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ORIGINAL RESEARCH article

Front. Microbiol.
Sec. Virology
Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1541943

Rapid Detection of FAdV-4 by One-tube RPA-CRISPR/Cas12a Assay

Provisionally accepted
Lei Ma Lei Ma 1Xueping Wang Xueping Wang 2*Mingliang Zhang Mingliang Zhang 1Mengjie Zhu Mengjie Zhu 1*
  • 1 Anyang Institute of Technology, Anyang, China
  • 2 Tarim University, Aral, Xinjiang Uyghur Region, China

The final, formatted version of the article will be published soon.

    Fowl adenovirus serotype 4 (FAdV-4) is a highly contagious viral pathogen of global significance that affects various avian species. It primarily infects poultry and wild birds, leading to avian inclusion body hepatitis (IBH) and hepatitis-hydropericardium syndrome (HHS). The development of rapid diagnostic tools for detecting FAdV-4 is crucial for effective disease control and eradication efforts. In this study, we developed a recombinase polymerase amplification (RPA) combined with CRISPR/Cas12a assay, specifically targeting the FAdV-4 Hexon gene.RPA and CRISPR/Cas12a reagents were added to the bottom and lid of the test tube at once, allowing the detection process to occur within a single reaction tube. This approach reduced contamination. The RPA-CRISPR/Cas12a detection method can identify as few as 10 copies of the genome per reaction, demonstrating 100% sensitivity comparable to that of fluorescence PCR (qPCR). This approach exhibits high specificity for FAdV-4, with no cross-reactivity observed with other FAdV serotypes or common avian pathogens. Additionally, the agreement rate between the results of RPA-CRISPR/Cas12a and qPCR for detecting clinical samples is as high as 97.5%. Therefore, the RPA-CRISPR/Cas12a assay presents a promising alternative for the simple, sensitive, and specific identification of FAdV-4.

    Keywords: Avian adenovirus type 4, Cas12a, Recombinase polymerase amplification, detection, Sensitivity

    Received: 10 Dec 2024; Accepted: 13 Jan 2025.

    Copyright: © 2025 Ma, Wang, Zhang and Zhu. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

    * Correspondence:
    Xueping Wang, Tarim University, Aral, 843300, Xinjiang Uyghur Region, China
    Mengjie Zhu, Anyang Institute of Technology, Anyang, China

    Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.