocytes, starting with (1) apical single symbiont endocytosis, (2) symbiont division, (3) symbiont storage, (4) and symbiont digestion within lysosomes, above the basal lamina. After 4-9 days starvation, endocytosis occurred with (5) empty blebbing, (6) the lysosomes increased in size, and the bacteriocytes lost their apical membrane, resulting in (7) a baso-apical return of the symbiont-containing lysosomes outside the gills, while the nucleus showed condensed chromatin, characteristic of apoptosis/necroptosis (8). Between the bacteriocytes, narrow intercalary cells appear to divide (9). Our hypothesis is that intercalary cells are stem cells that replace lost bacteriocytes. After 61 days there was no symbiont left, and the epidermis resembled those of the non-symbiotic filter-feeding mussel Mytilus edulis.

Deep-sea mussels Bathymodiolus spp. harbor high densities of chemosynthetic bacterial symbionts located within their gill epithelial cells. Compared to non-symbiotic coastal mussel relatives of similar size, Bathymodiolus gills are considerably larger, a feature often considered an adaptation to symbiosis because it is related to the presence of intracellular bacteria in epithelial cells located in the lateral zone. In order to document the mechanisms underlying these sizes differences, this study compares gill cell proliferation patterns in Bathymodiolus azoricus and Mytilus edulis using microscopy-based approaches. We used incubation experiments with a synthetic nucleotide (5-ethynyl 2′-deoxyuridine, EdU), detectable throughout novel cell divisions, and phosphohistone H3 immunolabeling, a marker of mitosis. The results revealed proliferation areas in the ciliated zone and in the bacteria-loaded bacteriocytes located close to the frontal zone of gill filaments, swept by the incurrent sea-waterflow, and also in the dorsal region of gills in B. azoricus. Cell proliferation seems far less intensive in M. edulis. This study overall suggests high cell turnover and fast tissue dynamics in symbiont-bearing mussels.

Symbiosis between Bathymodiolus and Gammaproteobacteria allows these deep-sea mussels to live in toxic environments such as hydrothermal vents and cold seeps. The quantity of endosymbionts within the gill-bacteriocytes appears to vary according to the hosts environment; however, the mechanisms of endosymbiont population size regulation remain obscure. We investigated the possibility of a control of endosymbiont density by apoptosis, a programmed cell death, in three mussel species. Fluorometric TUNEL and active Caspase-3-targeting antibodies were used to visualize and quantify apoptotic cells in mussel gills. To control for potential artefacts due to depressurization upon specimen recovery from the deep-sea, the apoptotic rates between mussels recovered unpressurised, versus mussels recovered in a pressure-maintaining device, were compared in two species from hydro thermal vents on the Mid-Atlantic Ridge: Bathymodiolus azoricus and B. puteoserpentis. Results show that pressurized recovery had no significant effect on the apoptotic rate in the gill filaments. Apoptotic levels were highest in the ciliated zone and in the circulating hemocytes, compared to the bacteriocyte zone. Apoptotic gill-cells in B. aff. boomerang from cold seeps off the Gulf of Guinea show similar distribution patterns. Deep-sea symbiotic mussels have much higher rates of apoptosis in their gills than the coastal mussel Mytilus edulis, which lacks chemolithoautotrophic symbionts. We discuss how apoptosis might be one of the mechanisms that contribute to the adaptation of deep-sea mussels to toxic environments and/or to symbiosis.

We provide the first detailed identification of Barents Sea cold seep frenulate hosts and their symbionts. Mitochondrial COI sequence analysis, in combination with detailed morphological investigations through both light and electron microscopy was used for identifying frenulate hosts, and comparing them to Oligobrachia haakonmosbiensis and Oligobrachia webbi, two morphologically similar species known from the Norwegian Sea. Specimens from sites previously assumed to host O. haakonmosbiensis were included in our molecular analysis, which allowed us to provide new insight on the debate regarding species identity of these Oligobrachia worms. Our results indicate that high Arctic seeps are inhabited by a species that though closely related to Oligobrachia haakonmosbiensis, is nonetheless distinct. We refer to this group as the Oligobrachia sp. CPL-clade, based on the colloquial names of the sites they are currently known to inhabit. Since members of the Oligobrachia sp. CPL-clade cannot be distinguished from O. haakonmosbiensis or O. webbi based on morphology, we suggest that a complex of cryptic Oligobrachia species inhabit seeps in the Norwegian Sea and the Arctic. The symbionts of the Oligobrachia sp. CPL-clade were also found to be closely related to O. haakonmosbiensis symbionts, but genetically distinct. Fluorescent in situ hybridization and transmission electron micrographs revealed extremely dense populations of bacteria within the trophosome of members of the Oligobrachia sp. CPL-clade, which is unusual for frenulates. Bacterial genes for sulfur oxidation were detected and small rod shaped bacteria (round in cross section), typical of siboglinid-associated sulfur-oxidizing bacteria, were seen on electron micrographs of trophosome bacteriocytes, suggesting that sulfide constitutes the main energy source. We hypothesize that specific, local geochemical conditions, in particular, high sulfide fluxes and concentrations could account for the unusually high symbiont densities in members of the Oligrobrachia sp. CPL-clade.

