Darren J Obbard

University of Edinburgh | UoE · Institute of Ecology and Evolution

Drosophila melanogaster is a valuable invertebrate model for viral infection and antiviral immunity, and is a focus for studies of insect-virus coevolution. Here we use a metagenomic approach to identify more than 20 previously undetected RNA viruses and a DNA virus associated with wild D. melanogaster. These viruses not only include distant relati...

Pathogens switching to new hosts can result in the emergence of new infectious diseases, and determining which species are likely to be sources of such host shifts is essential to understanding disease threats to both humans and wildlife. However, the factors that determine whether a pathogen can infect a novel host are poorly understood. We have e...

It is estimated that a large proportion of amino acid substitutions in Drosophila have been fixed by natural selection, and as organisms are faced with an ever-changing array of pathogens and parasites to which they must adapt, we have investigated the role of parasite-mediated selection as a likely cause. To quantify the effect, and to identify wh...

Large scale genomic resources can place genetic variation into an ecologically informed context. To advance our understanding of the population genetics of the fruit fly Drosophila melanogaster , we present an expanded release of the community-generated population genomics resource Drosophila Evolution over Space and Time (DEST 2.0; https://dest.bi...

We present a genome assembly from an individual male Drosophila helvetica (drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence spans 224.20 megabases. Most of the assembly is scaffolded into 6 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled and...

Long-read sequencing is driving rapid progress in genome assembly across all major groups of life, including species of the family Drosophilidae, a longtime model system for genetics, genomics, and evolution. We previously developed a cost-effective hybrid Oxford Nanopore (ONT) long-read and Illumina short-read sequencing approach and used it to as...

We present a genome assembly from an individual male Drosophila limbata (drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 233.5 megabases in span. Most of the assembly is scaffolded into 6 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 16.09 kilobases in length.

Drosophila remains a pre-eminent insect model system for host–virus interaction, but the host range and fitness consequences of the drosophilid virome are poorly understood. Metagenomic studies have reported approximately 200 viruses associated with Drosophilidae, but few isolates are available to characterize the Drosophila immune response, and mo...

Drosophila melanogaster is not only one of the most important models of antiviral immunity in invertebrates, but is also a powerful model for research of the gut microbiome. Although recent studies have continued to improve our knowledge of the fly gut microbiota, the viral component of the microbiome has remained unexplored. Here we explore the vi...

We present a genome assembly from an individual male Drosophila phalerata (drosophilid fruit fly, Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 223.9 megabases in span. Most of the assembly is scaffolded into 7 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled...

We present a genome assembly from an individual female Drosophila histrio (the drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 189.2 megabases in span. Most of the assembly is scaffolded into 5 chromosomal pseudomolecules, including the X sex chromosome. The mitochondrial genome has also been assembled an...

We present a genome assembly from an individual male Chymomyza fuscimana (drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 338.0 megabases in span. Most of the assembly is scaffolded into 5 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled...

We present a genome assembly from an individual male Drosophila funebris (drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 181.1 megabases in span. Most of the assembly is scaffolded into 7 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled...

Long-read sequencing is driving rapid progress in genome assembly across all major groups of life, including species of the family Drosophilidae, a longtime model system for genetics, genomics, and evolution. We previously developed a cost-effective hybrid Oxford Nanopore (ONT) long-read and Illumina short-read sequencing approach and used it to as...

Drosophila remains a pre-eminent insect model system for host-virus interaction, but the host range and fitness consequences of the drosophilid virome are poorly understood. Metagenomic studies have reported over 160 viruses associated with Drosophilidae, but few isolates are available to characterise the Drosophila immune response, and most charac...

Drosophila melanogaster has one of the best characterized antiviral immune responses among invertebrates. However, relatively few easily transmitted natural virus isolates are available, and so many Drosophila experiments have been performed using artificial infection routes and artificial host–virus combinations. These may not reflect natural infe...

We present a genome assembly from an individual female Hirtodrosophila cameraria (a drosophilid fruit fly; Arthropoda; Insecta; Diptera; Drosophilidae). The genome sequence is 214.5 megabases in span. Most of the assembly is scaffolded into 4 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 15.94 kilobases in len...

