Carbon fixation by sulfur-oxidizing chemosynthetic bacteria forms the base of the food chain in deep-sea (diffuse-flow) hydrothermal vent ecosystems. Temperature and the availability of oxygen and reduced sulfur are believed to be factors that contribute to the structure of hydrothermal vent communities. Sulfur concentration and speciation can change rapidly as highly reducing vent fluids mix with cold oxygenated seawater, thus the path followed by source water before passing over organisms hosting sulfur-oxidizing endosymbionts may have important implications for these animals. Here we show an apparent correlation between the zonation of symbiont-containing species and the sulfur chemical speciation (sulfide, polysulfides, thiosulfate) in the water bathing them. We also report in-situ measurements of thiosulfate (S₂O₃ ²⁻) in hydrothermal fluids, predominantly in the area inhabited by the mussel, Bathymodiolus brevior. These results provide field evidence of environmental levels of thiosulfate that may be capable of supporting thiosulfate-utilizing symbionts at hydrothermal vents. The three dominant shellfish species were arranged concentrically in a distinct bull's eye pattern at our study site. Alviniconcha sp. 1 snails inhabited the central, most reducing sulfide-rich zone, Ifremeria nautilei snails formed a thin band surrounding the Alviniconcha, and B. brevior mussels were farthest away from the source fluid in the most oxidized region. After removal of many of the Alviniconcha, some I. nautilei moved into the vacated space, whereas the B. brevior remained around the extreme periphery of the area impacted by the diffuse flow. These results suggest that diffuse flow chemistry is one of the key parameters affecting organism distribution.