Seven new endemic genera and ten new species of Tortricidae are described from the Hawaiian Islands; all are assigned to Archipini (Tortricinae). A further eleven new combinations and one restored status are proposed. Iliahia, gen. nov. (type species: Capuaflavopicta Walsingham), is described for six species which feed on the foliage of 'iliahi (Santalum spp.; Santalaceae): Iliahia flavocincta (Walsingham), comb. nov., Iliahia flavopicta (Walsingham), comb. nov., Iliahia lilinoe, sp. nov., Iliahia pahulu, sp. nov., Iliahia santalata (Swezey), comb. nov., reinst. stat., and Iliahia xanthogona (Walsingham), comb. nov.; Nomewaimea, gen. nov. (type species: Epagoge infaustana Walsingham), is described for four species which feed on the leaves, shoots, and stems of ōpuhe (Touchardia sandwicensis; Urticaceae) and māmaki (Pipturus spp.; Urticaceae): Nomewaimea alaea, sp. nov., Nomewaimea kupenuia, sp. nov., Nomewaimea infaustana (Walsingham), comb. nov., and Nomewaimea urerana (Swezey), comb. nov.; Kumakena, gen. nov. (type species: Capua cassia Swezey), is described for Kumakenacassia (Swezey), comb. nov., which feeds on the leaves of kolomona (Senna gaudichaudii; Fabaceae); Limua, gen. nov. (type species: Archips lichenoides Walsingham), is described for four species which feed on the leaves of olopua (Notelaea sandwicensis; Oleaceae) and kōpiko (Psychotria spp.; Rubiaceae): Limua fuscoviridis (Walsingham), comb. nov., Limua lichenoides (Walsingham), comb. nov., Limua pahole, sp. nov., and Limua trochilidanus (Walsingham), comb. nov.; Paalua, gen. nov. (type species: Panaphelix asteliana Swezey), is described for three species which feed on the leaves of pa'iniu (Astelia spp.; Asteliaceae): Paalua asteliana (Swezey), comb. nov., Paalua maunaloa, sp. nov., and Paalua leleole, sp. nov.; Aipoola kaumualii, gen. nov., sp. nov., is described for a new species which feeds on the leaves of po'olā (Claoxylon sandwicense; Euphorbiaceae); and Maneapakele, gen. nov. (type species: Maneapakelehapalua Austin & Rubinoff), is described for two species which feed in the fruits of pāpala kēpau (Ceodes spp.; Nyctaginaceae): Maneapakele hapalua, sp. nov., and Maneapakele kahaha, sp. nov. Lectotypes are designated for four species. With the exception of K. cassia and possibly I. pahulu and I.santalata, all species are believed to be single-island endemics. We assess the conservation status for all species treated. All previously described species transferred to these new genera are provided with a new diagnosis in the context of the new species. A generic key to Tortricidae in Hawai'i and a species checklist is provided.

Free-ranging domestic cats (Felis catus) are globally distributed invasive carnivores. While recognition of their impacts has focused on consumption of vertebrates, increasing evidence suggests that they also consume large numbers of invertebrate species. Given the ongoing concern over invertebrate population declines across the planet, we compiled and analyzed a global database of invertebrate species reported in studies of cat diet. Despite making up >90% of all terrestrial animal species, invertebrates constituted only 7% of the >2000 species we identified as eaten by cats. However, when invertebrates were recorded in cat dietary studies, few were identified to species-level. Four of the 148 invertebrate species we reported to be eaten by cats are considered threatened by the IUCN, but cat predation is not recognized as a threat in their IUCN accounts. IUCN accounts do report cat predation as a threat for 48 invertebrate species of conservation concern, however none of these appear in our database. Insects (especially beetles) constituted similar to 80% of the 148 invertebrate species reported in cat dietary studies, with crustacean, arachnid, centipede, snail and slug, and millipede species occurring less frequently. Our results add to the growing consensus that cats consume a wide variety of invertebrate species and that they depredate more invertebrate species than is currently recognized. We recommend more cat dietary studies using eDNA (complemented by a more comprehensive eDNA library of invertebrate species), more autecological studies of threatened invertebrate species, and studies of responses of invertebrates to eradication or exclusion of feral cats.

