Phylogenetic and Transcriptional Analyses of a Tetrachloroethene-Dechlorinating "Dehalococcoides" Enrichment Culture TUT2264 and Its Reductive-Dehalogenase Genes

A dechlorinating microbial enrichment culture designated TUT2264 was cultured with tetrachloroethene and then characterized for tetrachloroethene-dechlorination by culture-dependent and -independent methods. The fourth-transferred TUT2264 culture completely dechlorinated tetrachloroethene and trichloroethene, and accumulated more trans-1,2-dichloroethene than cis-1,2-dichloroethene. A real-time PCR analysis revealed that "Dehalococcoides" cells made up only 0.3% of the total. Eight distinct reductive-dehalogenase-homologous genes (rdh) were detected with degenerate primers. Phylogenetic analyses revealed 5 of the 8 RdhAs to be very similar to RdhAs reported previously but not to share 100% identity. Transcriptional levels were quantified as the number of transcripts per rdhA by combining the reverse transcription real-time PCR and exogenous internal reference mRNA methods. TUT2264 responded to all the chloroethenes tested. rdhA4 was transcribed with all chloroethenes except vinyl chloride, whereas rdhA8 was only transcribed on tetrachloroethene. Furthermore, multiple rdhAs were induced to express by a single chloroethene as a growth-supporting or non-supporting substrate. These results suggested that Rdhs are multi-functional and rdhAs are a powerful tool to evaluate the potential of contaminated sites and isolates to dechlorinate chloroethenes.