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Response of cassava cultivars to African cassava mosaic virus infection across a range of inoculum doses and plant ages.

ABSTRACT: Cassava production in Africa is constrained by cassava mosaic disease (CMD) that is caused by the Cassava mosaic virus (CMV). The aim of this study was to evaluate the responses of a range of commonly cultivated West African cassava cultivars to varying inoculum doses of African cassava mosaic virus (ACMV). We grafted 10 cultivars of cassava plants with different inoculum doses of CMV (namely two, four, or six CMD-infected buds) when the experimental plants were 8, 10, or 12 weeks old, using non-inoculated plants as controls. Three cultivars showed disease symptoms when grafted with two buds, and four cultivars showed disease symptoms when grafted with four or six buds. Most cultivars became symptomatic six weeks after inoculation, but one ('TMS92/0326') was symptomatic two weeks after inoculation, and two ('Ntollo' and 'Excel') were symptomatic after four weeks. Root weight tended to be lower in the six-bud than in the two-bud dose, and disease severity varied with plant age at inoculation. These results indicate that the level of CMD resistance in cassava cultivars varies with inoculum dose and timing of infection. This will allow appropriate cultivars to be deployed in each production zone of Africa in accordance with the prevalence of CMD.

SUBMITTER: Houngue JA

PROVIDER: S-EPMC6927654 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Response of cassava cultivars to African cassava mosaic virus infection across a range of inoculum doses and plant ages.

Houngue Jerome Anani JA Pita Justin S JS Ngalle Hermine Bille HB Zandjanakou-Tachin Martine M Kuate Apollin Fotso AF Cacaï Gilles Habib Todjro GHT Bell Joseph Martin JM Ahanhanzo Corneille C

PloS one 20191223 12

Cassava production in Africa is constrained by cassava mosaic disease (CMD) that is caused by the Cassava mosaic virus (CMV). The aim of this study was to evaluate the responses of a range of commonly cultivated West African cassava cultivars to varying inoculum doses of African cassava mosaic virus (ACMV). We grafted 10 cultivars of cassava plants with different inoculum doses of CMV (namely two, four, or six CMD-infected buds) when the experimental plants were 8, 10, or 12 weeks old, using non ...[more]

PMID: 31869401

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Project description:Background Cassava is a staple food for over 800 million people globally providing a cheap source of carbohydrate. However, the cultivation of cassava in the country is facing to viral diseases, particularly cassava mosaic disease (CMD) which can cause up to 95% yield losses. With aim to supply farmers demand for clean planting materials, there is need to accelerate the production of the elite cultivars by use of tissue culture in order to cope with the demand. Methods Nodal explants harvested from the greenhouse grown plants were sterilised using different concentrations of a commercial bleach JIK (3.85% NaOCl) and varying time intervals. Microshoots induction was evaluated using thidiazuron (TDZ), benzyl amino purine (BAP), and kinetin. Rooting was evaluated using different auxins (Naphthalene acetic acid NAA and Indole-3-butyricacid IBA). PCR-based SSR and SCAR markers were used to verify the presence of CMD2 gene in the regenerated plantlets. Results The highest level of sterility in explants (90%) was obtained when 20% Jik was used for 15?min. The best cytokinin for microshoots regeneration was found to be kinetin with optimum concentrations of 5, 10 and 20??M for Agric-rouge, Atinwewe, and Agblehoundo respectively. Medium without growth regulators was the best for rooting the three cultivars. A survival rate of 100, 98, and 98% was recorded in the greenhouse for Agric-rouge, Atinwewe, and Agblehoundo respectively and the plantlets appeared to be morphologically normal. The SSR and SCAR analysis of micropropagated plants showed a profile similar to that of the mother plants indicating that the regenerated plantlets retained the CMD2 gene after passing through in vitro culture, as expected with micropropagation. Conclusion The nodal explants was established to be 20% of Jik (3.85% NaOCl) with an exposure time of 15?min. Kinetin was proved to be the best cytokinins for microshoot formation with the optimum concentration of 5, 10 and 20??M for Agric-rouge, Atinwewe, and Agblehoundo respectively. The protocol developed during this study will be useful for mass propagation of the elite cassava cultivars.

