Diversity of carbofuran-degrading soil bacteria and detection of plasmid-encoded sequences homologous to the mcd gene

Abstract

Fifty-five bacterial isolates, from English and French soils with different histories of carbofuran field treatment, which hydrolysed the N-methylcarbamate insecticide carbofuran to carbofuran 7-phenol were characterised phenotypically and genetically. The isolates were compared by using 125 physiological tests and morphological features, plasmid profiles and restriction fragment length polymorphism (RFLP) patterns of total DNA using the rRNA operon of Escherichia coli as a DNA probe. Cluster analysis of both phenotypic characters and RFLP patterns showed a high degree of diversity amongst the bacteria. Ten distinct plasmid profiles with 2–4 plasmids ranging in size from 84 to about 438 kb were visualised in 50 isolates. The majority of isolates had one of two types of plasmid profiles. Plasmid profiles and EcoRI restricted total DNA patterns were hybridised with an internal fragment of the carbofuran hydrolase ( mcd) gene and 22 diverse soil isolates exhibited sequence homology with this gene probe. Our results indicate that sequences homologous to the mcd gene are located on a conserved EcoRI fragment (12 or 14 kb) of a plasmid (100, 105, 115 or 124 kb) found in diverse soil isolates from geographically distant areas. Thirty-three isolates did not exhibit detectable homology to the mcd gene probe and the hydrolase enzymes and genes in these isolates need further investigation.