AbstractBackgroundMeadow brown butterflies (Maniola jurtina) on the Isles of Scilly represent an ideal model in which to dissect the links between genotype, phenotype and long-term patterns of selection in the wild - a largely unfulfilled but fundamental aim of modern biology. To meet this aim, a clear description of genotype is required.FindingsHere we present the draft genome sequence of M. jurtina to serve as an initial genetic resource for this species. Seven libraries were constructed using DNA from multiple wild caught females and sequenced using Illumina, PacBio RSII and MinION technology. A novel hybrid assembly approach was employed to generate a final assembly with an N50 of 214 kb (longest scaffold 2.9 Mb). The genome encodes a total of 36,294 genes. 90.3% and 88.7% of core BUSCO (Benchmarking Universal Single-Copy Orthologs) Arthropoda and Insecta gene sets were recovered as complete single-copies from this assembly. Comparisons with 17 other Lepidopteran species placed 86.5% of the assembled genes in orthogroups.ConclusionsOur results provide the first high-quality draft genome and annotation of the butterfly M. jurtina.

Meadow brown butterflies (Maniola jurtina) on the Isles of Scilly represent an ideal model in which to dissect the links between genotype, phenotype and long-term patterns of selection in the wild - a largely unfulfilled but fundamental aim of modern biology. To meet this aim, a clear description of genotype is required. Here we present the draft genome sequence of M. jurtina to serve as a founding genetic resource for this species. Seven libraries were constructed using pooled DNA from five wild caught spotted females and sequenced using Illumina, PacBio RSII and MinION technology. A novel hybrid assembly approach was employed to generate a final assembly with an N50 of 214 kb (longest scaffold 2.9 Mb). The sequence assembly described here predicts a gene count of 36,294 and includes variants and gene duplicates from five genotypes. Core BUSCO (Benchmarking Universal Single-Copy Orthologs) gene sets of Arthropoda and Insecta recovered 90.5% and 88.7% complete and single-copy genes respectively. Comparisons with 17 other Lepidopteran species placed 86.5% of the assembled genes in orthogroups. Our results provide the first high-quality draft genome and annotation of the butterfly M. jurtina.

Host shifts can lead to ecological speciation and the emergence of new pests and pathogens. However, the mutational events that facilitate the exploitation of novel hosts are poorly understood. Here, we characterize an adaptive walk underpinning the host shift of the aphid Myzus persicae to tobacco, including evolution of mechanisms that overcame tobacco chemical defenses. A series of mutational events added as many as 1.5 million nucleotides to the genome of the tobacco-adapted subspecies, M. p. nicotianae, and yielded profound increases in expression of an enzyme that efficiently detoxifies nicotine, both in aphid gut tissue and in the bacteriocytes housing the obligate aphid symbiont Buchnera aphidicola. This dual evolutionary solution overcame the challenge of preserving fitness of a mutualistic symbiosis during adaptation to a toxic novel host. Our results reveal the intricate processes by which genetic novelty can arise and drive the evolution of key innovations required for ecological adaptation.

Many non-model organisms lack reference genomes and the sequencing and de novo assembly of an organisms transcriptome is an affordable means by which to characterize the coding component of its genome. Despite the advances that have made this possible, assembling a transcriptome without a known reference usually results in a collection of full-length and partial gene transcripts. The downstream analysis of genes represented as partial transcripts then often requires further experimental work in the laboratory in order to obtain full- length sequences. We have explored whether partial transcripts, encoding genes of interest present in de novo assembled transcriptomes of a model and non-model insect species, could be further extended by iterative mapping against the raw transcriptome sequencing reads. Partial sequences encoding cytochrome P450s and carboxyl/cholinesterase were used in this analysis, because they are large multigene families and exhibit significant variation in expression. We present an effective method to improve the contiguity of partial transcripts in silico that, in the absence of a reference genome, may be a quick and cost-effective alternative to their extension by laboratory experimentation. Our approach resulted in the successful extension of incompletely assembled transcripts, often to full length. We experimentally validated these results in silico and using real-time PCR and sequencing.

