CHARACTERIZATION OF A DEFINED 2,3,5,6-TETRACHLOROBIPHENYL-ORTHO-DECHLORINATING MICROBIAL COMMUNITY BY COMPARATIVE SEQUENCE-ANALYSIS OF GENES-CODING FOR 16S RIBOSOMAL-RNA

Defined microbial communities were developed by combining selective en richment with molecular monitoring of total community genes coding for 16S rRNAs (16S rDNAs) to identify potential polychlorinated biphenyl (PCB) dechlorinating anaerobes that ortho dechlorinate 2,3,5,6-tetrach lorobiphenyl. In enrichment cultures that contained a defined estuarin e medium, three fatty acids, and sterile sediment, a Clostridium sp, w as predominant in the absence of added PCB, but undescribed species in the delta subgroup of the class Proteobacteria, the low-G+C gram-posi tive subgroup, the Thermotogales subgroup, and a single species with s equence similarity to the deeply branching species Dehalococcoides eth enogenes were more predominant during active dechlorination of the PCB , Species with high sequence similarities to Methanomicrobiales and Me thanosarcinales archaeal subgroups were predominant in both dechlorina ting and nondechlorinating enrichment cultures. Deletion of sediment f rom PCB-dechlorinating enrichment cultures reduced the rate of dechlor ination and the diversity of the community. Substitution of sodium ace tate for the mixture of three fatty acids increased the rate of dechlo rination, further reduced the community diversity, and caused a shift in the predominant species that included restriction fragment length p olymorphism patterns not previously detected. Although PCB-dechlorinat ing cultures were methanogenic, inhibition of methanogenesis and elimi nation of the archaeal community by addition of bromoethanesulfonic ac id only slightly inhibited dechlorination, indicating that the archaea were not required for ortho dechlorination of the congener, Deletion of Clostridium spp, from the community profile by addition of vancomyc in only slightly reduced dechlorination, However, addition of sodium m olybdate, an inhibitor of sulfate reduction, inhibited dechlorination and deleted selected species from the community profiles of the class Bacteria. With the exception of one 16S rDNA sequence that had the hig hest sequence similarity to the obligate perchloroethylene-dechlorinat ing Dehalococcoides, the 16S rDNA sequences associated with PCB ortho dechlorination had high sequence similarities to the delta, low-G+C gr am-positive, and Thermotogales subgroups, which all include sulfur-, s ulfate-, and/or iron(III)-respiring bacterial species.