Visual detection and differentiation of porcine epidemic diarrhea virus wild−type strains and attenuated vaccine strains using CRISPR/Cas13a-based lateral flow strip

Abstract

Porcine epidemic diarrhea virus (PEDV) is an enteric coronavirus that causes acute watery diarrhea and vomiting in unweaned piglets. Infections result in high mortality and serious economic losses to the swine industry. PEDV attenuated vaccine does not completely protect against all mutant wild-type strains, and PEDV infection can periodically occur. A sensitive, accurate, and simple detection method for PEDV is needed to reduce the occurrence of the disease. In this study, the CRISPR/Cas13a system was combined with recombinase aided amplification to develop a rapid diagnostic method to distinguish PEDV wild-type strains from attenuated vaccine strains. The method is based on isothermal detection at 37°C. The results are used for visual readout. The assay had high sensitivity and specificity, with a detection limit of 101 copies/μL for the gene of interest, and no cross-reactivity with other pathogens. The Cas13a detection worked well with clinical samples. This visual, sensitive, and specific nucleic acid detection method based on CRISPR/Cas13a should be a powerful tool for detecting PEDV.

Rapid detection of mexX in Pseudomonas aeruginosa based on CRISPR-Cas13a coupled with recombinase polymerase amplification

“…In fluorescence assay of another Ebola and Lassa virus of LASV-IV in the same research, the LoD was 100 copies/μL, whereas the LFD had a LoD of 1,000 copies/μL ( Barnes et al, 2020 ). Similar phenomena were also found in the detection of dengue virus ( Gootenberg et al, 2018 ), Porcine Circovirus Type 4 ( Wang et al, 2023 ), SARS-CoV-2 ( Su et al, 2022 ), epidemic diarrhea virus (PEDV; Yin et al, 2022 ), and hepatitis B virus ( Tian et al, 2023 ). Even while LFD’s sensitivity is slightly lower, it can evaluate detection findings directly with the human eye and eliminates the need for fluorescence detection instruments, simplifying the equipment required.…”

Rapid detection of H5 subtype avian influenza virus using CRISPR Cas13a based-lateral flow dipstick

“…In recent years, an increasing number of scholars combined RAA with CRISPR to increase the detection limit of the assay. RAA and CRISPR-Cas13a’s detection methods have been successfully used to detect swine fever virus (SFV) ( Zhang et al, 2022 ), African swine fever virus (ASFV) ( Wei et al, 2022 ), Toxoplasma gondii ( Zhao et al, 2022 ), and porcine epidemic diarrhea virus (PEDV) ( Yin et al, 2022 ).…”