Complete genome sequence of a new member of the genus Badnavirus, Dioscorea bacilliform RT virus 3, reveals the first evidence of recombination in yam badnaviruses

Yams (Dioscorea spp.) host a diverse range of badnaviruses (genus Badnavirus, family Caulimoviridae). The first complete genome sequence of Dioscorea bacilliform RT virus 3 (DBRTV3), which belongs to the monophyletic species group K5, is described. This virus is most closely related to Dioscorea bacilliform SN virus (DBSNV, group K4) based on a comparison of genome sequences. Recombination analysis identified a unique recombination event in DBRTV3, with DBSNV likely to be the major parent and Dioscorea bacilliform AL virus (DBALV) the minor parent, providing the first evidence for recombination in yam badnaviruses. This has important implications for yam breeding programmes globally

Characterization of transposable elements within the Bemisia tabaci species complex

Background: Whiteflies are agricultural pests that cause negative impacts globally to crop yields resulting at times in severe economic losses and food insecurity. The Bemisia tabaci whitefly species complex is the most damaging in terms of its broad crop host range and its ability to serve as vector for over 400 plant viruses. Genomes of whiteflies belonging to this species complex have provided valuable genomic data; however, transposable elements (TEs) within these genomes remain unexplored. This study provides the first accurate characterization of TE content within the B. tabaci species complex. Results: This study identified that an average of 40.61% of the genomes of three whitefly species (MEAM1, MEDQ, and SSA-ECA) consists of TEs. The majority of the TEs identified were DNA transposons (22.85% average) while SINEs (0.14% average) were the least represented. This study also compared the TE content of the three whitefly genomes with three other hemipteran genomes and found significantly more DNA transposons and less LINEs in the whitefly genomes. A total of 63 TE superfamilies were identified to be present across the three whitefly species (39 DNA transposons, six LTR, 16 LINE, and two SINE). The sequences of the identified TEs were clustered which generated 5766 TE clusters. A total of 2707 clusters were identified as uniquely found within the whitefly genomes while none of the generated clusters were from both whitefly and non-whitefly TE sequences. This study is the first to characterize TEs found within different B. tabaci species and has created a standardized annotation workflow that could be used to analyze future whitefly genomes. Conclusion: This study is the first to characterize the landscape of TEs within the B. tabaci whitefly species complex. The characterization of these elements within the three whitefly genomes shows that TEs occupy significant portions of B. tabaci genomes, with DNA transposons representing the vast majority. This study also identified TE superfamilies and clusters of TE sequences of potential interest, providing essential information, and a framework for future TE studies within this species complex

Tissue culture and next-generation sequencing: A combined approach for detecting yam (Dioscorea spp.) viruses

In vitro culture offers many advantages for yam germplasm conservation, propagation and international distribution. However, low virus titres in the generated tissues pose a challenge for reliable virus detection, which makes it difficult to ensure that planting material is virus-free. In this study, we evaluated next-generation sequencing (NGS) for virus detection following yam propagation using a robust tissue culture methodology. We detected and assembled the genomes of novel isolates of already characterised viral species of the genera Badnavirus and Potyvirus, confirming the utility of NGS in diagnosing yam viruses and contributing towards the safe distribution of germplasm

In silico prediction of candidate gene targets for the management of African cassava whitefly (Bemisia tabaci, SSA1-SG1), a key vector of viruses causing cassava brown streak disease

