Development of primers for amplifying genes encoding CprA- and PceA-like reductive dehalogenases in anaerobic microbial consortia, dechlorinating trichlorobenzene and 1,2-dichloropropane

Gene sequence alignments of the reductive dehalogenases PceA (Dehalospirilh an multivorans) and CprA (Desulfitobacterium dehalogenans) were used to dev elop specific PCR primers binding to conserved regions of these sequences. These primers enabled us to amplify and subsequently sequence cprA-like gen e fragments from the chlororespiring species Dehalobacter restrictus, Dehal obacter restrictus, Desulfitobacterium sp. strain PCE1, and D. hafniense. N o specific amplicons were obtained from the chlororespiring species D, frap pieri, D. chlororespirans. and Desulfoloniletiedjei. Furthermore, we were a ble to amplify and sequence cprAlpceA-like gene fragments from both trichlo robenzene (TCB)- and I,2-dichloropropane (DCP)-dechlorinating microbial con sortia using the novel primers. Subsequent sequence analysis of the fragmen ts obtained fi-om the microbial consortia revealed a group of four clusters (I-IV). Of these, clusters I and II showed the highest similarities to the cprA-like gene of Dchalobacter restrietus (79.0 and 96.20% respectively). Cluster III comprised cprA-like sequences found in both the TCB- and the DC P-dechlorinating consortia, whereas sequences of cluster IV were most simil ar to the pceA gene of Dehalospirillium multivorans (97.8%). Our detection of genes encoding reductive dehalogenases, the key enzymes of chlororespira tion, supports the hypothesis that reductive dechlorination of TCB and DCP occurs via a respiratory pathway. (C) 2001 Federation of European Microbiol ogical Societies. Published by Elsevier Science B.V. All rights reserved.