Horst Felbeck

Microhabitat variation in the hydrothermal vent mussel, Bathymodiolus thermophilus, at the Rose Garden vent on the Galapagos Rift

Abstract Clumps of Bathymodiolus thermophilus were collected from three discrete areas at the ‘Rose Garden’ site on the Galapagos Rift using the deep submersible Alvin. Two mussel collections were made from the central Riftia mass, an area associated with very active venting, and three other collections were of two different peripheral mussel clumps. Before collection the clumps were extensively photographed and the water at two of the ‘microhabitats’ was analysed in situ for oxygen silica, sulfide and temperature. Sulfide levels of up to 300 μM were recorded at the central collection site, while the highest sulfide level recorded at the peripheral site assayed was 35 μM. Levels of RuBP carboxylase activity in the gills were significantly higher in mussels collected from the central ‘Riftia site’ than in either peripheral site. ATP sulfurylase was significantly higher in the gills of mussels from the central clump than in one of the peripheral clump collections. The chemical composition (% water, protein, carbohydrate, lipid and ash) and stable carbon isotope ratios (δ13C) of the mussels showed the same trends, with highest lipid and carbohydrate and the lowest water content and δ13C in the central site mussels. Similarly, the mussels from the central site were significantly depleted in stable nitrogen (δ15N) when compared with the peripheral site mussels. Variations between sites and tissues of the same animal may be indicative of differential utilization of inorganic or dissolved molecular nitrogen sources. The condition index (CI = soft tissue dry mass / internal shell volume) was similar for all animals collected at Rose Garden. The presence of a commensal polychaete, Branchipolynoe symmytilida, in the mantle cavity of the mussels was also correlated with the collection site, with the highest incidence of occurrence in the central clump. Levels of the enzyme RuBP carboxylase are quite variable in B. thermophilus and are on the average much lower (0.001 international units) than either Calyptogena magnifica (0.006 I.U.) or Riftia pachyptila (0.16 I.U.). We conclude that the mussels are able to thrive over a wider range of conditions than either C. magnifica or R. pachypila and that this is due to a lesser reliance on their symbiotic bacteria as a source of nutrition.

Sulfide oxidation and carbon fixation by the gutless clamSolemya reidi: an animal-bacteria symbiosis

SummarySolemya reidi, a protobranch clam lacking a digestive system and found exclusively in habitats rich in hydrogen sulfide (HS−), contains high densities of gram-negative bacteria within certain cells (“bacteriocytes”) of its large gills (Fig. 1). These bacteria are proposed to be responsible for the capacity ofS. reidi to oxidize HS− (Fig. 2) and produce sulfate. Some of the energy released during HS− oxidation could provide the necessary ATP and reducing power for the net fixation of CO2 via the reactions of the Calvin-Benson cycle, which have been found in the gills, but in no other tissues, of this clam.Solemya reidi exhibited a rapid fixation of H14CO3− compared to other clams and a fast transfer of label into a variety of metabolites, the labeling pattern varying with time of incubation (Tables 2–3). The CO2 fixation mechanism inS. reidi appears to involve an initial trapping of CO2 into a four-carbon compound (Table 2), which subsequently is decarboxylated to generate CO2 for the ribulose-1,5-bisphosphate carboxylase reaction of the Calvin-Benson cycle. Aspartate and malate were the major sites of14C during short-term incubations of intact clams or isolated gills; longer incubation periods led to appearance of radioactivity in a variety of amino acids and carboxylic acids. Of several carboxylating enzymes tested, only pyruvate carboxylase was found in gill tissue.Solemya reidi is capable of absorbing dissolved organic molecules from seawater (Tables 4 and 5). These findings indicate thatS. reidi may be nourished by reduced carbon and nitrogen compounds synthesized by symbiotic bacteria housed within its gills, and by dissolved organic material present in the muds in which the animal lives. The metabolic organization of this species is discussed in relation to the animal-bacteria symbioses recently discovered at the deep-sea hydrothermal vent sites where HS− may play an important role in driving primary productivity.

