Abstract：
Bovine coronavirus (BCoV) is one of the important causes of diarrhoea in cattle. The virus is responsible for the high fatality rate associated with acute diarrhoea in calves. Rapid and accurate tests need to be conducted to detect the virus and minimise economic losses associated with the disease. Nucleic acid-based detection assays including PCR is an accurate test for detecting pathogens. However, these tests need skilled personnel, time and expensive devices. In this study, we developed a novel assay for the detection of BCoV in clinical cases. This novel assay combined reverse transcription-recombinase polymerase amplification with CRISPR/Cas13 and conducted a rapid visualisation of cleavage activity using a Lateral Flow Device. A conserved sequence of the BCV M gene was used as a target gene and the assays were tested in terms of specificity, sensitivity and time consumption. The result showed the specificity of the assay as 100% with no false positives being detected. Ten copies of the input RNA were enough to detect the virus and perform the assay. It took up to forty minutes for reading the results. Conducted together, the assay should be used as a rapid test to clinically diagnose infectious pathogens including bovine coronavirus. However, the assay needed the RNA to be extracted from the clinical sample in order to detect the virus. Therefore, more studies are needed to optimise the assay to be able to detect the virus in the clinical sample without extracting the RNA.
摘要：
牛冠状病毒（BCoV）是牛腹泻的重要原因之一。该病毒是与小牛急性腹泻相关的高致死率的原因。需要进行快速准确的检测，以检测病毒并最大程度地减少与疾病相关的经济损失。包括PCR在内的基于核酸的检测测定法是用于检测病原体的准确测试。然而,这些测试需要熟练的人员，时间和昂贵的设备。在这项研究中,我们开发了一种在临床病例中检测BCoV的新方法。这种新的测定将逆转录-重组酶聚合酶扩增与CRISPR/Cas13组合,并使用侧流装置进行切割活性的快速可视化。BCVM基因的保守序列用作靶基因，并在特异性方面测试了测定，灵敏度和时间消耗。结果显示该测定的特异性为100%,没有检测到假阳性。十个拷贝的输入RNA足以检测病毒并进行测定。阅读结果花了40分钟。一起进行，该测定法应用作临床诊断包括牛冠状病毒在内的感染性病原体的快速测试。然而,该试验需要从临床样本中提取RNA以检测病毒.因此,需要更多的研究来优化检测方法，以便能够在不提取RNA的情况下检测临床样品中的病毒。
