Chen Lei, Zhang Qiaoli, Sun Wenbo, Mauk Michael G, Li Qingmei
Shandong Provincial Key Laboratory of Animal Resistance Biology, College of Life Sciences, Shandong Normal University, Jinan, Shandong, China.
Shandong Key Laboratory of Animal Disease Control and Breeding, Institute of Animal Science and Veterinary Medicine, Shandong Academy of Agricultural Sciences, Jinan, Shandong, China.
Poult Sci. 2025 May 1;104(8):105241. doi: 10.1016/j.psj.2025.105241.
Duck viral hepatitis is very common in China, causing significant impact and economic losses to the duck farming industry. Currently, DHAV-3 has become the main factor causing duck viral hepatitis in China. However, the existing DHAV-3 vaccines cannot completely rule out the potential risk of the vaccine strain becoming more virulent. Due to the high similarity in genomic sequences between wild strains and vaccine strains (with only a few base differences), traditional detection methods struggle to accurately differentiate them, severely interfering with disease control decisions. Therefore, simultaneously detecting both the virulent strain and the attenuated strain of DHAV-3 is crucial for evaluating vaccine efficacy, monitoring virus mutations, and optimizing control strategies. This study, using the DHAV-3 SD70 attenuated strain as an example, developed a highly sensitive and rapid detection method to identify and distinguish between the DHAV-3 virulent and SD70 attenuated strains, providing a new strategy for identifying both strains. Currently, there are no literature reports on the detection methods for the two strains. Therefore, we propose a single-base recognition system strategy based on RPA-CRISPR. DHAV-3 virulent and attenuated strains were specifically identified by this method based on only a few different base sequences. This method can detect two target genes as low as 10° copy/μL within 35 min. In addition, when this method was used for samples analysis, the results of this method, sequencing results, and the results provided by the company were compared and found to be consistent. This method has the advantages of fast speed, simple operation, high specificity and sensitivity, which can be used for the detection of DHAV-3 virulence strain and SD70 attenuated strain, and lays a technical foundation for disease control, vaccine evaluation and mutation monitoring.
鸭病毒性肝炎在中国非常普遍，给养鸭业造成了重大影响和经济损失。目前，DHAV - 3已成为中国引起鸭病毒性肝炎的主要因素。然而，现有的DHAV - 3疫苗不能完全排除疫苗株毒力增强的潜在风险。由于野生株与疫苗株的基因组序列相似度高（仅有少数碱基差异），传统检测方法难以准确区分它们，严重干扰疾病防控决策。因此，同时检测DHAV - 3的强毒株和弱毒株对于评估疫苗效力、监测病毒变异以及优化防控策略至关重要。本研究以DHAV - 3 SD70弱毒株为例，开发了一种高灵敏度、快速的检测方法，用于鉴定和区分DHAV - 3强毒株和SD70弱毒株，为两种毒株的鉴别提供了新策略。目前，尚无关于这两种毒株检测方法的文献报道。因此，我们提出了一种基于RPA - CRISPR的单碱基识别系统策略。基于仅有的少数不同碱基序列，该方法可特异性鉴定DHAV - 3强毒株和弱毒株。该方法能在35分钟内检测低至10°拷贝/μL的两个靶基因。此外，将该方法用于样本分析时，将其结果与测序结果以及公司提供的结果进行比较，发现结果一致。该方法具有速度快、操作简单、特异性和灵敏度高的优点，可用于DHAV - 3强毒株和SD70弱毒株的检测，为疾病防控、疫苗评估和变异监测奠定了技术基础。