Metabolic versatility of the Riftia pachyptila endosymbiont revealed through metagenomics

The facultative symbiont of Riftia pachyptila, named here Candidatus Endoriftia persephone, has evaded culture to date, but much has been learned regarding this symbiosis over the past three decades since its discovery. The symbiont population metagenome was sequenced in order to gain insight into its physiology. The population genome indicates that the symbionts use a partial Calvin-Benson Cycle for carbon fixation and the reverse TCA cycle (an alternative pathway for carbon fixation) that contains an unusual ATP citrate lyase. The presence of all genes necessary for heterotrophic metabolism, a phosphotransferase system, and dicarboxylate and ABC transporters indicate that the symbiont can live mixotrophically. The metagenome has a large suite of signal transduction, defence (both biological and environmental) and chemotaxis mechanisms. The physiology of Candidatus Endoriftia persephone is explored with respect to functionality while associated with a eukaryotic host, versus free-living in the hydrothermal environment.

Chemoautotrophic Symbiosis in a Hydrothermal Vent Gastropod

An undescribed gastropod species collected from recently discovered deep-sea hydrothermal vents in the western Pacific contains endosymbiotic bacteria within specialized gill cells. The snails inhabit rocky vent openings where they are exposed directly to warm (2-25°C) sulfide-rich (750 µM) water emitted from the vents. The gills of this snail contain elemental sulfur and high activities of enzymes catalyzing sulfide metabolism (sulfide oxidase, ATP-sulfurylase, APS-reductase, rhodanese) and autotrophic CO2 fixation (ribulose bisphosphate carboxylase) indicating that the bacteria function as sulfur oxidizing Chemoautotrophic endosymbionts—a symbiosis described previously only in vestimentiferan and pogonophoran tubeworms, oligocheate worms, and bivalve molluscs. This represents the first documentation of Chemoautotrophic potential among the numerous gastropod species found inhabiting the interface of reducing and oxidizing environments.

Chemoautotrophic, Sulfur-Oxidizing Symbiotic Bacteria on Marine Nematodes: Morphological and Biochemical Characterization

The marine, free-living Stilbonematinae (Nematoda: Desmodorida) inhabit the oxygen sulfide chemocline in marine sands. They are characterized by an association with ectosymbiotic bacteria. According to their ultrastructure the bacteria are Gram-negative and form morphologically uniform coats that cover the entire body surface of the worms. They are arranged in host-genus or host-species specific patterns: cocci form multilayered sheaths, rods, and crescent- or filament-shaped bacteria form monolayers. The detection of enzymes associated with sulfur metabolism and of ribulose-1,5 bisphosphate carboxylase oxygenase, as well as elemental sulfur in the bacteria indicate a chemolithoautotrophic nature of the symbionts. Their reproductive patterns appear to optimize space utilization on the host surface: vertically standing rods divide by longitudinal fission, whereas other bacteria form non-septate filaments of up to 100 μm length.

Variation in the hydrothermal vent clam, Calyptogen magnifica, at the Rose Garden vent on the Galapagos spreading center

Abstract Calyptogena magnifica occupy a relatively restricted habitat at the Rose Garden hydrothermal vent site on the Galapagos Rift. These clams are found in areas with very low flow of vent water and gain exposure to hydrogen sulfide by inserting their well-vascularized foot into cracks that contain this flow. Vent water is undetectable around the siphons of many of the individuals, and they therefore probably take up sulfide through their foot, and oxygen and inorganic carbon through their gills. Age estimates indicate that the bulk of the recruitment of C. magnifica occured between 1971 and 1976. Isotopic evidence indicates that symbionts are the main source of both nutritional carbon and nitrogen for the clams, and that the symbionts assimilate both of these substrates from inorganic sources. Carbohydrate and protein in the clam soft tissues, as well as the elemental sulfur content of their gills, decrease with increasing clam size. There is only slight variation in most of the parameters measured, and none of the parameters show nearly the variation seen in the other hydrothermal vent bivalve, Bathymodiolus thermophilus . However, several parameters, such as δ 13 C, condition index, and some bacterial enzyme activities, vary significantly with habitat.

