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Universal Primer‐Multiplex PCR Approach for Simultaneous Detection of Escherichia coli , Listeria monocytogenes, and Salmonella spp. in Food Samples
Author(s) -
Yuan Yanfang,
Xu Wentao,
Zhai Zhifang,
Shi Hui,
Luo Yunbo,
Chen Zhuojun,
Huang Kunlun
Publication year - 2009
Publication title -
journal of food science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.772
H-Index - 150
eISSN - 1750-3841
pISSN - 0022-1147
DOI - 10.1111/j.1750-3841.2009.01321.x
Subject(s) - listeria monocytogenes , salmonella , multiplex polymerase chain reaction , primer (cosmetics) , escherichia coli , microbiology and biotechnology , biology , listeria , food microbiology , polymerase chain reaction , virology , bacteria , chemistry , gene , genetics , organic chemistry
Escherichia coli , Listeria monocytogenes , and Salmonella spp. are 3 kinds of the most important food‐borne human pathogens. Traditional microbiological analysis is labor‐intensive, time‐consuming, and easily contaminated, thus producing false positive signals; it also involves much subjectivity judgments. Multiplex‐PCR could be applied to detect multiple target organisms simultaneously to save time and labor, but there is always disproportionate amplification resulting from the disparity of different primers. To gain a rapid and sensitive method, a universal primer‐multiplex PCR system (UP‐M‐PCR) was developed and applied for simultaneous detection of the 3 organisms. This method simplified traditional multiplex‐PCR reaction system and overcame its amplification disparities among different primers; moreover, it got a high specificity and sensitivity (85, 155, and 104 copies/reaction for E. coli O157, L. monocytogenes , and Salmonella spp., respectively). Compared with the time‐consuming and laborious microbiological analysis, UP‐M‐PCR had a lower risk of cross‐contamination without inoculation and incubation. Test results for 36 food samples showed that UP‐M‐PCR method got a relative accuracy of 91.77% when compared with traditional microbiological analysis. It could serve as a rapid screening method for pathogen detection and could detect target genes even in dead pathogenic cells. In addition, it has the potential to be performed in an automation mode and might find broader application in simultaneous detection of other multiple pathogens.