摘要

      抗生素耐药基因（ARGs）的传播已成为威胁全球健康的新问题。虽然水平基因转移（HGT）被认为是基因转移的主要途径之一，但更多的证据表明垂直基因转移（VGT）的参与是非常重要的。然而，传统的基于栽培的方法不能区分HGT和VGT，这往往导致矛盾的结论。本文利用单细胞微流控技术和延时成像技术，分析了质粒介导的HGT和VGT对ARG在环境中传播的影响。以抗甲氧苄啶的ARG编码质粒pKJK5大肠杆菌为供体，定量研究了三种代表性抗生素（甲氧苄啶、四环素和阿莫西林）对活性污泥菌群ARG转移过程的影响。结果表明，HGT受抗生素及其靶点（供者、受体单独或共同）的抑制机制影响，而VGT对转连接子的形成和ARG的扩散有重要作用。甲氧苄啶被供体和转接剂高度耐药，其存在显著增加了HGT和VGT的发生率。四环素和阿莫西林对供者均有抑制作用，但由于抑制机制不同，对HGT的抑制作用也不同。此外，我们还证明了HGT在社区中的传播动力学可以用流行病感染模型来描述，该模型结合HGT和VGT芯片上的定量测量，为研究和预测ARG在现实社区的传播动力学提供了一个有前途的工具。

       The spread of antibiotic resistance genes (ARGs) has become an emerging threat to the global health. Although horizontal gene transfer (HGT) is regarded as one of the major pathways, more evidence has shown the significant involvement of vertical gene transfer (VGT). However, traditional cultivation-based methods cannot distinguish HGT and VGT, resulting in often contradictory conclusions. Here, single-cell microfluidics with time-lapse imaging has been successfully employed to dissect the contribution of plasmid-mediated HGT and VGT to ARG transmission in an environmental community. Using Escherichia coli with an ARG-coded plasmid pKJK5 with trimethoprim resistance as the donor, we quantified the effects of three representative antibiotics (trimethoprim, tetracycline and amoxicillin) on the ARG transfer process in an activated sludge bacterial community. It was found that HGT was influenced by the inhibitory mechanism of an antibiotic and its targets (donor, recipient alone or together), whereas VGT contributes significantly to the formation of transconjugants and consequently ARG spreading. Trimethoprim is highly resisted by the donor and transconjugants, and its presence significantly increased both the HGT and VGT rates. Although tetracycline and amoxicillin both inhibit the donor, they showed different effects on HGT rate as a result of different inhibitory mechanisms. Furthermore, we show the kinetics of HGT in a community can be described using an epidemic infection model, which in combination with quantitative measure of HGT and VGT on chip provides a promising tool to study and predict the dynamics of ARG spread in real-world communities.

      https://www.sciencedirect.com/science/article/pii/S0160412019307871?via%3Dihub