DNA sequencing technology has undergone substantial improvements in recent years, to the extent that Third Generation Sequencing platforms are capable of massively generating long-reads. Amplicon sequencing has been among the most popular techniques due to its wide application in diverse fields of biological sciences. However, there is a lack of software specifically designed to analyse intra-individual genetic variation using amplicon long-read data. Here, we present CCS-consensuser, an end-to-end pipeline that generates consensus sequences from amplicon sequencing using high-fidelity reads produced by PacBio circular consensus sequencing (CCS). We evaluated the concordance of the results produced using CCS + CCS-consensuser and other sequencing platforms (Illumina and Sanger), as well as accuracy using a simulated dataset. This assessment showed that CCS amplicon data coupled with CCS-consensuser can produce high-quality sequences (PHRED > 30). The pipeline resulted in high proportions of identical sequence bins for real data, achieving up to 94.94% concordance with COI Sanger sequences and 92.61% with nuclear loci Illumina sequences (considering heterozygous loci), and 95.55% with a fully phased nuclear simulated dataset. Furthermore, our pipeline can be used to detect heteroplasmy in mtDNA, cross-contamination, resolve the phase of nuclear genes in diploid organisms, and conceivably for multi-copy gene systems such as rDNA. These results not only support its potential for application in studies using haploid data such as DNA barcoding, but also demonstrate its unique capacity to explore within individual haplotype variation. Therefore, our strategy shows promise for a broad range of applications in biology and medicine that have been challenging to assess using traditional techniques.

The species of Crocidosema Zeller occurring in Hawai'i are reviewed, resulting in the discovery of a previously unrecognized native species, described here as C. mawaena, Austin and Rubinoff, sp. nov., the removal of C. blackburnii (Butler), stat. rest. from synonymy with C. plebejana Zeller, and the recognition of two independent colonization events for the native Hawaiian Crocidosema. Crocidosema leprarum (Walsingham), a coastal species specializing on Sesuvium (Aizoaceae), appears to be the result of an independent colonization event from the other three native species of Crocidosema, all of which utilize various Malvaceae as their primary hosts. Despite being reported as present, the pest cotton tipworm (Crocidosema plebejana Zeller) does not occur in Hawai'i and should be a target species for biosecurity to prevent its establishment in the future. A fifth species, Crocidosema lantana Busck, deliberately introduced to control Lantana (Verbenaceae), is recorded from a native plant, Cryptocarya mannii (Lauraceae) for the first time, raising additional concerns about host specificity of early biological control introductions. All Hawaiian Crocidosema have multi-island distributions and often occur sympatrically; two species are known from the remote Northwestern Hawaiian Islands. All species in Hawai'i are redescribed and figured with a key to species provided. The conservation status for all native Hawaiian Crocidosema species is assessed.