Project description:BackgroundThe study of transient gene expression in cassava plants during virus infection using existing protocols is laborious and may take approximately fifteen weeks due to cassava's recalcitrance to transformation. The combination of a protoplast system with CRISPR-mediated gene editing promises to shorten the turnaround time from plant tissue culture to high-throughput gene expression screening for candidate genes. Here, we detail a protocol for screening genes associated with the response to South African cassava mosaic virus (SACMV) in cassava protoplasts, with reference to the ubiquitin E3 ligase gene, MeE3L.MethodsCassava protoplasts of model, and SACMV-susceptible and -tolerant genotypes, were transformed with SACMV infectious clones and/or a CRISPR-editing construct targeting the MeE3L using PEG4000-mediated transfection. DNA and RNA were extracted from transformed protoplasts at 24 h post-transfection. Relative SACMV DNA accumulation was determined via qPCR using DpnI-digested total DNA, MeE3L relative expression was determined via reverse transcriptase qPCR, and results were analysed using one-way ANOVA, Tukey's HSD test and the 2-ΔΔCTstatistical method. The MeE3L exonic region was sequenced on the ABI 3500XL Genetic Analyzer platform; and sequences were analysed for mutations using MAFTT and MEGA-X software. Construction of a phylogenetic tree was done using the Maximum Likelihood method and Jones-Taylor-Thornton (JTT) matrix-based model.ResultsThe differential expression of unedited and mutant MeE3L during SACMV infection of model, susceptible and tolerant cassava protoplasts was determined within 7 weeks after commencement of tissue culture. The study also revealed that SACMV DNA accumulation in cassava protoplasts is genotype-dependent and induces multiple mutations in the tolerant landrace MeE3L homolog. Notably, the susceptible cassava landrace encodes a RINGless MeE3Lwhich is silenced by SACMV-induced mutations. SACMV also induces mutations which silence the MeE3L RING domain in protoplasts from and tolerant cassava landraces.ConclusionsThis protocol presented here halves the turnaround time for high-throughput screening of genes associated with the host response to SACMV. It provides evidence that a cassava E3 ligase is associated with the response to SACMV and forms a basis for validation of these findings by in planta functional and interaction studies.

Project description:In susceptible plant hosts, co-evolution has favoured viral strategies to evade host defenses and utilize resources to their own benefit. The degree of manipulation of host gene expression is dependent on host-virus specificity and certain abiotic factors. In order to gain insight into global transcriptomic changes for a geminivirus pathosystem, South African cassava mosaic virus [ZA:99] (SACMV-ZA:99]) and Arabidopsis thaliana, 4 x 44K Agilent microarrays were adopted. After normalization, a 2-fold change filtering of data (p<0.05) identified 1,820 differentially expressed genes in apical leaf tissue. A significant increase in differential gene expression over time (451 genes at 14 dpi, 742 genes at 24 dpi, and 1011 genes at 36 dpi) was observed. This increase in expression, correlated with an increase in SACMV accumulation as virus copies were 5-fold higher at 24 dpi and 6-fold higher at 36 dpi than at 14 dpi (1.1x104 virus copies present at 14 dpi, 5.7x104 copies at 24 dpi, and 6.3x104 copies at 36 dpi). Many 2-fold genes were primarily involved in stress and defense responses, phytohormone signalling pathways, cellular transport, cell-cycle regulation, transcription, oxidation-reduction, and other metabolic processes. Forty-one genes (2.3%) were shown to be continuously expressed across the infection period, indicating that the majority of genes were transient and unique to a particular time point. Plant signalling networks were disrupted and manipulated by SACMV-[ZA:99] in order to affect homeostasis and antagonize host’s defense responses. At the same time, an adaptive response was initiated to reprogramme metabolism and divert energy from growth-related processes to defense, all leading to disruption of normal biological host processes. Comparisons between SACMV-[ZA:99] with plant-infecting RNA and DNA viruses revealed similarities and differences in expression patterns among viruses, showing either general defense or virus-specific responses. Within the Geminiviridae family in particular, similarities in cell-cycle regulation and gene expression patterns correlated between SACMV-[ZA:99] and Cabbage leaf curl virus (CaLCuV) but differences were also evident. For instance, CaLCuV showed antagonistic interactions between Salicyclic Acid (SA) and Jasmonic Acid (JA) pathways, whereas SACMV displayed synergism. Differences in gene induction, repression and outcome between the two geminiviruses clearly demonstrated host-specific interactions with SACMV-[ZA:99] leading to infection. To our knowledge this is the first geminivirus study identifying differentially expressed transcripts across 3 time points

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Response of cassava cultivars to African cassava mosaic virus infection across a range of inoculum doses and plant ages.

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Response of cassava cultivars to African cassava mosaic virus infection across a range of inoculum doses and plant ages.

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