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的最大威胁之一。有效实施创新的预防措施和治疗概念需要全面了解细菌耐药机制及其演变。然而，目前只有很少的信息可用于细菌种群内的社会微生物学相互作用的演变，使抗生素治疗期间的基因型敏感细菌的生存和生长。抗生素分解酶的产生不仅使产生抗生素的细菌产生耐药性，而且使其附近的非耐药性细菌产生耐药性。这种合作耐药的特殊情况被称为共享抗生素耐药（SHARE）。从进化的角度来看，SHARE是利他主义社会行为的一种表现，因为生产细胞消耗能量，易感细胞从公共产品中受益，而不分担健身成本。然而，人们认为，利他主义的抵抗特征只能在长期内建立，如果机制的发展，要么否定非生产者的进步，或通过社会依赖或强制稳定社会特征的发展。有许多迹象表明，水平基因转移（HGT），即移动的遗传元件的交换，在细菌种群内利他抗性性状的稳定中起着核心作用。然而，HGT的作用还没有最终阐明在共享的特定背景下。该研究项目旨在通过赋予β-内酰胺抗生素抗性的质粒的例子来研究HGT与细菌群体内利他抗性性状的稳定之间的相互作用。为此目的，选择的质粒将用可阻遏的绿色荧光蛋白（GFP）标签标记，并引入组成型表达相应阻遏蛋白的供体菌株中。构建的生物报告基因将用于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）在真实的时间中研究表达率和HGT频率。此外，定量实时PCR（qPCR）将用于测量质粒拷贝数。总体研究目标是提供新的见解的作用，质粒转移和质粒拷贝数的特定背景下的共享，从而作出重要贡献的新的预防措施和治疗概念的发展。

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.