AbstractThe amphipod Parhyale hawaiensis is a model organism of growing importance in the fields of evolutionary development and regeneration. A small, hardy marine crustacean that breeds year-round with a short generation time, it has simple lab culture requirements and an extensive molecular toolkit including the ability to generate targeted genetic mutant lines. Here we identify canonical core and regulatory clock genes using genomic and transcriptomic resources as a first step in establishing this species as a model in the field of chronobiology. The molecular clock of P. hawaiensis lacks orthologs of the canonical circadian genes cryptochrome 1 and timeless, in common with the mammalian system but in contrast to many arthropods including Drosophila melanogaster. Furthermore the predicted CLOCK peptide is atypical and CRY2 shows an extended 5’ region of unknown function. These results appear to be shared by two other amphipod species.

AbstractNeonicotinoids are effective insecticides used on many important arable and horticultural crops. They are nicotinic acetylcholine receptor agonists which disrupt the function of insect neurons and cause paralysis and death. In addition to direct mortality, there are numerous sublethal effects of low doses of neonicotinoids on bees. We hypothesize that some of these large array of effects could be a consequence of epigenetic changes in bees induced by neonicotinoids. We compared whole methylome (BS-seq) and RNA-seq libraries of the brains of buff tailed bumblebee Bombus terrestris workers exposed to field realistic doses of the neonicotinoid imidacloprid to libraries from control workers. We found numerous genes which show differential expression between neonicotinoid treated bees and control bees, but no differentially methylated cytosines in any context. We found CpG methylation to be focused mainly in exons and associated with highly expressed genes. We discuss the implications of our results for future legislation.

The Neotropical brown stink bug, Euschistus heros, is a major pest of soybean in South America. The importance of E. heros as a pest has grown significantly in recent times due to increases in its abundance and range, and the evolution of insecticide resistance. Recent work has begun to examine the genetic diversity, population structure, and genetic mechanisms of insecticide resistance in E. heros. However, to date, investigation of these topics has been hampered by a lack of genomic resources for this species. Here we address this need by assembling a high-quality draft genome for E. heros. We used a combination of short and long read sequencing to assemble an E. heros genome of 1.4 Gb comprising 906 contigs with a contig N50 of 3.5 MB. We leveraged this new genomic resource, in combination with genotyping by sequencing, to explore genetic diversity in populations of this species in Brazil and identify genetic loci in the genome which are under selection. Our genome-wide analyses, confirm that there are two populations of E. heros co-occurring in different geographical regions in Brazil, and that, in certain regions of the country these populations are hybridizing. We identify several regions of the genome as under selection, including markers associated with putative insecticide resistance genes. Taken together, the new genomic resources generated in this study will accelerate research into fundamental aspects of stinkbug biology and applied aspects relating to the sustainable control of a highly damaging crop pest.

Molecular phenotypes induced by environmental stimuli can be transmitted to offspring through epigenetic inheritance. Using transcriptome profiling, we show that the adaptation of Helicoverpa armigera larvae to soybean peptidase inhibitors (SPIs) is associated with large-scale gene expression changes including the upregulation of genes encoding serine peptidases in the digestive system. Furthermore, approximately 60% of the gene expression changes induced by SPIs persisted in the next generation of larvae fed on SPI-free diets including genes encoding regulatory, oxidoreductase, and protease functions. To investigate the role of epigenetic mechanisms in regulating SPI adaptation, the methylome of the digestive system of first-generation larvae (fed on a diet with and without SPIs) and of the progeny of larvae exposed to SPIs were characterized. A comparative analysis between RNA-seq and Methyl-seq data did not show a direct relationship between differentially methylated and differentially expressed genes, while trypsin and chymotrypsin genes were unmethylated in all treatments. Rather, DNA methylation potential epialleles were associated with transcriptional and translational controls; these may play a regulatory role in the adaptation of H. armigera to SPIs. Altogether, our findings provided insight into the mechanisms of insect adaptation to plant antiherbivore defense proteins and illustrated how large-scale transcriptional reprograming of insect genes can be transmitted across generations.

