Genetic diversity among 3-chloroaniline- and aniline-degrading strains of the Comamonadaceae

We examined the diversity of the plasmids and of the gene tdnQ, involved in the oxidative deamination of aniline, in five bacterial strains that are a ble to metabolize both aniline and 3-chloroaniline (3-CA). Three strains ha ve been described and identified previously, i.e., Comamonas testosteroni 1 2 and Delftia acidovorans CA28 and BN3.1. Strains LME1 and B8c were isolate d in this study from linuron-treated soil and from a wastewater treatment p lant, respectively, and were both identified as D. acidovorans. Both Delfti a and Comamonas belong to the family Comamonadaceae. All five strains posse ss a large plasmid of ca. 100 kb, but the plasmids from only four strains c ould be transferred to a recipient strain by selection on aniline or 3-CA a s a sole source of carbon and/or nitrogen. Plasmid transfer experiments and Southern hybridization revealed that the plasmid of strain 12 was responsi ble for total aniline but not 3-CA degradation, while the plasmids of strai ns LME1 and B8c were responsible only for the oxidative deamination of anil ine. Several transconjugant clones that had received the plasmid from strai n CA28 showed different degradative capacities: all transconjugants could u se aniline as a nitrogen source, while only some of the transconjugants cou ld deaminate 3-CA. For all four plasmids, the IS1071 insertion sequence of Tn5271 was found to be located on a 1.4-kb restriction fragment, which also hybridized with the tdnQ probe. This result suggests the involvement of th is insertion sequence element in the dissemination of aniline degradation g enes in the environment. By use of specific primers for the tdnQ gene from Pseudomonas putida UCC22, the diversity of the PCR-amplified fragments in t he five strains was examined by denaturing gradient gel electrophoresis (DG GE). With DGGE, three different clusters of the tdnQ fragment could be dist inguished. Sequencing data showed that the tdnQ sequences of 12, LME1, B8c, and CA28 were very closely related, while the tdnQ sequences of BN3.1 and P. putida UCC22 were only about 83% identical to the other sequences. North ern hybridization revealed that the tdnQ gene is transcribed only in the pr esence of aniline and not when only 3-CA is present.