Comparison of PCR-RFLP and PFGE for determining the clonality of enterohemorrhagic Escherichia coli strains.
We report here on a comparative evaluation of PCR-restriction fragment length polymorphism (PCR-RFLP) and pulsed-field gel electrophoresis (PFGE) assays, and ascertain the clonal relationship between 13 enterohemorrhagic Escherichia coli O157 : H7 strains isolated from fecal samples collected from three cows over a period of 2 months. PCR-RFLP analysis was carried out with either BglI or EcoRV digested LA-PCR amplicons, generated by targeting region V of the Stx-phage. While PCR-RFLP analysis placed these 13 strains into a single clonal type, pulsotyping analysis, as reported earlier, grouped these strains into four different PFGE subtypes of which three were closely related, while the other appeared to be different. The comparative analysis was extended further using two clonally different wild-type (3-0 and Sakai 215) strains and 17 derivative strains which had passed through an animal's gastrointestinal tract. The PCR-RFLP assay, which was not only able to differentiate the wild-type strains, but also placed the passaged derivative strains into their respective parental group, although PFGE patterns of the same set of strains resulted from different PFGE subtypes. These data indicate that PCR-RFLP is the more reliable and useful assay for a molecular epidemiological survey of enterohemorrhagic E. coli strains.
Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, Japan.
Kensuke
K
Shima
Noriyo
N
Yoshii
Masato
M
Akiba
Kazuhiko
K
Nishimura
Muneo
M
Nakazawa
Shinji
S
Yamasaki
D000818:Animals; D002417:Cattle; D002418:Cattle Diseases / Q000453:epidemiology* / Q000382:microbiology; D016521:Electrophoresis, Gel, Pulsed-Field / Q000379:methods*; D004927:Escherichia coli Infections / Q000453:epidemiology / Q000662:veterinary* / Q000821:virology; D019453:Escherichia coli O157 / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D005243:Feces / Q000382:microbiology; D017720:Molecular Epidemiology / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length