Involvement of RpoN in regulating stress response, biofilm formation and virulence of Vibrio mimicus

Abstract

RpoN, an alternative sigma factor, plays a crucial role in response to changes in environmental cues and pathogenicity. Vibrio mimicus, a serious pathogen in both fresh and seawater aquaculture industries, has garnered considerable attention. Nevertheless, reports of rpoN in V. mimicus remain extremely scarce. In this study, we constructed the rpoN-RNAi strain to investigate the role of rpoN in V. mimicus using phenotypic and RNA-seq analysis. The data indicated that rpoN positively regulated the resistance of V. mimicus to H2O2, ethanol and acid, while negatively regulating the resistance to alkali and heat, and had no influence on the resistance to NaCl. Over the period of 12 to 72 h, the biofilm formation ability of rpoN-RNAi strain was decreased by 0.29 to 0.84-fold compared to wild-type strain. Compared to wild strain, the siderophore production was decreased from 85.9% to 60.3% in rpoN-RNAi strain. Furthermore, the LD50 increased from 6.16 × 105 to 3.92 × 106 CFU/mL in Macrobrachium nipponense through artificial infection test after rpoN silencing. Transcriptome analysis revealed that several well-known virulence-related genes were down-regulated in rpoN-RNAi strain. These genes included the type VI secretion system genes (vipB, vipA, impI, and vasA), type IV pilus assembly protein (flpB-G), outer membrane proteins (ompU and ompA), and hemolysin. Additionally, stress-protective genes encoding heat shock protein, oxidoreductase, aldehyde dehydrogenase, quorum sensing system, etc., were also down-regulated by rpoN-RNAi. Transcriptome analysis was consistent with the observed phenotypes in rpoN-RNAi strain. These results revealed that rpoN is a critical determinant for bacterial fitness and pathogenicity, laying the groundwork for further characterization of RpoN regulatory networks.