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Characterisation of p-nitrophenylglycerol-resistant Proteus mirabilis super-swarming mutants
Shwu-Jen Liaw, H-C Lai1, S-W Ho1
1Graduate Institute of Microbiology, *School and Graduate Institute of Medical Technology, College of Medicine, National Taiwan University and †Department of Laboratory Medicine, National Taiwan University Hospital, Taipei, Taiwan, Republic of China.
Abstract:
p-Nitrophenylglycerol (PNPG) inhibits the co-ordinately regulated activities of swarming behaviour and virulence factor expression in Proteus mirabilis. The inhibitory action of PNPG was investigated by the isolation of Tn5 insertion mutants that could swarm, albeit with much reduced ability, in the presence of PNPG. The mutants exhibited a super-swarming phenotype in the absence of PNPG; i.e., they migrated further in a given time than did the wild-type cells. Cloning and sequence analysis of the mutants indicated that Tn5 was inserted into the rsbA gene, which may encode a membrane sensor histidine kinase of the bacterial two-component signalling system. In the absence of PNPG, the mutants exhibited several swarming-related phenotypes that were different from those of the wild type; they initiated swarming earlier and had a less conspicuous consolidation phase, they differentiated earlier and maintained a differentiated state for longer, they started to express virulence factors earlier and maintained high expression levels of these factors for longer, and they had higher cell invasion ability than the wild type. These mutant phenotypes could be complemented by a plasmid-borne copy of rsbA. Together, these data suggest that RsbA may act as a repressor of swarming and virulence factor expression. In the presence of PNPG, these rsbA-mutated mutants could still swarm, differentiate and express virulence factors, whereas the wild type could not, suggesting that PNPG may target RsbA or RsbA-regulated pathways to exert its inhibitory effect. Together, these data reveal a novel mechanism through which bacteria may negatively regulate swarming differentiation and virulence factor expression and identify a potential target of PNPG action.
Insights
p-Nitrophenylglycerol (PNPG) inhibits bacterial swarming and virulence. Mutants lacking the RsbA protein, a potential repressor, bypass PNPG inhibition, revealing RsbA
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- Swarming behavior and virulence factor expression are crucial for bacterial pathogenesis in Proteus mirabilis.
- p-Nitrophenylglycerol (PNPG) is known to inhibit these coordinated activities.
- The precise mechanism of PNPG inhibition and its molecular targets remain largely unelucidated.
Purpose of the Study:
- To investigate the mechanism by which p-Nitrophenylglycerol (PNPG) inhibits swarming and virulence in Proteus mirabilis.
- To identify the bacterial targets or pathways involved in PNPG's inhibitory action.
- To elucidate the role of the RsbA protein in regulating swarming and virulence.
Main Methods:
- Isolation and characterization of Tn5 insertion mutants exhibiting altered responses to PNPG.
- Phenotypic analysis of mutant swarming, differentiation, and virulence factor expression.
- Genetic complementation and sequence analysis to identify the mutated gene (rsbA).
Main Results:
- Tn5 insertion mutants in the rsbA gene displayed a super-swarming phenotype and initiated swarming and differentiation earlier.
- These rsbA mutants showed enhanced and prolonged expression of virulence factors and increased cell invasion ability.
- PNPG failed to inhibit the swarming and virulence of rsbA mutants, suggesting RsbA is a target or is in a targeted pathway.
Conclusions:
- RsbA acts as a repressor of swarming behavior and virulence factor expression in Proteus mirabilis.
- PNPG likely targets RsbA or its regulatory pathway to inhibit bacterial swarming and virulence.
- This study reveals a novel negative regulatory mechanism for bacterial differentiation and virulence, identifying RsbA as a key component and potential PNPG target.