Rapid Molecular Detection of Escherichia coli O157:H7 / 감염

ABSTRACT

BACKGROUND: Sorbitol fermenting Escherichia coli O157 were reported. And E. coli O157H7 produce various Shiga toxin (Stx) such as Stx1, Stx2, or variants of Stx2. In this study, we tried to establish laboratory methods that detect E. coli O157H7 quickly and precisely by analyzing sensitivity of colony hybridization test and PCR technique. METHODS: Stx1-producing E. coli ATCC 43890, Stx2-producing E. coli ATCC 43889, and Stx2vha- producing E. coli ATCC 51435 were tested. Three strains of E. coli were diluted with 0.1 g of diarrheal stools from 107 CFU to 101 CFU respectively. The stool samples were incubated overnight in MacConkey agar plates. A mean of 63 colonies were hybridized by stx1- and stx2-specific oligonucleotide probes. PCR for stx1 gene and stx2 gene was done after overnight- incubation of stool samples in the LB broth with vancomycin (6 ug/mL). Positive colonies by colony hybridization were confirmed by PCR for stx1 gene and stx2 gene. RESULTS: Colony hybridization test could detect Stx1-producing E. coli at 103 CFU per 0.1 g of stool, Stx2-producing E. coli at 105 CFU per 0.1 g of stool, and Stx2vha-producing E. coli at 104 CFU per 0.1 g of stool. PCR technique after enrichment in LB broth with vancomycin (6 ug/mL) could detect stx1-, stx2-, and stx2vha-containing E. coli at 10 CFU per 0.1 g of stool respectively. CONCLUSOIN A combination of colony hybridization and PCR after enrichment in broth with vancomycin (6 ug/mL) is useful for the rapid and precise diagnosis of infections of Shiga toxin-producing E. coli O157H7.