Vesicomyids live in endosymbiosis with sulfur-oxidizing bacteria and therefore need hydrogen sulfide to survive. They can nevertheless live in a wide range of sulfide and oxygen levels and depths, which may explain the exceptional diversity of this clam family in deep-sea habitats. In the Gulf of Guinea, nine species of vesicomyid clams are known to live in cold-seep areas with pockmarks from 600 to 3200m deep, as well as in the organic-rich sediments of the Congo deep-sea fan at 5000m deep. Our previous study showed that two species living in a giant pockmark have different oxygen carriers, suggesting different adaptations to hypoxia. Here, we studied the hemoglobin structure and oxygen affinity in three other species, Calyptogena valdiviae, Elenaconcha guiness and Abyssogena southwardae to determine whether the characteristics of their oxygen carriers contribute to their distribution in sulfide-rich sediments at a regional scale. Documenting pairwise species associations in various proportions, we give a semi-quantitative account of their local distribution and oxygen and sulfide measurements at seven sites. Mass spectrometry showed that each vesicomyid species has four intracellular monomeric hemoglobin molecules of 15–16kDa, all differing in their molecular mass. As expected, the monomers showed no cooperativity in oxygen binding. Their oxygen affinities were very high (below 1Torr), but differed significantly. C. valdiviae had the highest affinity and was dominant in the Harp pockmark, the site with the lowest oxygen content (half the value of fully oxygenated water). A. southwardae dominated in the Congo Lobe area, the site with the deepest sulfides. We discuss how hemoglobin may favor an active, vertical distribution of vesicomyids in sulfide-rich sediments.

The diet of Pomatoschistus microps has been studied using both gut content and stable isotope analyses. In the Roscoff Aber Bay (Brittany, France), this fish is commonly found on sandy muddy intertidal flats. Gut content analyses were also interpreted using trophic indices. Owing to the large diversity of prey consumed, these indices emphasised the opportunistic feeding behaviour of P. microps. Here, this species fed mainly on endofauna with meiofauna being of high relative importance. The main biotic components of its trophic habitat, characterized by δ13C and δ15N, provided evidence of a major trophic pathway based on drift Enteromorpha sp. Trophic positions estimated by both diet analyses and isotopic analyses led to similar results. In this bay, P. microps is a first-order predator with a low degree of omnivory. Despite a preferential consumption of the amphipod Corophium arenarium, we assumed that this goby behaves as a generalist feeding on a uniform variety of endofauna taxa.

Bathymodiolus mussels associated with deep-sea hydrothermal vents and cold seeps harbor chemosynthetic endosymbiotic bacteria in bacteriocytes located in the gill epithelium. Two distinct morphotypes of γ-proteobacteria, sulfur- and methane-oxidizing, have been identified and form a dual symbiosis in B. azoricus mussels from the Mid-Atlantic Ridge and in B. aff. boomerang from cold seeps in the Gulf of Guinea. Thiotrophic bacteria (SOX) are capable of fixing CO2 in the presence of sulfide or thiosulfate and methanotrophic bacteria (MOX) use methane both as a carbon and an energy source. In this study we used quantitative real-time PCR to test whether symbiont abundance and gene expression varied between the two mussel species. Results showed that B. azoricus from two hydrothermal sites had similar ratios and gene expression pattern for both symbiont types. In B. aff. boomerang , SOX ratio and ATP sulfurylase gene expression show differences between specimens collected on the different sites. Analysis of symbiont ratios in both species indicated a clear dominance of MOX symbionts in B. aff. boomerang and SOX symbionts in B. azoricus . We also evidenced that the species from the deeper sites ( B. aff. boomerang ) and mussels collected from sulfur and methane rich habitats showed higher symbiont ratio suggesting that environmental parameters may have significant impacts on the symbiont ratios in Bathymodiolus mussels.

