Single-fly assemblies fill major phylogenomic gaps across the Drosophilidae Tree of Life.

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 assemble 101 drosophilid genomes from laboratory cultures, greatly increasing the number of genome assemblies for this taxonomic group. The next major challenge is to address the laboratory culture bias in taxon sampling by sequencing genomes of species that cannot easily be reared in the lab. Here, we build upon our previous methods to perform amplification-free ONT sequencing of single wild flies obtained either directly from the field or from ethanol-preserved specimens in museum collections, greatly improving the representation of lesser studied drosophilid taxa in whole-genome data. Using Illumina Novaseq X Plus and ONT P2 sequencers with R10.4.1 chemistry, we set a new benchmark for inexpensive hybrid genome assembly at US $150 per genome while assembling genomes from as little as 35 ng of genomic DNA from a single fly. We present 183 new genome assemblies for 179 species as a resource for drosophilid systematics, phylogenetics, and comparative genomics. Of these genomes, 62 are from pooled lab strains and 121 from single adult flies. Despite the sample limitations of working with small insects, most single-fly diploid assemblies are comparable in contiguity (>1Mb contig N50), completeness (>98% complete dipteran BUSCOs), and accuracy (>QV40 genome-wide with ONT R10.4.1) to assemblies from inbred lines. We present a well-resolved multi-locus phylogeny for 360 drosophilid and 4 outgroup species encompassing all publicly available (as of August 2023) genomes for this group. Finally, we present a Progressive Cactus whole-genome, reference-free alignment built from a subset of 298 suitably high-quality drosophilid genomes. The new assemblies and alignment, along with updated laboratory protocols and computational pipelines, are released as an open resource and as a tool for studying evolution at the scale of an entire insect family.

Bacterial and fungal components of the gut microbiome have distinct, sex-specific roles in Hawaiian Drosophila reproduction.

Gut microbiomes provide numerous physiological benefits for host animals. The role of bacterial members of microbiomes in host physiology is well-documented. However, much less is known about the contributions and interactions of fungal members of the microbiome even though fungi are significant components of many microbiomes, including those of humans and insects. Here, we used antibacterial and antifungal drugs to manipulate the gut microbiome of a Hawaiian picture-wing Drosophila species, D. grimshawi, and identified distinct, sex-specific roles for the bacteria and fungi in microbiome community stability and reproduction. Female oogenesis, fecundity and mating drive were significantly diminished when fungal communities were suppressed. By contrast, male fecundity was more strongly affected by bacterial but not fungal populations. For males and females, suppression of both bacteria and fungi severely reduced fecundity and altered fatty acid levels and composition, implicating the importance of interkingdom interactions on reproduction and lipid metabolism. Overall, our results reveal that bacteria and fungi have distinct, sexually-dimorphic effects on host physiology and interkingdom dynamics in the gut help to maintain microbiome community stability and enhance reproduction.

Identification and Bioassays of Sex-Specific Compounds From a Nuisance Net-Spinning Caddisfly Smicridea fasciatella (Trichoptera: Hydropsychidae).

Municipalities in Arizona and Nevada along the Colorado River are subject to seasonal mass emergences of a nuisance net-spinning caddisfly, Smicridea fasciatella McLachlan (Trichoptera: Hydropsychidae). Here, we describe the characterization and field testing of S. fasciatella extracts to evaluate their potential as lures in baited traps. Solvent extracts of external (i.e., full body-cuticular hydrocarbon, abdominal hexane washes) and internal (i.e., crushed abdomen) parts of adult S. fasciatella were prepared from both sexes, and analyzed by coupled gas chromatography-mass spectrometry (GCMS). Several sex-specific compounds were identified, including (6Z,9Z)-6,9-nonadecadiene and (3Z,6Z,9Z)-3,6,9-nonadecatriene from males, and 2-undecanone, 2-tridecanone, and a heptadecene isomer from females. Extracts from both sexes were also analyzed by coupled gas chromatography-electroantennographic detection (EAD) using antennae of males for detection. Antennae of males weakly responded to 2-undecanone and 2-tridecanone, and to their corresponding alcohols, 2-undecanol and 2-tridecanol, which were included in field tests. Extracts of adult males did not elicit a response from male antennae, suggesting that males do not produce aggregation pheromones attractive to other males. The synchronized, dense populations of lekking males and other possible mating signals (e.g., visual recognition) may have contributed to the minimal attraction seen to test lures deployed in PHEROCON 1C traps. Overall, our results suggest that for this species, attractant pheromones have minimal or no role in bringing the sexes together for mating.

Two new day-flying species of Agrotis Ochsenheimer (Lepidoptera: Noctuidae) from the alpine summit of the Maunakea Volcano.

Two new endemic Hawaiian species of Agrotis Ochsenheimer (Noctuidae) are described: A. helela and A. kuamauna. Both species are day-flying and occur at high-elevations. Observations of adult and larval morphology and biology are included, as well as illustrations of adult moths and genitalia for both sexes.

Two new endemic Hawai'ian Lepidoptera: a new species of Pseudoschrankia (Erebidae) from O'ahu, and a new species of Thyrocopa (Xyloryctidae) from Moloka'i.

Two new endemic Hawai'ian Lepidoptera species are herein described. The first, Pseudoschrankia brevipalpis sp. nov., is a pollinator of the rare endemic plant Schiedea kaalae Wawra. The second, Thyrocopa keliae sp. nov., is endemic to Moloka'i island. Observations of undescribed parasitic mites that attack Hawai'ian Lepidoptera are presented as well.

Thermal stability and muscle efficiency in hovering orchid bees (Apidae: Euglossini).

To test whether variation in muscle efficiency contributes to thermal stability during flight in the orchid bee, Euglossa imperialis, we measured CO2 production, heat loss and flight kinematics at different air temperatures (Ta). We also examined the relationship between wingbeat frequency (WBF) and Ta in five additional species of orchid bees. Mean thoracic temperature (Tth) for Eg. imperialis hovering in a screened insectary and in the field was 39.3+/-0.77 degrees C (mean +/- 95% C.I.), and the slope of Tth on Ta was 0.57. Head and abdominal temperature excess ratios declined with Ta, indicating that Eg. imperialis were not increasing heat dissipation from the thorax at high Ta. Elevation of Tth above Ta was correlated with WBF, but Tth alone was not. Estimates of heat production from both respirometry and heat loss experiments decreased 33% as Ta rose from 24 to 34 degrees C. Mean muscle efficiency over this temperature range was 18% assuming perfect elastic energy storage and 22% assuming zero elastic energy storage. Both efficiency estimates increased significantly as Ta rose from 24 to 34 degrees C. In all six species examined, WBF declined significantly with Ta. These data indicate that hovering orchid bees regulate heat production through changes in wingbeat kinematics and consequent changes in energy conversion by the flight motor. Temperature-dependent variation in elastic energy storage or muscle contraction efficiency or both may contribute to the observed trends.