Establishment and Application of Mismatch Amplification Mutation Assay-PCR for Rapid Detection and Differentiation of Duck Hepatitis A Virus-1 Attenuated Vaccine and Wild Strains

• Published in: Animals (Basel) Vol. 14; no. 18; p. 2733
• Main Authors: Yu, Cheng-Dong, Choi, Yu-Ri, Park, Jong-Yeol, Kim, Sang-Won, Cha, Se-Yeoun, Jang, Hyung-Kwan, Kang, Min, Wei, Bai
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 21.09.2024; MDPI
• Subjects: attenuated vaccine; Design; DHAV-1; differentiation; Ducks; Embryos; Epidemiology; Farms; Genetic aspects; Genomes; Hepatitis A; Hepatitis A virus; Laboratory animals; MAMA-PCR; Mutation; Pathogens; Poultry industry; Proteins; Single nucleotide polymorphisms; Taxonomy; Vaccination; Vaccines; Virulence; Viruses; wild-type; South Korea; United States > US; DHAV-1; differentiation; attenuated vaccine; MAMA-PCR; wild-type
• ISSN: 2076-2615; 2076-2615
• DOI: 10.3390/ani14182733

Duck hepatitis A virus type 1 (DHAV-1) is the main pathogen causing viral hepatitis in ducks, marked by high contagion and acute mortality. Live attenuated DHAV-1 vaccines are widely used to control the disease. This study aims to develop a mismatch amplification mutation assay (MAMA)-PCR for the rapid detection and differentiation of Korean DHAV-1 wild-type strains from vaccine strains. A MAMA primer was designed to target a single nucleotide polymorphism (SNPs) at position 2276 within the VP1 gene, allowing differentiation in a single PCR reaction. The MAMA-PCR accurately identified both strains, with detection limits of 10 ELD /mL and 10 ELD /mL, respectively. The MAMA-PCR demonstrated specificity, showing no cross-reactivity with 12 other viral and bacterial pathogens. The MAMA-PCR was applied to 89 farms, yielding results consistent with nested-PCR and sequence determination, identifying four positive farms for DHAV-1 vaccine strains. In conclusion, this study is the first to employ the MAMA-PCR method to distinguish between DHAV-1 wild-type and vaccine strains. The developed method is rapid, simple, specific, and sensitive, thereby serving as an effective tool for clinical diagnostics in identifying and differentiating between Korean DHAV-1 wild-type and vaccine strains.