Coronaviruses (Co. Vs) are critical human and animal pathogens because of their potential to cause severe epidemics of respiratory or enteric diseases. In pigs, the newly emerged porcine deltacoronavirus (PDCoV) and re-emerged porcine epidemic diarrhea virus (PEDV) reported in the US and Asia, as well as the discovery of novel Co. Vs in wild bats or birds, has necessitated development of improved detection and control measures for these Co. Vs. Because the previous pancoronavirus (panCoV) RT-PCR established in our laboratory in 2007–2011 did not detect deltacoronaviruses (δ-Co. Vs) in swine fecal and serum samples, our goal was to develop a new panCoV RT-PCR assay to detect known human and animal Co. Vs, including δ-Co. Vs. In this study, we designed a new primer set to amplify a 668 bp-region within the RNA-dependent RNA polymerase (RdRP) gene that encodes the most conserved protein domain of α-, β-, γ-, and δ-Co. Vs. We established a one-step panCoV RT-PCR assay and standardized the assay conditions. The newly established panCoV RT-PCR assay was demonstrated to have a high sensitivity and specificity. Using a panel of 60 swine biological samples (feces, intestinal contents, and sera) characterized by PEDV, PDCoV and transmissible gastroenteritis virus-specific RT-PCR assays, we demonstrated that sensitivity and specificity of the newly established panCoV RT-PCR assay were 100%. 400 avian fecal (RNA) samples were further tested simultaneously for CoV by the new panCoV RT-PCR and a one-step RT-PCR assay with the δ-CoV nucleocapsid-specific universal primers. Four of 400 avian samples were positive for CoV, three of which were positive for δ-CoV by the conventional RT-PCR. Pan. CoV RT-PCR fragments for 3 of the 4 Co. Vs were sequenced. Phylogenetic analysis revealed the presence of one γ-CoV and two δ-CoV in the sequenced samples. The newly designed panCoV RT-PCR assay should be useful for the detection of currently known Co. Vs in animal biological samples.
year ⏰ 2018
issn 🗄 18790984 01660934
volume 256
number
page 116-122
citedbycount 5
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