Improving disease resistance in plants by editing the epigenome

Pathogens rely on expression of host susceptibility (S) genes to promote infection and disease. As DNA methylation is an epigenetic modification that affects gene expression, blocking access to S genes through targeted methylation could increase disease resistance. Xanthomonas axonopodis pv. manihotis, the causal agent of cassava bacterial blight (CBB), uses transcription activator-like20 (TAL20) to induce expression of the S gene MeSWEET10a. We directed methylation to the TAL20 effector binding element within the MeSWEET10a promoter using a synthetic zinc-finger DNA binding domain fused to a component of the RNA-directed DNA methylation pathway. We demonstrate that this methylation prevents TAL20 binding, blocks transcriptional activation of MeSWEET10a in vivo and that these plants display increased resistance to CBB while maintaining normal growth and development. This work establishes epigenome editing as a new strategy for crop improvement.

Resequencing of 388 cassava accessions identifies valuable loci and selection for variation in heterozygosity

Background Heterozygous genomes are widespread in outcrossing and clonally propagated crops. However, the variation in heterozygosity underlying key agronomic traits and crop domestication remains largely unknown. Cassava is a staple crop in Africa and other tropical regions and has a highly heterozygous genome. Results We describe a genomic variation map from 388 resequenced genomes of cassava cultivars and wild accessions. We identify 52 loci for 23 agronomic traits through a genome-wide association study. Eighteen allelic variations in heterozygosity for nine candidate genes are significantly associated with seven key agronomic traits. We detect 81 selective sweeps with decreasing heterozygosity and nucleotide diversity, harboring 548 genes, which are enriched in multiple biological processes including growth, development, hormone metabolisms and responses, and immune-related processes. Artificial selection for decreased heterozygosity has contributed to the domestication of the large starchy storage root of cassava. Selection for homozygous GG allele in MeTIR1 during domestication contributes to increased starch content. Selection of homozygous AA allele in MeAHL17 is associated with increased storage root weight and cassava bacterial blight (CBB) susceptibility. We have verified the positive roles of MeTIR1 in increasing starch content and MeAHL17 in resistance to CBB by transient overexpression and silencing analysis. The allelic combinations in MeTIR1 and MeAHL17 may result in high starch content and resistance to CBB. Conclusions This study provides insights into allelic variation in heterozygosity associated with key agronomic traits and cassava domestication. It also offers valuable resources for the improvement of cassava and other highly heterozygous crops.

Seasonal Variation on the Incidence and Severity of Major Folia Diseases of Cassava in Sierra Leone

A diagnostic survey was conducted in the rainy and dry seasons from 2014 to 2015 to determine the incidence and severity of major diseases of cassava in Sierra Leone. At least three chiefdoms and five villages per district were targeted. The survey was carried out in fourteen districts of the country with geo references using a GPS. On the spot assessment was conducted in all fields. Prevalence, severity and incidence were calculated. The most dominant diseases included the cassava mosaic disease and the cassava bacterial blight. The prevalence of cassava mosaic disease was high with 69.1% and 61.5% in the rainy season and dry season, respectively. The prevalence of cassava bacterial blight was 100% and 92% in the rainy season and dry season, respectively. Diseases of less importance included white spot and brown spot diseases as well as cassava anthracnose disease. The study provides bases for the deployment of improved varieties and provides information on the seasonal prevalence, incidence and severity of cassava diseases in Sierra Leone.

Reaction of Sweet Cassava Genotypes to Xanthomonas phaseoli pv. manihotis From Three Regions of Brazil

Xanthomonas phaseoli pv. manihotis (Xpm) is the causal agent of Cassava Bacterial Blight (CBB), one of the most important cassava diseases. The aim of this study was to evaluate the susceptibility of sweet cassava genotypes to strains of Xpm from three different geographic regions of Brazil in greenhouse conditions. The inoculation of 14 genotypes of cassava was made by cutting leaflets using scissors previously soaked in a bacterial suspension with 1 × 108 UFC ml-1 and also by inserting a soaked toothpick into the bud of the oldest leaf. The results showed significant differences when the cassava genotypes were individually evaluated in relation to the Xpm strains used; however, the relationship between cassava genotypes and Xpm strains was significant for wilt symptom. The UnB 1111 strain was more aggressive than the UnB 1386 strain based on the average value of the reaction grade, showing the variation that exists between the bacterial isolates from different regions. Considering the reaction of cassava germplasm’s resistance to the three strains used in the study, the BGMC 434 genotype was the only one classified as resistant based on the average reaction grade. The genotypes BGMC 753, BGMC 1289, BGMC 982 and the elite clones BRS 396, BRS 397, BRS 398, 259/08 and BRS 399 were classified as moderately resistant, which indicates the possibility of recommending them for disease favorable regions.

