Profiling of biodegradation and bacterial 16S rRNA genes in diverse contaminated ecosystems using 60-mer oligonucleotide microarray

Abstract

We have developed an oligonucleotide microarray for the detection of biodegradative genes and bacterial diversity and tested it in five contaminated ecosystems. The array has 60-mer oligonucleotide probes comprising 14,327 unique probes derived from 1,057 biodegradative genes and 880 probes representing 110 phylogenetic genes from diverse bacterial communities, and we named it as BiodegPhyloChip. The biodegradative genes are involved in the transformation of 133 chemical pollutants. Validation of the microarray for its sensitivity specificity and quantitation were performed using DNA isolated from well-characterized mixed bacterial cultures also having non-target strains, pure degrader strains, and environmental DNA. Application of the developed array using DNA extracted from five different contaminated sites led to the detection of 186 genes, including 26 genes unique to the individual sites. Hybridization of 16S rRNA probes revealed the presence of bacteria similar to well-characterized genera involved in biodegradation of various pollutants. Genes involved in complete degradation pathways for hexachlorocyclohexane (lin), 1,2,4-trichlorobenzene (tcb), naphthalene (nah), phenol (mph), biphenyl (bph), benzene (ben), toluene (tbm), xylene (xyl), phthalate (pht), Salicylate (sal), and resistance to mercury (mer) were detected with highest intensity. The most abundant genes belonged to the enzyme hydroxylases, monooxygenases, and dehydrogenases which were present in all the five samples. Thus, the array developed and validated here shall be useful in assessing not only the biodegradative potential but also the composition of environmentally useful bacteria, simultaneously, from hazardous ecosystems.