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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]