Oxidative Stress,Motility And Virulence Of TrxB In Edwardsiella Piscicida

Edwardsiella piscicida（formerly known as Edwardsiella tarda）is an important fish pathogen that can infect various economic fish including flounder and tilapia.However,its pathogenic mechanism remains to be revealed and many new virulence factors remain to be identified.Thioredoxin reductase（TrxB/Trx R）is important for bacterial resistance to oxidative stress,but its function is largely unknown in E.piscicida.In this study,bioinformatics analysis showed that E.piscicida TrxB(TrxB_Ep)shares high sequence homology with thioredoxin reductase in other species and contains a conserved redox-active CATC motif.Although TrxB_Ep is predicted to be extracellular by Protter illustration,Western blot analysis confirmed TrxB_Ep is an intracellular protein.In addition,the thioredoxin reductase activity test showed that TrxB_Ep had thioredoxin reductase activity.The above results indicated that TrxB_Ep is a thioredoxin reductase.By constructing the mutant strain TX01ΔtrxB and the complementary strain TX01ΔtrxBC,the role of TrxB_Ep in the physiological activity of E.piscicida was explored,including biofilm formation ability,motility and flagella formation ability,and some novel findings were obtained.The results showed that the deletion of TrxB_Ep had no effect on the biofilm formation ability of E.piscicida but led to the loss of swimming ability and flagella formation ability,and significantly weakened the swimming ability.It indicates that TrxB_Epplays an important role in the motility and flagella formation of E.piscicida.To explore the role of TrxB_Ep in stress adaptation of E.piscicida,we tested the stress resistance of TX01 and TX01ΔtrxB.The experimental results showed that compared with TX01,TX01ΔtrxB exhibited more resistance against H₂O₂ stress but more sensitivity to thiol-specific oxidizing agent diamide and metal ion oxidants.TX01ΔtrxB also displayed weaker growth than TX01 under acid condition and high-temperature condition.This indicated that TrxB_Ep played an important role in the stress resistance of E.piscicida.To further explore the role of TrxB_Ep in the virulence of E.piscicida,we tested the ability of TX01,TX01ΔtrxB and TX01ΔtrxBC to infect cells in vitro and tissues in vivo,kill tilapia and resist serum sterilization.The results showed that the deletion of TrxB_Ep had no effect on the replication ability of E.piscicida in host cells and had no effect on the survival ability of tilapia,but it significantly weakened the adhesion and invasion of E.piscicida to host cells,and greatly reduced the resistance of bacteria to non-immune host serum,thus weakening the transmission ability of E.piscicida in host tissues.It indicated that TrxB_Ep played an important role in the virulence of E.piscicida.In summary,this study identified the thioredoxin reductase TrxB_Ep of E.piscicida and found that it played an important role in motility,flagella formation,stress resistance,and virulence,providing new insights into the pathogenesis of E.piscicida.