Day-night gene expression reveals circadian gene disco as a candidate for diel-niche evolution in moths.

Temporal ecological niche partitioning is an underappreciated driver of speciation. While insects have long been models for circadian biology, the genes and circuits that allow adaptive changes in diel-niches remain poorly understood. We compared gene expression in closely related day- and night-active non-model wild silk moths, with otherwise similar ecologies. Using an ortholog-based pipeline to compare RNA-Seq patterns across two moth species, we find over 25 pairs of gene orthologs showing differential expression. Notably, the gene disco, involved in circadian control, optic lobe and clock neuron development in Drosophila, shows robust adult circadian mRNA cycling in moth heads. Disco is highly conserved in moths and has additional zinc-finger domains with specific nocturnal and diurnal mutations. We propose disco as a candidate gene for the diversification of temporal diel-niche in moths.

Linking Colony Size with Foraging Behavior and Brain Investment in Odorous Ants (Formicidae: Dolichoderinae).

Superorganisms represent a unique level of biological organization in which the phenotype of the reproductive unit, the colony, results from traits expressed at the level of individual workers. Because body size scaling has important consequences for cell diversity and system complexity in solitary organisms, colony size is a trait of particular interest in superorganism evolution. In some instances, division of labor and worker polymorphism scale with colony size, but in general little is known about how colony size drives differences in individual-level behavior or neural traits. Ants represent the greatest diversity of superorganisms and provide a manner of natural experiment to test trends in trait evolution across multiple instances of colony size expansion. In this study, we control for environmental differences and worker size polymorphism to test if colony size correlates with measures of foraging behavior and brain size in dolichoderine ants. We present data from 3 species ranked by colony size. Our results suggest colony size correlates with measures of exploratory behavior and brain investment, with small-colony ants showing higher exploratory drive and faster exploration rate than the larger colony species, and greater relative investment in the primary olfactory brain region, the antennal lobe, than the larger colony species.

Brain evolution in social insects: advocating for the comparative approach.

Sociality is classified as one of the major transitions in the evolution of complexity and much effort has been dedicated to understanding what traits predispose lineages to sociality. Conversely, studies addressing the role of sociality in brain evolution (e.g., the social brain hypothesis) have not focused on particular traits and instead relied largely on measurements of relative brain composition. Hymenoptera range from solitary to advanced social species, providing enticing comparisons for studying sociality and neural trait evolution. Here we argue that measuring the role of sociality in brain evolution will benefit from attending to recent advances in neuroethology and adopting existing phylogenetic comparative methods employed in analysis of non-neural traits. Such analyses should rely on traits we expect to vary at the taxonomic level used in comparative analyses and include phylogenetic structure. We outline the limits of brain size and volumetric interpretation and advocate closer attention to trait stability and plasticity at different levels of organization. We propose neural traits measured at the cellular, circuit, and molecular levels will serve as more robust variables for evolutionary analyses. We include examples of particular traits and specific clades that are well-suited to answer questions about the role of sociality in nervous system evolution.

Long read genome assembly of Automeris io (Lepidoptera: Saturniidae) an emerging model for the evolution of deimatic displays.

Automeris moths are a morphologically diverse group with 135 described species that have a geographic range that spans from the New World temperate zone to the Neotropics. Many Automeris have elaborate hindwing eyespots that are thought to deter or disrupt the attack of potential predators, allowing the moth time to escape. The Io moth (Automeris io), known for its striking eyespots, is a well-studied species within the genus and is an emerging model system to study the evolution of deimatism. Existing research on the eyespot pattern development will be augmented by genomic resources that allow experimental manipulation of this emerging model. Here, we present a high-quality, PacBio HiFi genome assembly for Io moth to aid existing research on the molecular development of eyespots and future research on other deimatic traits. This 490 Mb assembly is highly contiguous (N50 = 15.78 mbs) and complete (benchmarking universal single-copy orthologs = 98.4%). Additionally, we were able to recover orthologs of genes previously identified as being involved in wing pattern formation and movement.

A long-read draft assembly of the Chinese mantis (Mantodea: Mantidae: Tenodera sinensis) genome reveals patterns of ion channel gain and loss across Arthropoda.

Praying mantids (Mantodea: Mantidae) are iconic insects that have captivated biologists for decades, especially the species with cannibalistic copulatory behavior. This behavior has been cited as evidence that insects lack nociceptive capacities and cannot feel pain; however, this behaviorally driven hypothesis has never been rigorously tested at the genetic or functional level. To enable future studies of nociceptive capabilities in mantids, we sequenced and assembled a draft genome of the Chinese praying mantis (Tenodera sinensis) and identified multiple classes of nociceptive ion channels by comparison to orthologous gene families in Arthropoda. Our assembly-produced using PacBio HiFi reads-is fragmented (total size = 3.03 Gb; N50 = 1.8 Mb; 4,966 contigs), but is highly complete with respect to gene content (BUSCO complete = 98.7% [odb10_insecta]). The size of our assembly is substantially larger than that of most other insects, but is consistent with the size of other mantid genomes. We found that most families of nociceptive ion channels are present in the T. sinensis genome; that they are most closely related to those found in the damp-wood termite (Zootermopsis nevadensis); and that some families have expanded in T. sinensis while others have contracted relative to nearby lineages. Our findings suggest that mantids are likely to possess nociceptive capabilities and provide a foundation for future experimentation regarding ion channel functions and their consequences for insect behavior.

