摘要

      最近越来越多的证据表明，重金属可以刺激细菌之间的抗生素耐药基因（ARGs）转移。然而，以往的研究大多集中在纯菌株上，重金属对细菌群落尤其是活性污泥中ARGs转移的影响尚未得到明确的探索。本研究采用计算机培养箱（Bioscreen C）和流式细胞仪相结合的高通量方法，研究了不同浓度的重金属对污泥细菌群落中ARGs迁移的影响。以大肠杆菌MG1655作为大范围IncP-1质粒pKJK5的供体，发现0.5 mmol/L铅、0.1 mmol/L砷和0.005 mmol/L汞均能明显促进污泥细菌群落中ARGs的转移。此外，微流控芯片上的交配实验也证明，在上述重金属胁迫下，附着群落的转移频率较高。分离了铅、砷、汞胁迫下的转录结合子，并进行了系统发育描述。砷和汞以假单胞菌属为主，分别占88%和96%。铅胁迫下，以气单胞菌属和肠杆菌属为主，分别占56%和32%。ABC转运蛋白和氨基酸代谢与重金属转运和细胞代谢有关，在预测转运蛋白的微生物代谢功能中占主导地位。本研究有助于污水处理厂ARGs扩散的风险评估和控制。

        Recent growing evidence suggests that heavy metals can stimulate the transfer of antibiotic resistance genes (ARGs) between bacteria. However, most previous studies focused on pure strains, the effect of heavy metals on ARG transfer in bacterial communities, especially in activated sludge, has not been clearly explored. In this study, a high-throughput method, combining computerized incubator (Bioscreen C) and flow cytometry, was developed to evaluate different concentrations of heavy metals influencing ARG transfer in sludge bacteria communities. By using Escherichia coli MG1655 as the donor of broad-host range IncP-1 plasmid pKJK5, it was found that 0.5 mmol/L Pb, 0.1 mmol/L As and 0.005 mmol/L Hg could obviously promote ARG transfer in sludge bacteria communities. Furthermore, mating assays on microfluidic chips also proved higher transfer frequencies in attached communities under the above heavy metal stresses. Transconjugants under Pb, As and Hg stresses were isolated and phylogenetically described. For As and Hg, the dominant genus was Pseudomonas, accounting for 88% and 96%, respectively. While under Pb stress, the genera Aeromonas and Enterobacter were the main transconjugants, accounting for 56% and 32% respectively. Moreover, ABC transporters and Amino acid metabolism, which were related to heavy metal transport and cellular metabolism, were dominant in the prediction of microbial metabolic function of transconjugants. This study can be helpful for risk assessment and control of ARG spreading in WWTPs.

        https://linkinghub.elsevier.com/retrieve/pii/S0048-9697(19)33810-0