Identification and characterization of integron mediated antibiotic resistance in pentachlorophenol degrading bacterium isolated from the chemostat

A bacterial consortium was developed by continuous enrichment of microbial population isolated from sediment core of pulp and paper mill effluent in mineral salts medium (MSM) supplemented with pentachlorophenol (PCP) as sole source of carbon and energy in the chemostat. The consortia contained three bacterial strains. They were identified as Escherichia coli, Pseudomonas aeruginosa and Acinetobacter sp. by 16S rRNA gene sequence analysis. Acinetobacter sp. readily degraded PCP through the formation of tetrachloro-p-hydroquinone (TecH), 2-chloro-1,4-benzenediol and products of ortho ring cleavage detected by gas chromatograph/mass spectrometer (GC-MS). Out of the three acclimated PCP degrading bacterial strains only one strain, Acinetobacter sp. showed the presence of integron gene cassette as a marker of its stability and antibiotic resistance. The strain possessed a 4.17 kb amplicon with 22 ORF's. The plasmid isolated from the Acinetobacter sp. was subjected to shotgun cloning through restriction digestion by BamHI, HindIII and SalI, ligated to pUC19 vector and transformed into E. coli XLBlue1α, and finally selected on MSM containing PCP as sole source of carbon and energy with ampicillin as antibiotic marker. DNA sequence analysis of recombinant clones indicated homology with integron gene cassette and multiple antibiotic resistance genes.