Characterization of Edwardsiella tarda rpoN: roles in σ70 family regulation, growth, stress adaption and virulence toward fish
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Edwardsiella tarda EIB202, a Gram-negative pathogen with strong virulence, is an opportunistic pathogen capable of causing edwardsiellosis with high mortality to fish. Alternative sigma factor 54 (RpoN) is an important regulator of virulence and stress resistance genes in many bacterial species and mainly responsible for transcription of genes in nitrogen utilization. In this study, the in-frame rpoN deletion mutant was constructed to analyze the function of RpoN in Edwardsiella tarda firstly. Compared to the wild-type and complemented strain rpoN +, the ΔrpoN was impaired in terms of the ability to survive under oxidative stress, osmotic stress and acid resistance, as well as the growth in Luria–Bertani medium, demonstrating essential roles of RpoN in stress resistance and nitrogen utilization. In addition, the ΔrpoN displayed markedly decreased biofilm formation and chondroitinase activity and was attenuated in virulence reflected in the increased median lethal dose value and extended infection cycle. Real-time polymerase chain reaction revealed that the expression levels of σ70 class changed in varying degrees in the rpoN mutant. Especially, the expression levels of rpoS and fliA were down-regulated 4.1-fold and 7.9-fold in stationary phase in comparison with the wild type, respectively. Furthermore, two differential expression genes, znuA and flhC, were detected in the wild type and ΔrpoN using the method of differential display reverse transcription PCR.
KeywordsEdwardsiella tarda rpoN Biofilm formation Virulence
We thank Dr. Zhaolan Mo (Institute of Oceanology, Chinese Academy of Sciences, China) for kindly sending strains and plasmids used for the mutant construction in this study. This work was supported by the National Special Fund for State Key Laboratory of Bioreactor Engineering.
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