Tree Lab: Portable genomics for early detection of plant viruses and pests in Sub-Saharan Africa

Abstract

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 effective point-of-need field diagnostic system. The PDQeX extracts DNA using a cocktail of thermophilic proteinases and cell wall degrading enzymes, thermo-responsive extractor cartridges and a temperature control unit. This single-step 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 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 set up.To evaluate and validate capability of the system for unbiased pathogen identification by realtime 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 completely destroy cassava crops. 800 million people worldwide depend on cassava for food and yearly income, and viral diseases are a significant constraint on its production (https://cassavavirusactionproject.com). 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 effective 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 4 hours. 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.

GCRTcall: a Transformer based basecaller for nanopore RNA sequencing enhanced by gated convolution and relative position embedding via joint loss training

“…This technique enables rapid single-molecule sequencing with significantly increased read lengths, reaching hundreds of kilobases or even magabases. It holds immense potential in various omics sequencing studies such as genomics, transcriptomics, epigenomics, and proteomics [1][2][3][4][5][6][7][8][9][10][11] .…”

New variants in the Chloroplast Genome Sequence of two Colombian individuals of the Cedar timber species (Cedrela odorataL.), using long read Oxford Nanopore Technology

“…Wang et al, 2018). Long read Oxford Nanopore Technology (ONT) have been use in a diverse range of applications, including genome assembly (Michael et al, 2018), building new reference genomes (Jain et al, 2016;Scott et al, 2020), rapid diagnoses of plant viruses (Boykin et al, 2019), among others. The MinION, which uses the ONT technology, is a portable, pocket-sized device, ideal for species with few or null genomic information (Y.…”