Physiological and immunological evidence for two distinct C-1-utilizing pathways in Bathymodiolus puteoserpentis (Bivalvia : Mytilidae), a dual endosymbiotic mussel from the Mid-Atlantic Ridge

The existence of endosymbiotic sulfur-oxidizing chemoautotrophic and methan otrophic bacteria associating with marine mytilid mussels has previously be en inferred by 16S rDNA analysis in Bathymodiolus puteoserpentis Von Cosel et al. 1994, a hydrothermal vent mussel from a site on the Mid-Atlantic Rid ge. In mussels collected in June 1993, we found evidence of enzymes diagnos tic of two distinct C-1 assimilation pathways in this symbiosis. Assays for the utilization of radiolabelled methane and for immunodetection of methan ol dehydrogenase were positive, indicating that oxidation and incorporation of this substrate are occurring in this symbiosis. Sulfide or thiosulfate had no detectable stimulatory effect on CO2 incorporation, and assays for t he enzyme ribulose-l,5-bisphosphate carboxylase/oxygenase (RubisCO), an enz yme diagnostic for the Calvin-Benson cycle, were negative. RubisCO was dete cted in all samples examined by immunoblot analysis, indicating this enzyme is expressed in the B. puteoserpentis symbiosis. Stable isotope data showe d that carbon isotope Values were in agreement with previously reported val ues, and nitrogen isotope values were among the most depleted ever reported for bivalve symbioses. The carbon isotope values do not preclude the utili zation of vent-derived methane. These data could be explained by the presen ce of two metabolically distinct bacterial symbionts or a Type X methanotro phic symbiont.