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Background Flight can drastically enhance dispersal capacity and is a key trait defining the potential of exotic insect species to spread and invade new habitats. The phytophagous European spongy moths (ESM, Lymantria dispar dispar ) and Asian spongy moths (ASM; a multi–species group represented here by L. d. asiatica and L. d. japonica ), are globally invasive species that vary in adult female flight capability—female ASM are typically flight capable, whereas female ESM are typically flightless. Genetic markers of flight capability would supply a powerful tool for flight profiling of these species at any intercepted life stage. To assess the functional complexity of spongy moth flight and to identify potential markers of flight capability, we used multiple genetic approaches aimed at capturing complementary signals of putative flight–relevant genetic divergence between ESM and ASM: reduced representation genome–wide association studies, whole genome sequence comparisons, and developmental transcriptomics. We then judged the candidacy of flight–associated genes through functional analyses aimed at addressing the proximate demands of flight and salient features of the ecological context of spongy moth flight evolution. Results Candidate gene sets were typically non–overlapping across different genetic approaches, with only nine gene annotations shared between any pair of approaches. We detected an array of flight–relevant functional themes across gene sets that collectively suggest divergence in flight capability between European and Asian spongy moth lineages has coincided with evolutionary differentiation in multiple aspects of flight development, execution, and surrounding life history. Overall, our results indicate that spongy moth flight evolution has shaped or been influenced by a large and functionally broad network of traits. Conclusions Our study identified a suite of flight–associated genes in spongy moths suited to exploration of the genetic architecture and evolution of flight, or validation for flight profiling purposes. This work illustrates how complementary genetic approaches combined with phenotypically targeted functional analyses can help to characterize genetically complex traits.

Background: Detecting and controlling the movements of invasive species, such as insect pests, relies upon rapid and accurate species identification in order to initiate containment procedures by the appropriate authorities. Many species in the tussock moth genus Lymantria are significant forestry pests, including the gypsy moth Lymantria dispar L., and consequently have been a focus for the development of molecular diagnostic tools to assist in identifying species and source populations. In this study we expand the taxonomic and geographic coverage of the DNA barcode reference library, and further test the utility of this diagnostic method, both for species/subspecies assignment and for determination of geographic provenance of populations. Methodology/Principal Findings: Cytochrome oxidase I (COI) barcodes were obtained from 518 individuals and 36 species of Lymantria, including sequences assembled and generated from previous studies, vouchered material in public collections, and intercepted specimens obtained from surveillance programs in Canada. A maximum likelihood tree was constructed, revealing high bootstrap support for 90% of species clusters. Bayesian species assignment was also tested, and resulted in correct assignment to species and subspecies in all instances. The performance of barcoding was also compared against the commonly employed NB restriction digest system (also based on COI); while the latter is informative for discriminating gypsy moth subspecies, COI barcode sequences provide greater resolution and generality by encompassing a greater number of haplotypes across all Lymantria species, none shared between species. Conclusions/Significance: This study demonstrates the efficacy of DNA barcodes for diagnosing species of Lymantria and reinforces the view that the approach is an under-utilized resource with substantial potential for biosecurity and surveillance. Biomonitoring agencies currently employing the NB restriction digest system would gather more information by transitioning to the use of DNA barcoding, a change which could be made relatively seamlessly as the same gene region underlies both protocols.

Anoplophora chinensis (Forster) is an invasive species that can damage many tree species in orchard, urban, and forested habitats. Adult survival, reproduction, and egg hatch of A. chinensis from Italy and China are evaluated at eight constant temperatures (5, 10, 15, 20, 25, 30, 35, and 40 °C) under laboratory conditions. The estimated Tmax for longevity was 42 and 33 °C for females and 42 and 39 °C for males from China and Italy, respectively. The estimated Tmax, Tmin, and optimum temperature for fecundity were 35, 9, and 29 °C, respectively. Females laid eggs at 15–30 °C and eggs hatched at 15–35 °C. Days to first oviposition increased exponentially from 13 days at 30 °C to >300 days near 10 °C. The estimated Tmin for egg hatch was 13 °C, the Tmax at 38 °C, and the optimum 29 °C. Percentage hatch was estimated to be highest at 26 °C and have a Tmax of 31 °C and Tmin of 10 °C. These results indicate that summer temperatures over a wide range of latitudes should support beetle survival and reproduction, but at temperatures ≥35 °C, oviposition ceases, and adult survivorship declines. In addition, females may survive into the fall, but lay fewer eggs that may not hatch. These responses of A. chinensis to temperature can be used for developing phenological models to predict the timing of stages for management or eradication efforts.

We studied the mating behavior and reproductive biology of three members of the genus Agrilus: the bronze birch borer, Agrilus anxius Gory; the twolined chestnut borer, Agrilus bilineatus (Weber); and the emerald ash borer, Agrilus planipennis Fairmaire. All three species share a highly stereotyped mating behavior. However, the copulation duration of A. planipennis was 90% longer than that of its two congeners. Female reproductive tracts of the three species were anatomically similar, as were the spermatophores. Within the spermatophores, sperm were single in A. anxius and A. bilineatus, while in A. planipennis, sperm were bundled in groups of approximately 20 in a hyaline sheath. We found that field-caught A. anxius and A. bilineatus had higher rates of female insemination than A. planipennis. In additional studies with A. planipennis and A. anxius, we found that mating duration was related to mating success, and fecundity for A. planipennis, but not for A. anxius. For both A. planipennis and A. anxius, the spermatophore was passed to the female toward the end of the copulatory period. Sperm were found in the spermatheca immediately after copulation ended in A. planipennis and 30 min after copulation ended in A. anxius. We present possible explanations for these differences.