Development of a Luminex Assay for the Simultaneous Detection of Human Enteric Viruses in Sewage and River Water

Real time PCR (qPCR) is increasingly being used for viral detection in aquatic environments because it enables high specificity and sensitivity of detection. However, the limited number of fluorescent reporter dyes restricts its multiplex application. In this study, a multiplex Luminex assay was established for the simultaneous detection of human adenovirus (HAdV), human polyomavirus (HPyV), enterovirus (EV), rotavirus (RoV), norovirus GI (NoVGI) and norovirus GII (NoVGII). Different river water and wastewater samples were tested for the viruses using both qPCR and the multiplex Luminex xMAP assay. HAdV and HPyV were the most abundant in all environmental samples. HAdV was detected in all river water and wastewater samples, and HPyV was detected in 79% of river water and 95.8% of wastewater samples. The multiplex xMAP assay revealed high specificity and no cross-reactivity. Using the multiplex Luminex assay, the viral detection rates in river water samples were lower than the rates obtained by qPCR for all viruses. Conversely, in wastewater samples, the viral detection rates were the same for both methods. In addition, the analytical sensitivity of the monoplex Luminex assay was comparable to or lower than qPCR. Results suggest that the multiplex Luminex assay could be a reliable method for the simultaneous detection of viral pathogens in wastewater.