Detection of Escherichia coli O157: H7 vt and rfb(O157) by multiplex polymerase chain reaction.
A rapid method for detection of Escherichia coli O157: H7 using multiplex PCR was developed. Two oligonucleotide primer pairs were used for simultaneously detection of vt encoding verotoxin genes for virulence factor and rfb(O157) encoding the O-antigen specific for E. coli O157: H7. Multiplex PCR generated two products of 215 bp and 420 bp for vt and rfb(O157), respectively. Multiplex PCR detected reference strain O157: H7 (NF-7777) with a sensitivity of 10(5) CFU per ml with no amplification of other 15 pathogenic bacteria. After incubation of 10(2) CFU/25 gram raw meat in tryptic soy broth at 37 degrees C for 8 hours, multiplex PCR conducted with the addition of 100 mg bovine serum albumin produced the two specific PCR products for E. coli O157: H7. This modified multiplex PCR is a rapid, sensitive, and specific technique for detecting and differentiating E. coli O157: H7 and has the potential to be used as an alternative to conventional methods for the screening of O157: H7 strains isolated from raw meat.
Visetsripong
Apirak
A
Department of Microbiology, Faculty of Science, Chulalongkorn University, Bangkok, Thailand.
Pattaragulwanit
Kobchai
K
Thaniyavarn
Jiraporn
J
Matsuura
Ryosuke
R
Kuroda
Akio
A
Sutheinkul
Orasa
O
D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics* / Q000302:isolation & purification; D017931:DNA Primers; D019453:Escherichia coli O157 / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D005516:Food Microbiology; D008460:Meat / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013785:Thailand; D037521:Virulence Factors
D001427:Bacterial Toxins; D017931:DNA Primers; D037521:Virulence Factors