An isothermal recombinase polymerase assay coupled with lateral flow dipstick for differentiation of pseudorabies virus wild isolates and gE-deleted vaccine strains.

Pseudorabies virus (PRV) is one of the most important infectious diseases which leads to significant economic losses in the global swine industry. The gE-deleted vaccine is widely used to prevent susceptible pigs from PRV infection. There is no report of the differentiation of PRV wild strain and vaccine strain by recombinase polymerase amplification (RPA) coupled with a lateral flow dipstick (LFD) method. In the present study, the gD and gE gene-targeted primer-probe sets were designed. The RPA-LFD assay could discriminate between the PRV wild strain and the vaccine strain. The RPA reaction conditions were also evaluated. The optimal reaction temperature and reaction time for the RPA-LFD assay were 37℃ and 20 min. The detection limit was 10 genome copies per reaction for PRV wild strain and gE-deleted vaccine strain. The assay did not have cross-reaction with other common swine viral pathogens. The effectiveness of the RPA-LFD assay for detecting the clinical samples was evaluated by testing 80 samples. The result of the assay was compared with that of the conventional PCR. The positive rate of PRV wild strain by the RPA-LFD assay was 20%, whereas the positive rate of PRV wild strain by the PCR assay was 18.8%. The assay therefore provides a novel alternative for differentiation of PRV wild strain and vaccine strain.