East African Cassava Mosaic virus-Uganda Research Articles

An Alarming Eastward Front of Cassava Mosaic Disease in Development in West Africa

Begomoviruses are a major threat to cassava production in Africa. Indeed, during the 1990s, the emergence of a recombinant begomovirus (East African cassava mosaic virus-Uganda, EACMV-Ug) resulted in crop devastation and severe famine in Uganda. In 2023, during a pre-survey of cassava farms at Forécariah, South-West Guinea, 22 samples showing peculiar cassava mosaic disease (CMD) symptoms were collected, and subsequent laboratory analysis confirmed the presence of EACMV-Ug in the samples. Deep analysis of DNA-A and DNA-B of the EACMV-Ug isolates from Guinea indicated that they are similar to those associated with the severe CMD epidemic in Uganda in the 1990s. Therefore, a country-wide survey was conducted throughout Guinea in April 2024 to evaluate the extent of the spread of EACMV-Ug in the country and to collect critical CMD epidemiological data. Findings showed a high whitefly population in Lower Guinea averaging 17 per plant; however, the data suggest a spread of EACMV-Ug via infected cuttings. High CMD incidence was found in Lower Guinea and Forest Guinea, whereas the highest CMD severity was observed in Forest Guinea (2.70 ± 0.06) and the lowest CMD severity was found in Middle Guinea (2.20 ± 0.05). Several cases of double and triple infections involving African cassava mosaic virus, East African cassava mosaic virus, East African cassava mosaic Cameroon virus, and EACMV-Ug were observed. EACMV-Ug was detected throughout Guinea, as well as from samples collected in 2022 in Kambia (Sierra Leone). The 63 accessions cultivated in Guinea that were assessed in this study were found susceptible to at least one of the viruses cited above. This study alerts us to an alarming situation in development in West Africa and provides scientific evidence to guide the rapid response needed to contain and stop the progression of EACMV-Ug in West Africa.

Application of Integrated Computational Approaches in Prediction of Plant Virus Encoded miRNAs and Their Targeted Plant Genes.

This chapter presents a comprehensive approach to predict novel miRNAs encoded by plant viruses and identify their target plant genes, through integration of various ab initio computational approaches. The predictive process begins with the analysis of plant viral sequences using the VMir Analyzer software. VMir Viewer software is then used to extract primary hairpins from these sequences. To distinguish real miRNA precursors from pseudo miRNA precursors, MiPred web-based software is employed. Verified real pre-miRNA sequences with a minimum free energy of< -20 Kcal/mol, arefurther analyzedusingthe RNAshapes software. Validation of predictions involves comparing them with available Expressed Sequence Tags (ESTs) from the relevant plant using BlastN. Short sequences with lengths ranging from 19 to 25 nucleotides and exhibiting <5 mismatches are prioritized for miRNA prediction. The precise locations of these short sequences within pre-miRNA structures generated using RNAshapes are meticulously identified, with a focus on those situated on the 5' and 3' arms of the structures, indicating potential miRNAs. Sequences within the arms of pre-miRNA structures are used to predict target sites within the ESTs of the specific plant, facilitated by psRNA Target software, revealing genes with potential regulatory roles in the plant. To confirm the outcome of target prediction, results are individually submitted to the RNAhybrid web-based software. For practical demonstration, this approach is applied to analyze African cassava mosaic virus (ACMV) and East African cassava mosaic virus-Uganda (EACMV-UG) viruses, as well as the ESTs of Jatropha and cassava.

Cassava begomovirus species diversity changes during plant vegetative cycles.

