Research and Development Centre, Indian Immunologicals Limited, Gachibowli, Hyderabad 500032, India; Department of Biotechnology, Acharya Nagarjuna University, Guntur 522510, India.
J Virol Methods. 2012 May;181(2):197-201. doi: 10.1016/j.jviromet.2012.02.004. Epub 2012 Feb 21.
The antigenic types of canine parvovirus (CPV) are defined based on differences in the amino acids of the major capsid protein VP2. Type specificity is conferred by a limited number of amino acid changes and in particular by few nucleotide substitutions. PCR based methods are not particularly suitable for typing circulating variants which differ in a few specific nucleotide substitutions. Assays for determining SNPs can detect efficiently nucleotide substitutions and can thus be adapted to identify CPV types. In the present study, CPV typing was performed by single nucleotide extension using the mini-sequencing technique. A mini-sequencing signature was established for all the four CPV types (CPV2, 2a, 2b and 2c) and feline panleukopenia virus. The CPV typing using the mini-sequencing reaction was performed for 13 CPV field isolates and the two vaccine strains available in our repository. All the isolates had been typed earlier by full-length sequencing of the VP2 gene. The typing results obtained from mini-sequencing matched completely with that of sequencing. Typing could be achieved with less than 100 copies of standard plasmid DNA constructs or ≤10¹ FAID₅₀ of virus by mini-sequencing technique. The technique was also efficient for detecting multiple types in mixed infections.
犬细小病毒（CPV）的抗原型是基于主要衣壳蛋白 VP2 的氨基酸差异来定义的。型特异性由少数氨基酸变化赋予，特别是由少数核苷酸取代赋予。基于 PCR 的方法不太适合对在少数特定核苷酸取代上存在差异的循环变体进行分型。用于确定单核苷酸多态性（SNP）的检测方法可以有效地检测核苷酸取代，因此可以适应于鉴定 CPV 型。在本研究中，通过使用迷你测序技术的单核苷酸延伸进行 CPV 分型。为所有四种 CPV 型（CPV2、2a、2b 和 2c）和猫泛白细胞减少症病毒建立了迷你测序特征。使用迷你测序反应对 13 个 CPV 田间分离株和我们库中可用的两种疫苗株进行 CPV 分型。所有分离株均已通过 VP2 基因全长测序进行了分型。从迷你测序获得的分型结果与测序结果完全匹配。通过迷你测序技术，使用少于 100 个标准质粒 DNA 构建体或≤10¹ FAID₅₀ 的病毒即可实现分型。该技术对于检测混合感染中的多种类型也非常有效。