Aprostocetus nitens Prinsloo & Kelly (Hymenoptera: Eulophidae) was identified as one of four hymenopteran ectoparasitoids utilizing three erythrina gall wasps, Quadrastichus bardus, Q. erythrinae, and Q. gallicola) (Hymenoptera: Eulophidae) in the native eastern Africa. In Hawai'i, the eurytomid wasp, Eurytoma erythrinae Gates & Delvare (Hymenoptera: Eurytomidae), was introduced and approved for statewide release in 2008 to control the erythrina gall wasp (EGW) Q. erythrinae Kim. EGW has devastated the wiliwili trees, Erythrina sandwicensis Degener (Fabaceae), an ecologically and culturally important native Hawaiian tree species. However, the parasitoid's impact on the galled inflorescences and shoots was not adequate to ensure adequate seed set and maturation for successful tree recruitment. Aprostocetus nitens was thus evaluated as a prospective natural enemy to enhance the biological control of EGW to further protect the wiliwili trees in Hawai'i. Both choice and no-choice host specificity tests were conducted on seven non-target gall formers in the Hawaii Department of Agriculture, Insect Containment Facility, and showed that the parasitoid was extremely specific to EGW. The potential for competition between this parasitoid and the established E. erythrinae was also investigated, showing that the release of a second parasitoid will potentially complement the success of the eurytomid wasp for control of EGW. Unlike what was found in the native region, the Hawaiian laboratory colony is thelytokous, producing only female offspring. The life cycle took 20.1 ± 0.28 days under the laboratory conditions. Non-ovipositing female survived for 102.5 ± 2.9 days when fed honey and laid eggs for 25.1 ± 2.3 days with average fecundity of 156.7 ± 22.3 offspring/female. This value is 3.9-fold higher than offspring produced by E. erythrinae. Aprostocetus nitens, host specificity, competition with E. erythrinae, and its tri-trophic association with 15 Erythrina host plants and 5 gall wasp assemblages in the native African regions were defined. Implications to reduce frequent galls on the native Erythrina plants and likely domination over E. erythrinae, are discussed.

Patterns of divergence and speciation on islands have long been of interest in the broader study of evolution. Hawaiʻi’s endemic Kamehameha butterfly (Vanessa tameamea) is experiencing population decline, but because of its high vagility and assumed genetic homogeneity as a species, its population structure has not been investigated. To evaluate V. tameamea genetic variation across the Hawaiian Islands, we assembled a reference quality genome assembly for the species using HiFi and HiC reads and performed range-wide population genetic analyses using ddRAD sequencing data. A discriminant analysis of principal components (DAPC) revealed that, contrary to prior assumptions, V. tameamea populations appear to be diverging based on geography, in a pattern similar to other native Hawaiian terrestrial arthropods. Specifically, through demographic history analyses, we find that the distinct population on Kauaʻi is likely to be ancestral, the central islands of Maui, Molokaʻi, and Oʻahu comprise another population, and Hawaiʻi Island forms a third population, with likely more gene exchange with the central islands. Finally, we investigate the SNPs driving differences between groups and find that many are associated with genes that may be relevant to local adaptation to environmental chemicals such as host plant defenses or chemicals introduced by human activity, notably to do with metabolism and detoxification. While much field work remains to investigate any cryptic or phenotypic patterns as well as quantify effective migration, we hope that this work will inform refinement of conservation plans for one of Hawaiʻi’s two native butterflies.

Fruit flies of the tribe Dacini (Tephritidae) include many agricultural pests, but also crucial pollinators of orchids and other plants. Surveys for Dacini fruit flies were conducted using methyl eugenol, cue lure, and zingerone as male attractants across Sabah, Borneo, Malaysia in 2018 and 2019, in habitats ranging from primary forest in highly protected Conservation Areas, selectively logged forest, secondary forest, and highly disturbed sites. Because 2019 was a mast year with mass fruiting, our surveys of that year collected more than 33,000 flies, compared to just more than 500 flies in 2018 – with similar trapping efforts. Our work adds 46 species to the 43 previously known from Borneo, bringing the total for the island to 89. Three new species are described: Bactrocera (Bactrocera) melanobivittata Doorenweerd, sp. nov., Dacus (Mellesis) danumensis Doorenweerd, sp. nov., and Zeugodacus (Zeugodacus) cataracta Doorenweerd, sp. nov. The new species are only found in conservation areas; B. melanobivittata is attracted to methyl eugenol and D. danumensis and Z. cataracta are attracted to zingerone. A discussion on how biogeographic affinities of the species in the checklist support a strong biogeographic boundary across Wallacea is provided, significant records are highlighted, and the relevance of these fly species for protecting agriculture as well as native ecosystems is discussed.