Drosophila melanogaster has one of the best characterized antiviral immune responses among invertebrates. However, relatively few easily–transmitted natural virus isolates are available, and so many Drosophila experiments have been performed using artificial infection routes and artificial host–virus combinations. These may not reflect natural infe...

Mutation is the ultimate source of all genetic variation, and over the last 10 years the ready availability of whole-genome sequencing has permitted direct estimation of mutation rate for many non-model species across the tree of life. In this meta-analysis, we make a comprehensive search of the literature for mutation rate estimates in eukaryotes,...

The rates of mutation, recombination, and transposition are core parameters in models of evolution. They impact genetic diversity, responses to ongoing selection, and levels of genetic load. However, even for key evolutionary model species such as Drosophila melanogaster and D. simulans, few estimates of these parameters are available, and we have...

Mutation is the ultimate source of all genetic variation, and over the last ten years the ready availability of whole-genome sequencing has permitted direct estimation of mutation rate for many non-model species across the tree of life. In this meta-analysis we make a comprehensive search of the literature for mutation rate estimates in eukaryotes,...

Mutation, recombination, and transposition occurring during meiosis provide the variation on which natural selection can act and the rates at which they occur are important parameters in models of evolution. The de novo mutation rate determines levels of genetic diversity, responses to ongoing selection, and levels of genetic load. Recombination br...

Daphnia are well-established ecological and evolutionary models, and the interaction between D. magna and its microparasites is widely considered a paragon of the host-parasite coevolutionary process. Like other well-studied arthropods such as Drosophila melanogaster and Anopheles gambiae, D. magna is a small, widespread, and abundant species that...

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome data sets from natural populations of this species have been published over the last years. A major challenge is the integration of disparate data sets, often generated using different sequencing technologies and bioinformatic pipeli...

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome datasets from natural populations of this species have been published over the last years. A major challenge is the integration of disparate datasets, often generated using different sequencing technologies and bioinformatic pipeline...

Drosophila melanogaster is an important model for antiviral immunity in arthropods, but very few DNA viruses have been described from the family Drosophilidae. This deficiency limits our opportunity to use natural host-pathogen combinations in experimental studies, and may bias our understanding of the Drosophila virome. Here we report fourteen DNA...

Viruses are key population regulators, but we have limited knowledge of the diversity and ecology of viruses. This is even the case in wild host populations that provide ecosystem services, where small fitness effects may have major ecological impacts in aggregate. One such group of hosts are the bumblebees, which have a major role in the pollinati...

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome datasets from natural populations of this species have been published over the last 20 years. A major challenge is the integration of these disparate datasets, often generated using different sequencing technologies and bioinformatic...

Viruses are a key regulator of natural populations. Despite this, we have limited knowledge of the diversity and ecology of viruses that lack obvious fitness effects on their host. This is even the case in wild host populations that provide ecosystem services, where small fitness effects may have major ecological and financial impacts in aggregate....

Drosophila melanogaster is an important model for antiviral immunity in arthropods, but very few DNA viruses have been described in association with the Drosophilidae. This has limited the opportunity to use natural host-pathogen combinations in experimental studies, and may have biased our understanding of the Drosophila virome. Here we describe f...

Genetic variation is the fuel of evolution, with standing genetic variation especially important for short-term evolution and local adaptation. To date, studies of spatio-temporal patterns of genetic variation in natural populations have been challenging, as comprehensive sampling is logistically difficult, and sequencing of entire populations cost...

[This corrects the article DOI: 10.1371/journal.pgen.1007533.].

Metagenomic sequencing has revolutionised our knowledge of virus diversity, with new virus sequences being reported faster than ever before. However, virus discovery from metagenomic sequencing usually depends on detectable homology: without a sufficiently close relative, so-called 'dark' virus sequences remain unrecognisable. An alternative approa...

Background RNA interference (RNAi) related pathways provide defense against viruses and transposable elements, and have been implicated in the suppression of meiotic drive elements. Genes in these pathways often exhibit high levels of adaptive substitution, and over longer timescales show gene duplication and loss—most likely as a consequence of th...

