Quick detection of Carassius auratus herpesvirus (CaHV) by recombinase-aid amplification lateral flow dipstick (RAA-LFD) method

Abstract

Crucian carp (Carassius auratus) is one of the major freshwater species and is also a common food fish in China. Recently, Carassius auratus herpesvirus (CaHV) could induce fatal viral disease with high mortality of crucian carp, which had caused huge economic losses. In this study, we described a rapid and simple recombinase-aid amplification (RAA) assay coupled with lateral flow dipstick (LFD), which could achieve sensitive diagnosis of tumor necrosis factor receptor (TNFR) of CaHV within 35 min at 40°C. Our RAA-LFD method had a satisfactory detection limit of 100 gene copies per reaction, which was 100-fold more sensitive than traditional PCR. In addition, no cross-reaction was observed with other viral pathogens, including koi herpesvirus (KHV), cyprinid herpesvirus 2 (CyHV-2), infectious hematopoietic necrosis virus (IHNV), spring viremia of carp virus (SVCV) and grass carp reovirus (GCRV). Furthermore, the overall cost of the method was cut in half compared to previous studies. In conclusion, RAA-LFD assay is therefore, a promising alternative for point-of-care testing (POCT) of CaHV, which is feasible and of certain value in application of aquatic disease control.

Development of real‐time recombinase polymerase amplification (RPA) and RPA combined with lateral flow dipstick (LFD) assays for the rapid and sensitive detection of cyprinid herpesvirus 3

“…CyHV-3 which was designed by Cano I et al was 100 viral copies (Cano et al, 2021). The detection limit of RPA-LFD method which was developed by Gui L et al (Gui et al, 2022) for the detection of Carassius auratus herpesvirus was 100 gene copies per reaction. When our RPA-LFD method had a detection limit of 13 copies.…”

Advances in the application of recombinase-aided amplification combined with CRISPR-Cas technology in quick detection of pathogenic microbes

“…At present, loop-mediated isothermal amplification (LAMP), recombinase polymerase amplification (RPA), and RAA are several mature isothermal nucleic acid detection technologies (Yan et al, 2014). They are isothermal, specific, and efficient, among which RAA is the only backup technology that is possible to carry out in vivo nucleic acid amplification technology (Gui et al, 2022).…”

Development and application of a recombinase‐aided amplification combined with a lateral flow dipstick assay for rapid visual detection of anguillid herpesvirus 1

“…When combined with a lateral flow dipstick (LFD) assay, the amplicon produced in the presence of the probe will include two labels on one DNA sequence, which can be easily visualized in a sandwich assay by antibodies or antibody/streptavidin (Posthuma‐Trumpie et al., 2009). The RAA method has been developed for the detection of aquatic pathogens, including parasites (Wu et al., 2019), bacteria (Li et al., 2023), and viruses (Gui et al., 2022). In this study, we developed a rapid, effective, and convenient RAA‐LFD assay for detecting AngHV‐1.…”