Development and evaluation of a one-step multiplex real-time TaqMan ® RT-qPCR assay for the detection and genotyping of equine G3 and G14 rotaviruses in fecal samples

摘要: Equine rotavirus A (ERVA) is the leading cause of diarrhea in neonatal foals and has a negative impact on equine breeding enterprises worldwide. Among ERVA strains infecting foals, genotypes G3P[12] G14P[12] are most prevalent, while infections by with other genomic arrangements infrequent. The identification circulating critical for diagnostic surveillance purposes, as well to understand their molecular epidemiology. Current genotyping methods available rotaviruses affecting animal species rely Sanger sequencing significantly time-consuming, costly labor intensive. Here, we developed first one-step multiplex TaqMan® real-time reverse transcription polymerase chain reaction (RT-qPCR) assay targeting NSP3 VP7 genes G3 G14 rapid detection G-typing directly from fecal specimens. RT-qPCR was designed. analytical sensitivity assessed using serial dilutions vitro transcribed RNA containing target sequences specificity determined DNA derived panel group along viruses bacteria. clinical performance this evaluated 177 samples compared VP7-specific standard RT-PCR sequencing. Limits (LOD), sensitivity, specificity, agreement were determined. assays demonstrated high efficiency, perfect linearity. 100-fold difference observed when singleplex assays; however, did not have an performance. Clinical that had sensitivity/specificity every (100% NSP3, > 90% and > 99% VP7, respectively) overall (> 98%) conventional This new constitutes useful, very reliable tool could aid field.