Enhanced Biodegradation Of Polychlorinated Biphenyls By Rhodococcus With Resuscitation-promoting Factor Supplementing And Its Application In Soil Pollution Bioremediation

Polychlorinated biphenyls?PCBs?are still one of the typical pollutants in the global soils.Microbial degradation has been considered a promising technology in the removal of PCBs in soils.However,its effectiveness could be limited partially because PCBs degraders are prone to enter a viable but nonculturable?VBNC?state under adverse conditions.Currently,there is very little research in this area.In this study,a highly efficient PCBs-degrading bacterium named Rhodococcus biphenylivorans TG9was induced to enter the VBNC state by the antibiotic norfloxacin.We investigated the mechanism of the resuscitation-promoting factor?Rpf?protein derived from Micrococcus lutues on the resuscitation,growth stimulation and enhanced PCBs biodegradation of strain TG9,and verified the role of Rpf in the bioremediation of simulated soils contaminated with PCBs.The main results and conclusions are as follows:?1?In the pure bacteria system of culture medium,the supplementation of exogenous Rpf promoted the resuscitation of TG9 cells in the VBNC state,and stimulated the proliferation of resuscitated TG9 cells.During the resuscitation process,the endogenous expression levels of four rpf gene orthologs of strain TG9 in the group with exogenous Rpf supplementing were significantly higher than those in the control group without Rpf added?P<0.05?.Exogenous Rpf stimulated the endogenous expression of rpf gene orthologs of strain TG9 might be an important mechanism by which Rpf promoted the resuscitation and stimulated the growth of TG9 cells.?2?In the pure bacterial degradation system,the degradation ability of resuscitated TG9 cells to biphenyl?BP?was significantly higher than that of cells in the exponential phase and VBNC state?P<0.05?.Supplementing Rpf enhanced biodegradation of PCBs by strain TG9.Under the condition of using 50 mg L^-1 PCB12 as the sole carbon source and energy source,the addition of Rpf promoted the resuscitation and stimulated the proliferation of strain TG9 in the VBNC state.After 72 h of incubation,the degradation efficiency of PCB12 reached 20.2%,which was significantly higher than that of the sterilized Rpf control group and the solvent control group?P<0.05?.Supplementing Rpf also stimulated the growth of strain TG9 in the exponential phase.The degradation efficiency of PCB12 at 72 h reached 87.6%,which was significantly higher than that of the sterilized Rpf control group and the solvent control group?P<0.05?.Additionally,the supplementation of Rpf did not stimulate the expression of key genes bph A and bph C involved in the PCBs degradation.Rpf promoted the resuscitation and stimulated the growth of strain TG9,thereby increasing the cell density of strain TG9,which might be the key mechanism to enhance PCBs degradation.?3?In the bioremediation of simulated soils contaminated with PCBs,the supplementation of Rpf promoted the removal efficiency of PCBs.For soil microcosms inoculated with exponential-phase and VBNC TG9 cells that originally contained 38.6mg kg^-1 PCBs,after 49 d of Rpf addition,the removal efficiency of PCBs reached 35.8%and 34.2%,respectively,which was significantly higher than the respective solvent control group and sterilized Rpf control group?P<0.05?.Additionally,supplementing Rpf improved the overall culturability of the soil bacteria,affected the soil bacterial community structure,and increased the relative abundance of Rhodococcus which played an important role in the biodegradation of PCBs.Furthermore,the addition of Rpf stimulated the growth of TG9-like bacteria and increased their activity.The expression levels of PCBs-degrading genes bph A,bph C,and rpf gene orthologs in per gram of soil increased with Rpf addition.It indicated that Rpf promoted the resuscitation and stimulated the growth of PCBs-degrading bacteria that responded to it,thereby enhancing the bioremediation of PCBs-contaminated soil.The results of this study further clarified the mechanism of Rpf on the resuscitation,growth stimulation and enhanced organic pollutants biodegradation of Rhodococcus.This research has certain significance for developing new technologies and preparations for the remediation of organic pollutants,which ultimately contributed to the improvement of microbial remediation efficiency in the practical application.