Metagenomic sequencing has revolutionised our knowledge of virus diversity, with new virus sequences being reported at a higher rate than ever before. However, virus discovery from metagenomic sequencing usually depends on detectable homology: without a sufficiently close relative, so-called 'dark' virus sequences remain unrecognisable. An alternat...

Circadian clocks in eukaryotes involve both transcriptional-translational feedback loops, post-translational regulation, and metabolic, non-transcriptional oscillations. We recently identified the involvement of circadian oscillations in the intracellular concentrations of magnesium ions (Mg2+i) that were conserved in three eukaryotic kingdoms. Mg2...

Coevolution of multicellular organisms and their natural viruses may lead to an intricate relationship in which host survival requires effective immunity and virus survival depends on evasion of such responses. Insect antiviral immunity and reciprocal virus immunosuppression tactics have been well studied in Drosophila melanogaster , primarily duri...

Why a pathogen associates with one host but not another is one of the most important questions in disease ecology. Here we use transcriptome sequencing of wild-caught bumblebees from 13 species to describe their natural viruses, and to quantify the impact of evolutionary history on the realised associations between viruses and their pollinator host...

RNA interference (RNAi) related pathways provide defense against viruses and transposable elements, and have been implicated in the suppression of meiotic drive elements. Genes in these pathways often exhibit high levels of adaptive substitution, and over longer timescales show frequent gene duplication and loss-most likely as a consequence of thei...

Metagenomic sequencing has led to a recent and rapid expansion of the animal virome. It has uncovered a multitude of new virus lineages from under-sampled host groups, including many that break up long branches in the virus tree, and many that display unexpected genome sizes and structures. Although there are challenges to inferring the existence o...

RNA interference (RNAi)-related pathways target viruses and transposable element (TE) transcripts in plants, fungi, and ecdysozoans (nematodes and arthropods), giving protection against infection and transmission. In each case, this produces abundant TE and virus-derived 20-30nt small RNAs, which provide a characteristic signature of RNAi-mediated...

Phylogenetic trees. Maximum likelihood phylogenetic trees. Support values (approximate likelihood ratio test) and NCBI accession identifiers are provided. Viruses newly identified here are highlighted in red, and unannotated virus-like sequences from publicly-available transcriptome datasets are denoted ‘TSA’. Clade names follow [100,104]. Alignmen...

Properties and repeatability of virus-derived small RNAs. Panels A-D are dog-whelk RNA viruses, panels E-H starfish DNA virus-like contigs, panel I is the anemone DNA virus, and panel J is the brown alga virus (note that only one library was made for this sample). In each panel, rows (top to bottom) represent each library: Library A, polyphosphatas...

Properties and repeatability of TE-derived small RNAs. Panels A-R show the small RNA properties of selected high-confidence TE-like contigs: starfish panels A-C, dog whelk D-F, sponge G-I, earthworms J-L, sea anemone M-O, brown alga P-R. Rows and columns are as in S6 Fig. The data required to plot the size distributions are provided in S5 Table. (P...

PCR primers and conditions. PCR primer names and sequences, thermocycler conditions, and PCR recipes for virus prevalence, and RT-negative (EVE detection), assays. (XLSX)

Sampling, tissue preparations and RNA extractions. Detailed description of the sampling, tissue preparations and RNA extractions techniques employed for each target taxon. (PDF)

Relative read counts from high- copy-number viruses. The bar plot shows the relative number of RNAseq reads that mapped to each of virus contigs, as a percentage relative to the read count of host COI reads (both normalised by contig length). Both positive and negative sense reads were included, from library ‘B’ only. Viruses with less than 0.01% o...

Small RNAs mapping to bacterial contigs. Columns show (left) the number and size distribution of small RNAs that mapped to contigs provisionally classified as bacterial by similarity search, and the hotspots and size distributions for the nominal bacterial contig that displayed the largest (centre) and second-largest (right) number of small RNA map...

