Wagaba Henry, Beyene Getu, Trembley Cynthia, Alicai Titus, Fauquet Claude M, Taylor Nigel J
Donald Danforth Plant Science Center, 975 N, Warson Rd, St Louis, MO, USA.
BMC Res Notes. 2013 Dec 6;6:516. doi: 10.1186/1756-0500-6-516.
Techniques to study plant viral diseases under controlled growth conditions are required to fully understand their biology and investigate host resistance. Cassava brown streak disease (CBSD) presents a major threat to cassava production in East Africa. No infectious clones of the causal viruses, Cassava brown streak virus (CBSV) or Ugandan cassava brown streak virus (UCBSV) are available, and mechanical transmission to cassava is not effective. An improved method for transmission of the viruses, both singly and as co-infections has been developed using bud grafts.
Axillary buds from CBSD symptomatic plants infected with virulent isolates of CBSV and UCBSV were excised and grafted onto 6-8 week old greenhouse-grown, disease-free cassava plants of cultivars Ebwanateraka, TME204 and 60444. Plants were assessed visually for development of CBSD symptoms and by RT-PCR for presence of the viruses in leaf and storage root tissues. Across replicated experiments, 70-100% of plants inoculated with CBSV developed CBSD leaf and stem symptoms 2-6 weeks after bud grafting. Infected plants showed typical, severe necrotic lesions in storage roots at harvest 12-14 weeks after graft inoculation. Sequential grafting of buds from plants infected with UCBSV followed 10-14 days later by buds carrying CBSV, onto the same test plant, resulted in 100% of the rootstocks becoming co-infected with both pathogens. This dual transmission rate was greater than that achieved by simultaneous grafting with UCBSV and CBSV (67%), or when grafting first with CBSV followed by UCBSV (17%).
The bud grafting method described presents an improved tool for screening cassava germplasm for resistance to CBSD causal viruses, and for studying pathogenicity of this important disease. Bud grafting provides new opportunities compared to previously reported top and side grafting systems. Test plants can be inoculated as young, uniform plants of a size easily handled in a small greenhouse or large growth chamber and can be inoculated in a controlled manner with CBSV and UCBSV, either singly or together. Disease symptoms develop rapidly, allowing better studies of interactions between these viral pathogens, their movement within shoot and root systems, and how they induce their destructive disease symptoms.
为全面了解植物病毒病的生物学特性并研究宿主抗性，需要在可控生长条件下研究植物病毒病的技术。木薯褐色条纹病（CBSD）对东非的木薯生产构成重大威胁。目前尚无致病病毒木薯褐色条纹病毒（CBSV）或乌干达木薯褐色条纹病毒（UCBSV）的感染性克隆，且机械传播至木薯的效果不佳。已开发出一种利用芽接来单独或同时传播这两种病毒的改进方法。
从感染了CBSV和UCBSV强毒株的CBSD症状植株上切下腋芽，嫁接到6 - 8周龄、在温室中生长的、无病的木薯品种Ebwanateraka、TME204和60444植株上。通过肉眼评估植株是否出现CBSD症状，并通过逆转录聚合酶链反应（RT-PCR）检测叶片和贮藏根组织中是否存在病毒。在重复实验中，接种CBSV的植株在芽接后2 - 6周有70 - 100%出现了CBSD叶片和茎部症状。在嫁接接种后12 - 14周收获时，受感染植株的贮藏根出现典型的严重坏死病变。先将感染UCBSV植株的芽嫁接，10 - 14天后再将携带CBSV的芽嫁接到同一试验植株上，结果100%的砧木同时感染了这两种病原体。这种双重传播率高于同时嫁接UCBSV和CBSV时的传播率（67%），也高于先嫁接CBSV再嫁接UCBSV时的传播率（17%）。
所描述的芽接方法为筛选木薯种质对CBSD致病病毒的抗性以及研究这种重要病害的致病性提供了一种改进工具。与先前报道的顶接和侧接系统相比，芽接提供了新的机会。试验植株可以作为幼小、均匀的植株接种，其大小便于在小型温室或大型生长室中操作，并且可以以可控的方式单独或同时接种CBSV和UCBSV。病害症状发展迅速，有利于更好地研究这些病毒病原体之间的相互作用、它们在地上部和根系中的移动以及它们如何引发破坏性病害症状。
