The glycolate-oxidizing, sulfate-reducing bacterium strain PerGlyS and the syntrophically glycolate-oxidizing bacterium strain FlGlyR were studied with respect to their phylogenetic relationships on the basis of in vitro amplification and direct sequencing of 16S rRNA-encoding DNA. Strain PerGlyS clustered with representatives of the delta subclass of the class Proteobacteria, close to "Desulforhopalus vacuolatus" but sufficiently distinct to preclude its assignment to this genus. These organisms, together with Desulfobulbus propionicus, represent a phylogenetic subgroup among members of the delta subclass of Proteobacteria. Strain FlGlyR was found to cluster with the gram-positive bacteria with low-G + C DNA, and Desulfitobacterium dehalogenans and Desulfotomaculum orientis are its closest relatives. Other species of the genus Desulfotomaculum are phylogenetically only moderately closely related to these organisms. These results necessitate the establishment of new genera and species for these two strains. Strain PerGlyS was designated the type strain of Desulfofustis glycolicus gen. nov., sp. nov., and strain FlGlyR was designated the type strain of Syntrophobotulus glycolicus gen. nov., sp. nov.