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Drosophila suzukii (Diptera: Drosophilidae) is a significant pest of wild and cultivated soft-skinned fruits. D. suzukii was first detected outside of its native range in 1983 on Mauna Ka‘ala in Wai‘anae, Hawai‘i, and has since spread throughout North America, South America, and Europe. While D. suzukii is not considered a crop pest in Hawai‘i, little data is available on the distribution of the species on a landscape scale on the archipelago. In this study, we document the distribution and abundance of D. suzukii and characterize its host use of ‘ōhelo (Vaccinium reticulatum) across an altitude gradient on the eastern slope of Mauna Loa, Hawai‘i. In total we collected 2,503 D. suzukii across 14 field sites over a four-month period in 2016. Endemic ‘ōhelo is a host for D. suzukii as we detected adult emergence across field sites with up to 1.88 flies per 1 mL of berries. Our preliminary population data shows that D. suzukii abundance is greater at higher altitudes and in forested habitats on Mauna Loa. Given the population abundance of D. suzukii and their ability to use at least one of the three endemic Vaccinium in Hawai‘i as a host, further research on host–use interactions with native and non-native insects is warranted.

In the context of evolutionary theory, invasion biology provides a fantastic enigma: how does a species with limited standing genetic variation survive and adapt to a novel environment? Reduced genetic diversity is typically associated with low fitness and evolutionary potential, yet some introduced species have proven to be successful invaders despite undergoing a genetic bottleneck during the early stages of colonization. Our goal in this study was to characterize population genomic and phenotype diversity of invasive Drosophila suzukii (Diptera: Drosophilidae) since colonizing the Hawaiian archipelago as early as the 1980s. Wing phenotype analysis revealed that high altitude populations possessed significantly larger wings than low altitude populations, supporting the hypothesis that insects cope with high altitude environments by developing larger wings. While we discovered low genetic diversity and differentiation in all Hawai‘i populations, three unique genetic clusters were detected with a model-free, multivariate statistical approach. We identified 23 candidate loci under selection using two complementary analyses to detect FST outliers across the genome. For 12 of these loci, predicted proteins are associated with Drosophila spp. chemosensation, amino acid and sodium ion transport, a Ras effector pathway, and cytidine deamination. Despite a genetic bottleneck, adventive D. suzukii populations are beginning to differentiate across the Hawaiian archipelago and selection for key behavioral and cellular processes are likely ongoing.

Interspecies hybrids can express phenotypic traits far outside the range of parental species. The atypical traits of hybrids provide insight into differences in the factors that regulate the expression of these traits in the parental species. In some cases, the unusual phenotypic traits of hybrids can lead to phenotypic dysfunction with hybrids experiencing reduced survival or reproduction. Cuticular hydrocarbons (CHCs) in insects are important phenotypic traits that serve several functions, including desiccation resistance and pheromones for mating. We used gas chromatography mass spectrometry to investigate the differences in CHC production between two closely related sympatric Hawaiian picture-wing Drosophila species, Drosophila heteroneura and D. silvestris, and their F1 and backcross hybrid offspring. CHC profiles differed between males of the two species, with substantial sexual dimorphism in D. silvestris but limited sexual dimorphism in D. heteroneura. Surprisingly, F1 hybrids did not produce three CHCs, and the abundances of several other CHCs occurred outside the ranges present in the two parental species. Backcross hybrids produced all CHCs with greater variation than observed in F1 or parental species. Overall, these results suggest that the production of CHCs was disrupted in F1 and backcross hybrids, which may have important consequences for their survival or reproduction.

Over 100 years of studies in Drosophila melanogaster and related species in the genus Drosophila have facilitated key discoveries in genetics, genomics, and evolution. While high-quality genome assemblies exist for several species in this group, they only encompass a small fraction of the genus. Recent advances in long-read sequencing allow high-quality genome assemblies for tens or even hundreds of species to be efficiently generated. Here, we utilize Oxford Nanopore sequencing to build an open community resource of genome assemblies for 101 lines of 93 drosophilid species encompassing 14 species groups and 35 sub-groups. The genomes are highly contiguous and complete, with an average contig N50 of 10.5 Mb and greater than 97% BUSCO completeness in 97/101 assemblies. We show that Nanopore-based assemblies are highly accurate in coding regions, particularly with respect to coding insertions and deletions. These assemblies, along with a detailed laboratory protocol and assembly pipelines, are released as a public resource and will serve as a starting point for addressing broad questions of genetics, ecology, and evolution at the scale of hundreds of species.

Determining the diet of arthropods can be difficult due to their small size and complex food webs, especially in Hawai‘i, where knowledge of arthropod predator–prey interactions is sparse. The diet of the Hawai‘i Island-endemic carabid beetle, Blackburnia hawaiiensis Sharp (Coleoptera: Carabidae) is of particular interest because of its peculiar arboreal behavior and metathoracic flight wings. Our study objective was to determine the diet of B. hawaiiensis in replicated, geographically separated locations by using two different yet complementary laboratory techniques: natural abundance stable isotope analysis (SIA) and high-throughput sequencing (HTS). Overall, B. hawaiiensis had a greater average δ15N and similar δ13C compared to the other arthropods sampled in this study and HTS data revealed Diptera and Lepidoptera sequences in the beetle's gut contents. These results are consistent with B. hawaiiensis being classified as a generalist predator. The combination of SIA and HTS are important methods for determining the diet of species within complex food webs, particularly for species that are difficult to observe in nature.

Genome-scale sequence data have invigorated the study of hybridization and introgression, particularly in animals. However, outside of a few notable cases, we lack systematic tests for introgression at a larger phylogenetic scale across entire clades. Here we leverage 155 genome assemblies, from 149 species, to generate a fossil-calibrated phylogeny and conduct multilocus tests for introgression across nine monophyletic radiations within the genus Drosophila. Using complementary phylogenomic approaches, we identify widespread introgression across the evolutionary history of Drosophila. Mapping gene-tree discordance onto the phylogeny revealed that both ancient and recent introgression has occurred across most of the nine clades that we examined. Our results provide the first evidence of introgression occurring across the evolutionary history of Drosophila and highlight the need to continue to study the evolutionary consequences of hybridization and introgression in this genus and across the Tree of Life.

