Vanderhallen H, Koenen F
Veterinary and Agrochemic Research Center (CODA/CERVA), B-1180 Ukkel, Belgium.
J Clin Microbiol. 1998 Dec;36(12):3463-7. doi: 10.1128/JCM.36.12.3463-3467.1998.
The objective of the present study was to gain a better understanding of the epidemiology of encephalomyocarditis virus (EMCV) infections in pigs by applying molecular techniques. The diagnostic potential of a reverse transcription-PCR (RT-PCR) targeting 286 nucleotides at the 3' end of the gene which encodes the viral polymerase was assessed with experimental and field samples. In addition, the use of the amplified sequences for an epidemiological study was evaluated. The heart was clearly shown to be the most suitable organ. The detection limit was determined to be 1 viral particle in 100 mg of heart tissue. The sensitivity and specificity of the assay on the basis of the results obtained in this study were 94 and 100%, respectively. Phylogenetic analysis of the amplified sequences classified EMCVs in two distinct lineages. Group A consists of the reference strain ATCC 129B, all isolates collected between 1991 and 1994 in Belgium in association with reproductive failure, and all Greek isolates. All Belgian isolates collected since the first isolation of EMCV in relation to myocardial failure in fatteners in Belgium group together with the isolates from Cyprus (1996 and 1997), Italy (1986 to 1996), and France (1995) in group B irrespective of their pathogenicity. The analyzed part of the 3D gene differed by 13.0% between Groups A and B. In contrast to the sequence homogeneity of the Belgian isolates collected between 1991 and 1994, molecular diversity, which ranged between 0 and 2%, was observed among the Belgian isolates collected in 1995 and 1996. Among all Greek isolates the diversity ranged between 1 and 8%. However, this diversity does not seem to reflect geographical links between the outbreaks. A RT-PCR for the rapid and specific diagnosis of EMCV in a variety of clinical samples followed by nucleotide sequence analysis proved to be valuable for molecular epidemiological studies.
本研究的目的是通过应用分子技术，更好地了解猪脑心肌炎病毒（EMCV）感染的流行病学情况。利用实验样本和现场样本，评估了针对编码病毒聚合酶基因3'端286个核苷酸的逆转录聚合酶链反应（RT-PCR）的诊断潜力。此外，还评估了扩增序列在流行病学研究中的应用。结果清楚地表明，心脏是最合适的器官。检测限确定为每100mg心脏组织中有1个病毒颗粒。根据本研究获得的结果，该检测方法的灵敏度和特异性分别为94%和100%。对扩增序列进行系统发育分析，将EMCV分为两个不同的谱系。A组包括参考菌株ATCC 129B、1991年至1994年在比利时收集的与繁殖失败相关的所有分离株以及所有希腊分离株。自比利时首次分离出与育肥猪心肌衰竭相关的EMCV以来收集的所有比利时分离株，与来自塞浦路斯（1996年和1997年）、意大利（1986年至1996年）和法国（1995年）的分离株一起归入B组，无论其致病性如何。A组和B组之间3D基因的分析部分差异为13.0%。与1991年至1994年收集的比利时分离株的序列同质性相反，在1995年和1996年收集的比利时分离株中观察到分子多样性，范围在0%至2%之间。在所有希腊分离株中，多样性范围在1%至8%之间。然而，这种多样性似乎并未反映疫情之间的地理联系。用于快速、特异性诊断各种临床样本中EMCV的RT-PCR，随后进行核苷酸序列分析，被证明对分子流行病学研究很有价值。
