Rapid Detection of Diarrheagenic Escherichia coli by a New Multiplex Real-Time Quantitative PCR Assay
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apid Detection of Diarrheagenic Escherichia coli by a New Multiplex Real-Time Quantitative PCR Assay J. Suna, b, Y. Shic, Y. Dub, Z. Wangd, Z. Liub, H. Wangb, G. Zhaoe, *, Y. Mab, **, and M. Zhengb, *** a
School of Life Sciences, University of Science and Technology of China, Hefei, Anhui, 230026 China Institute of Biomedical Engineering and Technology, Chinese Academy of Sciences, Suzhou, Jiangsu, 215163 China c The First Affiliated Hospital of Jilin University, Changchun, Jilin, 130021 China dSchool of Medical Technology, Xuzhou Medical University, Xuzhou, Jiangsu, 221004 China e Zhejiang University Kunshan Biotechnology Laboratory, Zhejiang University Kunshan Innovation Institute, Kunshan, Jiangsu, 215300 China *e-mail: [email protected] **e-mail: [email protected] ***e-mail: [email protected]
bSuzhou
Received January 1, 2020; revised March 3, 2020; accepted July 2, 2020
Abstract—Diarrheagenic Escherichia coli (DEC) is a group of important foodborne pathogens that can spread and cause infection in humans. As a significant public health issue, DEC infection may lead to a series of diseases such as abdominal pain, diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Therefore, its rapid and accurate detection has become an important concern for food safety issues and healthcare-associated infections. We have developed a multiplex real-time quantitative PCR assay (qPCR), DEC Sensor Test, to simultaneously recognize 5 major DEC pathotypes in a single tube, enterotoxigenic, enterohemorrhagic, enteropathogenic, enteroaggregative and enteroinvasive E. coli. In this assay, 13 DEC virulence genes, stp, sth, lt, stx1, stx2, eae, escV, bfpB, aggR, astA, pic, invE, and ipaH, can be simultaneously detected along with a spiked-in internal amplification control to avoid false negative results. All the constituent PCR for the 13 DEC virulence genes had high analytical sensitivity and specificity. Detection of all virulence genes by DEC Sensor Test can be much easier, cheaper and faster to perform than conventional tests. It also has added advantages of being able to distinguish live or dead bacteria as well as eliminating the possibility of switching partial results between different samples associated with multi-tube tests. Therefore, the application of this assay may significantly enhance our ability to prevent and fight DEC-associated diseases and to improve public health. Keywords: diarrheagenic Escherichia coli, multiplex real-time quantitative PCR, virulence gene, public health, food safety DOI: 10.1134/S0003683820060174
Diarrheagenic Escherichia coli (DEC) is a group of foodborne pathogens that can cause gastrointestinal diseases [1] and bring high disease burden to society [2]. Worldwide, there are nearly 1.7 billion cases of diarrheal diseases every year, and about 760000 children under 5 years old die from these diseases [3]. Among these, DEC is responsible for up to 30–40% of acute diarrheal episodes in children [4]. Foodborne diseases caused by DEC also pose a major economic risk, especially
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