Distribution And Transmission Risk Of Antibiotic Resistance Genes In Wastewater Treatment Systems

Antibiotic resistance genes（ARGs）have become the focus of environmental research in recent years.As an important repository of ARGs,wastewater treatment plant has attracted extensive attention.Understanding the distribution characteristics of ARGs in sewage treatment plants and the transmission risk of ARGs in effluent is of great significance for the control research and safety assessment of ARGs in sewage treatment plants in the future.In this study,the distribution characteristics of ARGs in different processes（AAO,MBR and MBBR）were studied based on metagenomics,and the co-occurrence relationship between bacteria and ARGs was discussed,so as to reveal the distribution rules of ARGs in different processes and the transmission risk of ARGs.Secondly,using Escherichia coli（E.coli）as model bacteria,microfluidic agarose chip（MAC）was used to quantify the ability of residual bacteria to receive ARGs under different disinfection conditions.The effect of different disinfection methods on gene expression of residual bacteria was studied by extracting RNA of residual bacteria.The main findings are as follows:（1）Proteobacteria,Actinobacteria and Bacteroidete are the dominant bacteria in all the processes.The diversity of bacteria in the wastewater treated by different wastewater treatment plants（D-type filter,MBR membrane filtration and Na Cl O disinfection）can be reduced,and the species of bacteria in the wastewater treated by the three wastewater treatment methods can be reduced by 0.52%,3.14%and 6.54%,respectively.（2）D-type filter can reduce the ARG subtypes in effluent（2.99%and 11.28%）,Na Cl O disinfection also reduced ARG subtypes in effluent by 13.58%,while MBR membrane filtration increased the ARG subtypes in effluent by 12.60%.Mac B is the ARG subtype with the largest relative abundance in all samples.（3）The proportion of Mycobacterium in water produced by AAO process increased significantly.Mycobacterium are potential hosts of rifampin phosphotransferase,ole C,ddr A and D-Ala-D-Ala-ligase.Nitrospira is the potential host of resistance genes of Staphylococcus mup,bcr A,bac A,efr A,rpo B2 and tet B（P）in MBR effluent and activated sludge.Nitrospira,the dominant bacteria of suspended filler biofilm in MBBR process,is the potential host of 20 ARG subtypes.Hyphomicrobium and Bradyrhizobium were the hosts of 11 and 7 subtypes of ARG,respectively,which were the dominant bacteria in both activated sludge and suspended packed biofilms.Further attention should be paid to the risk of ARGs transmission by these bacteria.（4）The ability of residual bacteria to receive ARGs increased after Na Cl O and Cl O₂ disinfection,but decreased after UV disinfection.The average transfer rate of ARGs in E.coli sensitive bacteria residual bacteria,kanamycin resistant residual bacteria and tetracycline resistant residual bacteria after Na Cl O disinfection were0.84%,1.10%and 0.18%,which were higher than those of the control group（0.28%,0.17%and 0）.The average transfer rates of E.coli sensitive residual bacteria,kanamycin resistant residual bacteria and tetracycline resistant residual bacteria after Cl O₂disinfection were 2.66%,0.96%and 0.55%,which were higher than those of the control group（0.28%,0.17%and 0）.The average transfer rates of E.coli sensitive bacteria residual bacteria and kanamycin resistant residual bacteria after UV disinfection were 0.04%and 0.08%,which were lower than those of the control group（0.28%and 0.17%）.Confocal laser Scanning microscope（CLSM）image analysis shows that disinfection affected the distribution characteristics of bacteria on the biofilm,and the growth of donor and recipient bacteria was conducive to the production of transconjugant.（5）Transcriptome sequencing showed that the three disinfection methods promoted the up regulated expression of reactive oxygen-related genes such as SODA,SODC,SOXR and SOXS in E.coli sensitive bacteria.Na Cl O and Cl O₂ disinfection promoted up-regulated expression of genes related to pili production in E.coli sensitive bacteria,while some genes related to pili production（fli D,fli L,fli M,fli N,fli S and fim H）were down-regulated after UV disinfection.