Short Francesca L, Maharjan Ram, Li Liping, Afrin Nusrat, Delgado Natasha, Boinett Christine J, Parkhill Julian, Cain Amy K, Paulsen Ian T
Department of Microbiology, Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.
ARC Centre of Excellence in Synthetic Biology, School of Natural Sciences, Macquarie University, Sydney, New South Wales, Australia.
mBio. 2025 Jun 27:e0096625. doi: 10.1128/mbio.00966-25.
Mutation rates in bacteria are an important determinant of adaptation to new environments and success in different niches. In some bacterial pathogens, "hypermutator" variants-most often associated with mutations in components of the DNA mismatch repair system-are associated with increased antibiotic resistance and poorer patient outcomes. We report the serendipitous finding of novel hypermutator genes in through genome-scale mutant fitness screening. Exposure of a transposon insertion mutant library of to extended weak antibiotic selection resulted in selection for mutations that directly increased fitness as expected, but also revealed genes where transposon insertion indirectly increased fitness due to elevated general mutation rates. Three novel hypermutator genes were confirmed in , encoding a transcription antiterminator; ABUW_0208, encoding a hypothetical protein; and ABUW_2121, which encodes a sulfite transporter. We find selection for hypermutator variants in transposon insertion sequencing (TIS) data sets from diverse bacteria under various antibiotic treatments. Our results expand the range of biological functions linked to hypermutator phenotypes in bacteria and provide a workflow for the identification of putative hypermutators by TIS.IMPORTANCEAll organisms have the capacity for evolution through mutation. Bacteria with high mutation rates have a survival advantage in some stressful environments because they generate beneficial mutations more frequently. "Hypermutators" are bacterial strains that carry gene inactivations that increase general mutation rates. These variants are important in chronic infections, as their increased genetic diversity allows higher drug resistance and prolonged survival in the host. Only a few different hypermutator genes are known, and there is no high-throughput method for their identification. We have made the serendipitous finding that hypermutator genes can be identified by genome-wide mutant fitness screening under specific selection conditions. We have identified novel hypermutator alleles in the notorious hospital pathogen and show that hypermutator variants can be detected in screens of a wide range of pathogens.
细菌中的突变率是其适应新环境及在不同生态位中取得成功的重要决定因素。在一些细菌病原体中，“高突变体”变体（最常见于DNA错配修复系统组件的突变）与抗生素耐药性增加及患者预后较差有关。我们通过全基因组规模的突变体适应性筛选意外发现了新的高突变体基因。将一个转座子插入突变体文库暴露于延长的弱抗生素选择条件下，如预期的那样，筛选出了直接提高适应性的突变，但同时也揭示了一些基因，由于总体突变率升高，转座子插入间接提高了适应性。在该文库中确认了三个新的高突变体基因，分别是编码转录抗终止子的基因；ABUW_0208，编码一种假定蛋白；以及ABUW_2121，它编码一种亚硫酸盐转运蛋白。我们发现在各种抗生素处理下，来自不同细菌的转座子插入测序（TIS）数据集中存在对高突变体变体的选择。我们的结果扩展了与细菌高突变体表型相关的生物学功能范围，并提供了一种通过TIS鉴定假定高突变体的工作流程。重要性所有生物体都有通过突变进行进化的能力。高突变率的细菌在某些压力环境中具有生存优势，因为它们更频繁地产生有益突变。“高突变体”是携带使总体突变率增加的基因失活的细菌菌株。这些变体在慢性感染中很重要，因为它们增加的遗传多样性使细菌具有更高的耐药性并能在宿主体内长期存活。目前已知的不同高突变体基因只有少数几种，并且没有用于鉴定它们的高通量方法。我们意外发现，在特定选择条件下通过全基因组突变体适应性筛选可以鉴定高突变体基因。我们在臭名昭著的医院病原体中鉴定出了新的高突变体等位基因，并表明在广泛的病原体筛选中可以检测到高突变体变体。
