Peter Sseruwagi

Colonization of non-cassava plant species by cassava whiteflies (Bemisia tabaci) in Uganda

Bemisia tabaci (Genn.) (Homoptera: Aleyrodidae) is the vector of cassava mosaic geminiviruses (CMGs), which are the main production constraint to cassava [Manihot esculenta Crantz (Euphorbiaceae)], both in Uganda and elsewhere in Africa. Two B. tabaci genotype clusters, Ug1 and Ug2, differentiated at 8% nucleotide (nt) divergence within the mitochondrial cytochrome oxidase I (mtCOI) gene, have been shown to occur on cassava in Uganda. However, the role of alternative hosts in the ecology of cassava B. tabaci genotypes and their possible involvement in the epidemiology of cassava mosaic disease (CMD) in Uganda remain unknown. In this study, we investigated the restriction of cassava B. tabaci genotypes to cassava and the colonization of alternative host species in select cassava‐growing areas of the country in 2003 and 2004. Bemisia tabaci adults and 4th instar nymphs were collected from cassava and 11 other cultivated and uncultivated species occurring adjacent to the sampled cassava fields. Phylogenetic analysis of mtCOI sequences revealed that only a single genotype cluster, Ug1, was present on both cassava and non‐cassava plant species sampled in this study. The Ug1 genotypes (n = 49) shared 97–99% nt identity with the previously described cassava‐associated B. tabaci populations in southern Africa, and were ∼8% and ∼13% divergent from Ug2 and the ‘Ivory Coast cassava’ genotypes in Uganda and Ivory Coast, respectively. The Ug1 genotypes occurred (as adults) on all 12 source‐plant species sampled. However, based on the presence of B. tabaci 4th instar nymphs, the Ug1 genotypes (n = 13) colonized cassava and five other non‐cassava plant species: Manihot glaziovii, Jatropha gossypifolia, Euphorbia heterophylla, Aspilia africana, and Abelmoschus esculentus, suggesting that cassava B. tabaci (Ug1 genotypes) are not restricted to cassava in Uganda. No Ug2‐like genotypes were detected on any of the plant species sampled, including cassava, in this study. The identification of additional hosts for at least one genotype cluster, Ug1, known also to colonize cassava, and which was hitherto thought to be ‘cassava‐restricted’ may have important epidemiological significance for the spread of CMGs in Uganda.

Analyses of Twelve New Whole Genome Sequences of Cassava Brown Streak Viruses and Ugandan Cassava Brown Streak Viruses from East Africa: Diversity, Supercomputing and Evidence for Further Speciation

Cassava brown streak disease is caused by two devastating viruses, Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) which are frequently found infecting cassava, one of sub-Saharan Africa’s most important staple food crops. Each year these viruses cause losses of up to

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

100 million USD and can leave entire families without their primary food source, for an entire year. Twelve new whole genomes, including seven of CBSV and five of UCBSV were uncovered in this research, doubling the genomic sequences available in the public domain for these viruses. These new sequences disprove the assumption that the viruses are limited by agro-ecological zones, show that current diagnostic primers are insufficient to provide confident diagnosis of these viruses and give rise to the possibility that there may be as many as four distinct species of virus. Utilizing NGS sequencing technologies and proper phylogenetic practices will rapidly increase the solution to sustainable cassava production.

Cassava brown streak virus has a rapidly evolving genome: implications for virus speciation, variability, diagnosis and host resistance

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.

African ancestry of New World, Bemisia tabaci-whitefly species

Cassava is a major staple food for about 800 million people in the tropics and sub-tropical regions of the world. Production of cassava is significantly hampered by cassava brown streak disease (CBSD), caused by Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV). The disease is suppressing cassava yields in eastern Africa at an alarming rate. Previous studies have documented that CBSV is more devastating than UCBSV because it more readily infects both susceptible and tolerant cassava cultivars, resulting in greater yield losses. Using whole genome sequences from NGS data, we produced the first coalescent-based species tree estimate for CBSV and UCBSV. This species framework led to the finding that CBSV has a faster rate of evolution when compared with UCBSV. Furthermore, we have discovered that in CBSV, nonsynonymous substitutions are more predominant than synonymous substitution and occur across the entire genome. All comparative analyses between CBSV and UCBSV presented here suggest that CBSV may be outsmarting the cassava immune system, thus making it more devastating and harder to control.

Two Novel DNAs that enhance symptoms and overcome CMD2 resistance to cassava mosaic disease

Bemisia tabaci whitefly species are some of the world’s most devastating agricultural pests and plant-virus disease vectors. Elucidation of the phylogenetic relationships in the group is the basis for understanding their evolution, biogeography, gene-functions and development of novel control technologies. We report here the discovery of five new Sub-Saharan Africa (SSA) B. tabaci putative species, using the partial mitochondrial cytochrome oxidase 1 gene: SSA9, SSA10, SSA11, SSA12 and SSA13. Two of them, SSA10 and SSA11 clustered with the New World species and shared 84.8‒86.5% sequence identities. SSA10 and SSA11 provide new evidence for a close evolutionary link between the Old and New World species. Re-analysis of the evolutionary history of B. tabaci species group indicates that the new African species (SSA10 and SSA11) diverged from the New World clade c. 25 million years ago. The new putative species enable us to: (i) re-evaluate current models of B. tabaci evolution, (ii) recognise increased diversity within this cryptic species group and (iii) re-estimate divergence dates in evolutionary time.