We examine the genetic history and population status of Hawaiian hoary bats (Lasiurus semotus), the most isolated bats on Earth, and their relationship to northern hoary bats (Lasiurus cinereus), through whole-genome analysis of single-nucleotide polymorphisms mapped to a de novo-assembled reference genome. Profiles of genomic diversity and divergence indicate that Hawaiian hoary bats are distinct from northern hoary bats, and form a monophyletic group, indicating a single ancestral colonization event 1.34 Ma, followed by substantial divergence between islands beginning 0.51 Ma. Phylogenetic analysis indicates Maui is central to the radiation across the archipelago, with the southward expansion to Hawai‘i and westward to O‘ahu and Kaua‘i. Because this endangered species is of conservation concern, a clearer understanding of the population genetic structure of this bat in the Hawaiian Islands is of timely importance.

Anthropogenic influences on global processes and climatic conditions are increasingly affecting ecosystems throughout the world.Hawaii Island’s native ecosystems are well studied and local long‐term climatic trends well documented, making these ecosystems ideal for evaluating how native taxa may respond to a warming environment.This study documents adaptive divergence of populations of a Hawaiian picture‐winged Drosophila, D. sproati, that are separated by only 7 km and 365 m in elevation.Representative laboratory populations show divergent behavioral and physiological responses to an experimental low‐intensity increase in ambient temperature during maturation. The significant interaction of source population by temperature treatment for behavioral and physiological measurements indicates differential adaptation to temperature for the two populations.Significant differences in gene expression among males were mostly explained by the source population, with eleven genes in males also showing a significant interaction of source population by temperature treatment.The combined behavior, physiology, and gene expression differences between populations illustrate the potential for local adaptation to occur over a fine spatial scale and exemplify nuanced response to climate change.

Stress in animals can have extensive repercussions on other life history traits. Male Drosophila melanogaster under long-term selection for starvation-resistance (starvation stress) had reduced wing-waving courtship behavior in comparison to control males. However, only control females required longer courtship time from this reduced courtship effort; starvation-selected females mated equally with starvation-selected and control males. This suggests that there are behavioral trade-offs with starvation stress including reduced courtship behavior in males and relaxation of female choosiness.

Whereas disruptive selection imposed by heterogeneous environments can lead to the evolution of extrinsic isolating barriers between diverging populations, the evolution of intrinsic postzygotic barriers through divergent selection is less certain. Long‐lived species such as trees may be especially slow to evolve intrinsic isolating barriers. We examined postpollination reproductive isolating barriers below the species boundary, in an ephemeral hybrid zone between two successional varieties of the landscape‐dominant Hawaiian tree, Metrosideros polymorpha, on volcanically active Hawai'i Island. These archipelago‐wide sympatric varieties show the weakest neutral genetic divergence of any taxon pair on Hawai'i Island but significant morphological and ecological differentiation consistent with adaptation to new and old lava flows. Cross‐fertility between varieties was high and included heterosis of F1 hybrids at the seed germination stage, consistent with a substantial genetic load apparent within varieties through low self‐fertility and a lack of self‐pollen discrimination. However, a partial, but significant, barrier was observed in the form of reduced female and male fertility of hybrids, especially backcross hybrids, consistent with the accumulation of genetic incompatibilities between varieties. These results suggest that partial intrinsic postzygotic barriers can arise through disruptive selection acting on large, hybridizing populations of a long‐lived species.

We assessed how assemblages of spiders were structured in small Hawaiian tropical forest fragments (Hawaiian, kipuka) within a matrix of previous lava flows, over both space (sampling kipuka of different sizes) and time (comparison with a similar study from 1998). Standardized hand-collection by night was carried out in May 2016. In total, 702 spiders were collected, representing 6 families and 25 (morpho-)species. We found that the number of individuals, but not species richness, was highly correlated with the area of sampled forest fragments, suggesting that kipuka act as separate habitat islands for these predatory arthropods. Species richness was significantly lower in the lava matrix outside the kipuka compared to the kipuka habitats, although there was no statistical difference in species composition between the two habitats, largely because of similarity of non-native species in both habitats. Over the last 20 years, the abundance of non-native spider species substantially increased in both kipuka and lava habitats, in marked contrast to the vegetation that has remained more intact. With endemicity of terrestrial arthropods reaching over 95% in native forests, non-native predatory species present a critical challenge to the endemic fauna.

Evidence is accumulating that evolutionary changes are not only common during biological invasions but may also contribute directly to invasion success. The genomic basis of such changes is still largely unexplored. Yet, understanding the genomic response to invasion may help to predict the conditions under which invasiveness can be enhanced or suppressed. Here we characterized the genome response of the spotted wing drosophilaDrosophila suzukiiduring the worldwide invasion of this pest insect species, by conducting a genome-wide association study to identify genes involved in adaptive processes during invasion. Genomic data from 22 population samples were analyzed to detect genetic variants associated with the status (invasive versus native) of the sampled populations based on a newly developed statistic, we calledC2, that contrasts allele frequencies corrected for population structure. This new statistical framework has been implemented in an upgraded version of the program BayPass. We identified a relatively small set of single nucleotide polymorphisms (SNPs) that show a highly significant association with the invasive status of populations. In particular, two genesRhoGEF64Candcpo, the latter contributing to natural variation in several life-history traits (including diapause) inDrosophila melanogaster, contained SNPs significantly associated with the invasive status in the two separate main invasion routes ofD. suzukii. Our methodological approaches can be applied to any other invasive species, and more generally to any evolutionary model for species characterized by non-equilibrium demographic conditions for which binary covariables of interest can be defined at the population level.

