Prevalence Of Heavy Metal Resistance Genes In Escherichia Coli And Salmonella And Whole Sequencing Of E.coli E308 Showing Co-resistance

In order to investigate the prevalence of heavy metal resistance genes in Escherichia coli and Salmonella,and the correlation between the prevalence of heavy metal resistance genes and the resistance of E.coli and Salmonella to antibiotics and quaternary ammonium compounds?QACs?disinfectant.79 strains of E.coli and 132strains of Salmonella isolated from broiler chicken farms,and 99 strains of E.coli and162 strains of Salmonella isolated from meat were detected for heavy metal resistance genes.The minimal inhibitory concentrations?MICs?of heavy metals suppressing strains with the different genotypes were tested.The correlations between heavy metal resistance genes and resistance of strains to antibiotics and disinfectants were anlyzed.A strain of E.coli E308,which is resistant to heavy metals,antibiotics and disinfectants,was sequenced,and the complete genome sequence of the strain was obtained.Nine heavy metal resistance genes were detected from E.coli and Salmonella.In E.coli,the prevalence rate of the gene zntA?encoding resistance to Zn?was highest?P=77.0%?,and the prevalence rate of zntB?encoding resistance to Zn?was the lowest?P=0.6%?.The zntA-rcnA genotype was the highest in E.coli?P=11.8%?,followed by arsB-zntA-rcnA genotype?P=10.7%?,and pcoR-zntA-rcnA genotype?P=8.4%?.The prevalence rates of arsB?encoding resistance to As?and merA?encoding resistance to Hg?in E.coli from meat were significantly higher than those in E.coli from chicken farms?p<0.05?.The MICs of heavy metals suppressing E.coli ranged from?12.5mg/L to 1600 mg/L.The MICs of Hg²⁺suppressing E.coli were lowest??12.5 mg/L?and the MICs of Zn²⁺and Mn²⁺suppressing E.coli were the highest?1600 mg/L?.In Salmonella,the prevalence rate of zntB gene was the highest?P=87.1%?,while the prevalence rate of zntA was the lowest?P=0.3%?.The genotype of chrA-zntB was highest in Salmonella?P=19.4%?,followed by zntB?P=14.6%?,and pcoR-chrA-zntB?P=6.5%?.27 strains of Salmonella?P=9.2%?were not detected any heavy metal resistance genes.The prevalence rate of arsB in Salmonella from chicken farms was significantly higher than that in Salmonella from meat?p<0.05?.The MICs of heavy metals suppressing Salmonella ranged from?12.5 mg/L to 1600 mg/L.The MICs of Hg²⁺suppressing Salmonella was lowest?<12.5 mg/L?,while the MICs of Cu²⁺,Zn²⁺,and Mn²⁺suppressing Salmonella were highest?1600 mg/L?.There was a significant correlation between prevalence rate of arsB and antibiotic resistance in E.coli?p<0.05?.There was no significant correlation between prevalence rates of antibiotic resistance genes and heavy metal resistance genes.There was a significant correlation between prevalence of of arsB and zntA and the resistance to Benzalkonium chloride?BC??p<0.01?,as well as arsB and pcoR genes and Cetylpyridinium chloride?CPC??p<0.01?.ArsB,merA,pcoR?encoding resistance to Cu?,zntA,and chrA?encoding resistance to Cr?were significantly correlated with the disinfectant resistance gene?p<0.05?.In Salmonella,there was a significant correlation between the prevalence rates of pcoR and pcoC?enconding resistance to Cu?and antibiotic resistance?p<0.05?.In Salmonella,the resistance genes of heavy metals were significantly associated with?-lactams,tetracyclines,sulfonamides,quinolones and aminoglycoside antibiotic resistance genes?p<0.05?.In Salmonella,the prevalence rates of arsB and chrA and the resistance to BC were correlated?p<0.05?.There was a significant correlation between the prevalence rates of arsB,chrA and the resistance to BC and CPC resistance?p<0.01?.In Salmonella,the prevalence of qacE?1 resistance gene was significantly associated with the prevalence of arsB,pcoR,and pcoA?p<0.001?.By assembling the whole genome-sequenced sequences of E308,10 contigs were finally obtained,and the total length of assembled contigs were 5,350,144 bp.5,155genes were annotated and the total length of annotated gene was 4,484,934 bp,accounting for 86.81%of the total sequence.A circular chromosomal sequence with a length of 4,786,360 bp with a GC content of 50.76%was assembled and 4,658 genes were predicted.Sequences of four circular plasmids were assembled,named pLKSZ01with 111,721 bp,pLKSZ02 with 113,645 bp,pLKSZ03 with 61,476 bp and pLKSZ04with 92,978 bp,and the plasmid types were IncI1,IncF2,IncI2,and p0111,respectively.A large number of resistance genes of antibiotics,QACs and heavy metals,and many mobile genetic elements?MGEs?were found in chromosomal sequences,which of them belong to the multidrug resistance efflux?MDR?pump system The plasmid pLKSZ01 sequence includes insertion sequences?ISs?,transposons,conjugation regions,and genes encoding colicin 1A,colicin 1A immune protein,and colicin M immune protein.The variable region of plasmid pLKSZ02 contains 12 antibiotic resistance genes?blaCTX-M-55,oqxA,oqxB,fosA3,floR,tetA,tetR,strA,strB,sul2,aph?3''?-II and blaTEM-1?and many ISs.The plasmid pLKSZ03 contains type IV pilus?T4Ps?assembly system,a type IV secretion system?T4SSs?,and mcr-1 gene?encoding resistance to colistin?.Plasmid pLKSZ04 carries 8 antibiotic resistance genes,one quaternary ammonium disinfectant resistance gene,and one heavy metal resistance gene to Cr?blaTEM-1,dfrA12,aadA2,qacE?1,sul1,chrA,mphA,mphR,tetR,and tetA?.The results showed that the heavy metal resistance genes are widespread in E.coli and Salmonella.There are significant correlations between heavy metal resistance genes,and antibiotic resistance and QACs resistance in E.coli and Salmonella.There are a large number of resistance genes of heavy metals,disinfectants and antibiotic on chromosome of E308.A large number of antibiotic resistance genes and several resistance genes of QACs,and Cr are carried on plasmids.It suggests that co-resistance to antibiotics,QACs and heavy metal may be spread between bacterias.Therefore,it is of great significance to strengthen contral heavy metals in livestock and poultry farming environment to prevent the emergence and spread of co-resistance strains to heavy metals,antibiotics,and disinfectants.