Marine symbioses are predominantly established through horizontal acquisition of microbial symbionts from the environment. However, genetic and functional comparisons of free-living populations of symbionts to their host-associated counterparts are sparse. Here, we assembled the first genomes of the chemoautotrophic gammaproteobacterial symbionts affiliated with the deep-sea snail Alviniconcha hessleri from two separate hydrothermal vent fields of the Mariana Back-Arc Basin. We used phylogenomic and population genomic methods to assess sequence and gene content variation between free-living and host-associated symbionts. Our phylogenomic analyses show that the free-living and host-associated symbionts of A. hessleri from both vent fields are populations of monophyletic strains from a single species. Furthermore, genetic structure and gene content analyses indicate that these symbiont populations are differentiated by vent field rather than by lifestyle. Together, this work suggests that, despite the potential influence of host-mediated acquisition and release processes on horizontally transmitted symbionts, geographic isolation and/or adaptation to local habitat conditions are important determinants of symbiont population structure and intra-host composition.