Identification of biphenyl-metabolising microbes in activated biosludge using cultivation-independent and -dependent approaches.

Microbes have important roles in removing organic pollutants in wastewater treatment plants (WWTPs), especially in mineralising recalcitrant persistent organic pollutants (POPs) such as polychlorinated biphenyls (PCBs). However, the majority of the microorganisms that metabolise these pollutants in situ remain elusive owing to barriers of traditional techniques in unravel yet-to-be-cultivated microbes. In this study, DNA stable-isotope probing (SIP) coupled with high-throughput sequencing was applied to identify the microbes responsible for PCB degradation in the activated biosludge of a WWTP using 13 C-labelled biphenyl (BP). Results of time-course SIP revealed different bacteria and archaea involved in BP metabolism, which dominated the BP-degrading community at different time points. BP degradation by the genera Spartobacteria, Alicyclobacillus, Flavobacterium and the order Cenarchaeales has not been reported previously. The abundance of biphenyl dioxygenase (bphA) genes increased over time and a novel bphA gene was identified from the 13 C-heavy DNA fraction. In addition, three cultivable BP degraders were isolated, but did not participate in BP degradation in situ or contain the identified bphA genes. Taken together, these data reveal the huge potential and important roles of yet-to-be-cultivated microbes responsible for PCB degradation in activated biosludge, providing fundamental knowledge on WWTP management to remove POPs.