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Rifampicin-Resistance Mutations in the rpoB Gene in Bacillus velezensis CC09 have Pleiotropic Effects.

ABSTRACT: Rifampicin resistance (Rifr) mutations in the RNA polymerase ? subunit (rpoB) gene exhibit pleiotropic phenotypes as a result of their effects on the transcription machinery in prokaryotes. However, the differences in the effects of the mutations on the physiology and metabolism of the bacteria remain unknown. In this study, we isolated seven Rifr mutations in rpoB, including six single point mutations (H485Y, H485C, H485D, H485R, Q472R, and S490L) and one double point mutation (S490L/S617F) from vegetative cells of an endophytic strain, Bacillus velezensis CC09. Compared to the wild-type (WT) strain (CC09), the H485R and H485D mutants exhibited a higher degree of inhibition of Aspergillus niger spore germination, while the H485Y, S490L, Q472R, and S490L/S617F mutants exhibited a lower degree of inhibition due to their lower production of the antibiotic iturin A. These mutants all exhibited defective phenotypes in terms of pellicle formation, sporulation, and swarming motility. A hierarchical clustering analysis of the observed phenotypes indicated that the four mutations involving amino acid substitutions at H485 in RpoB belonged to the same cluster. In contrast, the S490L and Q472R mutations, as well as the WT strain, were in another cluster, indicating a functional connection between the mutations in B. velezensis and phenotypic changes. Our data suggest that Rifr mutations cannot only be used to study transcriptional regulation mechanisms, but can also serve as a tool to increase the production of bioactive metabolites in B. velezensis.

SUBMITTER: Cai XC

PROVIDER: S-EPMC5303731 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Rifampicin-Resistance Mutations in the rpoB Gene in Bacillus velezensis CC09 have Pleiotropic Effects.

Cai Xun-Chao XC Xi Huan H Liang Li L Liu Jia-Dong JD Liu Chang-Hong CH Xue Ya-Rong YR Yu Xiang-Yang XY

Frontiers in microbiology 20170213

Rifampicin resistance (Rifr) mutations in the RNA polymerase β subunit (rpoB) gene exhibit pleiotropic phenotypes as a result of their effects on the transcription machinery in prokaryotes. However, the differences in the effects of the mutations on the physiology and metabolism of the bacteria remain unknown. In this study, we isolated seven Rifr mutations in rpoB, including six single point mutations (H485Y, H485C, H485D, H485R, Q472R, and S490L) and one dou ...[more]

PMID: 28243227

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Project description: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, rpoB. Different rpoB mutations are responsible for various levels of rifampicin resistance and for a range of secondary effects. rpoB 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 Vibrio vulnificus, even though rifampicin-resistant strains of V. vulnificus have been isolated previously. Moreover, spontaneous rifampicin-resistant strains of V. vulnificus have an important role in conjugation and mutagenesis protocols, with poor consideration of the effects of rpoB mutations. In this work, effects on growth, stress response and virulence of V. vulnificus were investigated using a set of nine spontaneous rifampicin-resistant derivatives of V. vulnificus 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-1], 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 rpoB mutations and indicate that rifampicin resistance is not an appropriate selectable marker for studies that aim to investigate phenotypic behaviour in this organism.

Dataset Information

Rifampicin-Resistance Mutations in the rpoB Gene in Bacillus velezensis CC09 have Pleiotropic Effects.

Publications

Rifampicin-Resistance Mutations in the <i>rpoB</i> Gene in <i>Bacillus velezensis</i> CC09 have Pleiotropic Effects.

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