Sample collection details. Detailed descriptions of the sample collection locations, dates and numbers of individuals sampled for each target taxon, along with sample pool information, including which extraction pools were included in sequencing pools, and which were excluded due to detection of suspected nematode contamination. (XLSX)

Detailed descriptions of putative viruses and virus-like contigs. Detailed descriptions of the candidate virus fragments identified by protein similarity search, including phylogenetic position, estimated prevalence, approximate coverage, ORF number and most similar viral proteins identified by BLASTp, detectability by RT-negative PCR, GenBank acce...

Virus prevalence. Estimated virus prevalence inferred by maximum likelihood (with 2 log-likelihood intervals) from an RT-PCR survey of pooled samples[methods in 149]. (XLSX)

RNA oxidation treatment. Protocol for sodium periodate (NaIO4) oxidation of RNA prior to library preparation, to enrich small RNA libraries for canonical piRNAs and viRNAs by reducing the relative ligation efficiency of metazoan miRNAs that lack 3′-Ribose 2′O-methylation. (PDF)

Taxonomic composition of contigs. For each of the six organisms, the coloured bars show (on a linear scale), the proportion of all Trinity contigs assigned to each major lineage using Diamond [97] and MEGAN6 [98] with ‘long reads’. (PDF)

Size distributions of small RNAs. Bar-plot size distributions of all small RNAs sequences. Columns correspond to species, rows to libraries. Panel A: All sRNAs from each library. B: sRNAs mapping to ribosomal sequences. Note that in most species read abundance decreases with size, indicative of degradation products, but that distinct peaks are visi...

Sources of RNAseq and small RNA reads. Cytochrome oxidase coverage relative to that of the target taxon, and virus coverage for the target viruses (positive and negative strand) relative to that of COI. (XLSX)

Size distribution of small RNAs. Raw counts necessary to plot Figs 3, 4 and 5, S4, S5 and S6 Figs. (XLSX)

Protein sequence alignments. Protein sequence alignments used for phylogenetic analyses are provided in compressed (gzipped) gapped fasta format, with regions of poor alignment (identified by eye) deleted. Sequence titles comprise the taxon name and NCBI accession identifier for the sequence. (TGZ)

Phylogenetic trees. Phylogenetic trees are provided in compressed (gzipped) newick format. Sequence titles comprise the taxon name and NCBI accession identifier for the original protein sequence. (TGZ)

Strand bias in RNA sequencing reads from viruses. Bars show the proportion of RNA sequencing reads (combined across libraries) derived from the positive (sense) strand of the virus, for all viruses represented by >150 read pairs. Error bars reflect 95% confidence intervals based on a likelihood ratio test, assuming a binomial distribution, and to c...

RNAseq and sRNA reads per metagenomic contig. For each metagenomic contig (pale grey) the ratio of sRNAs (20-31nt) to RNAseq reads is shown on the x-axis, and the ratio of 20-24nt sRNAs (expected viRNAs) to 25-31nt sRNAs (expected piRNAs) is shown on the y-axis. Contigs are only included if they are >0.75Kbp in length and produced at least 20 small...

RNAi related genes identified from organisms. Counts of key RNAi related genes identified in transcriptomes of target taxa along with GenBank accession numbers for sequences. (XLSX)

Putative virus-like contigs. Raw meta-transcriptomic contigs generated by Trinity that have detectable sequence similarity (using Diamond) to virus proteins in GenBank, provided in compressed (gzipped) fasta format. Contig titles are annotated using the species name of the top match, followed by the percentage identity of that match in the sequence...

Long high-confidence TE-like contigs. Selected meta-transcriptomic contigs generated by Trinity that have detectable sequence similarity (using Diamond) to TEs in Repbase [99], provided in compressed (gzipped) fasta format. Contig titles are annotated using the host species name and the top-match TE in Repbase. (GZ)

Interactions between the insect immune system and RNA viruses have been best studied in Drosophila, where RNA interference, NF-κB and JAK-STAT pathways underlie antiviral immunity. In response to these immune mechanisms, insect viruses have convergently evolved suppressors of RNA interference that act by diverse mechanisms to permit viral replicati...