Horizontal gene transfer events have played a major role in the evolution of microbial species, but their importance in animals is less clear. Here, we report horizontal gene transfer of cytolethal distending toxin B (cdtB), prokaryotic genes encoding eukaryote-targeting DNase I toxins, into the genomes of vinegar flies (Diptera: Drosophilidae) and aphids (Hemiptera: Aphididae). We found insect-encoded cdtB genes are most closely related to orthologs from bacteriophage that infect Candidatus Hamiltonella defensa, a bacterial mutualistic symbiont of aphids that confers resistance to parasitoid wasps. In drosophilids, cdtB orthologs are highly expressed during the parasitoid-prone larval stage and encode a protein with ancestral DNase activity. We show that cdtB has been domesticated by diverse insects and hypothesize that it functions in defense against their natural enemies.

Summary Lifetime reproductive capacity is a critical fitness component. In insects, female reproductive capacity is largely determined by the number of ovarioles, the egg-producing subunits of the ovary [e.g., 1 ]. Recent work has provided insights into ovariole number regulation in Drosophila melanogaster. However, whether mechanisms discovered under laboratory conditions explain evolutionary variation in natural populations is an outstanding question. We investigated potential effects of ecology on the developmental processes underlying ovariole number evolution among Hawaiian Drosophila, a large adaptive radiation wherein the highest and lowest ovariole numbers of the family have evolved within 25 million years. Previous studies proposed that ovariole number correlated with oviposition substrate [ 2 , 3 , 4 ] but sampled largely one clade of these flies and were limited by a provisional phylogeny and the available comparative methods. We test this hypothesis by applying phylogenetic modeling to an expanded sampling of ovariole numbers and substrate types and show support for these predictions across all major groups of Hawaiian Drosophila, wherein ovariole number variation is best explained by adaptation to specific substrates. Furthermore, we show that oviposition substrate evolution is linked to changes in the allometric relationship between body size and ovariole number. Finally, we provide evidence that the major changes in ovarian cell number that regulate D. melanogaster ovariole number also regulate ovariole number in Hawaiian drosophilids. Thus, we provide evidence that this remarkable adaptive radiation is linked to evolutionary changes in a key reproductive trait regulated at least partly by variation in the same developmental parameters that operate in the model species D. melanogaster.

Next-generation sequencing technologies have led to a decreased cost and an increased throughput in genome sequencing. Yet, many genome assemblies based on short sequencing reads have been assembled only to the scaffold level due to the lack of sufficient chromosome mapping information. Traditional ways of mapping scaffolds to chromosomes require a large amount of laboratory work and time to generate genetic and/or physical maps. To address this problem, we conducted a rapid technique which uses laser capture microdissection and enables mapping scaffolds of de novo genome assemblies directly to chromosomes in Hawaiian picture-winged Drosophila. We isolated and sequenced intact chromosome arms from larvae of D. differens. By mapping the reads of each chromosome to the recently assembled scaffolds from 3 Hawaiian picture-winged Drosophila species, at least 67% of the scaffolds were successfully assigned to chromosome arms. Even though the scaffolds are not ordered within a chromosome, the fast-generated chromosome information allows for chromosome-related analyses after genome assembling. We utilize this new information to test the faster-X evolution effect for the first time in these Hawaiian picture-winged Drosophila species.

Intraspecific hybrid zones involving long-lived woody species are rare and can provide insights into the genetic basis of early-diverging traits in speciation. Within the landscape-dominant Hawaiian tree, Metrosideros polymorpha, are morphologically distinct successional varieties, incana and glaberrima, that dominate new and old lava flows, respectively, below 1200 me on volcanically active Hawai'i Island, with var. glaberrima also extending to higher elevations and bogs. Here, we use morphological measurements on 86 adult trees to document the presence of an incana-glaberrima hybrid zone on the 1855 Mauna Loa lava flow on east Hawai'i Island and parent-offspring analysis of 1311 greenhouse seedlings from 71 crosses involving 72 adults to estimate heritabilities and genetic correlations among vegetative traits. Both the variation in adult leaf pubescence at the site and the consistency between adult and offspring phenotypes suggest the presence of two hybrid classes, F1s and var. incana backcrosses, as would be expected on a relatively young lava flow. Nine nuclear microsatellite loci failed to distinguish parental and hybrid genotypes. All four leaf traits examined showed an additive genetic basis with moderate to strong heritabilities, and genetic correlations were stronger for the more range-restricted var. incana. The differences between varieties in trait values, heritabilities and genetic correlations, coupled with high genetic variation within but low genetic variation between varieties, are consistent with a multi-million-year history of alternating periods of disruptive selection in contrasting environments and admixture in ephemeral hybrid zones. Finally, the contrasting genetic architectures suggest different evolutionary trajectories of leaf traits in these forms.

The Hawaiian Drosophila are an iconic example of sequential colonization, adaptive radiation and speciation on islands. Genetic and phenotypic analysis of closely related species pairs that exhibit incomplete reproductive isolation can provide insights into the mechanisms of speciation. Drosophila silvestris from Hawai'i Island and Drosophila planitibia from Maui are two closely related allopatric Hawaiian picture-winged Drosophila that produce sterile F1 males but fertile F1 females, a pattern consistent with Haldane's rule. Backcrossing F1 hybrid females between these two species to parental species gives rise to recombinant males with three distinct sperm phenotypes despite a similar genomic background: motile sperm, no sperm (sterile), and immotile sperm. We found that these three reproductive morphologies of backcross hybrid males produce divergent gene expression profiles in testes, as measured with RNA sequencing. There were a total of 71 genes significantly differentially expressed between backcross males with no sperm compared with those backcross males with motile sperm and immotile sperm, but no significant differential gene expression between backcross males with motile sperm and backcross males with immotile sperm. All of these genes were underexpressed in males with no sperm, including a number of genes with previously known activities in adult testis. An allele-specific expression analysis showed overwhelmingly more cis-divergent than trans-divergent genes, with no significant difference in the ratio of cis- and trans-divergent genes among the sperm phenotypes. Overall, the results indicate that the regulation of gene expression involved in sperm production likely diverged relatively rapidly between these two closely related species.