The impact of pesticides on the health of bee pollinators is determined in part by the capacity of bee detoxification systems to convert these compounds to less toxic forms. For example, recent work has shown that cytochrome P450s of the CYP9Q subfamily are critically important in defining the sensitivity of honey bees and bumblebees to pesticides, including neonicotinoid insecticides. However, it is currently unclear if solitary bees have functional equivalents of these enzymes with potentially serious implications in relation to their capacity to metabolise certain insecticides. To address this question, we sequenced the genome of the red mason bee, Osmia bicornis, the most abundant and economically important solitary bee species in Central Europe. We show that O. bicornis lacks the CYP9Q subfamily of P450s but, despite this, exhibits low acute toxicity to the N-cyanoamidine neonicotinoid thiacloprid. Functional studies revealed that variation in the sensitivity of O. bicornis to N-cyanoamidine and N-nitroguanidine neonicotinoids does not reside in differences in their affinity for the nicotinic acetylcholine receptor or speed of cuticular penetration. Rather, a P450 within the CYP9BU subfamily, with recent shared ancestry to the Apidae CYP9Q subfamily, metabolises thiacloprid in vitro and confers tolerance in vivo. Our data reveal conserved detoxification pathways in model solitary and eusocial bees despite key differences in the evolution of specific pesticide-metabolising enzymes in the two species groups. The discovery that P450 enzymes of solitary bees can act as metabolic defence systems against certain pesticides can be leveraged to avoid negative pesticide impacts on these important pollinators.

BACKGROUND: the glasshouse whitefly, Trialeurodes vaporariorum, is a damaging crop pest and an invasive generalist capable of feeding on a broad range of host plants. As such this species has evolved mechanisms to circumvent the wide spectrum of anti-herbivore allelochemicals produced by its host range. T. vaporariorum has also demonstrated a remarkable ability to evolve resistance to many of the synthetic insecticides used for control. RESULTS: to gain insight into the molecular mechanisms that underpin the polyphagy of T. vaporariorum and its resistance to natural and synthetic xenobiotics, we sequenced and assembled a reference genome for this species. Curation of genes putatively involved in the detoxification of natural and synthetic xenobiotics revealed a marked reduction in specific gene families between this species and another generalist whitefly, Bemisia tabaci. Transcriptome profiling of T. vaporariorum upon transfer to a range of different host plants revealed profound differences in the transcriptional response to more or less challenging hosts. Large scale changes in gene expression (> 20% of genes) were observed during adaptation to challenging hosts with a range of genes involved in gene regulation, signalling, and detoxification differentially expressed. Remarkably, these changes in gene expression were associated with significant shifts in the tolerance of host-adapted T. vaporariorum lines to natural and synthetic insecticides. CONCLUSIONS: Our findings provide further insights into the ability of polyphagous insects to extensively reprogram gene expression during host adaptation and illustrate the potential implications of this on their sensitivity to synthetic insecticides.

The development of insecticide resistance in insect pests of crops is a growing threat to sustainable food production, and strategies that slow the development of resistance are therefore urgently required. The insecticide synergist piperonyl butoxide (PBO) inhibits certain insect detoxification systems and so may delay the evolution of metabolic resistance. In the current study we characterized resistance development in the silverleaf whitefly, Bemisia tabaci, after selection with either a neonicotinoid (thiacloprid) or pyrethroid (alpha-cypermethrin) insecticide alone or in combination with PBO. Resistance development was significantly suppressed (> 60%) in the line selected with alpha-cypermethrin + PBO compared to the line selected with alpha-cypermethrin alone. RNA sequencing (RNAseq) analyses revealed an increase in frequency of a knock-down resistance mutation but no differentially expressed genes were identified that could explain the sensitivity shift. No significant difference was observed in the level of resistance between the thiacloprid and thiacloprid + PBO selected lines, and RNA sequencing (RNAseq) analyses revealed that the cytochrome P450 monooxygenase CYP6CM1, known to metabolize neonicotinoids, was significantly upregulated (>10-fold) in both lines. The findings of this study demonstrate that PBO used in combination with certain insecticides can suppress the development of resistance in a laboratory setting; however, the mechanism by which PBO supresses resistance development remains unclear.

A few diatom species produce toxins that affect human and animal health. Among these, members of the Pseudo-nitzschia genus were the first diatoms unambiguously identified as producer of domoic acid, a neurotoxin affecting molluscan shell-fish, birds, marine mammals, and humans. Evidence exists indicating the involvement of another diatom genus, Amphora, as a potential producer of domoic acid. We present a strategy for the detection of the diatom species Amphora coffeaeformis based on the development of species-specific oligonucleotide probes and their application in microarray hybridization experiments. This approach is based on the use of two marker genes highly conserved in all diatoms, but endowed with sufficient genetic divergence to discriminate diatoms at the species level. A region of approximately 450 bp of these previously unexplored marker genes, coding for elongation factor 1-a (eEF1-a) and silicic acid transporter (SIT), was used to design oligonucleotide probes that were tested for specificity in combination with the corresponding fluorescently labeled DNA targets. The results presented in this work suggest a possible use of this DNA chip technology for the selective detection of A. coffeaeformis in environmental settings where the presence of this potential toxin producer may represent a threat to human and animal health. In addition, the same basic approach can be adapted to a wider range of diatoms for the simultaneous detection of microorganisms used as biomarkers of different water quality levels.