Prevalence and Distribution of Cassava Bacterial Blight in the Kenyan Coast

Cassava (Manihot esculenta Crantz) is one of the staple food crops grown in Kenya. Diseases remain one of the major constraints for cassava production. Apart from other major viral diseaes Cassava mosaic and Cassava brown streak, Cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv manihotis and Xanthomonas axonopodis pv cassavae are a major constraint in cassava production in Kenya. This study was done to identify the prevalence, distribution, and farmers' knowledge of cassava bacterial blight in the coastal region of Kenya. A survey was conducted involving 250 farmers who were randomly selected from two regions of Kilifi and Taita Taveta counties. Among the 250 farmers interviewed, 61.6 % identified cassava bacterial blight symptoms in their farms. The main varieties found growing in the region were Tajirika, Karembo, Kibandameno, and Shibe which were all confirmed by farmers as susceptible to cassava bacterial blight. During the survey, plant samples were randomly collected in the field. Out of the 70 samples collected, 40 of them were confirmed positive with X.pv manihotis and X.pv cassavae which cause cassava bacterial blight. The study concluded that there is a widespread of cassava bacterial blight in Kilifi and Taita taveta counties. Kilifi County had the highest incidence of 22% with Taita Taveta having the lowest incidence of 13%. Kilifi County had a higher severity of 8% as compared to Taita Taveta which had 5% Severity. Therefore there is a need for a proper management program to be deployed in managing the disease to enhance cassava production in the region.

TAL Effector Repertoires of Strains of Xanthomonas phaseoli pv. manihotis in Commercial Cassava Crops Reveal High Diversity at the Country Scale

Transcription activator-like effectors (TALEs) play a significant role for pathogenesis in several xanthomonad pathosystems. Xanthomonas phaseoli pv. manihotis (Xpm), the causal agent of Cassava Bacterial Blight (CBB), uses TALEs to manipulate host metabolism. Information about Xpm TALEs and their target genes in cassava is scarce, but has been growing in the last few years. We aimed to characterize the TALE diversity in Colombian strains of Xpm and to screen for TALE-targeted gene candidates. We selected eighteen Xpm strains based on neutral genetic diversity at a country scale to depict the TALE diversity among isolates from cassava productive regions. RFLP analysis showed that Xpm strains carry TALomes with a bimodal size distribution, and affinity-based clustering of the sequenced TALEs condensed this variability mainly into five clusters. We report on the identification of 13 novel variants of TALEs in Xpm, as well as a functional variant with 22 repeats that activates the susceptibility gene MeSWEET10a, a previously reported target of TAL20Xam668. Transcriptomics and EBE prediction analyses resulted in the selection of several TALE-targeted candidate genes and two potential cases of functional convergence. This study provides new bases for assessing novel potential TALE targets in the Xpm–cassava interaction, which could be important factors that define the fate of the infection.

Gene tagging via CRISPR-mediated homology-directed repair in cassava

Research on a few model plant–pathogen systems has benefitted from years of tool and resource development. This is not the case for the vast majority of economically and nutritionally important plants, creating a crop improvement bottleneck. Cassava bacterial blight (CBB), caused by Xanthomonas axonopodis pv. manihotis (Xam), is an important disease in all regions where cassava (Manihot esculenta Crantz) is grown. Here, we describe the development of cassava that can be used to visualize one of the initial steps of CBB infection in vivo. Using CRISPR-mediated homology-directed repair (HDR), we generated plants containing scarless insertion of GFP at the 3’ end of CBB susceptibility (S) gene MeSWEET10a. Activation of MeSWEET10a-GFP by the transcription activator-like (TAL) effector TAL20 was subsequently visualized at transcriptional and translational levels. To our knowledge, this is the first such demonstration of HDR via gene editing in cassava.

Assessment of yield components of some cassava (Manihot esculenta Crantz) genotypes using multivariate analysis such as path coefficients

Cassava, which is a tropical storage root crop, serves as a veritable source of carbohydrate for people living in Sub-Saharan Africa. It is also an important source of industrial starch, biopolymers, animal feeds and ethanol. However, the genetic improvement of the crop is impeded by dearth in information on its genetic variability. A field experiment was carried out at the National Root Crops Research Institute, Umudike (05° 29′ N; 07° 33′ E; 122 m a.s.l.), Nigeria, aimed at assessing the interrelationship between fresh root yield and thirteen quantitative yield-related characters of twenty-eight (28) newly developed cassava genotypes. The rain-fed experiment was laid out in a randomized complete block design with dual replications in 2016/2017 cropping seasons. The results indicated that the newly developed cassava genotypes [2] NR110315 followed by [20] B1-5 and [1] NR110238 exhibited (P < 0.05) highest fresh root yield among the twenty-eight genotypes tested. The fresh root yield of [2] NR110315 (18.77 Mt ha−1) was relatively higher compared with [18] NR050080, which gave the least root yield of 5.38 Mt ha−1. The correlation coefficients obtained from the associations among the variables were positive and exhibited highly significant (P ≤ 0.01) association, especially between leaf area index (LAI) and root diameter as well as with fresh root yield of cassava. Among the nine variables subjected to the path analysis, LAI, starch content, above ground dry matter (AGDM), weight of fresh marketable roots per plant and root length that contributed positively and directly to higher fresh root yield require greater attention during selection and breeding in cassava studies. The direct effect of the variables indicating their magnitudes in decreasing order showed that LAI exhibited the greatest effect followed by starch content, AGDM, weight of fresh marketable roots plant−1, root length, % cassava bacterial blight incidence and % Cassava mosaic disease incidence, which had the lowest direct effect on root yield of cassava. Cyanide potential exhibited a direct, weak and negative effect on fresh root yield. The results further suggested that cassava improvement could be achieved through these characters that have positive and highly significant magnitude effect on fresh root yield.