The Hawaiian archipelago provides a natural arena for understanding adaptive radiation and speciation. The Hawaiian Drosophila are one of the most diverse endemic groups in Hawaiì with up to 1,000 species. We sequenced and analyzed entire genomes of recently diverged species of Hawaiian picture-winged Drosophila, Drosophila silvestris and Drosophila heteroneura from Hawaiì Island, in comparison with Drosophila planitibia, their sister species from Maui, a neighboring island where a common ancestor of all three had likely occurred. Genome-wide single nucleotide polymorphism patterns suggest the more recent origin of D. silvestris and D. heteroneura, as well as a pervasive influence of positive selection on divergence of the three species, with the signatures of positive selection more prominent in sympatry than allopatry. Positively selected genes were significantly enriched for functional terms related to sensory detection and mating, suggesting that sexual selection played an important role in speciation of these species. In particular, sequence variation in Olfactory receptor and Gustatory receptor genes seems to play a major role in adaptive radiation in Hawaiian pictured-winged Drosophila.

Cytolethal distending toxins (CDTs) are tripartite eukaryotic genotoxins encoded in diverse bacterial and phage genomes. The cdtB subunit is a DNAse that causes eukaryotic cell cycle arrest and apoptosis, and in one context, is associated with resistance against parasitoid wasp infections. Here we report the discovery of functional cdtB copies in the nuclear genomes of insect species from two distantly related insect orders, including fruit flies (Diptera: Drosophilidae) and aphids (Hemiptera: Aphididae). Insect cdtB copies are most closely related to bacteriophage copies, were horizontally transferred to insect genomes > 40 million years ago and encode a protein that retains ancestral DNase activity. This phage-derived toxin has been domesticated by diverse insects and we hypothesize that it is used as a defensive weapon against parasitoid wasps.One Sentence SummaryWe report horizontal transfer of the gene cytolethal distending toxin B, which encodes a DNase, into eukaryotic genomes from bacteriophage.SignificanceCytolethal distending toxins (CDTs) are secreted by diverse pathogenic bacterial species to kill animal cells. The cdtB subunit enters cell nuclei, damaging the DNA and leading to mitotic arrest and apoptosis. In the pea aphid, a bacterial endosymbiont provides protection against wasp attack, possibly via cdtB. We discovered that this same endosymbiont-encoded lineage of cdtB was transferred to the genomes of Diptera and Hemiptera species and retains ancestral DNase activity. This is the first report of cdtB outside of bacteria or phages. A toxin that first evolved to kill eukaryotic cells has been co-opted by insects, potentially to their benefit.

International audience; Aim The diversity and composition of species pools within oceanic archipelagos is determined by a combination of colonization, abiotic tolerance, in situ diversification, biotic interactions and extinction. The signature of biogeographic events and evolutionary processes, however, may be masked by recent coalescence and hybridization between closely related species. We used the species-rich plant genus Cyrtandra (Gesneriaceae) to investigate the roles of colonization and hybridization in shaping community assemblages on the geologically young Hawai'i Island. Location Hawaiian Islands. Taxon Cyrtandra (Gesneriaceae). Methods We sampled 29 Cyrtandra taxa and putative hybrids across the main Hawaiian Islands and generated single nucleotide polymorphisms (SNPs) from nine single-copy nuclear genes. Maximum likelihood and Bayesian inference were used to reconstruct phylogenetic relationships, divergence times were estimated using secondary calibrations and island ages, and ancestral area estimation was done using likelihood models. Lastly, we used a Bayesian population assignment test and principal components analysis to infer population genetic structure. Results Cyrtandra colonization of the main Hawaiian Islands appears to have followed the progression rule, whereby the oldest high Hawaiian Islands (Kaua'i and O'ahu) were colonized first, followed by colonization of the younger islands as habitat became available. Hawai'i Island was colonized four times, with two dispersal events from O'ahu and two from Maui Nui. The different colonization events gave rise to significantly uneven numbers of species, and hybridization among the incipient lineages was detected in the form of intermediate genotypes. Main conclusions Our investigation into community assembly in a species-rich plant genus on a geologically young oceanic island revealed a history of multiple colonizations and hybridization among colonizing lineages. The rapid diversification (3.5 species/My) of one of four Cyrtandra lineages on Hawai'i Island may be the result of hybridization between genetically diverse lineages that stem from independent colonization events. Multiple colonization events followed by a merging of lineages may be particularly common during early-stage community assembly on islands and, through the generation of genetic variation, may be especially important for species diversification.

Lifetime reproductive capacity, or the total number of offspring that an individual can give rise to in its lifetime, is a fitness component critical to the evolutionary process. In insects, female reproductive capacity is largely determined by the number of ovarioles, the egg-producing subunits of the ovary. Recent work has provided insights into the genetic and environmental control of ovariole number inDrosophila melanogaster. However, whether regulatory mechanisms discovered under laboratory conditions also explain evolutionary variation in natural populations is an outstanding question. Here we report, for the first time, insights into the mechanisms regulating ovariole number and its evolution among Hawai’ianDrosophila, a large adaptive radiation of fruit flies in which the highest and lowest ovariole numbers of the genus have evolved within 25 million years. Using phylogenetic comparative methods, we show that ovariole number variation among Hawai’ianDrosophilais best explained by adaptation to specific oviposition substrates. Further, we show that evolution of oviposition on ephemeral egg-laying substrates is linked to changes the allometric relationship between body size and ovariole number. Finally, we provide evidence that the developmental mechanism principally responsible for controlling ovariole number inD. melanogasteralso regulates ovariole number in natural populations of Hawai’ian drosophilids. By integrating ecology, organismal growth, and cell behavior during development to understand the evolution of ovariole number, this work connects the ultimate and proximate mechanisms of evolutionary change in reproductive capacity.

