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<p>The Chinese rice-field eel rhabdovirus (CrERV) is an emerging pathogen that causes hemorrhagic disease in Chinese rice-field eels (<i>Monopterus albus</i>), leading to epidemic outbreaks, mass mortality, and considerable economic losses in aquaculture. Thus, the development of rapid and reliable diagnostic tools for on-site detection is urgently needed to address this issue. In this study, we established an RPA-CRISPR/Cas12a-based assay for CrERV detection, which exhibited superior sensitivity, specificity, and stability. The assay achieved a detection limit of 10¹ copies/µL. Specificity testing confirmed the absence of cross-reactivity with five other major aquatic viruses, including Grass carp reovirus (GCRV-II), Spring viraemia of carp virus (SVCV), Largemouth bass virus (LMBV), Cyprinid herpesvirus 2 (CyHV-2), and White spot syndrome virus (WSSV). Reproducibility analysis showed intra- and inter-assay coefficients of variation below 10%. Analysis of the 26 clinical samples showed that the RPA‑CRISPR/Cas12a assay achieved a higher positivity rate (23.08%, 6/26) compared to qRT‑PCR (15.38%, 4/26), providing preliminary evidence for its diagnostic potential in detecting CrERV. Collectively, these findings indicate that the RPA-CRISPR/Cas12a platform is a highly sensitive, specific, and user-friendly tool for rapid CrERV surveillance in aquaculture settings.</p>

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Establishment and application of a detection method for chinese rice-field eels rhabdovirus (CrERV) using the RPA-CRISPR/Cas12a System

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年5月9日

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  • Na Su,
  • Chen Xu,
  • Wenzhi Liu,
  • Mingyang Xue,
  • Zhenyu Huang,
  • Nan Jiang,
  • Yan Meng,
  • Yong Zhou,
  • Yuding Fan,
  • Ya Zheng

摘要

The Chinese rice-field eel rhabdovirus (CrERV) is an emerging pathogen that causes hemorrhagic disease in Chinese rice-field eels (Monopterus albus), leading to epidemic outbreaks, mass mortality, and considerable economic losses in aquaculture. Thus, the development of rapid and reliable diagnostic tools for on-site detection is urgently needed to address this issue. In this study, we established an RPA-CRISPR/Cas12a-based assay for CrERV detection, which exhibited superior sensitivity, specificity, and stability. The assay achieved a detection limit of 10¹ copies/µL. Specificity testing confirmed the absence of cross-reactivity with five other major aquatic viruses, including Grass carp reovirus (GCRV-II), Spring viraemia of carp virus (SVCV), Largemouth bass virus (LMBV), Cyprinid herpesvirus 2 (CyHV-2), and White spot syndrome virus (WSSV). Reproducibility analysis showed intra- and inter-assay coefficients of variation below 10%. Analysis of the 26 clinical samples showed that the RPA‑CRISPR/Cas12a assay achieved a higher positivity rate (23.08%, 6/26) compared to qRT‑PCR (15.38%, 4/26), providing preliminary evidence for its diagnostic potential in detecting CrERV. Collectively, these findings indicate that the RPA-CRISPR/Cas12a platform is a highly sensitive, specific, and user-friendly tool for rapid CrERV surveillance in aquaculture settings.