Whiteflies (Bemisia tabaci sensu lato) have a wide host range and are globally important agricultural pests. In Sub-Saharan Africa, they vector viruses that cause two ongoing disease epidemics: cassava brown streak disease and cassava mosaic virus disease. These two diseases threaten food security for more than 800 million people in Sub-Saharan Africa. Efforts are ongoing to identify target genes for the development of novel management options against the whitefly populations that vector these devastating viral diseases affecting cassava production in Sub-Saharan Africa. This study aimed to identify genes that mediate osmoregulation and symbiosis functions within cassava whitefly gut and bacteriocytes and evaluate their potential as key gene targets for novel whitefly control strategies. The gene expression profiles of dissected guts, bacteriocytes and whole bodies were compared by RNAseq analysis to identify genes with significantly enriched expression in the gut and bacteriocytes. Phylogenetic analyses identified three candidate osmoregulation gene targets: two α-glucosidases, SUC 1 and SUC 2 with predicted function in sugar transformations that reduce osmotic pressure in the gut; and a water-specific aquaporin (AQP1) mediating water cycling from the distal to the proximal end of the gut. Expression of the genes in the gut was enriched 23.67-, 26.54- and 22.30-fold, respectively. Genome-wide metabolic reconstruction coupled with constraint-based modeling revealed four genes (argH, lysA, BCAT & dapB) within the bacteriocytes as potential targets for the management of cassava whiteflies. These genes were selected based on their role and essentiality within the different essential amino acid biosynthesis pathways. A demonstration of candidate osmoregulation and symbiosis gene targets in other species of the Bemisia tabaci species complex that are orthologs of the empirically validated osmoregulation genes highlights the latter as promising gene targets for the control of cassava whitefly pests by in planta RNA interference

Trimming and validation of Illumina short reads using Trimmomatic, trinity assembly, and assessment of RNA-Seq Data

Next-generation sequencing (NGS) technologies can generate billions of reads in a single sequencing run. However, with such high-throughput comes quality issues which have to be addressed before undertaking downstream analysis. Quality control on short reads is usually performed at default settings due to a lack of in-depth understanding of a particular software’s parameters and their effect if changed on the output. Here we demonstrate how to optimize read trimming using Trimmomatic. We highlight the benefits of trimming by comparing the quality of transcripts assembled using trimmed and untrimmed reads

BioNano genome mapping of individual chromosomes supports physical mapping and sequence assembly in complex plant genomes

The assembly of a reference genome sequence of bread wheat is challenging due to its specific features such as the genome size of 17 Gbp, polyploid nature and prevalence of repetitive sequences. BAC-by-BAC sequencing based on chromosomal physical maps, adopted by the International Wheat Genome Sequencing Consortium as the key strategy, reduces problems caused by the genome complexity and polyploidy, but the repeat content still hampers the sequence assembly. Availability of a high-resolution genomic map to guide sequence scaffolding and validate physical map and sequence assemblies would be highly beneficial to obtaining an accurate and complete genome sequence. Here, we chose the short arm of chromosome 7D (7DS) as a model to demonstrate for the first time that it is possible to couple chromosome flow sorting with genome mapping in nanochannel arrays and create a de novo genome map of a wheat chromosome. We constructed a high-resolution chromosome map composed of 371 contigs with an N50 of 1.3 Mb. Long DNA molecules achieved by our approach facilitated chromosome-scale analysis of repetitive sequences and revealed a ~800-kb array of tandem repeats intractable to current DNA sequencing technologies. Anchoring 7DS sequence assemblies obtained by clone-by-clone sequencing to the 7DS genome map provided a valuable tool to improve the BAC-contig physical map and validate sequence assembly on a chromosome-arm scale. Our results indicate that creating genome maps for the whole wheat genome in a chromosome-by-chromosome manner is feasible and that they will be an affordable tool to support the production of improved pseudomolecules

Genetic diversity of whitefly (Bemisia spp.) on crop and uncultivated plants in Uganda: implications for the control of this devastating pest species complex in Africa

