Yu Cheng-Dong, Park Jong-Yeol, Kim Sang-Won, Choi Yu-Ri, Cha Se-Yeoun, Jang Hyung-Kwan, Kang Min, Wei Bai
Department of Avian Diseases, College of Veterinary Medicine and Center for Avian Disease, Jeonbuk National University, Iksan 54596, Republic of Korea.
Bio Disease Control (BIOD) Co., Ltd., Iksan 54596, Republic of Korea.
Vet Sci. 2024 Dec 29;12(1):8. doi: 10.3390/vetsci12010008.
Duck hepatitis A virus type 3 (DHAV-3) is a viral pathogen that causes acute, high-mortality hepatitis in ducklings, and vaccination with attenuated live vaccines is currently the main preventive measure against it. However, differentiating infected from vaccinated animals (DIVA) is crucial for clinical diagnosis and effective disease control. This study aimed to develop a rapid mismatch amplification mutation assay PCR (MAMA-PCR) diagnostic method to simultaneously detect and differentiate between wild-type and vaccine strains. The method was specifically designed to target the critical single-nucleotide polymorphism (SNP) site (T→C at position 1143 in the VP0 gene) unique to the Korean vaccine strain AP04203-P100. MAMA-PCR demonstrated high sensitivity and specificity, with detection limits as low as 10 ELD/mL for wild strains and 10 ELD/mL for vaccine strains, and showed no cross-reactivity with 11 other common duck pathogens. The clinical sample results were completely consistent with those obtained using nested PCR detection and gold-standard sequencing. In summary, we successfully developed a rapid, one-step MAMA-PCR method that is more suitable for clinical diagnosis than traditional sequencing methods.
鸭甲型肝炎病毒3型（DHAV-3）是一种能在雏鸭中引发急性、高致死率肝炎的病毒病原体，目前针对该病毒的主要预防措施是接种减毒活疫苗。然而，区分感染动物和免疫动物（DIVA）对于临床诊断和有效的疾病防控至关重要。本研究旨在开发一种快速错配扩增突变分析PCR（MAMA-PCR）诊断方法，以同时检测和区分野生型和疫苗株。该方法专门针对韩国疫苗株AP04203-P100特有的关键单核苷酸多态性（SNP）位点（VP0基因第1143位的T→C）进行设计。MAMA-PCR表现出高灵敏度和特异性，对野生株的检测限低至10 ELD/mL，对疫苗株的检测限为10 ELD/mL，并且与其他11种常见鸭病原体无交叉反应。临床样本检测结果与巢式PCR检测及金标准测序结果完全一致。总之，我们成功开发了一种快速、一步法的MAMA-PCR方法，该方法比传统测序方法更适合临床诊断。
