PAKISTAN VETERINARY JOURNAL

Pak Vet J, 2026, 46(9) 2325-2327

One-Pot RPA-CRISPR/Cas12a assay for Rapid Visual Detection of Giardia duodenalis in Cattle and Goats

Xingang Yu^1*, Qiaoyu Li¹, Sifan Wang¹, Ding Dan², Xuanru Mu¹, Yilong Li¹, Ning Li³, Guanxi Lin¹, Xiaoying Peng¹ and Yang Hong^4*

¹School of Animal Science and Technology, Foshan University, Foshan 528231, China; ²Agro-Tech Extension Center of Guangdong Province, Guangzhou 510500, China; ³Macau University of Science and Technology, Faculty of Chinese Medicine Taipa, Macao 999078, PR China; ⁴National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute of Parasitic Diseases, Chinese Center for Diseases Control and Prevention (Chinese Center for Tropical Diseases Research), Key Laboratory of Parasite and Vector Biology, National Health Commission of the People's Republic of China (NHC), World Health Organization (WHO) Collaborating Center for Tropical Diseases, National Center for International Research on Tropical Diseases, Shanghai 200025, China.

*Corresponding author: yuxingang4525@163.com (XY); hongyang_7@126.com (YH)

Abstract

Giardia duodenalis is a widespread zoonotic protozoan causing giardiasis in humans and livestock. A one-pot RPA-CRISPR/Cas12a assay was developed for rapid, visual detection of G. duodenalis for the detection of a specific genomic segment (GenBank: CP110919.1). For this crRNA and RPA primers were designed and optimized. The assay completed the detection within 30min at 37°C in a single tube, which demonstrated high specificity with no cross-reactivity against six non-target pathogens. The limit of detection was 1 copy/μL for recombinant plasmids and 1 cyst per 10mL for cysts, surpassing nested PCR (10³ cysts/10mL). In 30 clinical fecal samples, the assay showed 100% concordance with nested PCR. This one-pot system eliminates aerosol contamination risk and provides intuitive visual readouts, offering a field-applicable tool for giardiasis screening in cattle and goats.

To Cite This Article: Yu X, Li Q, Wang S, Dan D, Mu X, Li Y, Li N, Lin G, Peng X and Hong Y, 2026. One-pot RPA-CRISPR/Cas12a assay for rapid visual detection of Giardia duodenalis in cattle and goats. Pak Vet J, 46(9): 2325-2327. http://dx.doi.org/10.29261/pakvetj/2026.214

Abstract

ISSN 0253-8318 (Print)ISSN 2074-7764 (Online)

ISSN 0253-8318 (Print)
ISSN 2074-7764 (Online)