Elijah Ateka

Genetic diversity and geographic distribution of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) genotypes associated with cassava in East Africa

The genetic variability of whitefly (Bemisia tabaci) species, the vectors of cassava mosaic begomoviruses (CMBs) in cassava growing areas of Kenya, Tanzania, and Uganda, was investigated through comparison of partial sequences of the mitochondria cytochrome oxidase I (mtCOI) DNA in 2010/11. Two distinct species were obtained including sub-Saharan Africa 1 (SSA1), comprising of two sub-clades (I and II), and a South West Indian Ocean Islands (SWIO) species. Among the SSA1, sub-clade I sequences shared a similarity of 97.8–99.7% with the published Uganda 1 genotypes, and diverged by 0.3–2.2%. A pairwise comparison of SSA1 sub-clade II sequences revealed a similarity of 97.2–99.5% with reference southern Africa genotypes, and diverged by 0.5–2.8%. The SSA1 sub-clade I whiteflies were widely distributed in East Africa (EA). In comparison, the SSA1 sub-clade II whiteflies were detected for the first time in the EA region, and occurred predominantly in the coast regions of Kenya, southern and coast Tanzania. They occurred in low abundance in the Lake Victoria Basin of Tanzania and were widespread in all four regions in Uganda. The SWIO species had a sequence similarity of 97.2–97.7% with the published Reunion sequence and diverged by 2.3–2.8%. The SWIO whiteflies occurred in coast Kenya only. The sub-Saharan Africa 2 whitefly species (Ug2) that was associated with the severe CMD pandemic in Uganda was not detected in our study.

Investigating the impact of rice blast disease on the livelihood of the local farmers in greater Mwea region of Kenya.

Rice is the most important cereal crop in Kenya coming third after maize and wheat. It forms a very important diet for a majority of families in Kenya. The demand for rice in Kenya has seen a dramatic increase over the last few years while production has remained low. This is because rice production has been faced by serious constraints notably plant diseases of which the most devastating is rice blast. Rice blast is known to cause approximately 60% -100% yield losses. It is caused by an Ascomycete fungus called Magnaporthe Oryzae. The aim of this study was to investigate the impact of rice blast disease on the livelihood of the local farmers in Greater Mwea region and develop a rice blast disease distribution map using GIS approach. The study methodology employed a questionnaire survey which were subjected to sample population of households in the 7 sections with 70 blocks within Mwea region. The collected data was analysed using SAS Version 9.1. Descriptive statistics were used to summarize the household characteristics, the farm characteristics and the farmers’ perceptions of rice blast disease. In the questionnaire, farmers’ response on whether they had been affected by rice blast disease and the total production per acreage was used to develop an attribute table with GPS points. The GPS points were interpolated to create a geographical distribution map of rice blast disease. From the research findings almost all the farmers’ had awareness and knowledge of rice blast disease, 98% of the farmers interviewed were aware of rice blast disease. Out of the 98% with knowledge and awareness 76% have been affected by the disease, while 24% have never been affected. Farmers attributed rice blast disease to a range of different causes, including excessive use of nitrogen fertilizer, water shortage, lack of proper drainage canal and due to climate change. Majority of the farmers interviewed (72%) did not engage themselves in any other socio-economic activity even after being affected by the rice blast disease. 15% opted to growing horticultural crops, 7% engaged in trading activities while 2% started livestock raring, wage earning and Boda boda business.

Differential response of cassava genotypes to infection by cassava mosaic geminiviruses

Highlights • Cassava genotypes respond differently to infection by cassava mosaic geminiviruses.• Cassava mosaic disease resistant loci prompt recovery from systemic infection.• CMD symptoms are directly correlated with contents of viral DNA and virus specific small RNAs.• CMD infected plants abundantly accumulate 21–24 nt of virus specific small RNAs.• VsRNAs heterogeneously map the entire virus genome in both polarities.

Diversity of Thrips Species and Vectors of Tomato Spotted Wilt Virus in Tomato Production Systems in Kenya

ABSTRACT Thrips have been recognized as primary vectors of tomato spotted wilt virus (TSWV) with Frankliniella occidentalis (Pergande) reported as the most important and efficient vector, while other species such as Thrips tabaci Lindeman also include populations that can vector the virus. A study was undertaken to establish the diversity of thrips and presence of vectors for TSWV in four major tomato production areas in Kenya. The cytochrome oxidase 1 (CO1) gene was used to generate sequences from thrips samples collected from tomatoes and weeds, and phylogenetic analysis done to establish the variation within potential vector populations. Ceratothripoides brunneus Bagnall was the predominant species of thrips in all areas. F. occidentalis and T. tabaci were abundant in Nakuru, Kirinyaga, and Loitokitok but not detected at Bungoma. Other vectors of tospoviruses identified in low numbers were Frankliniella schultzei (Trybom) and Scirtothrips dorsalis Hood. Variation was observed in T. tabaci, F. occidentalis, and F. schultzei. Kenyan specimens of T. tabaci from tomato belonged to the arrhenotokous group, while those of F. occidentalis clustered with the Western flower thrips G group. The detection of RNA of TSWV in both of these species of thrips supported the role they play as vectors. The study has demonstrated the high diversity of thrips species in tomato production and the occurrence of important vectors of TSWV and other tospoviruses.

