MOL-PCR and xMAP Technology: A Multiplex System for Fast Detection of Food- and Waterborne Viruses

Jakub Hrdy,Petra Vasickova,Michaela Nesvadbova,Jiri Novotny,Tomas Mati,Petr Kralik

doi:10.1016/j.jmoldx.2021.03.005

Abstract

Viruses are common causes of food- and waterborne diseases worldwide. Conventional identification of these agents is based on cultivation, antigen detection, electron microscopy, or real-time PCR. Because recent technological advancements in detection methods are focused on fast and robust analysis, a rapid multiplexing technology, which can detect a broad spectrum of pathogenic viruses connected to food or water contamination, was utilized. A new semiquantitative magnetic bead-based multiplex system has been designed for simultaneous detection of several targets in one reaction. The system includes adenoviruses 40/41 (AdV), rotavirus A (RVA), norovirus (NoV), hepatitis E virus (HEV), hepatitis A virus (HAV), and a target for external control of the system. To evaluate the detection system, interlaboratory ring tests were performed in four independent laboratories. Analytical specificity of the tool was tested on a cohort of pathogenic agents and biological samples with quantitative PCR as a reference method. Limit of detection (analytical sensitivity) of 5×100 (AdV, HEV, and RVA) and 5×101 (HAV and NoV) genome equivalents per reaction was reached. This robust, senstivie, and rapid multiplexing technology may be used to routinely monitor and manage viruses in food and water to prevent food and waterborne diseases.

Highlights

From the Department of Microbiology and Antimicrobial Resistance,* Veterinary Research Institute, Brno; the Department of Experimental Biology,y Faculty of Science, Masaryk University, Brno; the Department of Animal Origin Food and Gastronomic Sciences,z Faculty of Veterinary Hygiene and Ecology, University of Veterinary and Pharmaceutical Sciences, Brno; the Military Health Institute,x Military Medical Agency, Prague; and the Military Veterinary Institute,{ Hlucin, Czech Republic
The multiplex multiplex oligonucleotide ligation (MOL)-PCR system for detection of viruses connected to contaminated food and water was developed for the simultaneous detection of adenoviruses 40/41 (AdV), rotavirus A (RVA), noroviruses [genogroup I (NoV GI); genogroup II (NoV GII)], hepatitis E virus [genotype 1 (HEV-1); genotype 3 (HEV-3)], and hepatitis A virus (HAV)
The viruses used in this study for optimization and verification of the system were obtained either as cell culture products from the Collection of Animal Pathogenic Microorganisms at the Veterinary Research Institute (CAPM, Czech Republic) or as reference materials provided by Public Health England and by Paul-Ehrlich-Institut (Table 1)

Summary

Introduction

Limit of detection (analytical sensitivity) of 5 Â (AdV, HEV, and RVA) and 5 Â (HAV and NoV) genome equivalents per reaction was reached This robust, senstivie, and rapid multiplexing technology may be used to routinely monitor and manage viruses in food and water to prevent food and waterborne diseases. In the case of viruses, a more serious impact on public health is seen.6e8 Enteric and hepatic viruses can be transmitted through contaminated food and water, and cause disease outbreaks. The potential for these diseases to spread in today’s globalized world is measureless.[9,10]. Supported by the Ministry of Health of the Czech Republic grant NV1731921A (P.V.); the Ministry of the Interior of the Czech Republic grant VI20152020044 (P.K.)

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Conclusion