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Updated: Jun 29, 2025
Rapid and Specific Detection of Acinetobacter baumannii Infections Using a Recombinase Polymerase Amplification/Cas12a-based System
Published on: April 25, 2025
Agarose Hydrogel-Boosted One-Tube RPA-CRISPR/Cas12a Assay for Robust Point-of-Care Detection of Zoonotic Nematode
Lianjing Zhao1, Haolu Wang1, Xiuqin Chen1
1State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases, Key Laboratory for Zoonosis Research of the Ministry of Education, Institute of Zoonosis, and College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Abstract:
Rapid and accurate detection of the zoonotic nematode Anisakis is poised to control its epidemic. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas-associated assay shows great potential in the detection of pathogenic microorganisms. The one-tube method integrated the CRISPR system with the recombinase polymerase amplification (RPA) system to avoid the risk of aerosol pollution; however, it suffers from low sensitivity due to the incompatibility of the two systems and additional manual operations. Therefore, in the present study, the agarose hydrogel boosted one-tube RPA-CRISPR/Cas12a assay was constructed by adding the CRISPR system to the agarose hydrogel, which avoided the initially low amplification efficiency of RPA caused by the cleavage of Cas12a and achieved reaction continuity. The sensitivity was 10-fold higher than that of the one-tube RPA-CRISPR/Cas12a system. This method was used for Anisakis detection within 80 min from the sample to result, achieving point-of-care testing (POCT) through a smartphone and a portable device. This study provided a novel toolbox for POCT with significant application value in preventing Anisakis infection.
