Microdiversity of phenol hydroxylase genes among phenol-degrading isolates of Alcaligenes sp from an activated sludge system

Enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting classified 97 phenol-degrading isolates with identical amplified ribosomal DNA restriction analysis (ARDRA) patterns into six genotypic groups. The 16S rRNA gene of the representative isolate of each group had higher than 99.47% common identity with each other and higher than 98% identity with the type strain of Alcaligenes faecalis. PCR-TGGE (temperature gradient gel electrophoresis) analysis of the genes of the largest subunit of the multi-component phenol hydroxylase (LmPH) in each isolate followed with sequencing showed that isolates within each ERIC-PCR group had identical LmPH gene sequences. Among the six different ERIC-PCR groups, two were found to harbor two different LmPH genes encoding low- and high-Ks (affinity constants) phenol hydroxylases, and the low-Ks type LmPH was identical in sequence with one predominant LmPH of the parental activated sludge. Three ERIC-PCR groups had only the high-Ks type and one had no sequence similar to the known LmPHs. Our work suggests that there is no correlation between the phylogenetic groupings of phenol-degrading bacteria and their LmPH genotypes possibly due to extensive horizontal gene transfer of this functional gene. (C) 2004 Published by Elsevier B.V. on behalf of the Federation of European Microbiological Societies.