Cassava is a root crop important for global food security and the third biggest source of calories on the African continent. Cassava production is threatened by Cassava mosaic disease (CMD), which is caused by a complex of single-stranded DNA viruses (family: Geminiviridae, genus: Begomovirus) that are transmitted by the sweet potato whitefly (Bemisia tabaci). Understanding the dynamics of different cassava mosaic begomovirus (CMB) species through time is important for contextualizing disease trends. Cassava plants with CMD symptoms were sampled in Lake Victoria and coastal regions of Kenya before transfer to a greenhouse setting and regular propagation. The field-collected and greenhouse samples were sequenced using Illumina short-read sequencing and analyzed on the Galaxy platform. In the field-collected samples, African cassava mosaic virus (ACMV), East African cassava mosaic virus (EACMV), East African cassava mosaic Kenya virus (EACMKV), and East African cassava mosaic virus-Uganda variant (EACMV-Ug) were detected in samples from the Lake Victoria region, while EACMV and East African mosaic Zanzibar virus (EACMZV) were found in the coastal region. Many of the field-collected samples had mixed infections of EACMV and another begomovirus. After 3 years of regrowth in the greenhouse, only EACMV-like viruses were detected in all samples. The results suggest that in these samples, EACMV becomes the dominant virus through vegetative propagation in a greenhouse. This differed from whitefly transmission results. Cassava plants were inoculated with ACMV and another EACMV-like virus, East African cassava mosaic Cameroon virus (EACMCV). Only ACMV was transmitted by whiteflies from these plants to recipient plants, as indicated by sequencing reads and copy number data. These results suggest that whitefly transmission and vegetative transmission lead to different outcomes for ACMV and EACMV-like viruses.

The effect of competition on the control of invading plant pathogens.

New invading pathogen strains must compete with endemic pathogen strains to emerge and spread. As disease control measures are often non‐specific, that is, they do not distinguish between strains, applying control not only affects the invading pathogen strain but the endemic as well. We hypothesize that the control of the invasive strain could be compromised due to the non‐specific nature of the control.A spatially explicit model, describing the East African cassava mosaic virus‐Uganda strain (EACMV‐UG) outbreak, is used to evaluate methods of controlling both disease incidence and spread of invading pathogen strains in pathosystems with and without an endemic pathogen strain present.We find that while many newly introduced or intensified control measures (such as resistant cultivars or roguing) decrease the expected incidence, they have the unintended consequence of increasing, or at least not reducing, the speed with which the invasive pathogen spreads geographically. We identify the controls that cause this effect and methods in which these controls may be applied to prevent it.We found that the spatial spread of the invading strain is chiefly governed by the incidence at the wave front. Control can therefore be applied, or intensified, once the wave front has passed without increasing the pathogen's rate of spread.When trade of planting material occurs, it is possible that the planting material is already infected. The only forms of control in this study that reduces the speed of geographic spread, regardless of the presence of an endemic strain, are those that reduce the amount of trade and the distance over which trade takes place. Synthesis and applications. The best control strategy depends on the presence of competing endemic strains. Applying or intensifying the control can slow the rate of spread when absent but increase it if present. Imposing trade restrictions before the epidemic has reached a given area and intensifying other control methods only when the wave front has passed is the most effective way of both slowing down spread and controlling incidence when a competing endemic strain is present and is the safest approach when its presence is unknown.

First Report of East African Cassava Mosaic Virus-Uganda Infecting the Nile Tulip Tree in Western Kenya