The green peach aphid, Myzus persicae, is a highly damaging, globally distributed crop pest that has evolved multiple resistance to numerous insecticides. It is thus imperative that insecticides that are not strongly compromised by pre-existing resistance are carefully managed to maximise their effective life span. Sulfoxaflor is a sulfoximine insecticide that retains efficacy against M. persicae clones that exhibit resistance to older insecticides. In the current study we monitored the efficacy of sulfoxaflor against M. persicae populations collected in Western Australia, following reports of control failures in this region. We identified clones with low (4-23-fold across multiple independent bioassay experiments), but significant, levels of resistance to sulfoxaflor compared with a reference susceptible clone. Furthermore, we demonstrate that sulfoxaflor resistance can persist after many months of culturing in the laboratory in the absence of insecticide exposure. Resistance was not conferred by known mechanisms of resistance to neonicotinoid insecticides, that act on the same target-site as sulfoxaflor, i.e. the R81T mutation or overexpresssion of the P450 gene CYP6CY3. Rather, transcriptome profiling of multiple resistant and susceptible clones identified the P450 CYP380C40 and the UDP-glucuronosyltransferase UGT344P2 as highly overexpressed (21-76-fold and 6-33-fold respectively) in the resistant clones. Transgenic expression of these genes demonstrated that they confer, low, but significant, levels of resistance to sulfoxaflor in vivo. Taken together, our data reveal the presence of low-level resistance to sulfoxaflor in M. persicae populations in Australia and uncover two novel mechanisms conferring resistance to this compound. The findings and tools generated in this study provide a platform for the development of strategies that aim to slow, prevent or overcome the evolution of more potent resistance to sulfoxaflor.

BACKGROUND: Although around 1% of cytosines in bees' genomes are known to be methylated, less is known about methylation's effect on bee behavior and fitness. Chemically altered DNA methylation levels have shown clear changes in the dominance and reproductive behavior of workers in queen-less colonies, but the global effect of DNA methylation on caste determination and colony development remains unclear, mainly because of difficulties in controlling for genetic differences among experimental subjects in the parental line. Here, we investigated the effect of the methylation altering agent decitabine on the developmental rate of full bumblebee colonies. Whole genome bisulfite sequencing was used to assess differences in methylation status. RESULTS: Our results showed fewer methylated loci in the control group. A total of 22 CpG loci were identified as significantly differentially methylated between treated and control workers with a change in methylation levels of 10% or more. Loci that were methylated differentially between groups participated in pathways including neuron function, oocyte regulation and metabolic processes. Treated colonies tended to develop faster, and therefore more workers were found at a given developmental stage. However, male production followed the opposite trend and it tended to be higher in control colonies. CONCLUSION: Overall, our results indicate that altered methylation patterns resulted in an improved cooperation between workers, while there were no signs of abnormal worker dominance or caste determination.

. Neonicotinoids are effective insecticides used on many important arable and horticultural crops. They are nicotinic acetylcholine receptor agonists which disrupt the function of insect neurons and cause paralysis and death. In addition to direct mortality, there are numerous sublethal effects of low doses of neonicotinoids on bees. We hypothesize that some of these large array of effects could be a consequence of epigenetic changes in bees induced by neonicotinoids. We compared whole methylome (BS-seq) and RNA-seq libraries of the brains of buff-tailed bumblebee
. Bombus terrestris
. workers exposed to field-realistic doses of the neonicotinoid imidacloprid to libraries from control workers. We found numerous genes which show differential expression between neonicotinoid-treated bees and control bees, but no differentially methylated cytosines in any context. We found CpG methylation to be focused mainly in exons and associated with highly expressed genes. We discuss the implications of our results for future legislation.
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