Premise The drivers of isolation between sympatric populations of long-lived and highly dispersible conspecific plants are not well understood. In the Hawaiian Islands, the landscape-dominant tree, Metrosideros polymorpha, displays extraordinary phenotypic differences among sympatric varieties despite high dispersibility of its pollen and seeds, thereby presenting a unique opportunity to investigate how disruptive selection alone can maintain incipient forms. Stenophyllous M. polymorpha var. newellii is a recently evolved tree endemic to the waterways of eastern Hawai'i Island that shows striking neutral genetic differentiation from its ancestor, wet-forest M. polymorpha var. glaberrima, despite sympatry of these forms. We looked for evidence for, and drivers of, differential local adaptation of these varieties across the range of M. polymorpha var. newellii. Methods For paired populations of these varieties, we compared seedling performance under contrasting light conditions and a strong water current characteristic of the riparian zone. We also conducted a reciprocal transplant experiment and contrasted adult leaf anatomy. Results Results suggest that the riparian zone is harsh and that selection involving the mechanical stress of rushing water, and secondarily, light, led to significant reciprocal immigrant inviability in adjacent forest and riparian environments. The strongest adaptive divergence between varieties was seen in leaves and seedlings from the site with the sharpest ecotone, coincident with the strongest genetic isolation of M. polymorpha var. newellii observed previously. Conclusions These findings suggest that disruptive selection across a sharp ecotone contributes to the maintenance of an incipient riparian ecotype from within a continuous population of a long-lived and highly dispersible tree species.

PREMISE OF THE STUDY Recent reviews of reproductive isolation (RI) in plants propose that boundaries between closely related species are maintained predominantly through prezygotic mechanisms. However, few experimental studies have explored how boundaries are maintained in long-lived species. Hawaiian Cyrtandra presents an intriguing challenge to our understanding of RI, as it comprises 60 shrub or small tree species that are almost exclusively restricted to wet forests, where sympatry of multiple species is common. METHODS We assessed the relative strengths of pre- and postzygotic barriers among four species of Cyrtandra occurring at the extremes of the main Hawaiian Island's natural island-age gradient, Kaua'i (4.7 Myr) and Hawai'i Island (0.6 Myr), to contrast the strengths and stages of reproductive isolation among species at different stages of divergence. KEY RESULTS A combination of F1 seed germination, F1 seedling survival, and F1 seedling growth isolated (61-91%) three of the species from sympatric relatives. In contrast, the fourth species was isolated (59%) from its sympatric relative through phenological differences alone. Significant postzygotic barriers in between-island crosses were also observed in one species. CONCLUSIONS Results suggest that boundaries between sympatric Cyrtandra species in Hawaii are maintained predominantly through postzygotic barriers. Observations from between-island crosses indicate that postzygotic barriers can arise in allopatry, which may be important in the initial divergence of populations. Future studies of RI in Cyrtandra should include a broader range of species to determine if postzygotic isolating barriers are foremost in the maintenance of species boundaries in this large genus.

The Hawaiian picture wing Drosophila are a striking example of adaptive radiation in specialist saprophages on an island system. We use DNA sequences from five nuclear genes with a total of 4260 nucleotides to provide a comprehensive phylogeny and biogeographic analysis of 90 species in the Hawaiian Drosophila picture wing clade. The current analysis indicates that the evolution of the picture wing clade took place more recently than previously suggested. The relationships of several morphologically anomalous taxa are resolved with strong support. Biogeography and host plant analyses show two periods of rapid divergence occurred when Kauai and Oahu were the main high islands, indicating that a combination of complex topographical features of islands and development of novel host plant associations was key to the rapid diversification of these lineages. For the past 2 million years, host associations within lineages have been largely stable, and speciation has occurred primarily due to the establishment of populations on newer islands as they arose followed by divergence by isolation. The existence of several apparently relictual taxa suggests that extinction has also played a major role in assembly of the present Hawaiian Drosophila fauna.

The Hawaiian Drosophila are one of the most species-rich endemic groups in Hawaii and a spectacular example of adaptive radiation. Drosophila silvestris and D. heteroneura are two closely related picture-winged Drosophila species that occur sympatrically on Hawaii Island and are known to hybridize in nature, yet exhibit highly divergent behavioral and morphological traits driven largely through sexual selection. Their closest-related allopatric species, D. planitibia from Maui, exhibits hybrid male sterility and reduced behavioral reproductive isolation when crossed experimentally with D. silvestris or D. heteroneura. A modified four-taxon test for gene flow was applied to recently obtained genomes of the three Hawaiian Drosophila species. The analysis indicates recent gene flow in sympatry, but also, although less extensive, between allopatric species. This study underscores the prevalence of gene flow, even in taxonomic groups considered classic examples of allopatric speciation on islands. The potential confounding effects of gene flow in phylogenetic and population genetics inference are discussed, as well as the implications for conservation.

Irradiation is a postharvest quarantine treatment option for exported commodities such as stone fruits and small fruits to prevent movement of the new invasive pest spotted wing drosophila, Drosophila suzukii (Walker) (Diptera: Drosophilidae). The effects of irradiation on larval and pupal development and adult reproduction in D. suzukii were examined. Larvae (first, second, and third instars) and pupae (1-2-d-old, 3-5-d-old, and 7-8-d-old) on diet were irradiated at target doses of 20, 30, 40, and 50 Gy in replicated factorial experiments and survival to the adult stage was recorded. Tolerance to radiation increased with increasing age and developmental stage. Males and females were equally susceptible. A radiation dose of 40 Gy applied to first- and second-instar larvae prevented adult emergence. The late-stage pupa was the most radiation-tolerant stage that occurs in fruit, and individuals irradiated at this stage readily emerged as adults; therefore, prevention of F1 adults was the desired treatment response for large-scale validation tests with naturally infested fruit. In large-scale tests, a radiation dose of 80 Gy applied to late-stage pupae in sweet cherries or grapes resulted in no production of F1 adults in > 33,000 treated individuals, which meets the zero tolerance requirement for market access. A minimum absorbed dose of 80 Gy is recommended for quarantine control of D. suzukii.