Over the past three decades, highly increased whitefly (Bemisia tabaci) populations have been observed on the staple food crop cassava in eastern Africa and associated with ensuing viral disease pandemics and food insecurity. Increased whitefly numbers have also been observed in other key agricultural crops and weeds. Factors behind the population surges on different crops and their interrelationships are unclear, although in cassava they have been associated with specific populations within the Bemisia tabaci species complex known to infest cassava crops in Africa. This study carried out an in-depth survey to understand the distribution of B. tabaci populations infesting crops and uncultivated plant hosts in Uganda, a centre of origin for this pest complex. Whitefly samples were collected from 59 identified plant species and 25 unidentified weeds in a countrywide survey. Identities of 870 individual adult whiteflies were determined through mitochondrial cytochrome oxidase 1 sequences (651 bp) in the 3′ barcode region used for B. tabaci systematics. Sixteen B. tabaci and five related whitefly putative species were identified based on > 4.0% nucleotide divergence, of which three are proposed as novel B. tabaci putative species and four as novel closely related whitefly species. The most prevalent whiteflies were classified as B. tabaci MED-ASL (30.5% of samples), sub-Saharan Africa 1 (SSA1, 22.7%) and Bemisia Uganda1 (12.1%). These species were also indicated to be the most polyphagous occurring on 33, 40 and 25 identified plant species, respectively. Multiple (≥ 3) whitefly species occurred on specific crops (bean, eggplant, pumpkin and tomato) and weeds (Sida acuta and Ocimum gratissimum). These plants may have increased potential to act as reservoirs for mixed infections of whitefly-vectored viruses. Management of whitefly pest populations in eastern Africa will require an integration of approaches that consider their degree of polyphagy and a climate that enables the continuous presence of crop and uncultivated plant hosts

Molecular characterization of a new virus species identified in yam (Dioscorea spp.) by high-throughput sequencing

To date, several viruses of different genera have been reported to infect yam (Dioscorea spp.). The full diversity of viruses infecting yam, however, remains to be explored. High-throughput sequencing (HTS) methods are increasingly being used in the discovery of new plant viral genomes. In this study, we employed HTS on yam to determine whether any undiscovered viruses were present that would restrict the international distribution of yam germplasm. We discovered a new virus sequence present in 31 yam samples tested and have tentatively named this virus “yam virus Y” (YVY). Twenty-three of the samples in which YVY was detected showed mosaic and chlorotic leaf symptoms, but Yam mosaic virus was also detected in these samples. Complete genome sequences of two YVY viral isolates were assembled and found to contain five open reading frames (ORFs). ORF1 encodes a large replication-associated protein, ORF2, ORF3 and ORF4 constitute the putative triple gene block proteins, and ORF5 encodes a putative coat protein. Considering the species demarcation criteria of the family Betaflexiviridae, YVY should be considered as a novel virus species in the family Betaflexiviridae. Further work is needed to understand the association of this new virus with any symptoms and yield loss and its implication on virus-free seed yam production

Integrated physical map of bread wheat chromosome arm 7DS to facilitate gene cloning and comparative studies

Bread wheat (Triticum aestivum L.) is a staple food for a significant part of the world’s population. The growing demand on its production can be satisfied by improving yield and resistance to biotic and abiotic stress. Knowledge of the genome sequence would aid in discovering genes and QTLs underlying these traits and provide a basis for genomics-assisted breeding. Physical maps and BAC clones associated with them have been valuable resources from which to generate a reference genome of bread wheat and to assist map-based gene cloning. As a part of a joint effort coordinated by the International Wheat Genome Sequencing Consortium, we have con- structed a BAC-based physical map of bread wheat chromosome arm 7DS consisting of 895 contigs and covering 94% of its estimated length. By anchoring BAC contigs to one radiation hybrid map and three high resolution genetic maps, we assigned 73% of the assembly to a distinct genomic position. This map integration, inter- connecting a total of 1713 markers with ordered and sequenced BAC clones from a minimal tiling path, provides a tool to speed up gene cloning in wheat. The process of physical map assembly included the integration of the 7DS physical map with a whole-genome physical map of Aegilops tauschii and a 7DS Bionano genome map, which together enabled efficient scaffolding of physical-map contigs, even in the non-recombining region of the genetic centromere. Moreover, this approach facilitated a comparison of bread wheat and its ancestor at BAC-contig level and revealed a reconstructed region in the 7DS pericentromere