DNA primers, based on the ribosomal sequences of lethal yellowing-type disease (LYD) phytoplasmas, were used to analyse genetic variation within the lethal yellowing-type diseases of coconut in East Africa. Samples were collected from palms in Kenya, Mozambique and high, medium and low disease incidence areas of Tanzania. The mollicutespecific primer pair P1 and P6 amplified a 1·5 kbp product from all diseased palms and no product from symptomless palms, indicating that phytoplasmas were associated with all of these diseases. However, the Rohde forward and Rohde reverse primers (a second rRNA primer pair designed to detect East African LYD-associated phytoplasmas) only amplified products from Tanzanian and Kenyan diseased palms and not from those of Mozambique. Conversely, primers Ghana 813 and AK-SR, designed for specific detection of coconut-associated phytoplasmas in West Africa, amplified products only from the Mozambique palms, indicating that the phytoplasma associated with LYD in Mozambique is more closely related to those from West Africa. This was supported by restriction enzyme digestion of PCR products. DNA sequencing of PCR products from phytoplasmas within Tanzania revealed no detectable differences in the rDNA sequences of isolates from high, medium and low incidence areas.
The clock-like diversification of Rice yellow mottle virus (RYMV), a widespread RNA plant virus that infects rice in Africa, was tested following a three-step approach with (i) an exhaustive search of recombinants, (ii) a comprehensive assessment of the selective constraints over lineages, and (iii) a stepwise series of tests of the molecular clock hypothesis. The first evidence of recombination in RYMV was found in East Africa, in the region most favorable to co-infection. RYMV evolved under a pronounced purifying selection, but the selection pressure did vary among lineages. There was no phylogenetic evidence of transient deleterious mutations. ORF2b, which codes for the polymerase and is the most constrained ORF, tends to diversify clock-like. With the other ORFs and the full genome, the departure from the strict clock model was limited. This likely reflects the dominant conservative selection pressure and the clock-like fixation of synonymous mutations.
Lethal disease (LD) is a lethal yellowing-type disease of coconuts associated with phytoplasmas in Tanzania, but the insect vector for it has not yet been identified. In this study, the auchenorrynchous insects in LD-infected coconut fields were surveyed to determine potential vectors for the disease. No significant correlation was found between disease incidence and numbers of insects collected from the field, possibly reflecting the unknown incubation period for the disease. However, analysis of more than 5000 individual insects by the polymerase chain reaction (PCR), using LD-specific primers derived from the phytoplasma 16S rRNA gene, revealed PCR products of the correct size from eight individuals of Diastrombus mkurangai and four of Meenoplus spp. When digested with restriction endonucleases, fragments of the same size as the LD phytoplasma were obtained. No PCR products were detected in any of the other insect species tested. These results implicate D. mkurangai and Meenoplus spp. as probable vectors of the LD phytoplasma.
Tomato samples with typical symptoms of tomato yellow leaf curl virus (TYLCV) infection were collected from six tomato growing regions in Tanzania and dot-blotted on nylon membranes. The membranes were hybridised with nucleic acid probes synthesized to detect TYLCV from Israel and Sardinia. Viral DNA was extracted from the samples by phenol-chloform procedure and amplified by polymerase chain reaction using a primer pair (OTY 2 and OTY 6) designed to amplify a 1.2 kb fragment containing the coat protein, intergenic region and replicationassociated protein. The amplified DNA was ligated to pBluescript II KS + and transformed into Esherichia coli strain JM 83 cells by electroporation. Colonies containing the insert were sequenced using a Li-Cor DNA Semi-automatic Sequencer. The BLAST programme was used to search for viruses with similar sequences. Phylogenetic relationships with 20 geminiviruses were established using the CLUSTAL function of the Vector NT1.5 software. Amplified DNA was digested with Alu I and electrophoresed in polyacrylamide gel. Tomato yellow leaf curl virus samples from all the regions hybridised with both probes. Restriction analysis showed similar banding patterns for all the isolates. Both sequence comparison and phylogeny showed that TYLCV from Tanzania was closely related to TYLCV-Sar. The TYLCV isolates from the six regions were genetically the same.
A survey was initiated to detect tomato yellow leaf curl virus (TYLCV) and identify its reservoir weed hosts in six regions (Arusha, Morogoro, Dodoma, Iringa, Kilimanjaro and Dar es Salaam) in Tanzania. Three farms were randomly selected in each region. Assessment of TYLCV incidence was done by relating the number of infected tomato plants to the total number of plants assessed along a diagonal in five quadrants measuring 4 m × 4 m in size (one at each corner of the farm and one at the centre). Disease severity was scored on a scale of 0 to 4 (where 0 = no symptoms and 4 = very severe symptoms). Within and outside each farm, weeds showing TYLCV-like symptoms were collected and either squash-blotted, dot-blotted or both on nylon membranes. The membranes were hybridized with DIG-labelled probe synthesized for the detection of TYLCV from Sardinia (TYLCV-Sar) following standard protocols. Selected plant species were experimentally inoculated with screenhouse cultures of TYLCV representative isolates from the six regions using Bemisia tabaci to determine their host status. Results indicated that TYLCV incidence and severity were significantly higher (P = 0.05) in Dodoma region than the rest of the regions. In Iringa region, the incidence and severity of TYLCV were the lowest of all regions. TYLCV was detected in 12 of the 17 dot-blotted samples and in all the 21 squashed samples using the non-radioactively labelled riboprobes. Similarly, five plant species (Capsicum annuum, Datura stramonium, Lycopersicon esculentum, Nicotiana glutionsa and N. tabacum) tested in the screenhouse were infected by the six TYLCV isolates used. It is recommended that weeds within and outside tomato farms be removed to eliminate or reduce sources of virus inoculum. The dot and squash blot techniques are convenient for field detection of the virus, and are especially useful for the detection of early and latent infections so that management strategies can be initiated and implemented.
Tomato farms in the Dodoma region of Tanzania where a high incidence of Tomato yellow leaf curl Tanzania virus has been reported were visited to survey for weed hosts. Weeds exhibiting symptoms of infection by the virus were collected and identified. Lysates of leaf samples of the weed species were prepared and clarified by centrifugation. The clarified sap was dotted on nylon membranes and hybridised with a DIG-labelled probe. The putative viral DNA was extracted from the samples by phenol-chloroform procedures, and amplified by polymerase chain reaction methods using a primer pair designed to amplify a 1.2 kb fragment of the virus. Strong hybridisation signals were observed when sap from Ageratum conyzoides and Sida acuta were hybridised to the labelled probe. Similarly, the expected fragment size was obtained after amplification of DNA from both samples. It is concluded that these weeds are new hosts of Tomato yellow leaf curl Tanzania virus. An extensive search for yet undiscovered weed hosts is advocated, while the practice of farm sanitation is encouraged to eliminate reservoirs of the virus and vector.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.