Establishment Of A Multiplex PCR Assay For Detection Of Staphylococcus Hyicus And Proteomic Analysis Of ZLT-1Strain

Staphylococcus hyicus (S.hyicus) is an environment common bacteria, at the same time is the main pathogen of piglets exudative epidermitis(EE). In this study,6pairs of specific primers were designed and synthesized based on6sequences of exfoliative toxin genes which were reported on GenBank. A multiplex PCR assay for detected S.hyicus and its exfoliative toxin genes was established.The results of detection of9clinical isolations indicated that the23SrDNA(662bp) stripe were amplified in whole isolations and the ExhA(316bp),ExhC(525bp),ShetA(814bp) stripes were amplified in six isolations,each toxin gene were amplified from two different isolations,three others were none toxin genes amplification so that were identified avifulence strains. These results were matched with biochemical identify,nomal PCR and sequencing. This multiplex PCR assay were easy to operate,inexpensive,fast,it has the higher specificity,sensibility and better repeatablity,can be used for diagnosis of EE and fast identify of S.hyicus.The next study was performed for deeply researching on Metabolism, Growth characteristic, Role in host of S.hyicus. First,48strains of Staphylococcus were detected by the multiplex PCR assay which was established above, the result combined the clinical background and animal regression experiment so that a pathogenicity S.hyicus named ZLT-1was got. The whole bacteria and its culture supernatant protein were extracted. Then the whole protein were detected and digested in gel, separated by HPLC, finally analysed by LC-MS/MS. The results were analysed by GO,COG and pathway, indicated that919and111proteins were identified from bacteria and its culture supernatant of S.hyicus respectively,most of them partcipated in cell biological process, metabolism, adhesion and catalysis. Only exfoliative toxin, transacetylase, enolase and glyceraldehyde-3-phosphate dehydrogenase were found in33common virulence factors of Staphylococcus.11proteins in7pathways which related with disease, immunoregulation and apoptosis were analysed. Reaearching the S.hyicus proteomic is benefit for finding the potential virulence factors and its functions which provide the new scientific grounds to know the recessive impacts of S.hyicus to the host.