Drosophila melanogaster has played a key role in our understanding of invertebrate immunity. However, both functional and evolutionary studies of host-virus interaction in Drosophila have been limited by a dearth of native virus isolates. In particular, despite a long history of virus research, DNA viruses of D. melanogaster have only recently been...

KV-induced male mortality is not caused by contaminating RNA viruses. Chloroform-treatment is expected to inactivate enveloped viruses such as KV, but unenveloped viruses (including most +ssRNA viruses) are expected to retain infectivity. We confirmed mortality following KV infection was not caused by contaminating DAV by comparing injection of the...

Sex-by-genotype interaction significance manhattan plot. No polymorphism had a significant effect on sex-specific mortality. The blue line denotes prand = 0.05 and the red line is prand = 0.01. (TIF)

No enrichment of KV infection-associated SNPs in KV-responsive genes. Genes were split into KV-responsive and KV-unresponsive genes based on the RNA sequencing differential expression analysis (p < 0.05). The largest effect size (max βSNP) and lowest p-value was recorded for each gene in each GWAS, and compared between the KV-responsive and unrespo...

Differentially expressed genes. (XLSX)

RNA sequencing: Library quality and verification. (A) The first three principal components of read counts per gene in RNA-sequencing data, plotted such that each library is represented by a point. Males (blue) and females (red) are separated on PC1. Control-injected (green) and KV-injected (purple) are separated on PC3. For (B), we clustered librar...

Pathway analysis of genes identified by differential expression analysis or GWAS. Network analysis identified a pathway marginally enriched for genes identified in these studies (p = 0.03). Shown are interactions (edges) between genes identified in GWAS or RNA-sequencing studies (square nodes), allowing a single gene not included in the original ge...

Variable differential expression of the Tweedle gene family. A subset of KV-infected (purple) vials showed very high expression of Tweedle genes, whereas these were mostly unexpressed in control (green) adult flies. (TIF)

Expression of Kallithea virus genes. Most KV genes are expressed at 3 DPI. One KV-injected vial of flies had a lower level of infection. Control libraries also showed mapping to KV genes, most likely due to a low level (<0.5%) of barcode switching among libraries run together. The lower panel is a continuation of the upper panel. (TIF)

Significant GWAS hits. (XLSX)

Significant GO term enrichment for GWAS hits. (XLSX)

Significant GO terms for DE genes. (XLSX)

No effect of Wolbachia on KV growth or KV-induced mortality. Upper panels: Log-transformed relative viral titre in Wolbachia positive (green) or negative (orange) OreR female and male flies. Lower panels: mortality curves for gradient control-injected (circle) or KV-injected (triangle) OreR female and male flies, with or without Wolbachia. (TIF)

KV enriched in carcass relative to ovary. Females had higher viral titres in non-ovary tissues at 3 DPI. However, this could be affected by the route of infection, and the average ploidy of each sample. (TIF)

Virus titre in other confirmed GWAS knock-down lines. KV titre was measured in flies expressing a foldback hairpin targeting 18 genes identified in the GWAS, using GAL4 lines that knock each down in either the whole fly (tub-GAL4, green) or specifically in the gut (myo31DF-GAL4, purple). Only those causing a significant increase in titre relative t...

Kallithea virus assembly from RNA-seq reads. (FASTA)

Phylogeography places population genetics in an explicitly spatial context, and in doing so attempts to reconstruct the historical and contemporary evolutionary processes acting across a species range through space and time. Here we present the phylogeographical structure of Daphnia magna as determined for full mitochondrial genomes from samples of...

Metagenomic sequencing has led to a recent and rapid expansion in the animal virome. It has uncovered a multitude of new virus lineages from under-sampled host lineages, including many that break up long branches among previously known clades, and many with genomes that display unexpected sizes and structures. Although there are challenges to infer...

Genetic variation is the fuel of evolution. However, analyzing dynamics of evolutionary change in natural populations is challenging, genome sequencing of entire populations remains costly and comprehensive sample collection logistically challenging. To tackle this issue and to define relevant spatial and temporal scales of variation for a populati...