Multiplex real-time PCR assays for the detection of group C rotavirus, astrovirus, and Subgenus F adenovirus in stool specimens

Group C rotavirus (GCRV), astrovirus (AstV), and adenovirus (subgenus F AdenoV) are etiologic agents of acute nonbacterial gastroenteritis, which often represents community outbreaks. For the efficient detection of GCRV, AstV, and subgenus F AdenoV in stool specimens, a multiplex real-time PCR assay was developed to detect these three viruses simultaneously, with high sensitivity and specificity. In total, 8404 clinical specimens were collected between April 2008 and March 2011 and tested for GCRV, AstV, and subgenus F AdenoV by the multiplex real-time PCR, as well as for norovirus (NoV), sapovirus (SaV), and group A rotavirus (GARV) by non-multiplex real-time PCR. Forty-one specimens were positive for GCRV, AstV, or subgenus F AdenoV, including 15 specimens that were also positive for NoV, SaV, or GARV. Multiple viruses were detected simultaneously in 29 out of 4596 (0.63%) specimens infected with at least one virus. The association rates of AstV and subgenus F AdenoV with other viruses were significantly higher than those of NoV, SaV, GARV, or GCRV.

► Multiplex real-time PCR assays for GCRV, AstV, and AdenoV detection were designed.
► The sensitivity of our assay was higher than that of antigen-detection kits.
► Multiple viruses were detected simultaneously with our multiplex real-time PCR assay.