1.Genetic Characterization Of A Multidrug Resistant Plasmid In An ESBLs-producing Salmonella Typhi Isolate In China 2.Prevalence Of Polymyxin Resistance In Non-typhoidal Salmonella Isolates In China And The Exploring Of The Resistance Mechanism

Objective To describe the genomic characters of an ESBLs-producing Salmonella Typhi isolate from one typhoid fever patient.MethodsThe isolate was subjected to phenotypic antimicrobial susceptibility testing by broth dilution and ESBL confirmation tests.Conjugation experiment was applied to detect the transfer level of drug resistance and S1-PFGE was performed to reveal the size of the transferable plasmid.Polymerase chain reaction(PCR)and 2nd-generation whole-genome sequencing technology was adapted to illustrate the genotypes of ESBLs and the genome characteristics.Results The S.Typhi isolate was an ESBLs-producing strain which contained blaCTX-M-3 and blaTEM-1 genes and these genes were carried by a transferable plasmid pST1481.Theplasmid which belongs to the IncN incompatibility group was a closed circle DNA molecular with a size of～54 kb and was composed of 78 ORFs.The backbone region of the pST1481 showed more than 90%identity of nucleotide to three IncN plasmids(two of which were from E.coli and one from K.pneumoniae).Several mobile elements were found to be inserted into pST1481 together with antimicrobial resistance genes,including blaCTX-M-3,blaTEM-1 and aac(6’)-Ib-cr.Conclusion The first reported multidrug resistant S.Typhi simultaneously carried two groups ofβ-lactamases and quinolone resistance genes on one transferable plasmid.The multidrug resistantplasmid pST1481 was a chimera that acquired its multiple antimicrobial resistance determinants horizontally from different sources.It may evolve from an acestor plasimd from E.coli through the stepwise events of integration or recombination.The results indicated that antimicrobial resistance should be strengthened,and some resistance genes and elements and their transmission between different strains and species need to be monitored and studied.With the application of polymyxin in cinical practices,the resistance phonomenon is becoming more and more serious.Since the plasmid-mediated transferable resistace gene mcr-1 was reported,the polymyxin resistance was now becoming a hot spot.In order to investigate the prevalence of polymyxin resistance in non-typhoid Salmonella in China,404 strains from variant sources were selected through random sampling.Phenotypic antimicrobial susceptibility testing was applied by broth dilution and the resistant breakpoints for polymyxin B and colistin were presumed basing on MIC distribution data.The results showed that,among 404 strains,the polymyxin MICs ranged from ≤0.125μg/ml to>16μg/ml.The PMB MIC50 and MIC90 were 1μg/ml and 8μg/ml,respectively.While the MIC50 and MIC90 of PME were 2μg/ml and 8μg/ml,respectively.The distributions of MICs for different dominant serotypes including S.Typhimurium,S.Enteritidisand S.Derby,were different.The resistant breakpoints for polymyxin B and colistin were presumed as MIC=8μg/ml basing on their MIC frequency distribution.According to this criteria,the general resistance rates of PMB and colistin were 10.89%and 15.84%,respectively.The resistance rates of PMB were significantly different among different sources of Salmonella(food source 12.50%,animal source 7.16%and human 0.00%,respectively,P<0.01)as well as colistin resistance with food 8.30%,animal 27.94%and human 0.78%(P<0.01),respectively.To further study the prevalence and mechanism of polymyxin B resistance in NTS,we did research on all 5313 strains isolated from 2010 to 2015 in China.We chose PMB≥8 ug/mlstrains by broth dilution for further analysis.Antimicrobial susceptibility testing of another 16 antibiotics were tested on those trains.The most common serotypes were subjected to Xba I digestion and pulsed-fieldgelelectrophoresis(PFGE).The presence of mcr-1 gene and mutation of either pmrAB or phoPQ in polymyxin resistant Salmonella strains was detected by PCR technique.In this part,118 PMB≥8 ug/mlstrains were screened out.The rate of multi-drug resistance was 55.08%.12 Salmonella strains displayed concurrent resistance to polymyxin and third-generation cephalosporins.Among the most common serotypes,the rate of MDR in S.Typhimurium was higher than S.Enteritidis(χ2=8.729,P=0.003)with variant drug-resistant spectrum.Based on the PFGE analysis,the patterns of S.Enteritidis were relatively centralized while it showed high varianty in S.Typhimurium.Moreover,one Salmonella Typhimurium strain producing ESBLs isolated from human stool was mcr-1 gene positive and this gene was carried by a transferable plasmid.And the novel mutant loci were found in pmrAB.Although the resistance prevalence of polymyxin was low,but it is not negligible.The antimicrobial resistance surveillance of Salmonella from different resources should be strengthened.