Multiple trophic resources for a chemoautotrophic community at a cold water brine seep at the base of the Florida Escarpment

The biological community that surrounds the hypersaline cold water brine seeps at the base of the Florida Escarpment is dominated by two macrofaunal species: an undescribed bivalve of the family Mytilidac and a vestimentiferan worm, Escarpia laminata. These animals are apparently supported by the chemoautotrophic fixation of carbon via bacterial endosymbionts. Water column and sediment data indicate that high levels of both sulfide and methane are present in surface sediments around the animals but absent from overlying waters. Stable isotopic analyses of pore water indicate that there are two sources of sulfide: the first is geothermal sulfide carried in groundwater leaching from the base of the escarpment, and the second is microbial sulfide produced in situ. The vestimentiferan E. laminata, and the mytilid bivalve (seep mussel) live contiguously but rely on different substrates for chemoautotrophy. Enzyme assays, patterns of elemental sulfur storage and stable isotopic analyses indicate that E. laminata relies on sulfide oxidation and the seep mussel on methane oxidation for growth.

Primary production in deep-sea hydrothermal vent organisms: roles of sulfide-oxidizing bacteria

Abstract The dense aggregations of animal and bacterial life at the deep-sea hydrothermal vents are supported, at least in part, by sulfide-energy-based primary productivity. Sulfide-oxidizing bacteria are found free-living in the vent waters, on the surface of basaltic rocks, and as symbionts within certain tissues of the large vent tube worms and bivalve molluscs.

Sulphide mining by the superextensile foot of symbiotic thyasirid bivalves

In a symbiotic association between an invertebrate host and chemoautotrophic bacteria, each partner has different metabolic requirements, and the host typically supplies the bacteria with necessary reduced chemicals (sulphide or methane). Some combination of anatomical, physiological and behavioural adaptations in the host often facilitates uptake and transport of reduced chemicals to the symbionts. We have studied five species of bivalve molluscs of the family Thyasiridae (that is, thyasirids) three of which harbour chemoautotrophic bacteria. Here we show that the symbiotic bivalves extend their feet to form elongated and ramifying burrows in the sediment, most probably to gain access to reduced sulphur. Closely related bivalves (including some thyasirid species) without bacterial symbionts show no comparable foot extension behaviour. The length and number of burrows formed by chemosymbiotic thyasirids are related to the concentration of hydrogen sulphide in the sediment. The burrows are formed by the foot of each bivalve, which can extend up to 30 times the length of the shell, and may be the most extreme case of animal structure elongation documented to date.

Fate of nitrate acquired by the tubeworm Riftia pachyptila.

ABSTRACT The hydrothermal vent tubeworm Riftia pachyptila lacks a mouth and gut and lives in association with intracellular, sulfide-oxidizing chemoautotrophic bacteria. Growth of this tubeworm requires an exogenous source of nitrogen for biosynthesis, and, as determined in previous studies, environmental ammonia and free amino acids appear to be unlikely sources of nitrogen. Nitrate, however, is present in situ (K. Johnson, J. Childress, R. Hessler, C. Sakamoto-Arnold, and C. Beehler, Deep-Sea Res. 35:1723–1744, 1988), is taken up by the host, and can be chemically reduced by the symbionts (U. Hentschel and H. Felbeck, Nature 366:338–340, 1993). Here we report that at an in situ concentration of 40 μM, nitrate is acquired by R. pachyptila at a rate of 3.54 μmol g−1h−1, while elimination of nitrite and elimination of ammonia occur at much lower rates (0.017 and 0.21 μmol g−1 h−1, respectively). We also observed reduction of nitrite (and accordingly nitrate) to ammonia in the trophosome tissue. When R. pachyptila tubeworms are exposed to constant in situ conditions for 60 h, there is a difference between the amount of nitrogen acquired via nitrate uptake and the amount of nitrogen lost via nitrite and ammonia elimination, which indicates that there is a nitrogen “sink.” Our results demonstrate that storage of nitrate does not account for the observed stoichiometric differences in the amounts of nitrogen. Nitrate uptake was not correlated with sulfide or inorganic carbon flux, suggesting that nitrate is probably not an important oxidant in metabolism of the symbionts. Accordingly, we describe a nitrogen flux model for this association, in which the product of symbiont nitrate reduction, ammonia, is the primary source of nitrogen for the host and the symbionts and fulfills the associations nitrogen needs via incorporation of ammonia into amino acids.