A-293 Assessment of the Detection of Gastrointestinal Viruses Using Two Commercial Real-Time PCR Assays
Authors: Sánchez-Sánchez J , Fuentes C , Arrebola A , Costafreda M , Bosch A , Pintó R M , Guix S 
Published: September 2023

Abstract

Background
Acute Gastroenteritis (AGE) are the second most common infectious diseases, only overtaken by Respiratory Tract Infections. Sign and symptoms of AGE illness include nausea, diarrhoea, vomits, fever and abdominal pain. In infants and children, gastroenteritis is one of the most important causes of morbidity and mortality all around the Globe. Since their identification in the 1970s, enteric viruses are considered a leading cause of gastroenteritis worldwide. Among these, rotaviruses (RV) are considered to be the main cause of AGE in young children, whereas noroviruses (NoV) affect people of all ages. In addition, sapoviruses (SaV), human astroviruses (hAstV), human adenoviruses (AdV) are also relevant.
In the recent years, molecular methods, such as real-time PCR, have been widely used for detection of viral genome of enteric viruses due to their high sensitivity and specificity, becoming the gold standard. The aim of this study is the comparison of different commercial real-time PCR assays for the detection of human enteric viruses.

Material and methods
Three hundred and eighteen clinical stool samples were selected from different groups collected between 2016 and 2019, including specimens from symptomatic adults and symptomatic and asymptomatic children. Samples were stored at −20°C. The collection included a minimum of 25 samples positive for RV, 55 positives for AdV, 16 positives for AstV, 22 positives for SaV and 65 positives for NoV (31 for GI and 34 for GII). All specimens were extracted using the Maxwell®RSC Blood DNA Kit on the Maxwell RSC equipment (Promega). Samples were analysed using the following assays: RIDA®GENE Viral Stool Panel II, RIDA®GENE Sapovirus and RIDA®GENE Norovirus I & II (R-Biopharm), Vitassay qPCR kits for the detection of Rotavirus, Astrovirus, Norovirus GI, Norovirus GII, and Sapovirus. Results obtained with Vitassay were compared to R-Biopharm assays. Sensitivity and specificity were calculated using the software MetaDisc 1.4. Discrepant samples were tested in triplicate.

Results
After assessing the discrepant results, 29/114 samples were positive for RV in both RIDA®GENE and Vitassay kit. This latter reported 2 false negative (FN) and 4 false positive (FP) results when compared to RIDA®GENE. Sensitivity was 93.5% (78.6–99.2) and specificity was 95.2% (88.1–98.7). For AstV, 27/138 were positive in both kits. Vitassay qPCR reported 1 FN and 2 FP, obtaining sensitivity and specificity values of 96.4% (81.7–99.9) and 98.6% (94.9–99.8), respectively. Regarding SaV, Vitassay reported 100% (88.8%-100) of sensitivity and 97.8% (88.5–99.9) of specificity, as 1 FP was obtained. 30/89 samples tested for Norovirus GI were detected by Vitassay qPCR Norovirus GI and 25/89 RIDA®GENE Norovirus I & II (R-Biopharm). On the other hand, Vitassay qPCR Norovirus GII reported 31/81 positive samples for NoV I, whereas RIDA®GENE obtained 30 positive results. In both cases, sensitivity was of 100%, whereas specificity was over 92%.

Conclusions
Vitassay real-time PCR assays for the detection of different enteric viruses present high specificity and sensitivity, which is essential for a proper diagnosis and epidemiology assessment. In addition, Vitassay qPCR kit presents the advantage of being a ready-to-use lyophilised kit.