Biodegradation pathway of di-(2-ethylhexyl) phthalate by a novel Rhodococcus pyridinivorans XB and its bioaugmentation for remediation of DEHP contaminated soil

Highlights: • A novel Rhodococcus pyridinivorans XB capable of degrading DEHP was isolated. • Observation by atomic force microscope confirmed well growth of strain XB under DEHP stress. • DEHP metabolites with dioxygenase genes revealed its complete degradation pathway. • Bioaugmentation of this strain can enhance DEHP removal from contaminated soils. A novel bacterial strain designated as Rhodococcus pyridinivorans XB, capable of utilizing various endocrine disruptor phthalates or phthalic acid (PA) as sole source of carbon and energy, was isolated from activated sludge. Under the optimal culture conditions (pH 7.08, 30.4 °C, inoculum size (OD_{600 nm}) of 0.6) obtained by response surface methodology, di-(2-ethylhexyl) phthalate (DEHP, 200 mg/L) could be degraded by strain XB with a removal rate of 98% within 48 h. Under the observation of an atomic force microscope, it was confirmed that DEHP did not inhibit the growth of strain XB which might produce some extracellular polymeric substances as a response to DEHP stress, resulting in rapid degradation of DEHP. At initial concentrations of 50–800 mg/L DEHP, its degradation curves were well fitted with the first-order kinetic model, and the half-life of DEHP degradation varied from 5.44 to 23.5 h. The degradation intermediates of DEHP were identified by both GC–MS and high performance liquid chromatography–time of flight-mass spectrometry (HPLC–TOF-MS). Significant up-regulation was observed for the relative expression levels of genes (i.e., phthalate hydrolase, PA 3,4-dioxygenase, protocatechuate 3,4-α and 3,4-β dioxygenase) involved in DEHP degradation determined by real-time quantitative PCR (RT-qPCR). A DEHP biodegradation pathway by strain XB was proposed based on the identified intermediates and the degrading genes. Bioaugmentation of DEHP-contaminated soils with strain XB could efficiently promote DEHP removal, offering great potential in bioremediation of DEHP-contaminated environment.