Lepidoptera is the most herbivorous of all the insect orders, with predatory caterpillars globally comprising less than 0.13% of the nearly 200,000 moth and butterfly species. Here, we report a species in which caterpillars are carnivorous inhabitants of spider's webs, feeding on the arthropods that they find there. This Hawaiian lineage also boasts an unprecedented and macabre practice of decorating its portable larval home with the body parts of the spider prey it harvests from the web where it resides. Phylogenomic data suggest that the origin of this unique spider cohabitant is at least six million years old, more than one million years older than Hawaii's current high islands. After decades of searching, only one species has been discovered, and it is restricted to 15 square kilometers of a single mountain range on the island of O'ahu, meaning that other members of the lineage have disappeared from older islands. Conservation action to save this globally unique lineage is imperative and overdue.

The species of Cryptophlebia occurring in Hawai'i are reviewed, resulting in the discovery of a previously unrecognized native species, described here as C. alaula, sp. nov. The discovery of C. alaula and its status as the putative sister species of C. illepida (Butler) supports the endemicity of C. illepida, a species whose status in Hawai'i has historically been contested, and is often treated as an invasive pest of unknown origin. Cryptophlebia alaula is hypothesized to be a specialist on fruits and seeds of 'a'ali'i ( Dodonaea viscosa). This raises interesting questions regarding the evolution of monophagy and polyphagy in Cryptophlebia and patterns of speciation in Hawaiian Tortricidae more broadly. The conservation status of C. alaula is briefly discussed. All species occurring in Hawai'i are described or redescribed, figured, with a key to species of both sexes provided.

Although insects make up the overwhelming majority of the described life on Earth, virtually nothing is known about the conservation status for all but the most charismatic taxa (i.e., butterflies, dragonflies) and many are at risk of imminent extinction. Given that island faunas are particularly vulnerable to extinction, we examined the conservation status of all 935 described species of native Lepidoptera in Hawaiʻi as a model to better understand levels of threat in poorly-known groups and how their rates of extinction might warrant actions to conserve them and inform future management. Using 100- and 50-year cut-offs for the time since each species was last recorded, we consider 269 species (28.8

The utility of a universal DNA 'barcode' fragment (658 base pairs of the Cytochrome C Oxidase I [COI] gene) has been established as a useful tool for species identification, and widely criticized as one for understanding the evolutionary history of a group. Large amounts of COI sequence data have been produced that hold promise for rapid species identification, for example, for biosecurity. The fruit fly tribe Dacini holds about a thousand species, of which 80 are pests of economic concern. We generated a COI reference library for 265 species of Dacini containing 5601 sequences that span most of the COI gene using circular consensus sequencing. We compared distance metrics versus monophyly assessments for species identification and although we found a 'soft' barcode gap around 2% pairwise distance, the exceptions to this rule dictate that a monophyly assessment is the only reliable method for species identification. We found that all fragments regularly used for Dacini fruit fly identification >450 base pairs long provide similar resolution. 11.3% of the species in our dataset were non-monophyletic in a COI tree, which is mostly due to species complexes. We conclude with recommendations for the future generation and use of COI libraries. We revise the generic assignment of Dacus transversus stat. rev. Hardy 1982, and Dacus perpusillus stat. rev. Drew 1971 and we establish Dacus maculipterus White 1998 syn. nov. as a junior synonym of Dacus satanas Liang et al. 1993.

