A novel multiplex PCR assay for rapid and simultaneous detection of five pathogenic bacteria: Escherichia coli O157:H7, Salmonella, Staphylococcus aureus, Listeria monocytogenes, and Vibrio parahaemolyticus.
Rapid and sensitive detection techniques for foodborne pathogens are important to the food industry. However, traditional detection methods rely on bacterial culture in combination with biochemical tests, a process that typically takes 4 to 7 days to complete. Thus, this study was conducted to address the issue of time lag inherent in traditional methods by developing a novel PCR assay for each of five foodborne pathogenic bacteria. This new system consists of a simultaneous screening method using multiplex PCR in a single reaction tube for the rapid and sensitive detection of each of the five bacteria. Specific primers for multiplex PCR amplification of the Shiga-like toxin (verotoxin type II), femA (cytoplasmic protein), toxR (transmembrane DNA binding protein), iap (invasive associative protein), and invA (invasion protein A) genes were designed to allow simultaneous detection of Escherichia coli O157:H7, Staphylococcus aureus, Vibrio parahaemolyticus, Listeria monocytogenes, and Salmonella, respectively. To confirm the specificity of each primer pair for the respective target gene, three types of experiments were carried out using boiled cell lysates and their DNAs. In the multiplex PCR with mixed DNA samples, specific bands for corresponding genes were simultaneously detected from a single reaction. The detection of all five foodborne pathogenic bacteria could be completed in less than 24 h with this novel PCR method.
Kim
Jeong Soon
JS
Samsung Everland Inc., Food Research & Development Center, 50 Mabuk-dong, Giheung-gu, Yongin 446-912, South Korea. jeongsoon.kim@samsung.com
Lee
Gang Gweon
GG
Park
Jong Seok
JS
Jung
Yong Hyun
YH
Kwak
Hyo Sun
HS
Kim
Soo Bok
SB
Nam
Young Suk
YS
Kwon
Suk-Tae
ST
D017931:DNA Primers; D019453:Escherichia coli O157 / Q000235:genetics / Q000302:isolation & purification; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D005784:Gene Amplification; D008089:Listeria monocytogenes / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012475:Salmonella / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013211:Staphylococcus aureus / Q000235:genetics / Q000302:isolation & purification; D013997:Time Factors; D014736:Vibrio parahaemolyticus / Q000235:genetics / Q000302:isolation & purification
D017931:DNA Primers