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Journal of Integrative Agriculture  2026, Vol. 25 Issue (4): 1736-1739    DOI: 10.1016/j.jia.2025.07.030
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A rapid and visual detection method for porcine deltacoronavirus with single-copy sensitivity based on the CRISPR/Cas12a assay

Zezhao Cao1*, Zi Li1*, Kaikai Jin1, Zanheng Huang1, Ruijie Hu1, Junchao Shi1, Huabo Yu1, Gaili Wang2, Wenqi He1, Hualei Wang1, Yungang Lan1#, Haili Zhang1#

1 State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases, Key Laboratory for Zoonosis Research of the Ministry of Education, Institute of Zoonosis, College of Veterinary Medicine, Jilin University, Changchun 130062, China

2 Jilin Academy of Animal Husbandry and Veterinary Medicine, Changchun 130062, China

 Highlights 
By integrating RT-RAA with CRISPR/Cas12a, a portable, one-pot visual detection system was engineered, significantly minimizing the potential for false positives caused by aerosol contamination.
RT-RAA-CRISPR/Cas12a assay can complete the detection within 40 min, with a sensitivity as high as 3.9 copies µL–1 for single copy.
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摘要  

猪德尔塔冠状病毒(PDCoV)感染可引发仔猪严重腹泻、脱水和死亡,是一种急性、高度接触性传染病,严重威胁养猪业的可持续发展。此外,PDCoV 还具有跨物种传播潜力,带来重大公共卫生安全风险。因此,建立快速、准确的诊断方法对于有效防控该病至关重要。本研究开发了一种将逆转录重组酶辅助扩增(RT-RAA)与 CRISPR/Cas12a 系统相结合的单管封闭式可视化检测方法。该方法全程在封闭系统内进行,可有效避免开盖操作导致的气溶胶污染及假阳性风险。该方法灵敏度高,在40分钟内可检测低至 3.9 拷贝/μL的阳性质粒及低至 100.8 TCID50/0.1 mL PDCoV。此外,该方法特异性良好,与猪血凝性脑脊髓炎病毒(PHEV)、伪狂犬病病毒(PRV)、猪圆环病毒 2 型(PCV2)和猪圆环病毒 3 型(PCV3)均无交叉反应。在临床样本检测中,其检测结果与 RT-qPCR 检测结果符合率为100%。综上所述,该方法具有高灵敏度、高特异性和快速简便的特点,有望成为 PDCoV 感染早期诊断的可靠手段,为 PDCoV 疫情的有效防控提供技术支持。



Received: 30 April 2025   Accepted: 26 June 2025 Online: 29 July 2025  
Fund: 

This work was financially supported by the National Key Research and Development Program of China (2021YFF0703600).

About author:  Zezhao Cao, E-mail: 599142457@qq.com; Zi Li, E-mail: lizi1227@jlu.edu.cn; #Correspondence Haili Zhang, E-mail: zhanghaili@jlu.edu.cn; Yungang Lan, E-mail: lanyungang@jlu.edu.cn * These authors contributed equally to this study.

Cite this article: 

Zezhao Cao, Zi Li, Kaikai Jin, Zanheng Huang, Ruijie Hu, Junchao Shi, Huabo Yu, Gaili Wang, Wenqi He, Hualei Wang, Yungang Lan, Haili Zhang. 2026. A rapid and visual detection method for porcine deltacoronavirus with single-copy sensitivity based on the CRISPR/Cas12a assay. Journal of Integrative Agriculture, 25(4): 1736-1739.

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