Browsing by Author "Savill, Anders"

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    Real time portable genome sequencing for global food security
    (F1000 Research, 2018) Boykin, Laura; Ghalab, Ammar; Rossitto De Marchi, Bruno; Savill, Anders; Wainaina, James M.; Kinene, Tonny; Lamb, Stephen; Rodrigues, Myriam; Kehoe, Monica; Ndunguru, Joseph; Tairo, Fred; Sseruwagi, Peter; Kayuki, Charles; Mark, Deogratius; Erasto, Joel; Bachwenkizi, Hilda; Alicai, Titus; Okao-Okuja, Geoffrey; Abridrabo, Phillip; Ogwok, Emmanuel; Osingada, John Francis; Akono, Jimmy; Ateka, Elijah; Muga, Brenda; Kiarie, Samuel
    Crop losses due to viral diseases and pests are major constraints on food security and income for millions of households in sub-Saharan Africa (SSA). Such losses can be reduced if plant diseases and pests are correctly diagnosed and identified early. Currently, accurate diagnosis for definitive identification of plant viruses and their vectors in SSA mostly relies on standard PCR and next generation sequencing technologies (NGS). However, it can take up to 6 months before results generated using these approaches are available. The long time taken to detect or identify viruses impedes quick, within-season decision-making necessary for early action, crop protection advice and disease control measures by farmers. This ultimately compounds the magnitude of crop losses and food shortages suffered by farmers. The MinION portable pocket DNA sequencer was used, to our knowledge globally for the first time, to sequence whole plant virus genomes. We used this technology to identify the begomoviruses causing the devastating cassava mosaic virus, which is ravaging smallholder farmers’ crops in sub-Saharan Africa.
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    Recombination detected in the Heat Shock Protein 90 (HSP90) of the Bemisia tabaci species complex
    (bioRxiv, 2019) Kinene, Tonny; Rossito De Marchi, Bruno; Alicai, Titus; Luboobi, Livingstone S.; Abu Omongo, Christopher; Savill, Anders; Boykin, Laura M.
    Bemisia tabaci (whiteflies) are a global insect pest causing billions of dollars in damage each year, leaving farmers with low yields. In East Africa, whiteflies are superabundant and present on cassava plants throughout the year. Whiteflies do not decrease in number in the hot dry seasons in East Africa, therefore, it has been suggested that the synthesis of Heat Shock Protein (HSP) may protect the whitefly from heat stress and other biotic factors. In this study we used data sequence generated from individual whiteflies to assess variability and recombination of the HSP90 gene in members of the B. tabaci species complex. Results A total of 21 samples were sequenced on Illumina Hiseq 2500 and Hiseq 4000. These included eight genetic groups of B. tabaci: 7 SSA1, 5 SSA2, 2 Australia I (AUSI), 2 New World Africa (NWAfrica), B. afer, Uganda, Mediterranean (MED), and Middle East Asia Minor 1 (MEAM1). An alignment of 21 HSP90 sequences was generated after mapping and de novo assembly. Recombination analysis was performed on an alignment of 27 HSP90 sequences (we added an additional 6 sequences from GenBank). There were 18 recombination events detected in the HSP90 gene of the B. tabaci species complex, 7 of which were regarded as events that could be caused by evolutionary mechanisms such as gene duplication other than recombination. The phylogenetic analysis carried out on dataset without recombination events revealed a tree pattern with short terminal branches. Conclusion Recombination events were detected for members of the B. tabaci species complex in the HSP90 gene. This could explain the variability in the HSP90 gene of the B. tabaci species complex and highlight the phenomenon of the increased chance of survival and reproductive abundance of whiteflies in hot conditions in East Africa, since recombination is a major driving force of evolution.
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    Tree Lab: Portable Genomics for Early Detection of Plant Viruses and Pests in Sub-Saharan Africa
    (Genes, 2019) Boykin, Laura M.; Sseruwagi, Peter; Alicai, Titus; Ateka, Elijah; Umar Mohammed, Ibrahim; Stanton, Jo-Ann L.; Kayuki, Charles; Mark, Deogratius; Fute, Tarcisius; Erasto, Joel; Bachwenkizi, Hilda; Muga, Brenda; Mumo, Naomi; Mwangi, Jenniffer; Abidrabo, Phillip; Okao-Okuja, Geofrey; Omuut, Geresemu; Akol, Jacinta; Apio, Hellen B.; Osingada, Francis; Kehoe, Monica A.; Eccles, David; Savill, Anders; Lamb, Stephen; Kinene, Tonny; Rawle, Christopher B.; Muralidhar, Abishek; Mayall, Kirsty; Tairo, Fred; Ndunguru, Joseph
    In this case study we successfully teamed the PDQeX DNA purification technology developed by MicroGEM, New Zealand, with the MinION and MinIT mobile sequencing devices developed by Oxford Nanopore Technologies to produce an e ective point-of-need field diagnostic system. The PDQeX extractsDNAusing a cocktail of thermophilic proteinases and cell wall-degrading enzymes, thermo-responsive extractor cartridges and a temperature control unit. This closed system delivers purified DNA with no cross-contamination. The MinIT is a newly released data processing unit that converts MinION raw signal output into nucleotide base called data locally in real-time, removing the need for high-specification computers and large file transfers from the field. All three devices are battery powered with an exceptionally small footprint that facilitates transport and setup. To evaluate and validate capability of the system for unbiased pathogen identification by real-time sequencing in a farmer’s field setting, we analysed samples collected from cassava plants grown by subsistence farmers in three sub-Sahara African countries (Tanzania, Uganda and Kenya). A range of viral pathogens, all with similar symptoms, greatly reduce yield or destroy cassava crops. Eight hundred (800) million people worldwide depend on cassava for food and yearly income, and viral diseases are a significant constraint to its production. Early pathogen detection at a molecular level has great potential to rescue crops within a single growing season by providing results that inform decisions on disease management, use of appropriate virus-resistant or replacement planting. This case study presented conditions of working in-field with limited or no access to mains power, laboratory infrastructure, Internet connectivity and highly variable ambient temperature. An additional challenge is that, generally, plant material contains inhibitors of downstream molecular processes making e ective DNA purification critical. We successfully undertook real-time on-farm genome sequencing of samples collected from cassava plants on three farms, one in each country. Cassava mosaic begomoviruses were detected by sequencing leaf, stem, tuber and insect samples. The entire process, from arrival on farm to diagnosis, including sample collection, processing and provisional sequencing results was complete in under 3 h. The need for accurate, rapid and on-site diagnosis grows as globalized human activity accelerates. This technical breakthrough has applications that are relevant to human and animal health, environmental management and conservation.