肺炎克雷伯菌是全球抗菌素耐药性上升的主要关注病原体,并已被认为是物种之间抗性基因转移的储库。外排泵的上调是一种特别值得关注的阻力获取机制,在许多情况下,单点突变可以同时提供对一系列抗微生物剂和杀生物剂的抗性。目前的研究调查了oqxR的突变,它编码RND家族外排泵基因的负调节因子,OQXAB,天然存在于肺炎克雷伯菌的染色体中。从单次暴露于30µg/mL氯霉素中选择四种肺炎克雷伯菌菌株(KP6870155,NTUH-K2044,SGH10和ATCC43816)的抗性突变体,并选择12个突变体进行全基因组测序以鉴定与抗性相关的突变。通过单次暴露于氯霉素产生的抗性突变体,四环素,或环丙沙星在≥4×MIC被复制平铺到所有三种抗生素上,以观察对所有化合物的同时交叉抗性,表明多药耐药表型。各种新的突变,包括单点突变,删除,和插入,被发现会破坏OQXR,导致对氯霉素的抗性显着同时增加,四环素,还有环丙沙星.oqxAB-oqxR基因座已动员并分散在许多肠杆菌科物种的质粒上,并检查了这些基因座的多样性以评估作用于这些基因的进化压力。oqxR-oqxAB操纵子质粒拷贝中oqxR启动子区域的比较表明,一些构建体可能会产生oqxR转录本的截短版本,这可能会影响OqxAB的调控和表达。在某些情况下,在肺炎克雷伯菌中发现了编码oqxAB-oqxR的染色体和质粒的共同携带,这意味着有选择性的压力来维持和扩大外排泵。鉴于OqxR是OqxAB的阻遏物,任何影响其表达或功能的突变都可能导致多药耐药性。这与抗生素靶位点突变形成对比,所述抗生素靶位点突变必须发生在有限的序列空间中才能有效且不影响细胞的适应性。因此,oqxR可以作为简单的遗传开关通过OqxAB介导的外排促进抗性。
Klebsiella pneumoniae is a pathogen of major concern in the global rise of antimicrobial resistance and has been implicated as a reservoir for the transfer of resistance genes between species. The upregulation of efflux pumps is a particularly concerning mechanism of resistance acquisition as, in many instances, a single point mutation can simultaneously provide resistance to a range of antimicrobials and biocides. The current study investigated mutations in oqxR, which encodes a negative regulator of the RND-family efflux pump genes, oqxAB, natively found in the chromosome of K. pneumoniae. Resistant mutants in four K. pneumoniae strains (KP6870155, NTUH-K2044, SGH10, and ATCC43816) were selected from single exposures to 30 µg/mL chloramphenicol and 12 mutants were selected for whole genome sequencing to identify mutations associated with resistance. Resistant mutants generated by single exposures to chloramphenicol,
tetracycline, or ciprofloxacin at ≥4 X MIC were replica plated onto all three antibiotics to observe simultaneous cross-resistance to all compounds, indicative of a multidrug resistance phenotype. A variety of novel mutations, including single point mutations, deletions, and insertions, were found to disrupt oqxR leading to significant and simultaneous increases in resistance to chloramphenicol,
tetracycline, and ciprofloxacin. The oqxAB-oqxR locus has been mobilized and dispersed on plasmids in many Enterobacteriaceae species and the diversity of these loci was examined to evaluate the evolutionary pressures acting on these genes. Comparison of the promoter regions of oqxR in plasmid-borne copies of the oqxR-oqxAB operon indicated that some constructs may produce truncated versions of the oqxR transcript, which may impact on oqxAB regulation and expression. In some instances, co-carriage of chromosomal and plasmid encoded oqxAB-oqxR was found in K. pneumoniae, implying that there is selective pressure to maintain and expand the efflux pump. Given that OqxR is a repressor of oqxAB, any mutation affecting its expression or function can lead to multidrug resistance. This is in contrast to antibiotic target site mutations that must occur in limited sequence space to be effective and not impact the fitness of the cell. Therefore, oqxR may act as a simple genetic switch to facilitate resistance via OqxAB mediated efflux.