The symptom and genetic diversity of cassava brown streak viruses infecting cassava in East Africa

The genetic and symptom diversity of six virus isolates causing cassava brown streak disease (CBSD) in the endemic (Kenya, Mozambique, and Tanzania) and the recently affected epidemic areas (Uganda) of eastern Africa was studied. Five cassava varieties; Albert, Colombian, Ebwanateraka, TMS60444 (all susceptible) and Kiroba (tolerant) were graft inoculated with each isolate. Based on a number of parameters including the severity of leaf and root symptoms, and the extent of virus transmission by grafting, the viruses were classified as either severe or relatively mild. These results were further confirmed by the mechanical inoculation of 13 herbaceous hosts in which the virulent isolates caused plant death in Nicotiana clevelandii and N. benthamiana whereas the milder isolates did not. Phylogenetic analysis of complete coat protein gene sequences of these isolates together with sequences obtained from 14 other field-collected samples from Kenya and Zanzibar, and reference sequences grouped them into two distinct clusters, representing the two species of cassava brown streak viruses. Put together, these results did not suggest the association of a hypervirulent form of the virus with the current CBSD epidemic in Uganda. Identification of the severe and milder isolates, however, has further implications for disease management and quarantine requirements

Prevalence and genetic diversity of endosymbiotic bacteria infecting cassava whiteflies in Africa

Background: Cassava provides over half of the dietary requirement for more than 200 million poor in Africa. In recent years, cassava has been affected by an epidemic of a virus disease called cassava brown streak disease (CBSD) that is spreading in much of eastern and central Africa, affecting food security and the economic development of the poor. The viruses that cause CBSD are transmitted by the insect vector whitefly (Bemisia tabaci), which have increased to very high numbers in some African countries. Strains of endosymbiotic bacteria infecting whiteflies have been reported to interact specifically with different whitefly populations with varied effects on its host biology and efficiency of virus transmission. The main aim of this study was therefore to investigate the prevalence and diversity of the secondary endosymbiotic bacteria infecting cassava whiteflies with a view to better understand their role on insect population dynamics and virus disease epidemics. Results: The genetic diversity of field-collected whitefly from Tanzania, Malawi, Uganda and Nigeria was determined by mitochondrial DNA based phylogeny and restriction fragment length polymorphism. Cassava in these countries was infected with five whitefly populations, and each one was infected with different endosymbiotic bacteria. Incidences of Arsenophonus, Rickettsia, Wolbachia and Cardinium varied amongst the populations. Wolbachia was the most predominant symbiont with infection levels varying from 21 to 97%. Infection levels of Arsenophonus varied from 17 to 64% and that of Rickettsia was 0 to 53%. Hamiltonella and Fritschea were absent in all the samples. Multiple locus sequence typing identified four different strains of Wolbachia infecting cassava whiteflies. A common strain of Wolbachia infected the whitefly population Sub-Saharan Africa 1-subgroup 1 (SSA1-SG1) and SSA1-SG2, while others were infected with different strains. Phylogeny based on 16S rDNA of Rickettsia and 23S rDNA of Arsenophonus also identified distinct strains. Conclusions: Genetically diverse bacteria infect cassava whiteflies in Africa with varied prevalence across different host populations, which may affect their whitefly biology. Further studies are required to investigate the role of endosymbionts to better understand the whitefly population dynamics

Molecular identification and sequence analysis of bipartite Begomovirus infecting Horsegram legume in India

The molecular diversity of Begomovirus infecting Horsegram yellow Mosaic viruses (HgYMV1 and HgYMV2), from two main horsegram growing farms near Bangalore, Karnataka State, South India was investigated. The viral DNA was amplified from horsegram plants exhibiting mild and severe symptoms by polymerase chain reaction, and complete genome of the HgYMV were identified by their sequence analysis. Isolates of HgYMV1 and HgYMV2 were found to be associated with severe symptom phenotype from HgYMV. HgYMV was most closely related to Mungbean yellow mosaic indian viruse (MYMIV) and Mungbean yellow mosaic virus (MYMV) at 81.8 to 84.8 % nucleotide identity, based on DNA-A and DNA-B component sequences. HgYMV was distantly related to Dolicos yellow mosaic virus from Asia (DoYMV-Ban and DoYMV-DB) and partially to Cowpea golden mosaic virus from Nigeria (CPGMV-[NG]) at 64 and 62 % DNA nucleotide identity. Analysis of the DNA-B sequence of HgYMV revealed a DNA-B component identical to those of Bean golden yellow mosaic virus BGYMV isolates described. Furthermore, the DNA-B component for extant BGYMV isolates and strains were also the closest relatives for the HgYMV1 DNA-B components at 48.7 % nucleotide identity. Therefore, HgYMV could be considered to be a new species of the genus Begomovirus (family Geminiviridae).Keywords: Begomovirus, Horsegram, yellow Mosaic viruses, DNA sequencin

Testing the infectivity of a Begomovirus by particle bombardment method using a gene gun

This study was design to identify the causal agent of Horsegram yellow mosaic disease and to investigate the pathogenicity of Horsegram yellow mosaic viruses (HgYMV) infective clones. The samples were obtained using standard method from the two main horsegram growing areas of Bangalore, Karnataka State of India. The viral DNA from horsegram plants exhibiting severe symptoms was amplified by PCR. An isolate of HgYMV1 and HgYMV2 were associated with severe symptom phenotype from HgYMV. Full-length clones of DNA-A and DNA-B genomic components were constructed and attempts were made to introduce homologous (HgYMV1/HgYMV2) combinations of DNA-A and DNA-B genomic components into Nicotiana benthamiana plants. Inoculation of linearized constructs containing full-length clones or partial head-to-tail dimers of DNA-A and DNAB genomic components resulted in the introduction of DNA-A genomic components into the host plant. However, these combinations of genomic DNA component were not detected in the inoculated plants bombarded using the gene gun. Thus, this study was unable to confirm the pathogenicity of HgYMV infective clones using N. benthamiana as model plant.Keywords: Horsegram, Begomovirus, yellow mosaic viruses, Particle bombardment, Gene gu