Phylogenetic Relationships among Whiteflies in the Bemisia tabaci (Gennadius) Species Complex from Major Cassava Growing Areas in Kenya

Whiteflies, Bemisia tabaci (Gennadius) are major insect pests that affect many crops such as cassava, tomato, beans, cotton, cucurbits, potato, sweet potato, and ornamental crops. Bemisia tabaci transmits viral diseases, namely cassava mosaic and cassava brown streak diseases, which are the main constraints to cassava production, causing huge losses to many small-scale farmers. The aim of this work was to determine the phylogenetic relationships among Bemisia tabaci species in major cassava growing areas of Kenya. Surveys were carried out between 2013 and 2015 in major cassava growing areas (Western, Nyanza, Eastern, and Coast regions), for cassava mosaic disease (CMD) and cassava brown streak disease (CBSD). Mitochondrial cytochrome oxidase I (mtCOI-DNA) was used to determine the genetic diversity of B. tabaci. Phylogenetic trees were constructed using Bayesian methods to understand the genetic diversity across the study regions. Phylogenetic analysis revealed two B. tabaci species present in Kenya, sub-Saharan Africa 1 and 2 comprising five distinct clades (A–E) with percent sequence similarity ranging from 97.7 % to 99.5%. Clades B, C, D, and E are predominantly distributed in the Western and Nyanza regions of Kenya whereas clade B is dominantly found along the coast, the eastern region, and parts of Nyanza. Our B. tabaci clade A groups with sub-Saharan Africa 2-(SSA2) recorded a percent sequence similarity of 99.5%. In this study, we also report the identification of SSA2 after a 15 year absence in Kenya. The SSA2 species associated with CMD has been found in the Western region of Kenya bordering Uganda. More information is needed to determine if these species are differentially involved in the epidemiology of the cassava viruses.

Unusual occurrence of a DAG motif in the Ipomovirus Cassava brown streak virus and implications for its vector transmission

Cassava is the main staple food for over 800 million people globally. Its production in eastern Africa is being constrained by two devastating Ipomoviruses that cause cassava brown streak disease (CBSD); Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV), with up to 100% yield loss for smallholder farmers in the region. To date, vector studies have not resulted in reproducible and highly efficient transmission of CBSV and UCBSV. Most virus transmission studies have used Bemisia tabaci (whitefly), but a maximum of 41% U/CBSV transmission efficiency has been documented for this vector. With the advent of next generation sequencing, researchers are generating whole genome sequences for both CBSV and UCBSV from throughout eastern Africa. Our initial goal for this study was to characterize U/CBSV whole genomes from CBSD symptomatic cassava plants sampled in Kenya. We have generated 8 new whole genomes (3 CBSV and 5 UCBSV) from Kenya, and in the process of analyzing these genomes together with 26 previously published sequences, we uncovered the aphid transmission associated DAG motif within coat protein genes of all CBSV whole genomes at amino acid positions 52–54, but not in UCBSV. Upon further investigation, the DAG motif was also found at the same positions in two other Ipomoviruses: Squash vein yellowing virus (SqVYV), Coccinia mottle virus (CocMoV). Until this study, the highly-conserved DAG motif, which is associated with aphid transmission was only noticed once, in SqVYV but discounted as being of minimal importance. This study represents the first comprehensive look at Ipomovirus genomes to determine the extent of DAG motif presence and significance for vector relations. The presence of this motif suggests that aphids could potentially be a vector of CBSV, SqVYV and CocMov. Further transmission and ipomoviral protein evolutionary studies are needed to confirm this hypothesis.

