Search

Simple Summary: Mammalian orthoreovirus (MRV) infects many hosts, including humans and animals, in which it causes gastroenteritis and respiratory disease. In China, it has been found in domestic animals such as pigs, cattle, deer, and minks; however, MRV prevalence in small ruminants is still unknown, and a rapid, specific, quantification assay for clinical detection of animal samples is urgently needed. In this study, we established a TaqMan qRT-PCR method for MRV detection targeting the conserved L1 gene. The method was optimized, and the specificity, sensitivity, and repeatability were used to assess quality. We used this method for MRV detection in goat and sheep samples. The overall MRV prevalence was 8.2% (35/429), and the test showed a significantly higher sensitivity than RT-PCR and virus isolation. MRV clearly showed seasonal prevalence, with a high positive rate of infections in goats and sheep from September to April. Small ruminants under two months of age were susceptible to MRV infection. This study investigated the epidemiological characteristics of MRV infection and provides the first evidence of MRV infection in sheep and goats in China, broadening our knowledge of MRV hosts in this country. Mammalian orthoreovirus (MRV) infections are ubiquitous in multiple mammalian species including humans, and mainly causes gastroenteritis and respiratory disease. In this study, we developed a rapid and sensitive TaqMan qRT-PCR method for MRV detection based on the primers and probe designed within the conserved L1 gene. The qRT-PCR assay was evaluated for its sensitivity, specificity, efficiency and reproducibility. It was found that the detection sensitivity was equivalent to 10 DNA copies/μL, and the standard curves had a linear correlation of R2 = 0.998 with an amplification efficiency of 99.6%. The inter- and intra-assay coefficients of variation (CV%) were in the range of 0.29% to 2.16% and 1.60% to 3.60%, respectively. The primer sets specifically amplified their respective MRV segments and had the highest detection sensitivities of 100.25 TCID50/mL with amplification efficiencies of 99.5% (R2 = 0.999). qRT-PCR was used for MRV detection from samples of sheep, goats, and calves from four regions in China, and the overall MRV prevalence was 8.2% (35/429), whereas 17/429 (4.0%) were detected by RT-PCR and 14/429 (3.3%) by virus isolation. The qRT-PCR assay showed significantly higher sensitivity than RT-PCR and virus isolation. Results from an epidemiological survey indicated that the positive rate of MRV in rectal swabs from sheep and goats tested in Shaanxi, Jiangsu, and Xinjiang were 9/80 (11.3%), 12/93 (12.9%) and 14/128 (10.9%), respectively. In goats and sheep, MRV prevalence was obviously associated with season and age, with a high positive rate of more than 8% during September to April and approximately 13% in small ruminant animals under two months of age. This is the first instance of MRV infection in sheep and goats in China, thus broadening our knowledge of MRV hosts. Consequently, primer optimization for qRT-PCR should not only prioritize amplification efficiency and specificity, but also sensitivity. This assay will contribute to more accurate and rapid MRV monitoring by epidemiological investigation, viral load, and vaccination efficacy. [ABSTRACT FROM AUTHOR]

Porcine deltacoronavirus (PDCoV) is an emerging swine coronavirus that causes severe diarrhea to pigs of all ages, especially the suckling piglets under one-week-old. We previously isolated a highly pathogenic PDCoV strain, CZ2020, from a diarrheal piglet and have passaged it for over 100 passages. The adaptability of the CZ2020 increased gradually in vitro as the passage increased. Amino acid mutations were observed in pp1a, pp1ab, spike, envelop, and membrane proteins, and the spike protein accounts for 66.7% of all amino acid mutations. Then, the high passage strains, CZ2020-F80 and CZ2020-F100, were selected for evaluation of the pathogenicity in three-day-old piglets to examine whether these amino acid changes affected their virulence. At 2 days postchallenge (DPC), 2/5 piglets started to show typical diarrhea, and at 4 DPC, severe diarrhea was observed in the CZ2020-challenged piglets. Viral RNA could be detected at 1 DPC in rectal swabs and reached its highest at 4 DPC in the CZ2020-challenged group. CZ2020-F80- and CZ2020-F100-challenged groups have one piglet exhibiting mild diarrhea at 4 and 6 DPC, respectively. Compared with the CZ2020-challenged group, the piglets in CZ2020-F80- and F100-challenged groups had lower viral loads in rectal swabs, intestines, and other organs. No obvious histopathological lesions were observed in the intestines of CZ2020-F80- and F100-challenged piglets. Virulent PDCoV infection could also induce strong interferons and proinflammatory cytokines in vitro and in vivo. These data indicate that the strains, CZ2020-F80 and CZ2020-F100, were significantly attenuated via serial passaging in vitro and have the potential for developing attenuated vaccine candidates. [ABSTRACT FROM AUTHOR]

Bovine coronavirus (BCoV), a persistent threat to global cattle industry, has caused significant economic losses worldwide. In this study, a viral strain was isolated from the intestinal content of a diseased calve, and identified by cytopathic effects observation, indirect immunofluorescence assay and electron microscopy. Results showed that BCoV NXWZ2310 belonging to the GIIb genotype and has a three-amino-acid deletion in the serine-rich region of the N gene. Importantly, the BCoV NXWZ2310 strain exhibited strong pathogenicity, causing nasal discharge and watery diarrhea in calves for 8 and 10 days, respectively. Viral shedding was detected in nasal, throat and rectal swabs at levels reaching 10 6.228 copies/mL, 10 5.0 copies /mL and 10 6.692 copies/mL, respectively. Pathological examination showed that NXWZ2310 resulted in parenchymal lesions of the pulmonary lobe and significant intestinal lesions. Both the lungs and intestines displayed marked microscopic lesions with clear viral antigens present. BCoV NXWZ2310 strain with N-gene deletion mutations, caused severe respiratory and digestive disease in calves. Therefore, effective strategies are needed for the prevention and control of this isolate., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Li, Yuan, Mao, Cai, Xu, Li and Li.)