Direct detection of Shigella flexneri and Salmonella typhimurium in human feces by real-time PCR.
We have established a SYBR Green-based realtime PCR method using AnyDirect solution, which enhances PCR from whole blood, for direct amplification of the virA gene of Shigella flexneri and the invA gene of Salmonella typhimurium from human feces without prior DNA purification. When we compared the efficiency of conventional or realtime PCR amplification of the virA and invA genes from the supernatant of boiled feces supplemented with S. flexneri and S. typhimurium in the presence or absence of AnyDirect solution, amplification products were detected only in reactions to which AnyDirect solution had been added. The detection limit of real-time PCR was 1 x 10(4) CFU/g feces for S. flexneri and 2 x 10(4) CFU/g feces for S. typhimurium this sensitivity level was comparable to other studies. Our real-time PCR assay with AnyDirect solution is simple, rapid, sensitive, and specific, and allows simultaneous detection of S. flexneri and S. typhimurium directly from fecal samples without prior DNA purification.
D017931:DNA Primers; D005243:Feces / Q000382:microbiology*; D005798:Genes, Bacterial; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012486:Salmonella typhimurium / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D012763:Shigella flexneri / Q000302:isolation & purification*
D017931:DNA Primers