Resolving species relationships within recent radiations requires analysis at the interface of phylogenetics and population genetics, where coalescence and hybridization may confound our understanding of relationships. We developed 18 new primer pairs for nuclear loci in Cyrtandra (Gesneriaceae), one of the largest plant radiations in the Pacific Islands, and tested the concordance of 14 loci in establishing the phylogenetic relationships of a small number of Hawaiian species. Four genes yielded tree topologies conflicting with the primary concordance tree, suggesting plastid capture and horizontal transfer via hybridization. Combining all concordant genes yielded a tree with stronger support and a different topology from the total-evidence tree. We conclude that a small number of genes may be insufficient for accurate reconstruction of the phylogenetic relationships among closely related species. Further, the combination of genes for phylogenetic analysis without preliminary concordance tests can yield an erroneous tree topology. It seems that the number of genes needed for phylogenetic analysis of closely related species is significantly greater than the small numbers commonly used, which fail to isolate coalescence, introgression and hybridization.

The Hawaiian picture-winged flies in the genus Drosophila are a spectacular example of rapid evolutionary diversification in which sexual selection is considered an important mechanism for reproductive isolation and speciation. We investigated the behavioral reproductive isolation of two closely related and sympatric Hawaiian picture-winged Drosophila species, D. silvestris and D. heteroneura, which are known to hybridize in nature and produce viable and fertile hybrids. We compared the mating success of parental, F1 and backcross males in pairings with D. heteroneura females. The F1 males were produced by mating D. heteroneura males with D. silvestris females, and the backcross males were produced by mating F1 females with D. heteroneura males. The mating success of backcross males paired with D. heteroneura females were significantly reduced relative to that of parental and F1 males. This reduced mating success occurred primarily at a late stage of courtship where female choice of mate may be important. Two- and three-gene models demonstrate that epistasis involving a few genes could account for the observed variation in male mating success. These results are consistent with negative epistasis in the backcross generation and support the importance of sexual selection and negative epistasis in the evolution and maintenance of these species.

The Island of Hawai'i is a dynamic assemblage of five volcanoes with wet forest habitat currently existing in four distinct natural regions that vary in area, age and geographical isolation. In this complex landscape, alternative assumptions of the relative importance of specific habitat characteristics on evolutionary and ecological processes predict strikingly different general patterns of local diversity and regional similarity. In this study, we compare alternative a priori hypotheses against observed patterns within two distinct biological systems and scales: community composition of wet forest vascular plant species and mitochondrial and nuclear genes of Drosophila sproati, a wet-forest-restricted endemic. All observed patterns display strong and similar regional structuring, with the greatest local diversity found in Kohala and the windward side of Mauna Loa, the least in Ka'ū and Kona, and a distinctive pattern of regional similarity that probably reflects the historical development of this habitat on the island. These observations largely corroborate a biogeographical model that integrates multiple lines of evidence, including climatic reconstruction, over those relying on single measures, such as current habitat configuration or substrate age. This method of testing alternative hypotheses across biological systems and scales is an innovative approach for understanding complex landscapes and should prove valuable in diverse biogeographical systems.

It has been hypothesized that protecting herbivorous fishes within Marine Reserves (MRs) will help these areas to resist algal overgrowth of corals. However, we lack empirical studies demonstrating the validity of key assumptions underpinning this concept, including that herbivorous fishes (1) are permanently resident within MR boundaries, (2) routinely consume the invasive algae of concern at significant levels and (3) are not significant mobile vectors for propagules of the invasive algae. A 46-year-old MR (Hawaii Marine Laboratory Refuge, 21°27′35″N, 157°48′15″W) in waters off Oahu provided a natural setting to examine these factors with the herbivorous bluespine unicornfish (Naso unicornis) and the invasive rhodophyte, Gracilaria salicornia. We acoustically tracked five unicornfish to quantify their residency and habitat use within the MR, conducted field and laboratory observations to confirm N. unicornis consume G. salicornia, and tested the viability of G. salicornia fragments recovered from unicornfish feces. Unicornfish were resident within the MR where they spent most time in reef crest habitat with occasional, brief forays into reef flat habitat. We confirmed unicornfish consume G. salicornia and found a significant positive correlation between algal canopy height and distance from the reef crest, presumably because grazing intensity is lower in less frequently utilized reef flat habitat. We demonstrated that unicornfish egest viable fragments of G. salicornia that resume vegetative growth after several weeks. We conclude that N. unicornis may act as both a natural control agent and a mobile vector for invasive alien algae.

The little fire ant, Wasmannia auropunctata (Roger) (Hymenoptera: Formicidae), is an invasive ant that forms supercolonies when it successfully invades new areas. W. auropunctata was first reported in Hawaii in 1999, and it has since invaded a variety of agricultural sites, including nurseries, orchards, and pastures. Amdro (hydramethylnon; in bait stations), Esteem (pyriproxyfen; broadcast bait), and Conserve (spinosad; ground spray) were tested for their efficacy against W. auropunctata in a rambutan, Nephelium lappaceum L. and mangosteen, Garcinia mangostana L., orchard by making treatments every 2 wk for 16 wk. Relative estimates of ant numbers in plots was determined by transect sampling using peanut butter-baited sticks. Significant treatment effects were observed on weeks 13-17, with reductions in ant counts occurring in the Amdro and Esteem treatments. During this period, the reduction in ant numbers from pretreatment counts averaged 47.1 and 92.5% in the Amdro and Esteem plots, respectively, whereas ant numbers in the untreated control plots increased by 185.9% compared with pretreatment counts. Conserve did not cause a reduction in ant counts as applied in our experiment. No plots for any of the treatments achieved 100% reduction. Pseudococcidae were counted on branch terminals at 4-wk intervals. The two predominant species, Nipaecoccus nipae (Maskell) and Nipaecoccus viridis (Newstead) were significantly lower in the Amdro and Esteem treatments on week 16 compared with controls. Many W. auropunctata were found nesting in protected sites in the orchard trees, which may have compromised the ground-based control methods. Absolute density estimates from shallow core samples taken from the orchard floor indicated the W. auropunctata supercolony exceeded 244 million ants and 22.7 kg wet weight per ha.

