Molecular Typing Of Staphylococcus Aureus, The Clone And Expression Of Panton-Valentine Leukocidin Gene

Purpose:To investigate the molecular epidemiological characteristics of Staphylococcus aureus associated with bloodstream infection in our hospital. To investigate the prevalence of Panton—Valentine leukocidin(pvl) gene in these Staphylococcus aureus strains.To construct the recombinant vectors of lukS-PV and lukF-PV, and express the protein in Escherichia coli.Materials and Methods:47 Staphylococcus aureus strains isolated from bloodstream in our hospital from January 2006 to December 2008 were collected. Their susceptibility to 11 antimicrobial angents was tested and the homology of their chromosomal DNA was analyzed with Rep-PCR DiversiLabTM Microbial Typing system. pvl gene was determined by PCR. For MRSA strains, the genotypes of SCCmec and pulse-field electrophoresis (PFGE) were determined and ST239 clone was screened with multiplex PCR. Multilocus sequence typing (MLST) was used to determine the sequence types of the selected strains. lukS-PV and lukF-PV were cloned and expressed by molecular biology techniques.Results:Out of the 47 Staphylococcus aureus,2 strains showed pvl gene positive. Methicillin-resistant strains accounted for 51.1%, all belonging to SCCmecⅢtype, and 22 MRSA strains were ST239.12 different patterns (A-L) were found among 47 strains with Rep-PCR DiversiLabTM Microbial Typing system. All MRSA strains clustered in the A and B subtypes. PFGE typing results showed 6 types for MRSA. LukS-PV and lukF-PV were cloned respectively. The expression vectors, lukF-PV-pET28a(+) and lukS-PV-pET28a(+), were constructed successfully. Expression of gene in BL21 cell got initial success.Conclusion:Most MRSA strains isolated from blood in our hospital belong to MRSA-ST239-SCCmecⅢclone. These MRSA strains have close genetic relationship. Epidemic outbreak of MRSA may exist in some departments. The clone and expression of lukS-PV and lukF-PV provide the precondition for establishment of new immunochemical method to detect Panton—Valentine leukocidin(PVL) from Staphylococcus aureus.