Discipline of Microbiology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
Institute of Microbiology, University of Greifswald, D-17489 Greifswald, Germany.
Microbiology (Reading). 2020 Dec;166(12):1160-1170. doi: 10.1099/mic.0.000991.
Rifampicin is a broad-spectrum antibiotic that binds to the bacterial RNA polymerase (RNAP), compromising DNA transcription. Rifampicin resistance is common in several microorganisms and it is typically caused by point mutations in the gene encoding the β subunit of RNA polymerase, . Different mutations are responsible for various levels of rifampicin resistance and for a range of secondary effects. mutations conferring rifampicin resistance have been shown to be responsible for severe effects on transcription, cell fitness, bacterial stress response and virulence. Such effects have never been investigated in the marine pathogen , even though rifampicin-resistant strains of have been isolated previously. Moreover, spontaneous rifampicin-resistant strains of have an important role in conjugation and mutagenesis protocols, with poor consideration of the effects of mutations. In this work, effects on growth, stress response and virulence of were investigated using a set of nine spontaneous rifampicin-resistant derivatives of CMCP6. Three different mutations (Q513K, S522L and H526Y) were identified with varying incidence rates. These three mutant types each showed high resistance to rifampicin [minimal inhibitory concentration (MIC) >800 µg ml], but different secondary effects. The strains carrying the mutation H526Y had a growth advantage in rich medium but had severely reduced salt stress tolerance in the presence of high NaCl concentrations as well as a significant reduction in ethanol stress resistance. Strains possessing the S522L mutation had reduced growth rate and overall biomass accumulation in rich medium. Furthermore, investigation of virulence characteristics demonstrated that all the rifampicin-resistant strains showed compromised motility when compared with the wild-type, but no major effects on exoenzyme production were observed. These findings reveal a wide range of secondary effects of mutations and indicate that rifampicin resistance is not an appropriate selectable marker for studies that aim to investigate phenotypic behaviour in this organism.
利福平是一种广谱抗生素，它与细菌 RNA 聚合酶（RNAP）结合，从而破坏 DNA 转录。利福平耐药在几种微生物中很常见，通常是由于编码 RNA 聚合酶β亚基的基因发生点突变引起的。不同的突变导致不同程度的利福平耐药，并产生一系列次要影响。已证实赋予利福平耐药性的突变会导致转录、细胞适应性、细菌应激反应和毒力产生严重影响。尽管以前已经分离出耐利福平的 菌株，但这种影响在海洋病原体 中从未被研究过。此外，耐利福平的 自发突变株在接合和诱变方案中具有重要作用，但对 突变的影响考虑不足。在这项工作中，使用一组 9 种自发耐利福平的 CMCP6 衍生株研究了对生长、应激反应和毒力的影响。鉴定出三种不同的突变（Q513K、S522L 和 H526Y），其发生率不同。这三种突变型均表现出对利福平的高度耐药性[最小抑菌浓度（MIC）＞800μg/ml]，但具有不同的次要影响。携带 H526Y 突变的菌株在丰富培养基中有生长优势，但在高 NaCl 浓度下耐盐应激能力严重降低，以及乙醇应激抗性显著降低。携带 S522L 突变的菌株在丰富培养基中的生长速度和总体生物量积累都降低了。此外，对毒力特征的研究表明，与野生型相比，所有耐利福平的菌株的运动性都受到了损害，但没有观察到对外酶产生的主要影响。这些发现揭示了 突变的广泛的次要影响，并表明利福平耐药性不是研究该生物表型行为的合适选择标记。