Over the last several decades many picture-winged Drosophila have become less common in both geographical distribution and local population size (pers. obs., Foote pers. comm., Montgomerey pers. comm.). Here we report on a study of two Hawaiian Drosophila species, D. engyochracea, and D. hawaiiensis, to determine the impact that changes in population sizes over the past thirty years have had on the genetic diversity of these species. D. engyochracea is known from only two locations on the Island of Hawai'i (Kipuka Ki and Kipuka Pua'ulu), while D. hawaiiensis is currently more wide spread across Hawai'i Island. We collected 65 D. hawaiiensis and 66 D. engyochracea from two forest patches (kipuka) isolated by a 400 year old volcanic ash deposit. DNA sequence data for 515 bases of the mitochondrial gene COII was analyzed for both species to estimate relative total genetic diversity as well as inter-kipuka gene flow. The more wide spread species, D. hawaiiensis, has more genetic diversity (23 vs. 11 unique haplotypes) than the rarer species, D. engyochracea. The distribution of haplotypes in the kipuka is consistent with more gene flow in D. engyochracea than in D. hawaiiensis. Phylogenetic analysis indicates a small number of individuals morphologically identified as one species but have DNA sequence diagnostic for the other species. These results are consistent with these individuals being descendant from hybrids between species.

Aim Understanding how ecological and evolutionary processes together determine patterns of biodiversity remains a central aim in biology.Guided by ecological theory, we use data from multiple arthropod lineages across the Hawaiian archipelago to explore the interplay between ecological (population dynamics, dispersal, trophic interactions) and evolutionary (genetic structuring, adaptation, speciation, extinction) processes. Our goal is to show how communities develop from the dynamic feedbacks that operate at different temporal and spatial scales. Location The Hawaiian islands (19–22° N, 155–160° W). Methods We synthesize genetic data from selected arthropods across the Hawaiian archipelago to determine the relative role of dispersal and in situ differentiation across the island chronosequence. From four sites on three high islands with geological ages ranging from 1 Myr. Herbivore–plant communities only transiently achieve statistical steady state during assembly, presumably due to incomplete assembly from dispersal in the early stages, and the increasing influence of island ontogeny on older islands.

Two Hawaiian picture-wing Drosophila differ in their temperature tolerances with the ecologically rare species, D. silvestris, showing reduced survival, reduced sperm mobility and greater gene expression changes at high temperatures compared to the common D. sproati. Thus the rare species may have reduced capacity to adapt to future climate changes., Climate change is compounding the threats to the future of biodiversity, already impacted by habitat loss, invasive species and diseases. In the Hawaiian Islands, many of the endemic species have narrow habitat ranges that make them especially vulnerable to climate change. The Hawaiian Drosophila, a remarkably diverse group of species with 11 listed as federally endangered, are thought to be sensitive to temperature changes. To examine the species differences in sensitivity of Hawaiian picture-wing Drosophila to temperature changes, wild populations of Drosophila sproati, a relatively common species, and Drosophila silvestris, a rare species, were collected from two locations on Hawaii Island and bred in common laboratory conditions. Adult flies were exposed to hot and cold temperatures and compared with adult flies at control temperatures. Drosophila silvestris adults were less tolerant to heat stress than D. sproati for both survival and sperm mobility. In contrast, D. silvestris adults were more tolerant to cold stress than D. sproati for adult survival. The expression of 4950 Gene Ontology annotated gene transcripts was also analysed in high-temperature-treated and control males to identify candidate genes related to heat tolerance. There were more than twice as many transcripts differentially expressed after high temperature treatment for D. silvestris (246 transcripts) as for D. sproati (106 transcripts), with 13 Gene Ontology terms enriched between temperatures for D. silvestris and merely three in D. sproati. The combined results are consistent with D. sproati occurring more widely today as well as occurring at lower elevations than D. silvestris and with a genetically based temperature response, which is more severe in D. silvestris at high temperatures than that in D. sproati. These experiments demonstrate the potential for different capacities of species to adapt to future climate change conditions as well as providing an explanation for historical changes in the distribution of species.

We investigate the age and sex distribution of genetic fitness under mutation-selection balance by means of simple one-locus two-allele models. We find that the extent of age and sex variation in the mutation load is very dependent on the average effect of new mutations. If the average heterozygote selective effect of new mutations is large, then age and sex differences may constitute a significant fraction of the total load, and be significant as compared to standing genetic variation. Whether the mutation load will increase or decrease with age depends on the age- and sex-specific effects of the new mutations, and on the rate of accumulation of mutations in the germ line as individuals age. We argue that the load will most likely increase with age in animals with continuous germ-cell division throughout life, and that this will occur even when mutations have unconditionally deleterious effects. We show that a male-biased mutation rate is likely to result in both a male-biased mutation load and a load that increases with male age.

We examined the relationship of genetic quality to age in male Drosophila melanogaster to test two contrasting hypotheses. The traditional hypothesis is that older males have proven their viability and therefore produce offspring of superior genetic quality. This hypothesis is often evoked as an explanation for female preference for older mates. In contrast, we have recently argued that older fathers may produce offspring of inferior genetic quality. Here, we present results from an experiment designed to measure the genetic quality of offspring produced by 2 day old, 2 week old and 5 week old male D. melanogaster. We found a statistically significant small reduction in larval viability and a similar but statistically non-significant reduction in son mating ability among the offspring of the 5 week old males. Daughter fecundity showed no apparent trend for a reduction nor an increase in performance with increasing age of the fathers. There was no evidence of a difference between the 2 day old and the 2 week old males for any of these three fitness components. These results are in somewhat better accordance with our alternative hypothesis, but the relatively weak and late occurring effects indicate that mate choice based on age may not be a viable strategy in this population.

