Insights into molecular mechanism of plasticizer biodegradation in Dietzia kunjamensis IITR165 and Brucella intermedia IITR166 isolated from a solid waste dumpsite.

Abstract

Isolation of phthalate esters (PAEs) degrading bacteria from a solid waste dumpsite could degrade many plasticizers efficiently and to investigate their degrading kinetics, pathways, and genes. Based on their 16S rRNA gene sequence the strains were identified as Dietzia kunjamensis IITR165 and Brucella intermedia IITR166, which showed a first-order degradation kinetic model under lab conditions. The quantification of phthalates and their intermediate metabolites identification were done by using UHPLC and GC-MS/MS respectively. Both the bacteria utilized>99% dibutyl phthalate at a high concentration of 100-400mg L-1 within 192hrs as monitored by UHPLC. GC-MS/MS revealed the presence of metabolites dimethyl phthalate (DMP), phthalic acid (PA), and benzoic acid (BA) during DBP degradation by IITR165 while monobutyl phthalate (MBP) and PA were identified in IITR166. Phthalate esters degrading gene cluster in IITR165 comprised two novel genes coding for carboxylesterase (dkca1) and mono-alkyl phthalate hydrolase (maph), having only 37.47% and 47.74% homology respectively with reported phthalate degradation genes, along with terephthalate dioxygenase system (tphA1, A2, A3, and B). However, IITR166 harboured different gene clusters comprising di-alkyl phthalate hydrolase (dph_bi), and phthalate dioxygenase (ophA, B, and C) genes. Two novel bacterial strains, Dietzia kunjamensis IITR165 and Brucella intermedia IITR166, were isolated and found to efficiently degrade DBP, at high concentrations. The degradation followed first-order kinetics, and both strains exhibited a removal efficiency of over 99%. Metabolite analysis revealed that both bacteria utilized de-methylation, de-esterification, and decarboxylation steps during degradation.