Richness and resilience in the Pacific: DNA metabarcoding enables parallelized evaluation of biogeographic patterns.

Islands make up a large proportion of Earth's biodiversity, yet are also some of the most sensitive systems to environmental perturbation. Biogeographic theory predicts that geologic age, area, and isolation typically drive islands' diversity patterns, and thus potentially impact non-native spread and community homogenization across island systems. One limitation in testing such predictions has been the difficulty of performing comprehensive inventories of island biotas and distinguishing native from introduced taxa. Here, we use DNA metabarcoding and statistical modelling as a high throughput method to survey community-wide arthropod richness, the proportion of native and non-native species, and the incursion of non-natives into primary habitats on three archipelagos in the Pacific - the Ryukyus, the Marianas and Hawaii - which vary in age, isolation and area. Diversity patterns largely match expectations based on island biogeography theory, with the oldest and most geographically connected archipelago, the Ryukyus, showing the highest taxonomic richness and lowest proportion of introduced species. Moreover, we find evidence that forest habitats are more resilient to incursions of non-natives in the Ryukyus than in the less taxonomically rich archipelagos. Surprisingly, we do not find evidence for biotic homogenization across these three archipelagos: the assemblage of non-native species on each island is highly distinct. Our study demonstrates the potential of DNA metabarcoding to facilitate rapid estimation of biogeographic patterns, the spread of non-native species, and the resilience of ecosystems.

Collective and harmonized high throughput barcoding of insular arthropod biodiversity: Toward a Genomic Observatories Network for islands.

Current understanding of ecological and evolutionary processes underlying island biodiversity is heavily shaped by empirical data from plants and birds, although arthropods comprise the overwhelming majority of known animal species, and as such can provide key insights into processes governing biodiversity. Novel high throughput sequencing (HTS) approaches are now emerging as powerful tools to overcome limitations in the availability of arthropod biodiversity data, and hence provide insights into these processes. Here, we explored how these tools might be most effectively exploited for comprehensive and comparable inventory and monitoring of insular arthropod biodiversity. We first reviewed the strengths, limitations and potential synergies among existing approaches of high throughput barcode sequencing. We considered how this could be complemented with deep learning approaches applied to image analysis to study arthropod biodiversity. We then explored how these approaches could be implemented within the framework of an island Genomic Observatories Network (iGON) for the advancement of fundamental and applied understanding of island biodiversity. To this end, we identified seven island biology themes at the interface of ecology, evolution and conservation biology, within which collective and harmonized efforts in HTS arthropod inventory could yield significant advances in island biodiversity research.

Categorization of species as native or nonnative using DNA sequence signatures without a complete reference library.

New genetic diagnostic approaches have greatly aided efforts to document global biodiversity and improve biosecurity. This is especially true for organismal groups in which species diversity has been underestimated historically due to difficulties associated with sampling, the lack of clear morphological characteristics, and/or limited availability of taxonomic expertise. Among these methods, DNA sequence barcoding (also known as "DNA barcoding") and by extension, meta-barcoding for biological communities, has emerged as one of the most frequently utilized methods for DNA-based species identifications. Unfortunately, the use of DNA barcoding is limited by the availability of complete reference libraries (i.e., a collection of DNA sequences from morphologically identified species), and by the fact that the vast majority of species do not have sequences present in reference databases. Such conditions are critical especially in tropical locations that are simultaneously biodiversity rich and suffer from a lack of exploration and DNA characterization by trained taxonomic specialists. To facilitate efforts to document biodiversity in regions lacking complete reference libraries, we developed a novel statistical approach that categorizes unidentified species as being either likely native or likely nonnative based solely on measures of nucleotide diversity. We demonstrate the utility of this approach by categorizing a large sample of specimens of terrestrial insects and spiders (collected as part of the Moorea BioCode project) using a generalized linear mixed model (GLMM). Using a training data set of known endemic (n = 45) and known introduced species (n = 102), we then estimated the likely native/nonnative status for 4,663 specimens representing an estimated 1,288 species (412 identified species), including both those specimens that were either unidentified or whose endemic/introduced status was uncertain. Using this approach, we were able to increase the number of categorized specimens by a factor of 4.4 (from 794 to 3,497), and the number of categorized species by a factor of 4.8 from (147 to 707) at a rate much greater than chance (77.6% accuracy). The study identifies phylogenetic signatures of both native and nonnative species and suggests several practical applications for this approach including monitoring biodiversity and facilitating biosecurity.