Whitefly-transmitted cassava mosaic begomoviruses (CMBs; family Geminiviridae; genus Begomovirus) are viruses with bipartite genomes (components referred to as DNA-A and DNA-B) containing a conserved (between the two components) noncoding sequence known as the common region (CR) required for DNA replication and transcription (Fondong 2013). CMBs cause cassava mosaic disease (CMD), a major limitation to cassava production in sub-Saharan Africa. In Kenya, CMD is associated with the begomoviruses East African cassava mosaic virus (EACMV), African cassava mosaic virus (ACMV), East African cassava mosaic Kenya virus, and East African cassava mosaic Zanzibar virus, as well as several recombinant viruses, such as the virulent EACMV-Uganda (EACMV-UG) that so far occurs only in western Kenya (Nyongesa Were et al. 2016). Within the CMB-cassava pathosystem, various noncassava plant species may play an epidemiological role as virus reservoirs for the whitefly vector Bemisia tabaci (Alabi et al. 2008). In May 2017, leaf samples were collected from three plants exhibiting virus-like symptoms including mosaic, mottling, twisted leaves, and reduction in leaf surface area as well as from two nonsymptomatic plants belonging to the family Bignoniaceae growing in three cassava fields in western Kenya. Total genomic DNA was extracted from leaf samples using the ZR Plant/Seed DNA MiniPrep kit as per the manufacturer’s instructions (Zymo Research Corp., U.S.A.). The identity of the plant species was confirmed as Markhamia lutea by phenotypic characteristics and by polymerase chain reaction (PCR) amplification, Sanger sequencing, and BLASTn analysis of the chloroplast rubisco large-chain gene using the primer pair RbcLa-F (5′-ATGTCACCACAAACAGAGACTAAAGC-3′)/RbcLa-R (5′-GTAAAATCAAGTCCACCRCG-3′). Samples were screened by PCR with the diagnostic primer pairs JSP001/JSP002 for ACMV, JSP001/JSP003 for EACMV (Fondong et al. 2000), and UV AL1-F1/ACMV CP-R3 for EACMV-UG (Zhou et al. 1997). Samples from nonsymptomatic plants did not yield amplification products, confirming absence of viral infection. A 1,650-bp fragment of the expected size was amplified from one symptomatic sample with the primer pair UV AL1-F1/ACMV CP-R3, suggesting the presence of EACMV-UG. To confirm the positive PCR result, the genome of the virus was enriched by rolling-circle amplification (RCA) using a TempliPhi 100 RCA Kit (GE Healthcare, U.S.A.). The library was prepared using the NEBNext Ultra II DNA library prep kit for Illumina (New England Biolabs, U.S.A.) according to the manufacturer’s instructions and sequenced using an Illumina MiSeq platform with the MiSeq Reagent Kit version 3 (600 cycles) (Illumina, U.S.A.). Following de novo sequence assembly, 344 and 89 paired-end reads aligned to generate two contigs identified by BLASTn search as the DNA-A and DNA-B of a bipartite virus. The nucleotide sequences were deposited in GenBank as accession numbers MF974568 and MH379641. The 2,798-nucleotide (nt) DNA-A and 2,774-nt DNA-B had an approximately 160-nt EACMV-like CR with 53%-nt sequence identity. DNA-A shared 99% nucleotide sequence identity with the DNA-A of an EACMV-UG isolate (AJ717522) from western Kenya, whereas the DNA-B was closely related to another EACMV-UG isolate (AJ704962) from western Kenya at 97% nucleotide sequence identity. To the best of our knowledge, this is the first report of occurrence of EACMV-UG in a species of the family Bignoniaceae. Further investigations are required to determine the epidemiological significance of M. lutea in the CMB-cassava pathosystem.

Spatial Analysis of Temporal Changes in the Pandemic of Severe Cassava Mosaic Disease in Northwestern Tanzania.

To improve understanding of the dynamics of the cassava mosaic disease (CMD) pandemic front, geospatial approaches were applied to the analysis of 3 years' data obtained from a 2-by-2° (approximately 222-by-222 km) area of northwestern Tanzania. In total, 80 farmers' fields were assessed in each of 2009, 2010, and 2011, with 20 evenly distributed fields per 1-by-1° quadrant. CMD-associated variables (CMD incidence, CMD severity, vector-borne CMD infection, and vector abundance) increased in magnitude from 2009 to 2010 but showed little change from 2010 to 2011. Increases occurred primarily in the two westernmost quadrants of the study area. A pandemic "front" was defined by determining the values of CMD incidence and whitefly abundance where predicted disease gradients were greatest. The pandemic-associated virus (East African cassava mosaic virus-Uganda) and vector genotype (Bemisia tabaci sub-Saharan Africa 1-subgroup 1) were both present within the area bounded by the CMD incidence front but both also occurred ahead of the front. The average speed and direction of movement of the CMD incidence front (22.9 km/year; southeast) and whitefly abundance front (46.6 km/year; southeast) were calculated, and production losses due to CMD were estimated to range from US$4.3 million to 12.2 million.