Cassava mosaic disease and associated viruses in Zambia: occurrence and distribution

ABSTRACT Cassava mosaic begomoviruses (CMBs) cause cassava mosaic disease (CMD) across Africa and the Indian subcontinent. Like all members of the geminivirus family, CMBs have small, circular single-stranded DNA genomes. We report here the discovery of two novel DNA sequences, designated SEGS-1 and SEGS-2 (for sequences enhancing geminivirus symptoms), that enhance symptoms and break resistance to CMD. The SEGS are characterized by GC-rich regions and the absence of long open reading frames. Both SEGS enhanced CMD symptoms in cassava (Manihot esculenta Crantz) when coinoculated with African cassava mosaic virus (ACMV), East African cassava mosaic Cameroon virus (EACMCV), or East African cassava mosaic virus-Uganda (EACMV-UG). SEGS-1 also overcame resistance of a cassava landrace carrying the CMD2 resistance locus when coinoculated with EACMV-UG. Episomal forms of both SEGS were detected in CMB-infected cassava but not in healthy cassava. SEGS-2 episomes were also found in virions and whiteflies. SEGS-1 has no homology to geminiviruses or their associated satellites, but the cassava genome contains a sequence that is 99% identical to full-length SEGS-1. The cassava genome also includes three sequences with 84 to 89% identity to SEGS-2 that together encompass all of SEGS-2 except for a 52-bp region, which includes the episomal junction and a 26-bp sequence related to alphasatellite replication origins. These results suggest that SEGS-1 is derived from the cassava genome and facilitates CMB infection as an integrated copy and/or an episome, while SEGS-2 was originally from the cassava genome but now is encapsidated into virions and transmitted as an episome by whiteflies. IMPORTANCE Cassava is a major crop in the developing world, with its production in Africa being second only to maize. CMD is one of the most important diseases of cassava and a serious constraint to production across Africa. CMD2 is a major CMD resistance locus that has been deployed in many cassava cultivars through large-scale breeding programs. In recent years, severe, atypical CMD symptoms have been observed occasionally on resistant cultivars, some of which carry the CMD2 locus, in African fields. In this report, we identified and characterized two DNA sequences, SEGS-1 and SEGS-2, which produce similar symptoms when coinoculated with cassava mosaic begomoviruses onto a susceptible cultivar or a CMD2-resistant landrace. The ability of SEGS-1 to overcome CMD2 resistance and the transmission of SEGS-2 by whiteflies has major implications for the long-term durability of CMD2 resistance and underscore the need for alternative sources of resistance in cassava.

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

Cassava mosaic disease (CMD) is caused by cassava mosaic begomoviruses (CMBs), which are transmitted by the whitefly (Bemisia tabaci) in sub-Saharan Africa. CMD and the pathogenic viruses associated with cassava were assessed in a country-wide survey in seven provinces of Zambia between April and May 2009. CMD incidence was highest in Northwestern (71.2%) and lowest in Western (34.3%) provinces. Disease symptoms were severe in Eastern (3.94) and Lusaka (3.88), moderate in Central (3.54), Luapula (3.48) and Northern (3.31) and mild in Northwestern (3.01) and Western (2.50) provinces. In addition, the numbers of adult whitefly were assessed on cassava fields, and found to be highest in Lusaka (2.12) and lowest in Central (0.02) province. Polymerase chain reaction detected two virus species: African cassava mosaic virus (ACMV) and East African cassava mosaic virus (EACMV), that occurred as single and dual infections in 65.4% (ACMV), 25% (EACMV) and 9.6% (ACMV + EACMV) of the positive reactions. None of the samples were positive for EACMV-Ug. This is the first comprehensive report of CMD and the pathogenic viruses infecting cassava in Zambia.

Review and guide to a future naming system of African Bemisia tabaci species: African Bemisia tabaci species

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.

Cassava whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) in East African farming landscapes: A review of the factors determining abundance

Once a pest has been correctly identified, its genus and species name can provide a link to valuable indications of its ecology, biology and life history that are critical for developing control strategies. Importantly, this link should exist even when the pest was known under other names (synonyms), or was not considered a pest at all (National Research Council, 1968). Many examples have shown that incorrect identification or classification of a pest has led to fruitless searches for biocontrol agents in the native range, incorrect assignments as disease vectors, and costly, yet misdirected, suppression measures. As new approaches for delimiting species based on molecular information become more widely used, the process of correctly identifying a species has become even more complex. Fortunately, we have good systematic frameworks and nomenclatural systems that are able to cope with these challenges. Here we review challenges associated with classification and identification within the Bemisia tabaci (Gennadius) species complex. These pests and the viruses they transmit have emerged in the past few decades as among the most damaging to food and fibre crops globally (Varma & Malathi, 2003; Pimental et al., 2005; Seal et al., 2006), especially in sub‐Saharan Africa (SSA). The systematics of the B. tabaci species group has been a highly debated topic for years (Boykin, 2014). Putative species are indistinguishable morphologically, so other biological data have been collected to investigate the species in the complex. Based on genetic differences (Colvin et al., 2004; Sseruwagi et al., 2005; Boykin et al., 2007; Boykin et al., 2013; Hsieh et al., 2014) and mating incompatibility (Colvin et al., 2004; Liu et al., 2007; Xu et al., 2010), B. tabaci is now recognized as a species complex that consists of at least 34 putative species (Boykin et al., 2012). The rapid discovery of significant species diversity has led to many changes in the informal names used over the last 10 years (Boykin, 2014), creating confusion in the literature.