The Oriental fruit fly Bactrocera dorsalis (Hendel), a global pest that can decimate regional fruit industries and elicit international quarantines, has been the subject of considerable taxonomic confusion. Previous phylogenetic work revealed that B. dorsalis is part of a monophyletic clade containing 12 species. We present restriction site‐associated DNA sequencing (RAD‐seq) genomic data for 2,292 specimens, which unequivocally supports the delimitation of two new species, here described as Bactrocera borneoensis sp. n. Doorenweerd & San Jose and B. incognita sp. n. Doorenweerd & San Jose. We additionally obtained 1,985 Cytochrome C oxidase I (COI) sequences for a subset of the specimens to see which species can be diagnosed with this mtDNA marker and conclude that B. dorsalis , B. incognita , B. carambolae Drew & Hancock, B. raiensis Drew & Hancock, B. occipitalis (Bezzi) and B. kandiensis Drew & Hancock cannot be identified reliably using COI due to introgression—but the newly described species B. borneoensis can be identified using COI. The supposed innocuous species B. raiensis distribution is underestimated in Asia and Africa. Bactrocera kandiensis COI genotypes occur in African flies, but RAD‐seq data confirm that these are B. dorsalis with introgressed B. kandiensis COI. The phylogenomic dataset brings new light to the extent of the B. dorsalis s.l. clade and the morphological and molecular confusion based on COI. This will have ramifications for ecological data—including host and distribution ranges—associated with B. dorsalis s.l. clade species, pest identification protocols and our understanding of the economic importance of the various species in the clade.

Invasive species are an increasing source of economic loss, costing nations billions of dollars annually. Significant financial resources are spent to manage invasive species, but few comprehensive syntheses of the economic expenditures associated with this management effort exist. As a relatively affluent developed country, the United States should serve as a model of how to both manage invasive species and, more critically, understand the economic costs of doing so. To begin understanding the scale of expenditures on invasive species in the U.S., our goal was to quantify spending on invasive species management at the state level. We contacted natural resource management officials from all 50 states following a standardized protocol. While 47 of the 50 states provided expenditures for at least one of the five years requested (2017–2021), the distribution of expenditures by state varied dramatically, suggesting that actual expenditures might be much higher than those reported. While most states shared annual expenditures, they varied by an order of magnitude from 28,370 for Connecticut to118,695,389 for Washington. Specifically, a widespread lack of careful and consistent expense tracking and coordination within and between states made clear and correct evaluation difficult. While the expenditures we obtained are almost certainly a significant underestimate, they also represent a serious lack of accounting at a state level. Hence, better tracking and coordination, within and between states, will be critical to handle the ongoing invasive species crisis.

The economic and ecological consequences of new insect invasions can be devastating and far-reaching. For example, the economic impact of the Oriental fruit fly (Bactrocera dorsalis [Hendel]) outbreak and subsequent eradication efforts in Florida (United States) in 2015 are estimated to have cost 41.2 million US dollars from direct costs, grower’s losses, and missed industry output from quarantine measures respectively, on top of hundreds of lost jobs (Steck et al., 2019). In Japan, the costs of eradicating several invasions of melon fly (Zeugodacus cucurbitae [Coquillett]) and Oriental fruit fly are estimated to have cost over 200 million US dollars between 1950 and 1998 (Kiritani, 1998). The potential losses caused by spongy moth (Lymantria dispar [Linnaeus]) and nun moth (Lymantria monacha [Linnaeus]) invasions in the United States between 1990 and 2003 are estimated to be 28–46 billion US dollars (Cock et al., 2003). The growing biosecurity risk from invasive pest insects is a wellestablished and inevitable consequence of globalization (Suckling et al., 2019). Luckily, the tools to combat these invasions are expanding and new genomic technologies are facilitating the development and implementation of better diagnostic markers and protocols. Thus, molecular species identification can play an especially important role in the prevention or rapid recognition of new pest invasions. Speedy recognition of a new invasion is crucial for eradication of the pest before establishment. Eradication is often expensive but is almost always orders of magnitude cheaper than suffering the long-term economic consequences to agriculture and trade of an established pest (Cantrell et al., 2002; Liebhold et al., 2016). Molecular species identification offers two significant advantages over conventional morphology-based identifications: (1) it does not rely on limited available taxonomic expertise for each identification, and (2) it can be applied to any insect life stage, sex, body part, or sometimes even traces left in the environment (eDNA) (e.g., Lopez-Vaamonde et al., 2012; Mlynarek et al., 2017; Rees et al., 2014). Therefore, molecular identification methods have the potential to become a critical part of biosecurity protocols around the world. However, although some molecular diagnostics methods have been around for decades, many require substantial improvement before they can be reliably applied in the prevention or early recognition of pest insect invasions. We break down the necessary steps for the development of robust molecular diagnostic tools into four major areas: (1) establishing accurate reference taxonomy, (2) facilitating multi-marker approaches, (3) ensuring sufficient intraspecific sampling of reference material, and (4) developing identification reliability metrics.