Brain size scaling through development in the whitelined sphinx moth (Hyles lineata) shows mass and cell number comparable to flies, bees, and wasps.

Factors regulating larval growth and determinants of adult body size are described for several holometabolous insects, but less is known about brain size scaling through development. Here we use the isotropic fractionation ("brain soup") method to estimate the number of brain cells and cell density for the whitelined sphinx moth (Lepidoptera: Hyles lineata) from the first instar through the adult stage. We measure mass and brain cell number and find that, during the larval stages, body mass shows an exponential relationship with head width, while the total number of brain cells increases asymptotically. Larval brain cell number increases by a factor of ten from nearly 8000 in the first instar to over 80,000 in the fifth instar. Brain cell number increases by another factor of 10 during metamorphosis, with the adult brain containing more than 900,000 cells. This is similar to increases during development in the vinegar fly (Drosophila melanogaster) and the black soldier fly (Hermetia illucens). The adult brain falls slightly below the brain-to-body allometry for wasps and bees but is comparable in the number of cells per unit brain mass, indicating a general conservation of brain cell density across these divergent lineages.

Evolutionary genomics of three agricultural pest moths reveals rapid evolution of host adaptation and immune-related genes.

BACKGROUND: Understanding the genotype of pest species provides an important baseline for designing integrated pest management (IPM) strategies. Recently developed long-read sequence technologies make it possible to compare genomic features of nonmodel pest species to disclose the evolutionary path underlying the pest species profiles. Here we sequenced and assembled genomes for 3 agricultural pest gelechiid moths: Phthorimaea absoluta (tomato leafminer), Keiferia lycopersicella (tomato pinworm), and Scrobipalpa atriplicella (goosefoot groundling moth). We also compared genomes of tomato leafminer and tomato pinworm with published genomes of Phthorimaea operculella and Pectinophora gossypiella to investigate the gene family evolution related to the pest species profiles. RESULTS: We found that the 3 solanaceous feeding species, P. absoluta, K. lycopersicella, and P. operculella, are clustered together. Gene family evolution analyses with the 4 species show clear gene family expansions on host plant-associated genes for the 3 solanaceous feeding species. These genes are involved in host compound sensing (e.g., gustatory receptors), detoxification (e.g., ABC transporter C family, cytochrome P450, glucose-methanol-choline oxidoreductase, insect cuticle proteins, and UDP-glucuronosyl), and digestion (e.g., serine proteases and peptidase family S1). A gene ontology enrichment analysis of rapid evolving genes also suggests enriched functions in host sensing and immunity. CONCLUSIONS: Our results of family evolution analyses indicate that host plant adaptation and pathogen defense could be important drivers in species diversification among gelechiid moths.

A high-quality, long-read genome assembly of the whitelined sphinx moth (Lepidoptera: Sphingidae: Hyles lineata) shows highly conserved melanin synthesis pathway genes.

The sphinx moth genus Hyles comprises 29 described species inhabiting all continents except Antarctica. The genus diverged relatively recently (40-25 MYA), arising in the Americas and rapidly establishing a cosmopolitan distribution. The whitelined sphinx moth, Hyles lineata, represents the oldest extant lineage of this group and is one of the most widespread and abundant sphinx moths in North America. Hyles lineata exhibits the large body size and adept flight control characteristic of the sphinx moth family (Sphingidae), but it is unique in displaying extreme larval color variation and broad host plant use. These traits, in combination with its broad distribution and high relative abundance within its range, have made H. lineata a model organism for studying phenotypic plasticity, plant-herbivore interactions, physiological ecology, and flight control. Despite being one of the most well-studied sphinx moths, little data exist on genetic variation or regulation of gene expression. Here, we report a high-quality genome showing high contiguity (N50 of 14.2 Mb) and completeness (98.2% of Lepidoptera BUSCO genes), an important first characterization to facilitate such studies. We also annotate the core melanin synthesis pathway genes and confirm that they have high sequence conservation with other moths and are most similar to those of another, well-characterized sphinx moth, the tobacco hornworm (Manduca sexta).

Evolution of five environmentally responsive gene families in a pine-feeding sawfly, Neodiprion lecontei (Hymenoptera: Diprionidae).

A central goal in evolutionary biology is to determine the predictability of adaptive genetic changes. Despite many documented cases of convergent evolution at individual loci, little is known about the repeatability of gene family expansions and contractions. To address this void, we examined gene family evolution in the redheaded pine sawfly Neodiprion lecontei, a noneusocial hymenopteran and exemplar of a pine-specialized lineage evolved from angiosperm-feeding ancestors. After assembling and annotating a draft genome, we manually annotated multiple gene families with chemosensory, detoxification, or immunity functions before characterizing their genomic distributions and molecular evolution. We find evidence of recent expansions of bitter gustatory receptor, clan 3 cytochrome P450, olfactory receptor, and antimicrobial peptide subfamilies, with strong evidence of positive selection among paralogs in a clade of gustatory receptors possibly involved in the detection of bitter compounds. In contrast, these gene families had little evidence of recent contraction via pseudogenization. Overall, our results are consistent with the hypothesis that in response to novel selection pressures, gene families that mediate ecological interactions may expand and contract predictably. Testing this hypothesis will require the comparative analysis of high-quality annotation data from phylogenetically and ecologically diverse insect species and functionally diverse gene families. To this end, increasing sampling in under-sampled hymenopteran lineages and environmentally responsive gene families and standardizing manual annotation methods should be prioritized.

