Comparative evaluation of three WOAH-recommended RT-PCR assays for the detection of yellow head virus genotype 1

다국어 초록 (Multilingual Abstract)

Yellow head virus genotype 1 (YHV-1), the causative agent of yellow head disease, is a major viral pathogen causing rapid and high mortality in shrimp aquaculture. The World Organisation for Animal Health recommends three RT-PCR methods for YHD diagnosis, however, simultaneous application of all three methods requires considerable time, labor, and reagent costs. This study evaluated the analytical and diagnostic performances of the three WOAH-recommended assays using YHV-1-positive samples. Analytical sensitivity showed that method 1 detected YHV-1 down to 1.8×103 copies/ µL, whereas methods 2 and 3 with nested PCR improved the detection to 1.8×102 copies/µL. Analytical specificity with YHV-1-positive and negative shrimp tissues mixture (1:0 to 1:7, w/w) showed that method 1 detected RNA at a 1:3 dilution (2.4×103 copies/µL). In contrast, methods 2 and 3 detected RNA at the 1:4 dilution (7.0×102 copies/µL) in the first PCR and extended detection to a 1:6 dilution (3.0×102 copies/µL) in the nested PCR. Evaluation of 30 positive and 30 negative shrimp samples revealed that all assays achieved 100.0±0.0% diagnostic specificity. Diagnostic sensitivity was 80.0±0.0% for method 1, 86.7±3.4% in the first PCR and 100.0±0.0% in the nested PCR for method 2, and 77.8±3.9% in the first PCR and 100±0.0% in the nested PCR for method 3. Overall, our findings provide practical guidance for selecting appropriate diagnostic methods for YHV-1 detection and support reliable disease surveillance and control in shrimp aquaculture.