Detection of Some SalmonellaEnteritidisVirulence Genes by Multiplex-PCR Assay Using Two Different DNA Extraction Methods
Keywords: Salmonella virulence genes, multiplex-PCR, Boiling method
AbstractThe aim of this study was screeningsome Salmonella genus-specific virulence genes by multiplex-PCR technique using a group of primers targeting InvA,SipB,SpiA,CdtB,PefA genes using two different DNA extraction methods. A total of fifteen Salmonella isolates from patient's stool samples were collected. Suspected colonies onHiChromeRajhans (HCR) Medium and XLD medium were selected, and biochemical and serological tests were then performed for identification of Salmonella. Identification of Salmonella serotypes was done in Central Public Health Laboratory (CPHL) with O and H antisera. Extraction of bacterial genome was down by boiling method and salting out method using commercial kit (WizardÂ® Genomic DNA purification Kit). The results showed that all screened genes (chromosomally and plasmid-mediated) were found in all tested SalmonellaEnteritidis strains except cdtB gene which is thought to be limited only to certain SalmonellaTyphi andS.Paratyphi A strains. Moreover,the presence of pefA gene could be depending on host-adapted serovars. Boiling extraction method and commercial kit both gave a good result in multiplex-PCR technique. In conclusion, the results from this study of occurrence of SPIs genes support previous studies suggesting that these virulence genes are widely distributed among Salmonella and required for full Salmonella virulence, in addition, the boiling method was good, easy and specific method for multiplex-PCR technique.