Studies on the role of horizontal gene transfer in the stabilisation of shared antibiotic resistance within bacterial populations

抗生素耐药性被认为是当今全球健康面临的最大威胁之一。创新的预防措施和治疗理念的有效实施需要对细菌耐药机制及其演变有全面的了解。然而，目前关于细菌群体中社会微生物相互作用的进化信息很少，这些相互作用使基因型易感细菌在抗生素治疗期间存活和生长。使抗生素失活的酶的产生不仅使产生抗生素的细菌产生耐药性，而且也使其附近的非耐药细菌产生耐药性。这种特殊的合作耐药性病例被称为共享抗生素耐药性（SHARE）。从进化的角度来看，共享是一种利他的社会行为，因为生产细胞消耗能量，而易感细胞从公共物品中获益，而不分担适应度成本。然而，人们认为，只有通过社会依赖或强制机制来否定非生产者的进步或稳定社会特征的发展，利他性抵抗特征才能在更长期内建立起来。有许多迹象表明，水平基因转移（HGT），即移动遗传元素的交换，在细菌种群中利他抗性性状的稳定中起着核心作用。然而，HGT的作用尚未在SHARE的特定背景下得到最终阐明。本研究项目旨在通过对β -内酰胺类抗生素产生抗性的质粒的例子，研究HGT与细菌种群中利他抗性性状稳定之间的相互作用。为此，选择的质粒将被标记上可抑制的绿色荧光蛋白（GFP）标签，并引入到组成性表达相应抑制蛋白的供体菌株中。构建的生物报告将使用荧光激活细胞分选（FACS）和高分辨率共聚焦激光扫描显微镜（CLSM）实时研究表达率和HGT频率。此外，定量实时PCR （qPCR）将用于测量质粒拷贝数。总体研究目标是为质粒转移和质粒拷贝数在SHARE特定背景下的作用提供新的见解，从而为新的预防措施和治疗概念的发展做出重要贡献。

Antibiotic resistance is considered one of the biggest threats to global health today. The effective implementation of innovative prevention measures and therapy concepts requires a comprehensive understanding of bacterial resistance mechanisms and their evolution. However, there is currently only little information available on the evolution of sociomicrobiological interactions within bacterial populations that enable survival and outgrowth of genotypically susceptible bacteria during antibiotic therapy. The production of enzymes that inactivate antibiotics confers resistance not only to the producing bacteria but also to non-resistant bacteria in their immediate vicinity. This particular case of cooperative resistance is referred to as shared antibiotic resistance (SHARE). From the evolutionary perspective, SHARE is a manifestation of altruistic social behaviour, because the producing cell spends energy and the susceptible cell benefits from the public goods without sharing the fitness cost. However, it is believed that altruistic resistance traits can only be established in the longer term if mechanisms are developed that either negate the advancement of non-producers or stabilize the development of social traits through social dependency or enforcement. There are many indications that horizontal gene transfer (HGT), i.e. the exchange of mobile genetic elements, plays a central role in the stabilisation of altruistic resistance traits within bacterial populations. However, the role of HGT has not yet been conclusively elucidated in the particular context of SHARE. This research project aims to study the interplay between HGT and the stabilisation of altruistic resistance traits within bacterial populations by the example of plasmids conferring resistance to beta-lactam antibiotics. For this purpose, selected plasmids will be marked with a repressible green fluorescent protein (GFP) tag and introduced into donor strains that constitutively express the corresponding repressor protein. The constructed bio-reporters will be used to study expression rates and HGT frequencies in real time using fluorescence-activated cell sorting (FACS) and high-resolution confocal laser scanning microscopy (CLSM). Moreover, quantitative real-time PCR (qPCR) will be used to measure plasmid copy numbers. The overall research objective is to provide new insights on the role of plasmid transfer and plasmid copy number in the particular context of SHARE and thus making an important contribution to the development of new prevention measures and therapy concepts.