Rapid Detection of Food-Borne Escherichia coli O157:H7 with Visual Inspection by Crossing Priming Amplification (CPA)
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Rapid Detection of Food-Borne Escherichia coli O157:H7 with Visual Inspection by Crossing Priming Amplification (CPA) Zhenbo Xu 1,2,3 Junyan Liu 1
&
Yuting Luo 1 & Thanapop Soteyome 4 & Chii-Wann Lin 5 & Xingyong Xu 1 & Yuzhu Mao 1 & Jianyu Su 1 &
Received: 1 July 2019 / Accepted: 22 September 2019 # Springer Science+Business Media, LLC, part of Springer Nature 2019
Abstract Escherichia coli O157:H7 is an important food-borne pathogen and has ability to contaminate food, such as water, meat products, and milk. Moreover, E. coli O157:H7 is the main toxin-producing serotype which can produce Shiga toxin type 1 and type 2 and cause intestinal disease. The strong pathogenicity and lethality of Escherichia coli O157:H7 pose a serious threat to human. This study aims to develop a rapid and visual detection assay of E. coli for rfbE, stx1, and stx2 genes by crossing priming amplification (CPA). The limit of detection of CPA assay for rfbE, stx1, and stx2 genes was 3.20 fg/μl, 320 fg/μl, and 320 fg/μl in genomic DNA, while that of in artificially contaminated food samples was 103 cfu/ml, 105 cfu/ml, and 105 cfu/ml, respectively, which was distinctly higher than that of PCR methods. And the specificity of CPA assay was tested by 22 different bacterial strains and except for E. coli O157:H7 ATCC43895 and other E. coli O157:H7 isolated from eggs, milk, and beef, all strains showed negative results. The visible detection assay was conducted by the addition of calcein in the reaction solutions. The CPA assay showed a successful detection of E. coli O157:H7 (Shiga toxin–producing and non-Shiga toxin–producing) within 60 min under 63 °C with high sensitivity and specificity. These results indicated that the CPA assays with calcein can provide an advanced method to achieve the rapid and visual detection of food-borne E. coli O157:H7. Keywords Escherichia coli O157:H7 . Shiga toxin . Crossing Priming Amplification (CPA) . Rapid detection
Introduction Food-borne illness is a leading concern to public food safety. According to the WHO, there are about 2.2 million people worldwide suffering from diarrheal diseases each year. Diarrheal diseases constitute more than half of the global food-borne diseases, causing 550 million people to * Zhenbo Xu [email protected] 1
School of Food Science and Engineering, Guangdong Province Key Laboratory for Green Processing of Natural Products and Product Safety, South China University of Technology, Guangzhou 510640, People’s Republic of China
2
Department of Microbial Pathogenesis, School of Dentistry, University of Maryland, Baltimore, MD 21201, USA
3
Overseas Expertise Introduction Center for Discipline Innovation of Food Nutrition and Human Health (111 Center), Guangzhou, China
4
Home Economics Technology, Rajamangala University of Technology Phra Nakhon, Bangkok, Thailand
5
Institute of Biomedical Engineering, National Taiwan University, Taipei 10617, China
suffer from illness and 230,000 deaths each year (Davies and Chaib 2015). Escherichia coli has been regarded as a compone
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