Zientara S, Sailleau C, Plateau E, Moulay S, Mertens P P, Crucière C
Centre National d'Etudes Vétérinaires et Alimentaires, Laboratoire Central de Recherches Vétérinaires, Maisons Alfort, France.
Arch Virol Suppl. 1998;14:221-34. doi: 10.1007/978-3-7091-6823-3_20.
This paper describes a method to rapidly identify African horse sickness virus (AHSV), using a single tube reverse transcription polymerase chain reaction (PCR). This method was used to amplify cDNA copies of genome segments 7 and 10 from several different AHSV strains, of different serotypes, which were then analysed by sequencing and/or endonuclease digestion. AHSV VP7 (encoded by genome segment 7) is one of the two major capsid proteins of the inner capsid layer, forming the outer surface of the core particle. VP7 is highly conserved and is the major serogroup specific antigen common to all nine AHSV serotypes. Digestion of the 1179 bp cDNA with restriction enzymes, allowed differentiation of several strains of different serotypes and identified six distinct groups containing AHSV-1, 3, 6 and 8; AHSV-2; AHSV-4; AHSV-5; AHSV-7; and AHSV-9. Differences were detected between wild type viruses and vaccine strains that had been attenuated by multiple passage in suckling mouse brain or in tissue cultures. RFLP analysis was also used to study variation the 758 bp cDNA copies of AHSV genome segment 10, which encodes the two small non-structural membrane proteins NS3 and NS3a. In this way it was possible to distinguish each of the strains tested, except AHSV 4 (USDA) and AHSV 9 (USDA). However, these isolates could be distinguished by RFLP analysis of genome segment 7 cDNA. Using sequence analysis of genome segment 10 we were able to classify the virus isolates into three groups: AHSV-1, 2 and 8; AHSV-3 and 7; AHSV 4, 5, 6 and 9. These studies confirmed that the virus which first appeared in central Spain in July 1987, subsequently spread into northern Morocco in October 1989.
本文描述了一种使用单管逆转录聚合酶链反应（PCR）快速鉴定非洲马瘟病毒（AHSV）的方法。该方法用于从几种不同血清型的不同AHSV毒株中扩增基因组片段7和10的cDNA拷贝，然后通过测序和/或核酸内切酶消化进行分析。AHSV VP7（由基因组片段7编码）是内衣壳层的两种主要衣壳蛋白之一，形成核心颗粒的外表面。VP7高度保守，是所有九种AHSV血清型共有的主要血清群特异性抗原。用限制性内切酶消化1179 bp cDNA，可区分几种不同血清型的毒株，并鉴定出六个不同的组，分别包含AHSV-1、3、6和8；AHSV-2；AHSV-4；AHSV-5；AHSV-7；以及AHSV-9。在野生型病毒和通过在乳鼠脑或组织培养物中多次传代而减毒的疫苗毒株之间检测到了差异。限制性片段长度多态性（RFLP）分析还用于研究AHSV基因组片段10的758 bp cDNA拷贝的变异，该片段编码两种小的非结构膜蛋白NS3和NS3a。通过这种方式，可以区分除AHSV 4（美国农业部）和AHSV 9（美国农业部）之外的每个测试毒株。然而，这些分离株可以通过基因组片段7 cDNA的RFLP分析来区分。使用基因组片段10的序列分析，我们能够将病毒分离株分为三组：AHSV-1、2和8；AHSV-3和7；AHSV 4、5、6和9。这些研究证实，1987年7月首次出现在西班牙中部的病毒，随后于1989年10月传播到摩洛哥北部。