Fitness costs associated with infections of secondary endosymbionts in the cassava whitefly species Bemisia tabaci

We investigated the dual effects of bacterial infections and diseased cassava plants on the fitness and biology of the Bemisia tabaci infesting cassava in Africa. Isofemale B. tabaci colonies of sub-Saharan Africa 1-subgroup 3 (SSA1-SG3), infected with two secondary endosymbiotic bacteria Arsenophonus and Rickettsia (AR+) and those free of AR infections (AR−), were compared for fitness parameters on healthy and East African cassava mosaic virus-Uganda variant (EACMV-UG)-infected cassava plants. The whitefly fecundity and nymph development was not affected by bacterial infections or the infection of cassava by the virus. However, emergence of adults from nymphs was 50 and 17% higher by AR− on healthy and virus-infected plants, respectively, than AR+ flies. Development time of adults also was 10 days longer in AR+ than AR−. The whiteflies were further compared for acquisition and retention of EACMV-UG. Higher proportion of AR− acquired (91.8%) and retained (87.6%) the virus than AR+ (71.8, 61.2%, respectively). Similarly, the AR− flies retained higher quantities of virus (~ninefold more) than AR+. These results indicated that bacteria-free whiteflies were superior and better transmitters of EACMV-UG, as they had higher adult emergence, quicker life cycle and better virus retention abilities than those infected with bacteria.

Virus versus host plant microRNAs: who determines the outcome of the interaction?

Considering the importance of microRNAs (miRNAs) in the regulation of essential processes in plant pathogen interactions, it is not surprising that, while plant miRNA sequences counteract viral attack via antiviral RNA silencing, viruses in turn have developed antihost defense mechanisms blocking these RNA silencing pathways and establish a counter-defense. In the current study, computational and stem-loop Reverse Transcription – Polymerase Chain Reaction (RT-PCR) approaches were employed to a) predict and validate virus encoded mature miRNAs (miRs) in 39 DNA-A sequences of the bipartite genomes of African cassava mosaic virus (ACMV) and East African cassava mosaic virus-Uganda (EACMV-UG) isolates, b) determine whether virus encoded miRs/miRs* generated from the 5′/3′ harpin arms have the capacity to bind to genomic sequences of the host plants Jatropha or cassava and c) investigate whether plant encoded miR/miR* sequences have the potential to bind to the viral genomes. Different viral pre-miRNA hairpin sequences and viral miR/miR* length variants occurring as isomiRs were predicted in both viruses. These miRNAs were located in three Open Reading Frames (ORFs) and in the Intergenic Region (IR). Moreover, various target genes for miRNAs from both viruses were predicted and annotated in the host plant genomes indicating that they are involved in biotic response, metabolic pathways and transcription factors. Plant miRs/miRs* from conserved and highly expressed families were identified, which were shown to have potential targets in the genome of both begomoviruses, representing potential plant miRNAs mediating antiviral defense. This is the first assessment of predicted viral miRs/miRs* of ACMV and EACMV-UG and host plant miRNAs, providing a reference point for miRNA identification in pathogens and their hosts. These findings will improve the understanding of host- pathogen interaction pathways and the function of viral miRNAs in Euphorbiaceous crop plants.