Transcriptional response of virus-infected cassava and identification of putative sources of resistance for cassava brown streak disease

Cassava (Manihot esculenta) is a major food staple in sub-Saharan Africa, which is severely affected by cassava brown streak disease (CBSD). The aim of this study was to identify resistance for CBSD as well as to understand the mechanism of putative resistance for providing effective control for the disease. Three cassava varieties; Kaleso, Kiroba and Albert were inoculated with cassava brown streak viruses by grafting and also using the natural insect vector the whitefly, Bemisia tabaci. Kaleso expressed mild or no disease symptoms and supported low concentrations of viruses, which is a characteristic of resistant plants. In comparison, Kiroba expressed severe leaf but milder root symptoms, while Albert was susceptible with severe symptoms both on leaves and roots. Real-time PCR was used to estimate virus concentrations in cassava varieties. Virus quantities were higher in Kiroba and Albert compared to Kaleso. The Illumina RNA-sequencing was used to further understand the genetic basis of resistance. More than 700 genes were uniquely overexpressed in Kaleso in response to virus infection compared to Albert. Surprisingly, none of them were similar to known resistant gene orthologs. Some of the overexpressed genes, however, belonged to the hormone signalling pathways and secondary metabolites, both of which are linked to plant resistance. These genes should be further characterised before confirming their role in resistance to CBSD

Genotype by environment interactions in identifying cassava (Manihot esculenta Crantz) resistant to cassava brown streak disease

Cassava landraces were evaluated for resistance to cassava brown streak disease (CBSD) for two cropping seasons at a disease hotspot area in Naliendele, Tanzania. Based on reactions to CBSD, several landraces including Chimaje, Mfaransa and Supa B were considered to be resistant to the disease while Kikwada, Mbuyu, and Nyoka were tolerant. ANOVA revealed that the largest sum of squares (SS) (41.9–86.7%) was attributed to the genotype of the cassava landraces, while a smaller proportion of SS (8.1–38.2%) was due to genotype by environment interactions for all traits tested, which included disease symptoms, root weight, number of roots per plant and dry matter content. Environment accounted for the smallest effect (0.01–26.3%), however, the mean squares was nonetheless significant for a few genotypes, which indicated that their disease expression was indeed influenced by the environment. Increased CBSD severity was associated with low temperatures and rainfall. Increased rainfall towards harvesting led to higher root weight but lower dry matter content in the first cropping season. Correlation analysis showed that the presence of CBSD symptoms reduces the amount of usable roots, total root weight, and root dry matter content. Many resistant/tolerant landraces also had high root weight and dry matter content, and they can be used by farmers to reduce CBSD losses. The landraces described here form novel sources of CBSD resistance that can be used for breeding disease-resistant cassava varieties with superior agronomic characteristics

Cassava whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae), in sub-Saharan African farming landscapes: a review of the factors determining abundance

Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a pest species complex that causes widespread damage to cassava, a staple food crop for millions of smallholder households in Sub-Saharan Africa. Species in the complex cause direct feeding damage to cassava and are the vectors of multiple plant viruses. Whilst significant work has gone into developing virus-resistant cassava cultivars, there has been little research effort aimed at understanding the ecology of these insect vectors. In this review we critically assess the knowledge base relating to factors that may lead to high population densities of Sub-Saharan African (SSA) Bemisia tabaci species in cassava production landscapes of East Africa. We focus first on empirical studies that have examined biotic or abiotic factors that may lead to high populations. We then identify knowledge gaps that need to be filled to deliver long-term sustainable solutions to manage both the vectors and the viruses that they transmit. We found that whilst many hypotheses have been put forward to explain the increases in abundance witnessed since the early 1990s, there are little available published data and these tend to have been collected in a piecemeal manner. The most critical knowledge gaps identified were: (i) understanding how cassava cultivars and alternative host plants impact B. tabaci population dynamics and its natural enemies; (ii) the impact of natural enemies in terms of reducing the frequency of outbreaks and (iii) the use and management of insecticides to delay or avoid the development of resistance. In addition, there are several fundamental methodologies that need to be developed and deployed in East Africa to address some of the more challenging knowledge gaps

A newly recorded Rickettsia of the Torix group is a recent intruder and an endosymbiont in the whitefly Bemisia tabaci

The bacterium Rickettsia is found widely in phytophagous insects and often exerts profound effects on the phenotype and fitness of its hosts. Here, we decrypt a new, independent, phylogenetically ancient Torix Rickettsia endosymbiont found constantly in a laboratory line of an economically important insect Asia II 7, a putative species of the Bemisia tabaci whitefly complex (Hemiptera: Aleyrodidae), and occasionally in field whitefly populations. This new Rickettsia distributes throughout the body of its whitefly host. Genetically, compared to Rickettsia_bellii_MEAM1 found earlier in whiteflies, the new Rickettsia species has more gene families and pathways, which may be important factors in shaping specific symbiotic relationships. We propose the name ‘Candidatus Rickettsia_Torix_Bemisia_tabaci (RiTBt)’ for this new endosymbiont associated with whiteflies. Comparative genomic analyses indicate that RiTBi may be a relatively recent intruder in whiteflies given its low abundance in the field and relatively larger genome compared to Rickettsia_bellii_MEAM1

Bemisia tabaci and geminivirus variability in relation to cassava mosaic disease

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