Stream flow alone does not predict population structure of diving beetles across complex tropical landscapes.

Recent theoretical advances have hypothesized a central role of habitat persistence on population genetic structure and resulting biodiversity patterns of freshwater organisms. Here, we address the hypothesis that lotic species, or lineages adapted to comparably geologically stable running water habitats (streams and their marginal habitats), have high levels of endemicity and phylogeographic structure due to the persistent nature of their habitat. We use a nextRAD DNA sequencing approach to investigate the population structure and phylogeography of a putatively widespread New Guinean species of diving beetle, Philaccolilus ameliae (Dytiscidae). We find that P. ameliae is a complex of morphologically cryptic, but geographically and genetically well-differentiated clades. The pattern of population connectivity is consistent with theoretical predictions associated with stable lotic habitats. However, in two clades, we find a more complex pattern of low population differentiation, revealing dispersal across rugged mountains and watersheds of New Guinea up to 430 km apart. These results, while surprising, were also consistent with the original formulation of the habitat template concept by Southwood, involving lineage-idiosyncratic evolution in response to abiotic factors. In our system, low population differentiation might reflect a young species in a phase of range expansion utilizing vast available habitat. We suggest that predictions of life history variation resulting from the dichotomy between lotic and lentic organisms require more attention to habitat characterization and microhabitat choice. Our results also underpin the necessity to study fine-scale processes but at a larger geographical scale, as compared to solely documenting macroecological patterns, to understand ecological drivers of regional biodiversity. Comprehensive sampling especially of tropical lineages in complex and threatened environments such as New Guinea remains a critical challenge.

Phylogeography and population genomics of a lotic water beetle across a complex tropical landscape.

The habitat template concept applied to a freshwater system indicates that lotic species, or those which occupy permanent habitats along stream courses, are less dispersive than lentic species, or those that occur in more ephemeral aquatic habitats. Thus, populations of lotic species will be more structured than those of lentic species. Stream courses include both flowing water and small, stagnant microhabitats that can provide refuge when streams are low. Many species occur in these microhabitats but remain poorly studied. Here, we present population genetic data for one such species, the tropical diving beetle Exocelina manokwariensis (Dytiscidae), sampled from six localities along a ~300 km transect across the Birds Head Peninsula of New Guinea. Molecular data from both mitochondrial (CO1 sequences) and nuclear (ddRAD loci) regions document fine-scale population structure across populations that are ~45 km apart. Our results are concordant with previous phylogenetic and macroecological studies that applied the habitat template concept to aquatic systems. This study also illustrates that these diverse but mostly overlooked microhabitats are promising study systems in freshwater ecology and evolutionary biology. With the advent of next-generation sequencing, fine-scale population genomic studies are feasible for small nonmodel organisms to help illuminate the effect of habitat stability on species' natural history, population structure and geographic distribution.

Museum specimen data reveal emergence of a plant disease may be linked to increases in the insect vector population.