Although trees comprise a primary component of terrestrial species richness, the drivers and temporal scale of divergence in trees remain poorly understood. We examined the landscape-dominant tree, Metrosideros polymorpha, for variation at nine microsatellite loci across 23 populations on young Hawai'i Island, sampling each of the island's five varieties throughout its full geographic range. For four varieties, principal coordinate analysis revealed strong clustering of populations by variety across the 10 430 km(2) island, indicating partitioning of the species into multiple evolutionarily significant units. The single island-endemic form, riparian var. newellii, showed especially strong differentiation from other varieties despite occurring in sympatry with other varieties and likely evolved from a bog form on the oldest volcano, Kohala, within the past 500 000 years. Along with comparable riparian forms on other Pacific Islands, var. newellii appears to represent parallel incipient ecological speciation within Metrosideros. Greater genetic distance among the more common varieties on the oldest volcano and an inverse relationship between allelic diversity and substrate age appear consistent with colonization of Hawai'i Island by older, partially diverged varieties followed by increased hybridization among varieties on younger volcanoes. This study demonstrates that broad population-level sampling is required to uncover patterns of diversification within a ubiquitous and long-lived tree species. Hawaiian Metrosideros appears to be a case of incipient radiation in trees and thus should be useful for studies of divergence and the evolution of reproductive isolating barriers at the early stages of speciation.

Zebra finch (Taeniopygia guttata) bills are sexually dichromatic with males possessing redder bills than females. Males with redder bills have been shown to be preferred as mates by females and to produce more offspring in laboratory experiments. In contrast, females with redder bills have suffered higher mortality and produced fewer offspring. In this paper, I report on a quantitative genetic study of male and female bill colour from a cross-fostering experiment. The heritability estimates for bill colour between parents and their biological offspring ranged from 0.23 to 0.58 and all but one of these estimates is significantly greater than zero. The heritability estimates between cross-fostered offspring and their non-biological parents were not significantly different from zero. Likewise, the genetic correlation between male and female bill colour was large (r$_{g}$ = 0.81) for parents and their biological offspring, but not for cross-fostered offspring and their non-biological parents. I also demonstrate the extent that the genetic correlation between the sexes can retard the predicted evolution of bill colour and create a selection load that reduces the expected reproduction of this laboratory population.

It has been suggested that female preference for older mates in species without parental care has evolved in response to an alleged higher genetic quality of older individuals. This is based on the widespread assumption that viability selection produces older individuals that are genetically superior to younger individuals. In contrast, we propose that the oldest individuals rarely are genetically superior. Quantitative genetic models of life history evolution indicate that young to intermediately aged individuals are likely to possess the highest breeding values of fitness. This conclusion is based on four arguments: 1) Viability selection on older individuals may decrease or at least not substantially increase breeding values of fitness, because there may exist negative genetic correlations between late-age and early-age life history parameters, 2) Fertility selection is likely to raise the fitness of gametes produced by young individuals more than those produced by old individuals, because the covariance between fertility and fitness often decreases with age, 3) The history of selection on their parents makes younger individuals more fit than older individuals, 4) Germ-line mutations, which are generally deleterious, significantly decrease the breeding value of fitness of an individual throughout its lifespan, especially in males. Therefore, females that mate with the oldest males in a population are doing so for reasons other than to obtain offspring of high genetic quality.

We investigated the role that courtship and aggressive interactions may have for the maintenance of reproductive isolation betweenDrosophila silvestris andD. heteroneura. We examined the behavioral bases of reproductive isolation between the parental species and we examined the courtship success of each sex of both reciprocal F1 hybrids when paired with the parental species. We found reduced copulation success among heterotypic parental pairs compared to homotypic pairs, which was primarily due to the lack of courtship initiation betweenD. silvestris males andD. heteroneura females. When hybrid males from both reciprocal crosses were paired with parental females their copulation successes were not significantly different from that of parental males. In contrast, hybrid females from both crosses had reduced copulation success withD. silvestris males, which in turn was primarily due to a reduced success of reaching later stages of courtship. The time spent in copulation by hybrid males was intermediate between the two parental males. We studied aggression by observing the interactions of males of heterotypic pairs, both between the parental species and between the hybrids and parental males. A lack of aggressive interactions betweenD. silvestris males andD. heteroneura males in addition to the lack of courtship suggests thatD. silvestris males do not respond toD. heteroneura individuals of either sex. Hybrid males were equally successful in winning fights with bothD. silvestris andD. heteroneura males. These results indicate that the behavioral isolation betweenD. silvestris andD. heteroneura may be largely a consequence of the earliest stages of interactions. The two species may differ either in activity levels or in morphological or chemical traits that are important for species and mate recognition. The relatively high copulation and aggressive success of hybrids indicates that sexual selection against hybrids alone is unlikely to be a sufficient force to reduce gene flow and maintain species distinctions.

We examined natural and sexual selection in a laboratory population of zebra finches (Taeniopygia guttata), with a principal focus on bill color. Previous studies indicated that females prefer males with redder bills, whereas males prefer females with less red bills. For male red bill color, both the selection differential and selection gradient coefficients were significantly positive with fitness (the number of offspring surviving to independence), which was primarily due to differences in the time to the first nest and in reproductive rate but not to differences in survival. In contrast, for female red bill color, there was significant negative selection, which was due to differences in most stages of reproductive and survival selection but not to differences in the time to the first nest. There were no significant correlations between male or female bill color and offspring weight or postfledging survival. All male plumage color traits (breast band, cheek patch, flank patch) showed nonsignificant sele...

The nēnē (Branta sandvicensis) is an endangered Hawaiian goose endemic to the Hawaiian Islands. The nēnē nearly went extinct in the mid-19.00s and the majority of the approximately 1300 individuals currently in Hawai'i are descendants from less than 30 birds. The low adult breeding success and juvenile survival is likely due, in part, to inbreeding depression in wild individuals. Thirty-eight microsatellite primer sets developed in nēnē, Canada goose, and waterfowl species provided 8 polymorphic loci. Four of these polymorphic loci exhibited only two alleles, which is likely a reflection of the high inbreeding in this species.