Studies on replication, pathogenicity and transmission of plant RNA viruses in the genus Furovirus

土传小麦花叶病毒(SBWMV)是福罗病毒属的模式种，其基因组为杆状病毒粒子。SBWMV侵染小麦和大麦植株，引起花叶和矮缩病。SBWMV是由土壤真菌传播并滞留在土壤中的，因此很难从侵染地控制该病毒病。在本研究中，我们对SBWMV的复制、致病性和传播进行了研究，以期控制该病。构建了SBWMV日本分离物、美国分离物和英国分离物的全长侵染性cDNA克隆。此外，还建立了大麦叶肉原生质体系统，并对其进行了侵染性测定。利用大麦原生质体对三个分离物进行的重组实验表明，RNA1和RNA2组合均支持病毒RNA复制，表明三个地理分离物之间的亲缘关系较近。小麦白粉病侵染需要20℃以下的温度。将感染的体外转录物接种大麦原生质体，在15~25℃的温度下培养，结果表明，大麦原生质体的最适复制温度为17℃。当病毒在17℃传代时，野生型RNA2发生频繁的缺失突变，缺失突变体对小麦植株的症状比野生型更严重。用全长感染性RNA2转录本分析了接种物中的缺失突变，发现上游衣壳蛋白基因发生了框内突变。症状的严重程度与缺失的时间长短相关。如上所述，为进一步详细分析呋喃病毒的复制、致病和传播提供了基本信息、材料和方法。

Soil-borne wheat mosaic virus (SBWMV) is the type species in the genus Furovirus with bipartite plus-sense RNA genome encapsidated in rod-shaped virions. SBWMV infects wheat and barley plants, causing mosaic and stunt diseaese. SBWMV is transmitted by and retained in soil-inhabiting fungus and therefore it is quite difficult to control this virus disease from the infested field. In this study, we investigated replication, pathogenicity and transmission of SBWMV for the purpose of controlling the disease.1. Three full-length infectious cDNA clones for Japanese, American and English isolates of SBWMV were constructed. Also, an infectivity assay, a barley mesophyl protoplasts system was developed for SBWMV. Reassortment experiments among the three isolates using barley protoplasts showed that all RNA1 and RNA2 combinations supported viral RNA replication, indicating a close relationship among the three geographical isolates.2. Infection of SBWMV to wheat plants requires temperatures lower than 20 ℃. Barley protoplasts inoculated with infectious in vitro transcripts were incubated at temperatures between 15 ℃ and 25 ℃, and it was proved that the optimal temperature for RNA replication is 17 ℃.3. Wild-type RNA2 of SBWMV undergoes frequent deletion mutation when the virus was passaged at 17 ℃ and deletion mutants cause more severe symptoms on wheat plants that the wild-type virus. Deletion mutation was analyzed using full-length infectious RNA2 transcripts in inoculum and found to occur in-frame to the upstream capsid protein gene. Symptom severities correlated with the length of deletion.As described above, the fundamental information, materials and methods were obtained for further detailed analysis on replication, pathogenicity and transmission of furoviruses.