Tobacco mosaic virus (TMV; genus Tobamovirus) is one of the most prevailing pathogens that seriously affects the quality and yield of tobacco (Nicotiana tabacum) leaves. Cross-protection using mild strains is a potential strategy for the biological prevention of plant viral diseases. Complementary mutations in attenuated strains may cause attenuated ones to suddenly evolve into virulent strains, which limits the application of cross-protection in practice. To data there has been no study on engineering the complementary mutation sites to generate stable attenuated mutants for cross-protection. In this study, we found that the substitution of the conserved arginine at position 88 (R88) in p126 protein with alanine (A) abolished the cell-to-cell movement and reduced the replication of TMV. However, a spontaneous complementary mutation of serine at position 114 (S114) to lysine (K) in p126 restored TMV virulence. Substitution of S114 with R in p126 restored the systemic infection but not the virulence of TMV, therefore, the mutant TMV-R88A/S114R was an attenuated one. Furthermore, our results showed that TMV-R88A/S114R was a stable attenuated mutant, and could effectively protect tobacco plants against the wild-type TMV infection. This study reports a promising TMV mild mutant for cross-protection in tobacco plants by modifying the complementary mutation site in p126.

烟草环斑病毒(Tobacco ringspot virus,TRSV)是豇豆花叶病毒科(Comoviridae)线虫传多面体病毒属(Nepovirus)的代表种,是我国公布的二类进境检疫有害生物.建立特异性强、灵敏性高的检测体系是有效控制TRSV传播的技术保障.本研究在分析引物的特异性、优化退火温度及dNTPs、引物、Taq DNA聚合酶浓度的基础上,建立了完善的TRSV RT-PCR检测体系.筛选出的TRSV特异性引物对为1-1 F/1-1 R,在退火温度为53℃、dNTPs终浓度为0.075 mmol/L、引物终浓度为0.30μmol/L、Taq DNA聚合酶终浓度为0.035 U/μL条件下检测效率最高,可从发病植物总RNA量为0.090 ng中检测到目的片段.研究结果对准确灵敏地检测TRSV具有重要意义.

马铃薯是我国重要粮食和经济作物.马铃薯Y病毒(potato virus Y,PVY)是危害马铃薯生产的重要病害.种植脱毒种薯是防治PVY最有效的途径.马铃薯种薯携带PVY问题严重,但种薯中PVY株系还不清楚.本研究利用PVY特异性抗体检测了7个马铃薯品种362个种薯,发现不同品种种薯带毒率差异较大,最高达12％.通过RT-PCR方法扩增获得了7个PVY分离物编码区全序列.重组分析发现7个分离物基因组均为重组型,根据重组位点的差异可以分为PVYNTN-NW(SYR-Ⅱ型)、Rec-l、Rec-2和Rec-3等4种重组类型,后3种为新重组类型.系统进化分析发现,分离物HQH18G3-10与PVYNTN-Nw(SYR-Ⅱ型)处于同一个大的分支,但与中国PVY大田分离物聚集在一起形成一个相对独立的组,命名为PVYNTN-Nw(CN型);其余6个分离物与数据库中的中国分离物聚集在PVYN.Wi组.这暗示PVY中国分离物具有相对独立的进化过程,PVY马铃薯大田分离物和种薯分离物进化上相近.所有分离物均能在珊西烟上引起典型叶脉坏死症状,HQH18G3-10引起的坏死症状最为严重.本研究首次报道了我国种薯内PVY发生情况,对分析病毒发生发展规律和防控具有借鉴作用.