Evolutionary insights into Bean common mosaic necrosis virus and Cowpea aphid borne mosaic virus using global isolates and thirteen new near complete genomes from Kenya

Plant viral diseases are one of the major limitations in legume production within sub Saharan Africa (SSA), as they account for up to 100 % in production losses within smallholder farms. In this study, field surveys were conducted in the western highlands of Kenya with viral symptomatic leaf samples collected. Subsequently, next-generation sequencing was carried out. The main aim was to gain insights into the selection pressure and evolutionary relationships of Bean common mosaic necrosis virus (BCMNV) and Cowpea aphid-borne mosaic virus (CABMV), within symptomatic common beans and cowpeas. Eleven near complete genomes of BCMNV and two for CABMV sequences were obtained from SSA. Bayesian phylogenomic analysis and tests for differential selection pressure within sites and across tree branches of the viral genomes was carried out. Three distinct well-supported clades were identified across the whole genome tree, and were in agreement with individual gene trees. Selection pressure analysis within sites and across phylogenetic branches suggested both viruses were evolving independently, but under strong purifying selection, with a slow evolutionary rate. These findings provide valuable insights on the evolution of BCMNV and CABMV genomes and their relationship to other viral genomes globally. These results will contribute greatly to the knowledge gap surrounding the phylogenomic relationship of these viruses, particularly for CABMV, for which there are few genome sequences available, and support the current breeding efforts towards resistance for BCMNV and CABMV.

Cassava brown streak disease in Rwanda, the associated viruses and disease phenotypes

Cassava brown streak disease (CBSD) was first observed on cassava (Manihot esculenta) in Rwanda in 2009. In 2014 eight major cassava‐growing districts in the country were surveyed to determine the distribution and variability of symptom phenotypes associated with CBSD, and the genetic diversity of cassava brown streak viruses. Distribution of the CBSD symptom phenotypes and their combinations varied greatly between districts, cultivars and their associated viruses. The symptoms on leaf alone recorded the highest (32.2%) incidence, followed by roots (25.7%), leaf + stem (20.3%), leaf + root (10.4%), leaf + stem + root (5.2%), stem + root (3.7%), and stem (2.5%) symptoms. Analysis by RT‐PCR showed that single infections of Ugandan cassava brown streak virus (UCBSV) were most common (74.2% of total infections) and associated with all the seven phenotypes studied. Single infections of Cassava brown streak virus (CBSV) were predominant (15.3% of total infections) in CBSD‐affected plants showing symptoms on stems alone. Mixed infections (CBSV + UCBSV) comprised 10.5% of total infections and predominated in the combinations of leaf + stem + root phenotypes. Phylogenetic analysis and the estimates of evolutionary divergence, using partial sequences (210 nt) of the coat protein gene, revealed that in Rwanda there is one type of CBSV and an indication of diverse UCBSV. This study is the first to report the occurrence and distribution of both CBSV and UCBSV based on molecular techniques in Rwanda.

Lanai: A small, fast growing tomato variety is an excellent model system for studying geminiviruses

Highlights • Florida Lanai is a tomato variety suitable for virus-host interaction studies.• Florida-Lanai was infected by geminiviruses delivered by different methods.• Florida-Lanai shows distinct measurable symptoms for different geminiviruses.• Florida-Lanai has a small size, rapid growth and is easy to maintain.• Florida-Lanai is an excellent choice for comparing geminivirus infections.

Efficacy of chemotherapy and thermotherapy in elimination of East African cassava mosaic virus from Tanzanian cassava landrace

Abstract Cassava mosaic disease is caused by cassava mosaic begomoviruses (CMBs) and can result in crop losses up to 100% in cassava (Manihot esculenta) in Tanzania. We investigated the efficacy of chemotherapy and thermotherapy for elimination of East African cassava mosaic virus (EACMV) of Tanzanian cassava. In vitro plantlets from EACMV‐infected plants obtained from coastal Tanzania were established in the greenhouse. Leaves were sampled from the plants and tested to confirm the presence of EACMV. Plantlets of plants positive for EACMV were initiated in Murashige and Skoog (MS) medium. On the second subculture, they were subjected into chemical treatment in the medium containing salicylic acid (0, 10, 20, 30 and 40 mg/L) and ribavirin (0, 5, 10, 15 and 20 mg/L). In the second experiment, EACMV‐infected plantlets were subjected to temperatures between 35 and 40°C with 28°C as the control. After 42 days of growth, DNA was extracted from plant leaves and PCR amplification was performed using EACMV specific primers. It was found that plant survival decreased with increasing levels of both salicylic acid and ribavirin concentrations. In general, plants treated with salicylic acid exhibited a lower plant survival % than those treated with ribavirin. However, the percentage of virus‐free plants increased with an increase in the concentration of both ribavirin and salicylic acid. The most effective concentrations were 20 mg/L of ribavirin and 30 mg/L of salicylic acid; these resulted in 85.0% and 88.9% virus‐free plantlets, respectively. With regard to thermotherapy, 35°C resulted in 79.5% virus‐free plantlets compared to 69.5% at 40°C. Based on virus elimination, ribavirin at 20 mg/L, salicylic acid 30 mg/L and thermotherapy at 35°C are recommended for production of EACMV free cassava plantlets from infected cassava landraces.