Genomic evolution of bacterial populations under co-selection by antibiotics and phage

Bacteria live in dynamic systems where selection pressures can alter rapidly, forcing adaptation to the prevailing conditions. In particular, bacteriophages and antibiotics of anthropogenic origin are major bacterial stressors in many environments. We previously observed that populations of the bacterium Pseudomonas fluorescens SBW25 exposed to the lytic bacteriophage SBW25Φ2 and a non-inhibitive concentration of the antibiotic streptomycin (co-selection) achieved higher levels of phage resistance compared to populations exposed to the phage alone. In addition, the phage became extinct under co-selection while remaining present in the phage alone environment. Further, phenotypic tests indicated that these observations might be associated with increased mutation rate under co-selection. In this study, we examined the genetic causes behind these phenotypes by whole-genome sequencing clones isolated from the end of the experiments. We were able to identify genetic factors likely responsible for streptomycin resistance, phage resistance and hypermutable (mutator) phenotypes. This constitutes genomic evidence in support of the observation that while the presence of phage did not affect antibiotic resistance, the presence of antibiotic affected phage resistance. We had previously hypothesized an association between mutators and elevated levels of phage resistance under co-selection. However, our evidence regarding the mechanism was inconclusive, since although with phage mutators were only found under co-selection, additional genomic evidence was lacking and phage resistance was also observed in non-mutators under co-selection. More generally, our study provides novel insights into evolution between univariate and multivariate selection (here two stressors), as well as the potential role of hypermutability in natural communities.

细菌栖息于动态系统中，选择压力可快速波动，迫使细菌适应当前的环境条件。具体而言，人为来源的噬菌体（bacteriophage）与抗生素是诸多环境中细菌面临的主要胁迫因子。我们此前观察到，与仅暴露于裂解性噬菌体的菌群相比，暴露于裂解性噬菌体SBW25Φ2与非抑制浓度链霉素（streptomycin）的共选择（co-selection）条件下的荧光假单胞菌（Pseudomonas fluorescens）SBW25菌群，其噬菌体抗性水平更高。此外，共选择条件下噬菌体最终消亡，而仅施加噬菌体胁迫的环境中噬菌体仍持续存在。进一步的表型实验表明，上述现象可能与共选择条件下突变率升高相关。本研究通过对实验末期分离得到的克隆株进行全基因组测序，探究了上述表型背后的遗传机制。我们成功鉴定出了可能与链霉素抗性、噬菌体抗性以及高突变性（mutator，突变株）表型相关的遗传因子。该结果为下述观测提供了基因组学证据：噬菌体的存在不会影响细菌的抗生素抗性，但抗生素的存在会提升细菌的噬菌体抗性。我们此前曾提出假设，认为共选择条件下突变株与噬菌体抗性水平升高存在关联。然而，我们关于该机制的证据尚不充分：尽管仅在共选择条件下才发现与噬菌体相关的突变株，但仍缺乏额外的基因组学证据，且共选择条件下非突变株中同样观测到了噬菌体抗性。更广泛而言，本研究为单变量选择与多变量选择（本研究中为两种胁迫因子）下的进化过程，以及高突变性在自然群落中的潜在作用提供了全新的见解。