Lu Xiaoman, Cao Yilin, Zhang Ping, Chen Xiaoyuan, Irwin David M, Shen Yongyi
Guangdong Laboratory for Lingnan Modern Agriculture, State Key Laboratory for Animal Disease Control and Prevention, Center for Emerging and Zoonotic Diseases, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China.
Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto M5S1A8, Canada; Banting and Best Diabetes Centre, University of Toronto, Toronto M5S1A8, Canada.
Vet J. 2025 Apr 12;312:106356. doi: 10.1016/j.tvjl.2025.106356.
Feline Coronavirus (FCoV) is responsible for causing intestinal lesions and potentially fatal systemic infections in cats, with a worldwide prevalence. Currently, there is no effective vaccine available, making early diagnosis essential for treatment. Although RT-PCR detection is known for its high specificity and accuracy, the method involves complex experimental procedures and necessitates costly equipment, which restricts its broader clinical use. In this study, we utilized Recombinase Aided Amplification (RAA) technology in conjunction with a nucleic acid-colloidal gold immunochromatographic test strip to establish a visual detection method that can rapidly and accurately identify FCoV and differentiate between its two serotypes, serotype I and serotype II. Primers and probes were designed based on conserved sequences from the N gene and the S1 gene. Under isothermal conditions at 39°C, results can be obtained in just 30 minutes. This method demonstrates no cross-reactivity with other prevalent infectious viruses in felines, such as feline calicivirus (FCV), feline panleukopenia virus (FPV), and feline herpesvirus type 1 (FHV-1). In comparison to qRT-PCR, which has a detection limit of approximately 100 copies/μL, the RAA method exhibits significantly enhanced sensitivity. The detection limits for the FCoV N gene and the S1 gene serotypes I and II were determined to be 15.5 copies/μL, 1.97 copies/μL, and 1.46 copies/μL, respectively. Preliminary clinical applications have shown that the results align perfectly with those obtained from RT-PCR detection, achieving 100 % consistency. In summary, this novel detection method offers high accuracy, a brief reaction time, robust specificity, elevated sensitivity, and is straightforward and convenient to implement.
猫冠状病毒（FCoV）可导致猫出现肠道病变，并可能引发致命的全身性感染，在全球范围内均有流行。目前尚无有效的疫苗，因此早期诊断对于治疗至关重要。尽管逆转录聚合酶链反应（RT-PCR）检测以其高特异性和准确性而闻名，但该方法涉及复杂的实验程序，且需要昂贵的设备，这限制了其在临床上的广泛应用。在本研究中，我们利用重组酶辅助扩增（RAA）技术结合核酸胶体金免疫层析试纸条，建立了一种可视化检测方法，该方法能够快速、准确地鉴定FCoV，并区分其两种血清型，即I型和II型。基于N基因和S1基因的保守序列设计了引物和探针。在39°C的等温条件下，只需30分钟即可获得结果。该方法与猫的其他常见传染性病毒，如猫杯状病毒（FCV）、猫泛白细胞减少症病毒（FPV）和猫疱疹病毒1型（FHV-1）无交叉反应。与检测限约为100拷贝/μL的qRT-PCR相比，RAA方法的灵敏度显著提高。FCoV N基因以及I型和II型S1基因血清型的检测限分别确定为15.5拷贝/μL、1.97拷贝/μL和1.46拷贝/μL。初步临床应用表明，结果与RT-PCR检测结果完全一致，一致性达到100%。总之，这种新型检测方法具有高准确性、短反应时间、强特异性、高灵敏度，且操作简单方便。
