| Pig associated methicillin-resistant Staphylococcus aureus(MRSA)has been considered a major concern for breeding industry,food safety and public health.However,reports of systematic assessment for MRSA in swine production in China remain limited,as are reports of the phylogenetics of epidemic clone.In this study,a total of 1485 samples from three swine farms,one slaughterhouse and one indoor market in Xiamen in 2015 were collected and screened for the presence of MRSA.Antibiotic susceptibilities of all isolates for 21 antimicrobial agents were determined by the disc agar diffusion method.All the MRSA isolates were screened for 38 antibiotic resistance genes,37 toxin genes and mobile genetic element(Tn558 and lsa(E)cluster).All the MRSA isolates were characterised by staphylococcal cassette chromosome mec(SCCmec),spa typing,multilocus Sequence Typing(MLST)and pulsed field gel electrophoresis(PFGE).At last,the phylogenetics and the genetic features of the epidemic clone were determined by whole genome sequences analysis.In this study,a total of 54 samples(3.6%,54/1485 samples)were positive for MRSA isolates.The carriage rate of MRSA differed among the different locations: swine farms(2.4%,13/542 samples),a slaughterhouse(4.0%,31/780 samples),and a market(6.1%,10/163 samples).Susceptibility assessment revealed that the percentage of MRSA isolates resistant to at least six classes of antibiotics was highest in samples of the swine farms(100.0%,13/13),followed by the slaughterhouse(80.6%,25/31)and the market(40.0%,4/10).The prevalence of lsa(E)gene cluster and Tn558 is much higher in MRSA isolates from the targeted swine farms(92.3%,12/13)and the desig-nated slaughterhouse(80.6%,25/31),than those from the market(10.0%,1/10).Genes coding for virulence factors(hla?hlb?seg?sei?sem?ebpS?fnbA and cap5)were more prevalent in MRSA isolates from the swine farms(84.6%,84.6%,84.6%,100.0%,100.0%,100.0%,100.0%,100.0%)than those from the slaughterhouse(77.4%,74.2%,64.5%,67.7%,90.3%,93.5%,90.3%,87.1%)and the market(70.0%,20.0%,0.0%,0.0%,70.0%,10.0%,90.0%,60.0%).The percentage of isolates that coagulated both bovine and caprine plasma was higher in MRSA isolates from the swine farms(100.0%,13/13)than those from the slaughterhouse(90.3%,28/31)and the market(20.0%,2/10).The results of molecular typing revealed that novel ST9-MRSA-SCCmec? was considerably prevalent among swine farm(84.6%,11/13 of MRSA isolates)and slaughterhouse(80.6%,25/31),but absent in the market isolates.Moreover,ST9-MRSA-t899-SCCmec? /PFGE J and its variants were detected in fourteen isolates that came from the swine farms and slaughterhouse,indicating the transmission of novel SCCmec? MRSA along the production-processing chain.We obtained whole-genome sequences of two ST9-MRSA-SCCmec? isolates from nasal swabs associated with live pigs in China,and compared them with 135 previously sequenced genomes of 78 human-associated,39 bovine and 18 porcine Staphylococcus aureus consisting of 11 MRSA of SCCmec?,62 MRSA of other SCCmec types and 62 MSSA.The distribution of diverse mobile genetic elements(MGEs),resistance genes and virulence determinants was investigated in relation to isolate phylogeny.Phylogenetic analysis revealed that both of our porcine ST9-MRSA-SCCmec? isolates clustered with porcine,bovine and human-associated ST9-MRSA-SCCmec?.Typically all of the available ST9-MRSA-SCCmec? isolates from porcine,human and bovine samples carried SCCmec??SaPIbov4?Tn552 and lsa(E),which were seldom found in other genome sequences.Both of our porcine ST9-MRSA-SCCmec? isolates possessed a novel type V pathogenicity island ?Sa? carrying von Willebrand binding protein gene vwb,immune evasion complex gene scn,aminoglycoside resistance gene aadE,staphylococcal superantigen-like genes(ssl1 – ssl11)and lpl tandem genes.Compared to bovine ST9-MRSA-SCCmec? BA01611,our porcine isolates contain non-synonymous nucleotide substitutions in genes encoding adhesins and insertion/deletions(indels)located in a phosphonate ABC transporter pseudogene.In summary,the novel ST9-MRSA-SCCmec? was considerably prevalent among isolates from swine farms and slaughterhouse.Notably,all ST9-MRSA-SCCmec? MRSA isolates were resistant to at least 6 classes of antibiotics,carried two mobile genetic elements(lsa(E)-containing multiresistance gene cluster and Tn558)and harbored multiple virulence genes.These multidrug resistant MRSA isolates could also coagulate both bovine and caprine plasma.The extraordinary success of the ST9-MRSA-SCCmec? group in colonization of various hosts is likely due to acquisition of many MGEs harboring functional antimicrobial resistance and virulence genes,including a novel type V pathogenicity island ?Sa?.Transmission of ST9-MRSA-SCCmec? between porcine and bovine hosts was accompanied by changes in binding profile and function in genes involved in metabolism. |
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