East African cassava mosaic virus‐Uganda (EACMV‐UG) and African cassava mosaic virus (ACMV) reported for the first time in Central African Republic and Chad

Cassava ( Manihot esculenta ) is a major staple food crop cultivated throughout Africa. It is affected most severely by cassava mosaic disease (CMD) at high incidence wherever it is grown on the continent. The disease may be caused by seven…

Occurrence and Distribution of Cassava Mosaic Begomovirus Related to Agro-ecosystems in the Sud-kivu Province, Democratic Republic of Congo

Cassava mosaic disease epidemiology and molecular characterization of the related viruses monitored during this study in the Sud-Kivu region, Democratic Republic of Congo. Collected epidemiologic data showed a negative correlation with the altitude (elevation) on the disease incidence and its vector population. Disease incidence was 9.5 to 37.8% associated to less than 3 insects per leaf when the elevation is higher than 1500 m while 65 to 100% and more than 10 insects per leaf were recorded where elevation was less than 1000 m. No impact of rainfall has been recorded. Associated molecular cassava begomovirus diagnostic focused on AC2 and AC4 genes revealed only two species occurring in this area, African Cassava Mosaic Virus (ACMV) and East African cassava mosaic virus-Uganda (EACMV-UG) with more EACMV-UG and mixed infections in low elevation area while ACMV were predominant in high elevation sites. Phylogeny analysis revealed a dual coexistence of indigenous and Ugandan spread among ACMV isolates while all EACMV isolates were restricted to Ugandan spread (95 to 100% of amino acids identity). No recombination was revealed in this study, isolates diversity was restricted to nucleotide substitution. © 2012 Academic Journals Inc.

Occurrence of African cassava mosaic virus (ACMV) and East African cassava mosaic virus – Uganda (EACMV-UG) in Jatropha curcas

Methods In this study we attempted to detect and molecularly characterize viruses infecting Jatropha in Eastern Africa (Kenya and Ethiopia). Detection methods will be valuable tools for early screening of plant viruses in order to make appropriate decisions and selection of planting material. A total of 127 Jatropha samples from Ethiopia and Kenya (districts: Kakamega, Siaya, Busia and Nakuru showing typical virus symptoms and symptomless plants were used in this study. ELISA was performed to detect the presence of three RNA viruses: CMV, Cassava common mosaic virus (CsCMV) and Cassava brown streak virus (CBSV). PCR was performed using newly designed primers based on multiple alignments of full length DNA A sequences of geminiviruses available in the NCBI Genbank, reported to infect either Jatropha or

Field Studies of Cross Protection with Cassava Mosaic Geminiviruses in Uganda

AbstractThe effect of cassava mosaic virus disease (CMD) was compared on plants grown from cuttings that were initially virus‐free at planting and those infected with a mild strain of East African cassava mosaic virus‐Uganda (EACMV‐UG). All initially healthy plants developed CMD symptoms within 5 months of planting (MAP) at both trial sites in Uganda, although spread was more rapid at Kamuli than at Serere. Significantly (P &lt; 0.001) higher symptom severity scores were recorded in initially healthy plants, which had average scores of 3.6 and 3.5 at Kamuli and Serere, respectively, compared with 2.8 for mildly diseased plants at each location. Severity scores of 4 and 5 were more frequent in initially healthy plants, accounting for 77 and 39% of the total infections recorded in comparison with 47 and 11% in mildly diseased plants at Kamuli and Serere, respectively. Mildly diseased plants were significantly taller than initially healthy plants 8 and 12 MAP at both locations. However, the converse was true 4 MAP although differences were significant at Serere but not at Kamuli. Mildly diseased plants yielded significantly more tuberous roots than initially healthy plants at Kamuli but not at Serere. Average total weights of tuberous roots per plant were 2.48 and 1.63 kg for mildly diseased and initially healthy plants at Kamuli and 4.46 and 4.61 kg at Serere, respectively. These results may help to explain the increased prevalence in recent years of mildly diseased plants of local CMD‐susceptible cultivars in eastern Uganda, from where these varieties virtually disappeared following the severe CMD epidemic in the 1990s. The results also provide the first field evidence of a cross protective effect of mild strains of a cassava mosaic geminivirus.