Regulators Of Two-component Signal Transduction Systems On Pathogenesis Of Salmonella Enterica Serovar Typhimurium

Salmonellosis is one of the main bacterial diseases in farming,livestock infected with Salmonella can cause the corresponding infectious diseases,such as S.choleraesuis,Salmonella pullorum.When contaminated meat food,the bacteria entered the human food chain and caused food poisoning in humans and had a great threat to human health.The purpose of this experiment is to study regulation of two-component signal transduction system cpx AR in Salmonella on typhimurium pathogenicity,to find a way to control the severe pathogenicity and and to lay the foundation for preparation of live attenuated vaccine.1.This study screened the epidemic strain JS_c(clinical isolate)by ERIC-PCR,the multi-locus sequence typing(MLST)and constructed the cpx R gene deletion mutant named as JS_c△cpx R by P22 phage transduction.Then the lethal dose(LD50)of JS_s(standard strains)、JS_s△cpx R、JS_c、JS_c△cpx R strains in mice were determined.The LD50 of JS_s△cpx R increased by about 1.36 times compared with that of its parent strain,while the LD50 of JS_c △cpx R increased by about 17.54 times compared with that of its parent strain,suggesting that the virulence of cpx R gene deletion strain become less virulent than that of the parent strain.In conclusion,cpx R gene is closely related to the pathogenicity of S.enterica Serovar Typhimurium and have important effects on the pathogenicity of S.enterica Serovar Typhimurium.2.Determining the ability of JS_s(standard strains),JS_c(clinical isolate)and constructing the cpx R gene deletion mutant named as JS_c△cpx R 、JS_s△cpx R on adhesion and invasion PK-15 cells.The results showed that the adhesion and invasion of JS_c were strong,and the adhesion and invasion ability of JS_c was higher than those of JS_s.the adhesion and invasion of cpx R gene deletion strain were significantly decreased than that of the parent strain,The adhesion amounts of JS_s△cpx R and JS_c △cpx R decreased respectively by about 1.35、2 times compared with that of their parent strains,while the invasion amounts of JS_s△cpx R and JS_c △cpx R decreased respectively by about 3.4 、 3 times compared with that of their parent strains,suggesting that cpx R gene is closely related to the adhesion and invasion of S.enterica Serovar Typhimurium.3.Real-time PCR analysis was applied to analyze m RNA expression levels of the 11 Virulence Related Genes of 7 strains and cpx R gene deletion strains.The results showed that m RNA expression levels of the gene related to virulence of Salmonella typhimurium,such as hil D,sop D,fur,inv A,sop E2,ssr A,spv C,hil A,inv F,sop B and sip A in JS_s△cpx R decreased totally compared to those of the JS_s(standard strains),but in clinical isolates cpx R gene deletion strains 6△cpx R、18△cpx R、25′△cpx R、35△cpx R、57△cpx R、69△cpx R、110△cpx R,the relative expression of strains 6,18,25,35,57,69’,110,fur and spv C decreased totally,while sip A,hil A,inv F,sop B,hil D,sop D,inv A,sop E2 and ssr A in 6△cpx R、18△cpx R、25′△cpx R、35△cpx R have a slightly declined and in 57△cpx R 、 69△cpx R 、110△cpx R significantly increased,as high as 354.6 times.The result shows variety.Point out cpx R gene and pathogenicity of S.enterica Serovar Typhimurium have close relationship.However,the virulence or pathogenicity had great differences in clinical isolated Salmonella typhimurium.Cpx R gene is not the only factor to regulate virulence gene of S.enterica Serovar Typhimurium expression.There might also be a complex regulatory network with the closely related to the cpx R regulation mechanism.