Vesicomyid bivalves form dense clam beds in both deep-sea cold seeps and hydrothermal vents. The species diversity within this family raises questions about niche separation and specific adaptations. To compare their abilities to withstand hypoxia, we have studied the structure and function of erythrocyte hemoglobin (Hb) and foot myoglobin (Mb) from two vesicomyid species, Christineconcha regab and Laubiericoncha chuni, collected from the Regab pockmark in the Gulf of Guinea at a depth of 3,000 m. Laubiericoncha chuni possesses three monomeric globins, G1 (15,361 Da), G2 (15,668 Da), and G3 (15,682 Da) in circulating erythrocytes (Hb), and also three globins, G1, G3, and G4 (14,786 Da) in foot muscle (Mb). Therefore, globins G2 and G4 appear to be specific for erythrocytes and muscle, respectively, but globins G1 and G3 are common. In contrast, C. regab lacks erythrocyte Hb completely and possesses only globin monomers G1′ (14,941 Da), G2′ (15,169 Da), and G3′ (15,683 Da) in foot muscle. Thus, these two vesicomyid species, C. regab and L. chuni, show a remarkable diversity in globin expression when examined by electrospray ionization mass spectrometry. Oxygen-binding affinities reveal extremely high oxygen affinities (P50 < 1 Torr, from 5° to 15°C at pH 7.5), in particular L. chuni globins, which might be an advantage allowing L. chuni to dig deeply for sulfides and remain buried for long periods in reduced sediments.

Hakon Mosby mud volcano (HMMV) is one of the most active and most studied seep sites in European waters. Many authors have described its thermal activity, dynamic of mud flows, and geochemical and microbial processes. It is characterised by a concentric zonation of successive biogenic habitats related to an activity and geochemical gradient from its centre to its periphery. Around the central area covered by mud flows, white and grey microbial mats occur among areas of bare sediment, whereas siboglinid tubeworm fields of Sclerolinum contortum and/or Oligobrachia haakonmosbiensis colonise the peripheral areas. The meiofaunal community is known to be structured among habitats, but the macrofauna has rarely been investigated and has never been sampled in situ. As part of the European project HERMES, using the ROVs Victor 6000 and Quest 4000, we sampled quantitatively the different habitats of the volcano for macrofauna sensus lato, retained on a 250- or 500-mu m sieve. We also sampled a newly discovered pockmark on Storegga slide (cne 5.6) and two pockmarks (G11, G12) in the Nyegga area. Macrofauna was identified and counted from phylum to family level. Our results on HMMV showed a gradient of increasing density and diversity from the volcano centre (13 taxa; 260 ind.m-2) to the peripheral siboglinid fields (814 taxa, 93,000 ind.m-2), with an intermediate situation for microbial mats. For macrofauna =500 mu m, non-siboglinid polychaetes dominated the communities of the central mud volcano area, white mats and S. contortum fields (83, 89 and 37% of the total, respectively), whereas gastropods dominated grey mats and O. haakonmosbiensis fields (89 and 44% of the total, respectively). Polychaete families followed the same pattern of diversity according to habitats within HMMV. Of 23 polychaete families identified, only one occurred in the centre, and three in the microbial mats. Capitellidae and Dorvilleidae (typical of organically and sulphide-enriched areas) occurred at remarkably high densities in white microbial mats and in O. haakonmosbiensis fields. The S. contortum fields were the most diverse habitat with 12 polychaete families. The 250-mu m fraction showed similar taxa dominating the habitats, but taking meiofauna into account, nematodes became the major taxon in white mats and in S. contortum fields, where they were particularly large in size, whereas copepods dominated in other habitats. Meiofauna and macrofauna did not show the same patterns of density according to habitats. Using principal components analysis the habitats at HMMV were clearly distinct, and clustered according to dominant species of siboglinids and type of microbial mats. Pockmarks at Nyegga showed a similar concentric pattern of habitats around fluid sources as on the volcano, which seemed similarly to influence macrofauna composition, but at a much smaller scale. Total taxa and polychaete diversity are high in the S. contortum fields in these pockmarks as well. Regional-scale comparisons including HMMV and Storegga suggested a higher influence of habitat-type than seep-site on the community structure.