ABSTRACT. Seven new species of endemic leaf-roller moths are described from the Hawaiian Islands: Eccoptocera hinanohomauna sp. nov. (O‘ahu), Eccoptocera ohiaha, sp. nov. (O‘ahu), Pararrhaptica kaiona, sp. nov. (O‘ahu), Pararrhaptica pilikuanani, sp. nov. (Kaua‘i), Spheterista huakunana, sp. nov. (O‘ahu), Spheterista hiwakakahi, sp. nov. (Kaua‘i), and Spheterista hakeaiki, sp. nov. (O‘ahu). With the possible exception of S. hakeaiki, all species appear to be single-island endemics. In addition, Pararrhaptica pycnomias (Meyrick), syn. nov. (O‘ahu), is treated as a junior synonym of Pararrhaptica notocosma (Meyrick) (O‘ahu), Macraesthetica Meyrick is transferred from Eucosmini to Olethreutini, and Macraesthetica semicinereana (Swezey), comb. nov. (Hawai‘i), is transferred from Nuritamburia Koçak & Kemal. A conservation status is proposed for each of the newly described species.

As genomic data proliferates, the prevalence of post-speciation gene flow is making species boundaries and relationships increasingly ambiguous. Although current approaches inferring fully bifurcating phylogenies based on concatenated datasets provide simple and robust answers to many species relationships, they may be inaccurate because the models ignore inter-specific gene flow and incomplete lineage sorting. To examine the potential error resulting from ignoring gene flow, we generated both a RAD-seq and a 500 protein-coding loci highly multiplexed amplicon (HiMAP) dataset for a monophyletic group of 12 species defined as the Bactrocera dorsalis sensu lato clade. With some of the world's worst agricultural pests, the taxonomy of the B. dorsalis s.l. clade is important for trade and quarantines. However, taxonomic confusion confounds resolution due to intra- and interspecific phenotypic variation and convergence, mitochondrial introgression across half of the species, and viable hybrids. We compared the topological convergence of our datasets using concatenated phylogenetic and various multispecies coalescent approaches, some of which account for gene flow. All analyses agreed on species delimitation, but there was incongruence between species relationships. Under concatenation, both datasets suggest identical species relationships with mostly high statistical support. However, multispecies coalescent and multispecies network approaches suggest markedly different hypotheses and detected significant gene flow. We suggest that the network approaches are likely more accurate because gene flow violates the assumptions of the concatenated phylogenetic analyses, but the data-reductive requirements of network approaches resulted in reduced statistical support and could not unambiguously resolve gene flow directions. Our study highlights the importance of testing for gene flow, particularly with phylogenomic datasets, even when concatenated approaches receive high statistical support.

As the amount of genomic data for nonmodel taxa grows, it is increasingly clear that gene flow across species barriers in insects is much more common than previously thought. In recent years, the decreased cost and increased accuracy of long-read sequencing has enabled the assembly of high-quality reference genomes and chromosome maps for nonmodel insects. With this long-read data, we can now not only compare variation across the genome among homologous genes between species, which has been the basis of phylogenetics for more than 30 years, but also tease apart evidence of ancient and recent hybridization and gene flow. The implications of hybridization for species adaptation may be more positive than previously considered, explaining its prevalence across many groups of insects. Unfortunately, due to anthropogenic actions, some pest species appear to be benefitting from hybridization and gene flow, facilitating future invasions.