The emergence rate of new plant diseases is increasing due to novel introductions, climate change, and changes in vector populations, posing risks to agricultural sustainability. Assessing and managing future disease risks depends on understanding the causes of contemporary and historical emergence events. Since the mid-1990s, potato growers in the western United States, Mexico, and Central America have experienced severe yield loss from Zebra Chip disease and have responded by increasing insecticide use to suppress populations of the insect vector, the potato psyllid, Bactericera cockerelli (Hemiptera: Triozidae). Despite the severe nature of Zebra Chip outbreaks, the causes of emergence remain unknown. We tested the hypotheses that (1) B. cockerelli occupancy has increased over the last century in California and (2) such increases are related to climate change, specifically warmer winters. We compiled a data set of 87,000 museum specimen occurrence records across the order Hemiptera collected between 1900 and 2014. We then analyzed changes in B. cockerelli distribution using a hierarchical occupancy model using changes in background species lists to correct for collecting effort. We found evidence that B. cockerelli occupancy has increased over the last century. However, these changes appear to be unrelated to climate changes, at least at the scale of our analysis. To the extent that species occupancy is related to abundance, our analysis provides the first quantitative support for the hypothesis that B. cockerelli population abundance has increased, but further work is needed to link B. cockerelli population dynamics to Zebra Chip epidemics. Finally, we demonstrate how this historical macro-ecological approach provides a general framework for comparative risk assessment of future pest and insect vector outbreaks.

Temporal dynamics influenced by global change: bee community phenology in urban, agricultural, and natural landscapes.

Urbanization and agricultural intensification of landscapes are important drivers of global change, which in turn have direct impacts on local ecological communities leading to shifts in species distributions and interactions. Here, we illustrate how human-altered landscapes, with novel ornamental and crop plant communities, result not only in changes to local community diversity of floral-dependent species, but also in shifts in seasonal abundance of bee pollinators. Three years of data on the spatio-temporal distributions of 91 bee species show that seasonal patterns of abundance and species richness in human-altered landscapes varied significantly less compared to natural habitats in which floral resources are relatively scarce in the dry summer months. These findings demonstrate that anthropogenic environmental changes in urban and agricultural systems, here mediated through changes in plant resources and water inputs, can alter the temporal dynamics of pollinators that depend on them. Changes in phenology of interactions can be an important, though frequently overlooked, mechanism of global change.

A range-wide genetic bottleneck overwhelms contemporary landscape factors and local abundance in shaping genetic patterns of an alpine butterfly (Lepidoptera: Pieridae: Colias behrii).

Spatial and environmental heterogeneity are major factors in structuring species distributions in alpine landscapes. These landscapes have also been affected by glacial advances and retreats, causing alpine taxa to undergo range shifts and demographic changes. These nonequilibrium population dynamics have the potential to obscure the effects of environmental factors on the distribution of genetic variation. Here, we investigate how demographic change and environmental factors influence genetic variation in the alpine butterfly Colias behrii. Data from 14 microsatellite loci provide evidence of bottlenecks in all population samples. We test several alternative models of demography using approximate Bayesian computation (ABC), with the results favouring a model in which a recent bottleneck precedes rapid population growth. Applying independent calibrations to microsatellite loci and a nuclear gene, we estimate that this bottleneck affected both northern and southern populations 531-281 years ago, coinciding with a period of global cooling. Using regression approaches, we attempt to separate the effects of population structure, geographical distance and landscape on patterns of population genetic differentiation. Only 40% of the variation in F(ST) is explained by these models, with geographical distance and least-cost distance among meadow patches selected as the best predictors. Various measures of genetic diversity within populations are also decoupled from estimates of local abundance and habitat patch characteristics. Our results demonstrate that demographic change can have a disproportionate influence on genetic diversity in alpine species, contrasting with other studies that suggest landscape features control contemporary demographic processes in high-elevation environments.

Pleistocene origin and population history of a neoendemic alpine butterfly.

