Antimicrobial Resistance Of Non-O157 Shiga Toxin-producing Escherichia Coli Isolates From Different Sources

Objective:Shiga toxin-producing Escherichia coli(STEC)are important zoonotic pathogens,which can cause infectious diarrhea,hemorrhagic colitis(HC),and even fatal hemolytic uremic syndrome(HUS).The treatment of STEC infections with antibiotics is still controversial,and the knowledge about antibiotic resistance of non-O157 STEC in China is very limited.Therefore,a surveillance and molecular epidemiological study of antimicrobial resistance(AMR)in non-O157 STEC from different sources will help us to understand the current status of drug resistance of STEC strains in humans and different host animals,which will provide a basis for the selection of specific antibiotics for patients with STEC infection,and provide a reference for the elimination of STEC in animals carrying the bacteria.Methods:Two hundred and ninety-eight non-O157 STEC isolates from cattle,sheep,pig and human obtained from active surveillance in 12 provinces/municipalities in China during 2009-2019 were used in this study.Minimum inhibitory concentration(MIC)of 19 antimicrobials belonging to 14 categories were determined by broth microdilution method using the BD PhoenixTM M50 Automated Microbiology System.Whole genome sequencing(WGS)and assembling was performed using Illumina NovaSeq platform and PacBio Sequel platform.The assembled sequences were compared with the local O:H serotype and Shiga toxin subtype databases by ABRICATE to obtain their corresponding O:H serotypes and Shiga toxin subtypes.Acquired drug resistance genes were obtained by searching against the CGE ResFinder and CARD databases.Plasmids were extracted from the completed genome sequences.Plasmid incompatibility was determined using PlasmidFinder,and genomic islands(GIs)in plasmids were predicted using IslandViewer4 and visualized by SnapGene and DNAplotter.The core genomes of 298 isolates were extracted by Snippy,followed by the removal of recombination using Gubbins;SNP loci were extracted and phylogenetic trees were constructed using IQTREE to study the population structure.Results:Out of the 298 isolates tested,115 strains showed resistance to at least one antimicrobial and 85 strains showed multidrug resistance(MDR).The highest resistance rate was to tetracycline(32.6%),followed by nalidixic acid(25.2%)and chloramphenicol and azithromycin(both 18.8%).However,ceftazidime-avibactam,imipenem,meropenem and ciprofloxacin were effective against all isolates.Antimicrobial resistance patterns varied among strains from different sources.Strains from pig,sheep,humans,and cattle showed resistance rates of 100.0%,46.9%,30.3%,and 6.3%to one or more antimicrobials,respectively.The antibiotic resistance among pig-derived isolates is the most prevalent.The MDR rate was 84.0%and was significantly higher than that of the other three host sources.A MDR stx1a-STEC O21:H25 clone resistant to ampicillin,azithromycin,chloramphenicol,nalidixic acid and tetracycline was identified from sheep-derived isolates.Forty-three genes belonging to 11 antimicrobial classes were identified among these strains.The positive rate of the tetracycline resistance gene tetA was the highest(30.2%,90/298),followed by the quinolone resistance gene qnrS1(18.5%,55/298)and the macrolide resistance gene mphA(17.1%,51/298).Class A β-lactam resistance gene spectrum is the most diverse,with 18 combinations.Followed by the combination of sulfonamides-related resistance genes(7 types)and the combination of chloramphenicol-related resistance genes(6 types).The colistin-resistance genes mcr-1 and mcr-3 were only carried by strains from pigs.A new fosfomycin-resistant gene,fosA7,was detected in strains from humans,cattle,and sheep.Among 27 non-O157 STEC complete genome sequences,58 plasmids were detected,of which drug-resistant plasmids accounted for 13.8%,including three broad host range(BHR)drug-resistant plasmids.Whole genome phylogenetic analysis showed that strains from the four sources were genetically diverse and scattered throughout the phylogenetic tree;however,some strains from the same source tended to cluster closely.Conclusion:This study explored the prevalence of antimicrobial resistant STEC strains among animals and humans.These results could guide the administration of antimicrobial drugs in STEC infections.With the emerging of MDR isolates and new AMR genes,it is necessary to persistently surveillance the AMR of non-O157 STEC from different sources.