Southward E. C., Andersen A. C. & Hourdez S. 2011. — Lamellibrachia anaximandri n. sp., a new vestimentiferan tubeworm (Annelida) from the Mediterranean, with notes on frenulate tubeworms from the same habitat. Zoosystema 33 (3): 245–279. ABSTRACT A new species of lamellibrachiid vestimentiferan, Lamellibrachia anaximandri n. sp., has been found in the Eastern Mediterranean, close to cold seeps of fluid carrying dissolved methane and sources of sulfide in superficial sediments. It occurs at about 1100 to 2100 m depth, on some of the mud volcanoes on the Anaximander Mountains, south of Turkey, on the Mediterranean Ridge, south of Crete, and on the Nile deep-sea fan. In addition, it has been obtained from rotting paper inside a sunken ship, torpedoed in 1915 and lying at 2800 m depth, southeast of Crete. Some frenulate pogonophores also occur on the mud volcanoes (including a species of Siboglinum resembling S. carpinei and tubes of other unidentified genera). The new Lamellibrachia is the first vestimentiferan species to be described from the Mediterranean. It differs from L. luymesi taken from the Gulf of Mexico population in the very weak development of collars on its tube and in having a smaller number of pairs of branchial lamellae in the branchial plume. Sequencing of the COI and the mt16S genes confirms a difference at the species level between the new species and L. luymesi, and a difference between these two species and four described species of Lamellibrachia from the Pacific Ocean. The largest individuals of L. anaximandri n. sp. may be many years old, but there are numerous young individuals at some sites, showing that favourable conditions are available for settlement and early growth. The development of the branchial plume in a series of young stages reveals that the sheath lamellae, which are characteristic of the genus Lamellibrachia, begin to form only after the establishment of several pairs of branchial lamellae. Examination of the adult trophosome by transmission electron microscopy shows Gram-negative bacteria without internal stacked membranes, indicating that the symbionts are most probably sulfide oxidizing.

Animals inhabiting hydrothermal vents and cold seeps face conditions that are challenging for survival. In particular, these two habitats are characterized by chronic hypoxia, sometimes reaching complete anoxia. The characteristics of the scaphognathite and gills were studied in four species of shrimp and three species of crabs from hydrothermal vents and cold seeps, in order to highlight potential adaptations that could enhance oxygen acquisition in comparison with shallow-water relatives. All the vent and seep species studied here exhibit significantly larger scaphognathites, likely allowing more water to flow over their gills per stroke of this appendage. This is probably more energetically efficient that prolonged hyperventilation. In contrast to annelids, vent and seep decapods usually do not possess enlarged gills, a phenomenon likely due to the physical limitations imposed by the size of the gill chamber. In the vent shrimp Rimicaris exoculata and the vent crab Bythograea thermydron, however, there is a significantly higher specific gill surface area linked to a higher number of lamellae per gram of gill. Again in contrast to annelids, the diffusion distance through the gills is not strikingly different between the vent shrimp Alvinocaris komaii and the shallow-water species Palaemon spp. This may indicate that the epithelium and cuticle of the decapod gills are already optimized for oxygen uptake and that reducing the thickness of these compartments is not physically possible without affecting the physical integrity of the gills.

Siboglinids are symbiotic polychete annelids having hemoglobins as essential oxygen- and sulfide-carriers for their endosymbiotic bacteria. We analyzed the structure of the hemoglobins from two species of siboglinids: the monilifera Sclerolinum contortum and the frenulata Oligobrachia webbi (i.e. haakonmosbiensis) from Norwegian cold seeps. Measured by Multi-Angle Laser Light Scattering (MALLS), Sclerolinum shows a 3190+/-50 kDa hexagonal bilayer hemoglobin (HBL-Hb) and a 461+/-46 kDa ring-Hb, just as vestimentifera, whereas Oligobrachia has a 409+/-3.7 kDa ring-Hb only. Electrospray Ionization-Mass Spectrometry (ESI-MS) showed Sclerolinum HBL-Hb composed of seven monomeric globins (15-16 kDa), three disulfide-bonded globin heterodimers and three linkers. The heterodimers always contain globin-b (15814.4+/-1.5 Da). Sclerolinum ring-Hb is composed of globins and dimers with identical masses as its HBL-Hb, but lacks linkers. Oligobrachia ring-Hb has three globin monomers (14-15 kDa) only, with no disulfide-bonded dimers. Comparison of Sclerolinum hemoglobins between Storegga and Haakon Mosby Mud Volcano, using the normalized height of deconvoluted ESI-MS peaks, shows differences in globin monomers abundances that could reflect genetic differences or differential gene expression between distinct seep populations. The discovery of HBL-Hb in Sclerolinum is a new element supporting the hypothesis of monilifera being phylogenetically more closely related to vestimentifera, than to frenulata.

During the European Commission's Framework Six Programme, HERMES, we investigated three main areas along the European margin, each characterized by the presence of seep-related structures exhibiting different intensity of activity and biological diversity. These areas are: (1) the Nordic margin with the Hakon Mosby mud volcano and many pockmarks, (2) the Gulf of Cadiz, and (3) the eastern Mediterranean with its hundreds of mud volcanoes and brine pool structures. One of the main goals of the HERMES project was to unravel the biodiversity associated with these seep-associated ecosystems, and to understand their driving forces and functions, using an integrated approach. Several multidisciplinary research cruises to these three areas provided evidence of high variability in ecosystem processes and associated biodiversity at different spatial scales, illustrating the "hotspot" nature of these deep water systems.