Effect Of Xylose On Proliferation And Virulence Of Salmonella Typhimurium

Most of the Salmonella are zoonotic pathogens,which have great safety hazards to humans and will bring huge economic losses to the breeding industry.Antibiotic treatment is commonly used for Salmonella infection.However,due to the long-term use and abuse of antibiotics,bacterial resistance is induced.Therefore,it is urgent to find new anti-infective strategies and drugs to prevent and treat Salmonella infection.In this paper,different concentrations of xylose were used to treat Salmonella Typhimurium,and the treated bacteria were incubated with pig intestinal epithelial cells(IPEC-J2).Analyzed bacterial adhesion rate,invasive rate,degree of cell damage,the mRNA expression levels of tight junction and inflammation-related genes to investigate the regulation of xylose on the proliferation and virulence of Salmonella Typhimurium.Experiment 1: Effect of xylose on proliferation and virulence gene expression of Salmonella TyphimuriumThe experiment was divided into 6 groups,with 4 replicates in each group.The xylose concentration in the experimental group was 0.625 mg/m L,1.25 mg/m L,2.5 mg/m L,5mg/m L and 10 mg/m L,respectively.Add equal volume of PBS to the control group.Determined the proliferation curve of Salmonella Typhimurium,the pH of the medium and the mRNA expression level of the virulence gene.The results showed:(1)0.625 mg/m L,1.25 mg/m L and 2.5 mg/m L xylose promoted significantly the proliferation of Salmonella Typhimurium(P<0.05);5 mg/m L and 10 mg/m L xylose significantly promoted the proliferation of Salmonella Typhimurium before 12 h(P<0.05),but inhibited its proliferation significantly after 24 h(P<0.05).Moreover,the concentration,time,interaction of xylose concentration and time had a significant effect on the proliferation of Salmonella Typhimurium(P<0.05).(2)When cultured until 12 h,0.625 mg/m L,1.25 mg/m L,2.5 mg/m L,5 mg/m L and10 mg/m L xylose significantly reduced the pH of Salmonella Typhimurium culture medium in logarithmic growth phase(P<0.05),and there was dose-dependent effection.(3)Salmonella Typhimurium was treated with 0.625 mg/m L and 1.25 mg/m L xylose for 12 h,which significantly reduced the mRNA expression levels of virulence genes Hil C,invF and SipA(P<0.05),and there was no significant effect on the mRNA expression of virulence genes HilD and Hil A(P>0.05);there was no significant effect on the mRNA expression of virulence genes,when Salmonella Typhimurium was treated with 2.5 mg/m L and 10 mg/m L xylose for 12 h(P>0.05);Salmonella Typhimurium was treated with 5mg/m L xylose for 12 h,significantly increased the mRNA expression of HilD,invF and SipA(P<0.05),and had no significant effect on the mRNA expression of Hil C and Hil A(P>0.05).When 0.625 mg/m L,1.25 mg/m L and 2.5 mg/m L xylose acted on Salmonella Typhimurium for 24 h,there was no significant effect on the mRNA expression of virulence genes of Salmonella Typhimurium(P>0.05);5 mg/m L and 10 mg/m L xylose could significantly increased the mRNA expression of virulence genes of Salmonella Typhimurium(P<0.05).Experiment 2: Effect of xylose treated Salmonella Typhimurium on IPEC-J2 injuryThere were 7 groups in this experiment,5 replicates in each group.The experimental group treated IPEC-J2 with Salmonella Typhimurium treated with different concentrations of xylose,the negative control group was treated with PBS-treated Salmonella Typhimurium,and the blank control group used an equal volume of PBS to determine the adhesion rate and invasive rate of Salmonella Typhimurium to IPEC-J2,LDH content of cell culture supernatant,mRNA expression level of tight junction and inflammation-related genes in IPEC-J2.The results showed:(1)Compared with the negative control group,0.625 mg/m L xylose treatment of Salmonella Typhimurium can significantly increase its adhesion to IPEC-J2(P<0.05),and was no significant effect on its invasion(P>0.05),treatment of Salmonella Typhimurium with 1.25 mg/m L xylose significantly increased its invasion to IPEC-J2(P<0.05),and had no significant effect on its adhesion(P>0.05),treatment of Salmonella Typhimurium with2.5 mg/m L,5 mg/m L and 10 mg/m L xylose didn’t significant effect on adhesion and invasion of IPEC-J2(P>0.05).(2)Compared with the negative control group,2.5 mg/m L xylose treatment of Salmonella Typhimurium can significantly increase the release of lactate dehydrogenase(LDH)from IPEC-J2(P<0.05),0.625 mg/m L,1.25 mg/m L,5 mg/m L and 10 mg/m L xylose treatment of Salmonella Typhimurium had no significant effect on the release of LDH from IPEC-J2(P>0.05).(3)Compared with the negative control group,0.625 mg/m L,1.25 mg/m L and 2.5mg/m L xylose treatment of Salmonella Typhimurium had no significant effect on the mRNA expression of Claudin-1,Occludin and ZO-1 in IPEC-J2(P>0.05);5 mg/m L xylose treatment of Salmonella Typhimurium significantly increased the expression of Occludin and ZO-1 in IPEC-J2(P<0.05),and had no significant effect on Claudin-1 mRNA expression(P>0.05);Salmonella Typhimurium treated with 10 mg/m L xylose significantly increased the mRNA expression of ZO-1 in IPEC-J2(P<0.05),and had no significant effect on the mRNA expression of Claudin-1 and Occludin(P>0.05).(4)Compared with the negative control group,0.625 mg/m L and 2.5 mg/m L xylose treatment of Salmonella Typhimurium had no significant effect on the mRNA expression of p38,p65 and IL-1β in IPEC-J2(P>0.05);1.25 mg/m L xylose treatment of Salmonella Typhimurium significantly increased the expression of IL-1β in IPEC-J2(P<0.05),and had no significant effect on the mRNA expression of p38 and p65(P>0.05);5 mg/m L xylose treatment of Salmonella Typhimurium significantly increased the mRNA expression levels of p38 and IL-1β in IPEC-J2(P<0.05),had no significant effect on the mRNA expression of p65(P>0.05);10 mg/m L xylose treatment of Salmonella Typhimurium significantly increased the mRNA expression levels of p38,p65 and IL-1β in IPEC-J2(P<0.05).In summary,xylose treatment for 12 h can promote the proliferation of Salmonella Typhimurium,and 5 mg/m L xylose treatment of Salmonella Typhimurium can increase its inflammatory damage to IPEC-J2.The possible reasion is that xylose up-regulat mRNA expression of virulence genes in Salmonella Typhimurium.