Rapid and simultaneous quantitation of Escherichia coli O157:H7 Salmonella, and Shigella in ground beef by multiplex real-time PCR and immunomagnetic separation
作者： LUXIN WANG , YONG LI , AZLIN MUSTAPHA
DOI: 10.4315/0362-028X-70.6.1366
关键词: Shigella 、 Immunomagnetic separation 、 Multiplex polymerase chain reaction 、 Microbiology 、 TaqMan 、 Multiplex 、 Enterobacteriaceae 、 Biology 、 Salmonella 、 Escherichia coli
摘要: The objective of this study was to establish a multiplex real-time PCR for the simultaneous quantitation Escherichia coli O157:H7, Salmonella, and Shigella. Genomic DNA extracted by boiling method. Three sets primers corresponding TaqMan probes were designed target these three pathogenic bacteria. Multiplex performed with Universal Master Mix in an ABI Prism 7700 Sequence Detection System. Final standard curves calculated each pathogen plotting threshold cycle value against bacterial number (log CFU per milliliter) via linear regression. With optimized conditions, quantitative detection range pure cultures 10(2) 10(9) CFU/ml E. 10(3) 10(1) 10(8) When established system applied artificially contaminated ground beef, limit 10(5) CFU/g 10(4) Immunomagnetic separation (IMS) further used separate O157:H7 Salmonella from beef samples. additional use IMS, both pathogens. Results showed that PCR, combined is potentially effective method rapid reliable 0157:H7, Shigella food.
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