In this study, a SYBR Green I-based real-time reverse transcription-polymerase chain reaction (RT-PCR) was developed for the clinical diagnosis of feline astroviruses (FeAstVs). Specific primers were designed based on the conserved region of the FeAstV ORF1b gene. Experiments for specificity, sensitivity, and repeatability of the assay were carried out. In addition, the assay was evaluated using clinical samples. Specificity analysis indicated that the assay showed negative results with samples of Feline Parvovirus, Feline Herpesvirus, Feline Calicivirus, Feline Bocavirus, and Feline Coronavirus, indicating good specificity of the assay. Sensitivity analysis showed that the SYBR Green I-based real-time RT-PCR method could detect as low as 3.72 × 10¹ copies/μL of template, which is 100-fold more sensitive compared to the conventional RT-PCR. Both intra-assay and inter-assay variability were lower than 1 %, indicating good reproducibility. Furthermore, an analysis of 150 fecal samples showed that the positive detection rate of SYBR Green I-based real-time RT-PCR was higher than that of the conventional RT-PCR, indicating the high reliability of the method. The assay is cheap and effective. Therefore, it could provide support for the detection of FeAstV in large-scale clinical testing and epidemiological investigation.

在这项研究中，基于SYBR Green I的实时逆转录聚合酶链反应（RT-PCR）被开发用于猫星状病毒（FeAstVs）的临床诊断。根据FeAstV ORF1b基因的保守区域设计特异性引物。进行了测定的特异性，敏感性和可重复性的实验。另外，使用临床样品评估该测定。特异性分析表明，对于猫细小病毒，猫疱疹病毒，猫杯状病毒，猫Bocavirus和猫冠状病毒样品，该试验显示阴性结果，表明该试验具有良好的特异性。敏感性分析表明，基于SYBR Green I的实时RT-PCR方法可检测到低至3.72×10 ¹模板拷贝数/μL，比常规RT-PCR灵敏度高100倍。批内和批间变异性均低于1％，表明重现性良好。此外，对150个粪便样品进行的分析表明，基于SYBR Green I的实时RT-PCR的阳性检出率高于常规RT-PCR，表明该方法具有很高的可靠性。该测定便宜且有效。因此，可以为大规模临床检测和流行病学调查中FeAstV的检测提供支持。