Development of a molecular-based detection tool for sweet potato leaf curl viruses and determination of their incidence levels in Tanzania

Sweet potato (Ipomoea batatas L.) is an important food security crop in sub-Saharan Africa, where its production is constrained by devastating diseases; including those caused by sweet potato leaf curl viruses (SPLCVs; Begomovirus; Geminiviridae). The objective of this study was to develop a molecular-based diagnostic detection tool for SPLCVs, as well as to generate information on their geographical distribution and incidence in Tanzania. A comprehensive survey of SPLCVs was done in all major sweet potato growing areas in Tanzania. Incidences of SPLCVs and their causative diseases were determined visually and by polymerase chain reaction (PCR), using primers designed, optimised and validated. DNA was extracted from 4166 sweet potato leaf samples and detection of SPLCVs was done. Visual incidence of disease symptoms ranged from 0 to 100%; while PCR-based incidence of SPLCVs ranged from 0 to 60%. The highest mean PCR-based incidence of SPLCVs was 32%. SPLCVs occurred in all sweet potato growing areas. There was a negative correlation between the visually assessed incidence of disease symptoms and PCR-based incidence of SPCLVs (r = -0.122 and R2 = 0.012). A weak positive correlation between altitude and PCR-based incidence of SPLCVs was also found. In ten-fold serially diluted sweet potato DNA samples, using our new primer pair d1-SPLCVF/d1-SPLCVR, the detection limit of SPLCVs was at the dilution of 10-3. The youngest fully expanded leaf of the sweet potato plant was the best for PCR detection of SPLCVs. These findings will be useful for strategic deployment of planting material and conducting sweet potato breeding experiments for resistance against SPLCVs. &nbsp

First report of banana bunchy top virus in banana and plantain (Musa spp.) in Tanzania

Open Access Article; Published online: 27 Sep 2021Banana (including plantain; Musa spp.) is an important vegetatively propagated food staple grown as a semi-perennial crop in fields and backyard gardens in Tanzania. Banana bunchy top disease (BBTD), caused by the banana bunchy top virus (BBTV, genus Babuvirus), is the most economically important viral disease of banana, infection of which results in severe stunting and reduction in fruit production by 90-100% within two seasons. The virus is spread by the banana aphid, Pentalonia nigronervosa, and through vegetative propagation of infected sources. BBTV is an introduced virus first reported in sub-Saharan Africa (SSA) in the 1960s in the Democratic Republic of Congo. Since then, BBTV spread was confirmed in 15 countries in Central, Southern, and Western African regions but was not detected in any previous surveys in the East African sub-region. During banana pests and disease surveys conducted in December 2020 – January 2021 in Buhigwe District in the Kigoma Region of Tanzania revealed banana plants with typical BBTV symptoms (severe stunting, leaves with shortened petioles, chlorotic streaks, and yellow leaf margins) in several banana fields in Muhinda (lon. 29.78662, lat. -4.53672) and Mwayaya (lon. 29.8218, lat. -4.49203) villages. Most of the affected plantations were 5 to 15 years old. Leaf samples (N=21) from symptomatic (N=6) and asymptomatic (N=15) banana plants were collected and used for total DNA extraction and BBTV detection by polymerase chain reaction (PCR) using the primer pair BBTV-1 and BBTV-2 to amplify ~240 bp sequence of DNA-R encoding for core master replication initiator protein gene. All samples from symptomatic plants tested positive and asymptomatic plants were negative. To further confirm the virus identity, four samples, each from symptomatic (PCR positive) and asymptomatic (PCR negative) plants from Muhinda and Mwayaya villages, were tested by Triple Antibody Sandwich-Enzyme-linked Immunosorbent Assay (TAS-ELISA) using BBTV ELISA reagent set (Cat. # SRA24700-1000, Agdia, France) following the manufacturer's protocol. Samples from symptomatic plants reacted positively in TAS-ELISA, and asymptomatic plants were negative. The 240-bp PCR product of two isolates was purified, and both strands were sequenced. A BLAST search of the nucleotide sequences (NCBI GenBank Acc.# MW711671 and MW711672) revealed 99% identity with DNA-R sequences of several other BBTV isolates from Africa (Acc. No# JF755994). Further analysis of the 240-bp nucleotide sequences with Maximum-likelihood phylogenetic analysis using MEGA-X software has grouped the two BBTV sequence isolates with the SSA sub-clade of the South-Pacific group. To our knowledge, this is the first report of BBTV infecting bananas in Tanzania, and East Africa endowed with rich banana diversity and popular East African Highland banana clone. BBTV presents a new threat to banana production in this sub-region due to the high risk of further spread through vegetative propagation, traditional planting material exchange practices, and the ubiquitous banana aphid vector. This study warrants delimitation surveys to assess the extent of spread, with simultaneous efforts to raise awareness about BBTD recognition and control measures among banana growers, including eradicating infected mats and replanting with healthy planting material to recover banana production

Evolution of cassava brown streak disease-associated viruses

Cassava brown streak disease (CBSD) has occurred in the Indian Ocean coastal lowlands and some areas of Malawi in East Africa for decades, and makes the storage roots of cassava unsuitable for consumption. CBSD is associated with Cassava brown streak virus (CBSV) and the recently described Ugandan cassava brown streak virus (UCBSV) [picorna-like (+)ssRNA viruses; genus Ipomovirus; family Potyviridae]. This study reports the first comprehensive analysis on how evolution is shaping the populations of CBSV and UCBSV. The complete genomes of CBSV and UCBSV (four and eight isolates, respectively) were 69.0–70.3 and 73.6–74.4% identical at the nucleotide and polyprotein amino acid sequence levels, respectively. They contained predictable sites of homologous recombination, mostly in the 39-proximal part (NIb-HAM1h-CP-39-UTR) of the genome, but no evidence of recombination between the two viruses was found. The CP-encoding sequences of 22 and 45 isolates of CBSV and UCBSV analysed, respectively, were mainly under purifying selection; however, several sites in the central part of CBSV CP were subjected to positive selection. HAM1h (putative nucleoside triphosphate pyrophosphatase) was the least similar protein between CBSV and UCBSV (aa identity approx. 55 %). Both termini of HAM1h contained sites under positive selection in UCBSV. The data imply an on-going but somewhat different evolution of CBSV and UCBSV, which is congruent with the recent widespread outbreak of UCBSV in cassava crops in the highland areas (.1000 m above sea level) of East Africa where CBSD has not caused significant problems in the past