Cycle 1 (2011 Deadline)
Characterization of cassava mosaic gemini viruses and their satellites in cassava at the cellular level
PI: Joseph Ndunguru, Mikocheni Agricultural Research Institute
US Partner: Linda Hanley-Bowdoin, North Carolina State University
Project Dates: May 2012 - April 2014
Cassava is an important staple crop in Africa and Asia, where it is eaten by more than 700 million people every day. It is grown by subsistence farmers in the poorest villages and is often the only food source when other crops fail or are destroyed by conflict. Cassava can grow under drought, high temperature, and poor soil conditions, but its production is severely limited by viral diseases. Cassava mosaic disease (CMD) is caused by a DNA virus complex consisting of seven geminivirus species that act synergistically to enhance disease severity. Recently, two satellite DNAs associated with the complex have been shown to break resistance and enhance symptoms.
Scientists from Mikocheni Agricultural Research Institute
(MARI) are being trained on to use the vibratome and
fluorescent microscope (Photo courtesy Joseph Ndunguru).
Linus Paul, who is in charge of microscopy
at MARI, is working with a vibratome (Photo
courtesy Joseph Ndunguru).
Cassava mosaic geminiviruses (CMGs) induce diverse symptoms in cassava depending on the host genotype, age of infection, amount of virus inoculum, virus strain, vector activity, and environmental and other host factors. Research on CMGs has generated extensive information about viral diversity, genome sequence, replication, transmission, disease epidemiology and disease control. In contrast, there are no reports that describe the changes that occur in cassava leaves at a cellular level in response to CMG infection. The goal of this proposal is to establish cell biology infrastructure and expertise at Mikocheni Agricultural Research Institute (MARI) in Tanzania and to use these resources to characterize CMG infection in cassava. A combination of light and fluorescent microcopy will be used to examine CMD processes at a cellular level in cassava leaves. Using in situ hybridization to detect CMG and satellite DNAs in cassava leaves, these researchers will seek to determine if different CMGs infect different leaf cell types and if the nature and number of the target cells change in mixed infections and/or in the presence of the satellites. The research team will also examine the cellular architecture of infected leaves as a first step toward understanding the physiological basis of the extreme leaf deformation phenotypes correlated with the presence of CMD-associated satellites. The application of cell biology to CMD represents a unique opportunity to study the interactions of different viruses with a common host and with each other and satellite DNAs. The increased knowledge to be gained through the project should contribute to understanding of this important plant virus and to the development of sustainable strategies to control it and thereby limit its economic and nutritional impact.
Summary of Recent Activities
With their new microscope facility operational, Dr. Ndunguru and his group spent the summer and early fall of 2013 focusing on objective 2 of the project, namely comparison of infection patterns of cassava mosaic geminiviruses in single or dual infection. For this purpose, the team designed digoxigenin-labeled probes specific to the Africa Cassava Mosaic Viruses (ACMV) and East Africa Cassava Mosaic Viruses (EACMV) species and strains and degenerate probes for both ACMV and EACMVs. All three probes have been ordered from the manufacturer and the team is currently awaiting their delivery. The researchers sub-cultured and raised young cassava plants (Kibandameno and TME-3) in the tissue culture laboratory, after which they acclimatized them in the screen house. Unfortunately, after acclimatization almost all the plants died due to temperature variations in the screen house, a situation that may be due to lack of sufficient acclimatization techniques. Therefore, the team has begun sub-culturing and raising plants again in the tissue culture lab to have them ready for bombardment and in situ hybridization once the appropriate probes are shipped.
Cassava plants maintained in a tissue culture lab ready for sub-culturing at MARI. (Photo courtesy Joseph Ndunguru).
A research student getting prepared for sub-culturing of cassava plants at MARI(Photo courtesy Joseph Ndunguru).
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