2016~2017年中国沿海省市虾类偷死野田村病毒(CMNV)分子流行病学调查

Molecular Epidemiological Survey of Covert Mortality Nodavirus (CMNV) in Cultured Crustaceans in China in 2016~2017

中文关键词: 偷死野田村病毒  分子流行病学调查  逆转录套式PCR  逆转录环介导等温扩增  TaqMan实时荧光定量RT-PCR  行动障碍野田村病毒

英文关键词: Covert mortality nodavirus (CMNV)  Molecular epidemiological survey  RT-nPCR  RT-LAMP  TaqMan RT-qPCR  Movement disorder nodavirus (MDNV)

中文摘要:

      偷死野田村病毒(CMNV)引起的病毒性偷死病(VCMD)使中国对虾养殖业遭受了严重的经济损失，为查明CMNV及其变异株的分子流行病学特征，本研究采用逆转录套式PCR (RT-nPCR)、逆转录环介导等温扩增(RT-LAMP)和TaqMan实时荧光定量PCR(TaqMan RT-qPCR) 3种方法，对2016~2017年中国沿海省市CMNV及一种新型野田村病毒——行动障碍野田村病毒(MDNV)的流行、分布和变异情况进行了分析。结果显示，养殖凡纳滨对虾、中国明对虾、日本囊对虾、斑节对虾和罗氏沼虾等种类中均可检测到CMNV阳性；在河北、山东、浙江、福建、广东和海南等省市采集的患病对虾中均存在CMNV。基于RT-nPCR、RT-LAMP和TaqMan RT-qPCR的检测结果显示，2016和2017年样品中CMNV的阳性检出率依次分别为11.8%和7.8%，6.7%和3.9%，17.7%和12.4%；基于上述3种方法检测结果计算得出，2016和2017年样品中CMNV的阳性检出率分别为26.8%和16.3%；基于RT-LAMP的分析显示，2016年样品中MDNV的阳性检出率为9.4%。本研究表明，中国沿海省市养殖虾类中VCMD的流行和危害仍不容忽视，RNA依赖的RNA聚合酶基因不断变异导致前期基于该基因开发的CMNV检测方法可靠性下降，检出假阴性风险升高；同时，发病对虾中出现了新的野田村病毒株系且具有较高的流行率，其传播危害风险值得高度关注。

英文摘要:

      Covert mortality nodavirus (CMNV), a recently emerged RNA virus, is the pathogen responsible for viral covert mortality disease, which has caused serious losses of shrimp aquaculture in China in recent years. In order to determine the transmission, prevalence, and evolution of CMNV in the coastal provinces and cities in China, national wide survey of the molecular epidemiology of the CMNV and it’s variants was conducted, based on analysis of shrimp samples collected from 2016 and 2017 by using the methods of RT-nPCR, RT-LAMP, and TaqMan RT-qPCR in present study. The analyses found the presence of CMNV-positive specimens in the major species of cultured crustaceans, including Litopenaeus vannamei, Marsupenaeus japonicus, Fenneropenaeus chinensis, Penaeus monodon, and Macrobrachium rosenbergii. CMNV-positive specimens appeared in almost all the coastal provinces, such as Hebei, Shandong, Zhejiang, Fujian, Guangdong, and Hainan. The results based on RT-nPCR assays showed that the prevalence rates of CMNV among the collected samples were 11.8% (30/254) and 7.8% (30/387) in 2016 and 2017, respectively. The results based on RT-LAMP assays showed that the prevalence rates of CMNV were 6.7% (17/254) and 3.9% (15/387) in 2016 and 2017, respectively. The results based on TaqMan RT-qPCR assays showed that the prevalence rates of CMNV were 17.7% (45/254) and 12.4% (48/387) in 2016 and 2017, respectively. The total prevalence rates of CMNV were 26.8% (68/254) and 16.3% (63/387) in 2016 and 2017, respectively, based on the above mentioned three methods. The results based on RT-LAMP assays showed that the prevalence rate of MDNV was 9.4% (24/254) in 2016. The high prevalence of CMNV in the major shrimp species and in the main farming areas revealed that this virus still threatened shrimp aquaculture in China during 2016~2017. Moreover, mutation of the RNA-dependent RNA polymerase (RdRp) gene of CMNV will raise the risk of appearing of false-negative result in CMNV molecular tests, which is of great concern. Meanwhile, the high prevalence rates of the emerging CMNV variant, MDNV, is a reminder that close attention needs to be paid to the high risk of MDNV transmission widely in farmed crustaceans.