Modelling TDP-43 proteinopathy in Drosophila uncovers shared and neuron-specific targets across ALS and FTD relevant circuits.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) comprise a spectrum of neurodegenerative diseases linked to TDP-43 proteinopathy, which at the cellular level, is characterized by loss of nuclear TDP-43 and accumulation of cytoplasmic TDP-43 inclusions that ultimately cause RNA processing defects including dysregulation of splicing, mRNA transport and translation. Complementing our previous work in motor neurons, here we report a novel model of TDP-43 proteinopathy based on overexpression of TDP-43 in a subset of Drosophila Kenyon cells of the mushroom body (MB), a circuit with structural characteristics reminiscent of vertebrate cortical networks. This model recapitulates several aspects of dementia-relevant pathological features including age-dependent neuronal loss, nuclear depletion and cytoplasmic accumulation of TDP-43, and behavioral deficits in working memory and sleep that occur prior to axonal degeneration. RNA immunoprecipitations identify several candidate mRNA targets of TDP-43 in MBs, some of which are unique to the MB circuit and others that are shared with motor neurons. Among the latter is the glypican Dally-like-protein (Dlp), which exhibits significant TDP-43 associated reduction in expression during aging. Using genetic interactions we show that overexpression of Dlp in MBs mitigates TDP-43 dependent working memory deficits, conistent with Dlp acting as a mediator of TDP-43 toxicity. Substantiating our findings in the fly model, we find that the expression of GPC6 mRNA, a human ortholog of dlp, is specifically altered in neurons exhibiting the molecular signature of TDP-43 pathology in FTD patient brains. These findings suggest that circuit-specific Drosophila models provide a platform for uncovering shared or disease-specific molecular mechanisms and vulnerabilities across the spectrum of TDP-43 proteinopathies.

Impacts of development and adult sex on brain cell numbers in the Black Soldier Fly, Hermetia illucens L. (Diptera: Stratiomyidae).

The Black Soldier Fly (Hermetia illucens, Diptera: Stratiomyidae) has been introduced across the globe, with numerous industry applications predicated on its tremendous growth during the larval stage. However, basic research on H. illucens biology (for example, studies of their central nervous system) are lacking. Despite their small brain volumes, insects are capable of complex behaviors; understanding how these behaviors are completed with such a small amount of neural tissue requires understanding processing power (e.g. number of cells) within the brain. Brain cell counts have been completed in only a few insect species (mostly Hymenoptera), and almost exclusively in adults. This limits the taxonomic breadth of comparative analyses, as well as any conclusions about how development and body size growth may impact brain cell populations. Here, we present the first images and cell counts of the H. illucens brain at four time points across development (early, mid, and late larval stages, and both male and female adults) using immunohistochemistry and isotropic fractionation. To assess sexual dimorphism in adults, we quantified the number of cells in the central brain vs. optic lobes of males and females separately. To assess if increases in body size during development might independently affect different regions of the CNS, we quantified the larval ventral nerve cord and central brain separately at all three stages. Together, these data provide the first description of the nervous system of a popular, farmed invertebrate and the first study of brain cell numbers using IF across developmental stages in any insect.

Opposing deer and caterpillar foraging preferences may prevent reductions in songbird prey biomass in historically overbrowsed forests.

Overbrowsing by ungulates decimates plant populations and reduces diversity in a variety of ecosystems, but the mechanisms by which changes to plant community composition influence other trophic levels are poorly understood. In addition to removal of avian nesting habitat, browsing is hypothesized to reduce bird density and diversity through reduction of insect prey on browse-tolerant hosts left behind by deer. In this study, we excluded birds from branches of six tree species to quantify differences in songbird prey removal across trees that vary in deer browse preference. Early in the breeding season, birds preyed on caterpillars at levels proportional to their abundance on each host. Combining these data with tree species composition data from stands exposed to experimentally controlled deer densities over 30 years ago, we tested whether overbrowsing by white-tailed deer reduces prey biomass long after deer densities are reduced. Our analysis predicts total prey availability in the canopy of regenerating forests is fairly robust to historic exposure to high deer densities, though distribution of prey available from host species changes dramatically. This predicted compensatory effect was unexpected and is driven by high prey abundance on a single host tree species avoided by browsing deer, Prunus serotina. Thus, while we confirm that prey abundance on host trees can act as a reliable predictor for relative prey availability, this study shows that quantifying prey abundance across host trees is essential to understanding how changes in tree species composition interact with ungulate browse preference to determine prey availability for songbirds.