Antibiotic And Disinfectant Resistance Of Salmonella Enterica Typhimurium And Disinfection Effect Of Benzalkonium Chloride On Eggs

This study isolated and identified Salmonella Typhimurium in retail pork and chicken collected from major supermarkets and farmers' markets in Wenjiang district,Chengdu city,Sichuan province during 2017-2018.According to the methods and standards recommended by the Clinical and Laboratory Standards Institute(CLSI),Kirby-Bauer disc diffusion method and broth microdilution method were used to determine the resistance phenotype of Salmonella Typhimurium to antibiotics and disinfectants,and disinfectant resistance genes were detected by polymerase chain reaction(PCR)amplification.Then,on the basis of resistance phenotype and genotype,constructing a resistance gene expression vector with a high detection rate of resistance genes to study the ability of resistance genes to mediate resistance.Finally,based on Salmonella Typhimurium disinfectant resistance and the requirements for the use of food disinfectants,benzalkonium chloride was selected to study the disinfection effect of Salmonella Typhimurium on the surface of eggs.(1)Antibiotic resistance of Salmonella TyphimuriumTotally,antibiotics susceptibility testing demonstrated 96.0%(290/302)S.typhimurium isolates were resistant to at least one antibiotic and 79.8%(241/302)were multidrug resistant(MDR).The resistance was highest to streptomycin(78.8%),followed by compound sulphonamides(78.2%),tetracycline(75.5%),ampicillin(71.9%),nalidixic acid(63.6%),chloramphenicol(59.9%)and trimethoprim/sulfamethoxazole(57.3%),and all isolates are sensitive to cefepime and imipenem.The antibiotic resistance rate of Salmonella Typhimurium from high to low was as follows:tetracyclines>quinolones>sulfonamides>aminoglycosides>?-lactams>carbapenems.(2)Phenotypic and genotypic disinfectant resistance of Salmonella TyphimuriumThe MIC of benzalkonium chloride(BC),triclosan(TCS),trichloroisocyanuric acid(TCCA)and povidone iodine(PVP-I)for S.Typhimurium isolates were 2-64,0.031251,32-1024 and 256->1024 mg/L,respectively.The MIC of the disinfectant against Salmonella Typhimurium from high to low were:PVP-I>TCCA>BC>TCS.The detection rates of disinfectant resistance genes were:qacE?1(56.95%,172/302),sugE(p)(20.53%,62/302),qacE(2.65%,8/302).(3)Resistance of qacE?1 and sugE(p)genes to disinfectantThe qacE?1 and sugE(p)genes were inserted into the pET30b(+)vector through these two cleavage sites.After pET30b(+)-qacE?1 and pET30b(+)-sugE(p)were transformed into E.coli BL21(DE3)and induced with different concentrations of IPTG,it was found that with the increase of IPTG induced concentration,benzalkonium chloride MIC also increased,compared with the control strain,MIC increased by 4-8 times,indicating that the qacE?1 and sugE(p)genes in this study could mediate the resistance of Salmonella to benzalkonium chloride.(4)Bactericidal effect of benzalkonium chloride to Salmonella Typhimurium on egg surfaceBenzalkonium chloride with concentrations of 0.5×MIC,1×MIC and 1.5×MIC were sprayed and disinfected on the surface of eggs inoculated with Salmonella typhimurium for 15 s,30 s and 45 s.Results showed that the treatment of 0.5×MIC/45 s,1×MIC/15 s,1×MIC/30 s,1×MIC/45 s,1.5×MIC/15 s,1.5×MIC/30 s and 1.5×MIC/45 s can reduce the number of residual colonies of Salmonella Typhimurium on the egg surface by more than 3.00 log10 CFU/mL.There was no significant difference in the sterilization logarithm between the treatment groups(p>0.05),in view of the use of sub-inhibitory concentration and high-inhibition concentration of benzalkonium chloride can promote the development and spread of bacterial resistance,combined with economic considerations,this paper considered that 1×MIC benzalkonium chloride treatment 15 s was the best combination to eliminate Salmonella Typhimurium on egg surface.