Alpine environments underwent dramatic transformation during glacial-interglacial cycles, with the consequence that geographical, ecological and demographic changes of alpine populations provided the opportunity for formation of neoendemic species. Several biogeographical models have been proposed to account for the unique history of alpine populations, with different expectations of genetic divergence and speciation. The expanding alpine archipelago model proposes that alpine populations expand spatially and demographically during glacial events, dispersing between mountain ranges. Under this model, alpine populations are unlikely to diverge in isolation due to substantial interpopulation gene flow. In contrast, the alpine archipelago refuge model proposes that gene flow during glacial phases is limited and populations expand demographically during interglacial phases, increasing genetic isolation and the likelihood of speciation. We assess these models by reconstructing the evolutionary history of Colias behrii, a morphologically and ecologically distinct alpine butterfly restricted to the California Sierra Nevada. C. behrii exhibits very low genetic diversity at mitochondrial and nuclear loci, limited population structure and evidence of population expansion. C. behrii and Rocky Mountain C. meadii share identical mitochondrial haplotypes, while in contrast, nuclear data indicate common ancestry between C. behrii and Cascades Range Colias pelidne. The conflict in gene genealogies may be a result of recent expansion in North American Colias, but an isolation with migration analysis indicates that genetic patterns in C. behrii might result from differential introgression following hybridization. Based on the timing of population expansion and gene flow between mountain ranges, the expanding alpine archipelago model is supported in C. behrii.

Evolutionary diversification of cryophilic Grylloblatta species (Grylloblattodea: Grylloblattidae) in alpine habitats of California.

BACKGROUND: Climate in alpine habitats has undergone extreme variation during Pliocene and Pleistocene epochs, resulting in repeated expansion and contraction of alpine glaciers. Many cold-adapted alpine species have responded to these climatic changes with long-distance range shifts. These species typically exhibit shallow genetic differentiation over a large geographical area. In contrast, poorly dispersing organisms often form species complexes within mountain ranges, such as the California endemic ice-crawlers (Grylloblattodea: Grylloblattidae: Grylloblatta). The diversification pattern of poorly dispersing species might provide more information on the localized effects of historical climate change, the importance of particular climatic events, as well as the history of dispersal. Here we use multi-locus genetic data to examine the phylogenetic relationships and geographic pattern of diversification in California Grylloblatta. RESULTS: Our analysis reveals a pattern of deep genetic subdivision among geographically isolated populations of Grylloblatta in California. Alpine populations diverged from low elevation populations and subsequently diversified. Using a Bayesian relaxed clock model and both uncalibrated and calibrated measurements of time to most recent common ancestor, we reconstruct the temporal diversification of alpine Grylloblatta populations. Based on calibrated relaxed clock estimates, evolutionary diversification of Grylloblatta occurred during the Pliocene-Pleistocene epochs, with an initial dispersal into California during the Pliocene and species diversification in alpine clades during the middle Pleistocene epoch. CONCLUSIONS: Grylloblatta species exhibit a high degree of genetic subdivision in California with well defined geographic structure. Distinct glacial refugia can be inferred within the Sierra Nevada, corresponding to major, glaciated drainage basins. Low elevation populations are sister to alpine populations, suggesting alpine populations may track expanding glacial ice sheets and diversify as a result of multiple glacial advances. Based on relaxed-clock molecular dating, the temporal diversification of Grylloblatta provides evidence for the role of a climate-driven species pump in alpine species during the Pleistocene epoch.

Male-killing bacteria trigger a cycle of increasing male fatigue and female promiscuity.

Sex-ratio distorters are found in numerous species and can reach high frequencies within populations. Here, we address the compelling, but poorly tested, hypothesis that the sex ratio bias caused by such elements profoundly alters their host's mating system. We compare aspects of female and male reproductive biology between island populations of the butterfly Hypolimnas bolina that show varying degrees of female bias, because of a male-killing Wolbachia infection. Contrary to expectation, female bias leads to an increase in female mating frequency, up to a point where male mating capacity becomes limiting. We show that increased female mating frequency can be explained as a facultative response to the depleted male mating resources in female biased populations. In other words, this system is one where male-killing bacteria trigger a vicious circle of increasing male fatigue and female promiscuity.

Post-colonization temporal genetic variation of an introduced fly, Rhagoletis completa.

Evolutionary biologists have been puzzled by the success of introduced species: despite founder effects that reduce genetic variability, invasive species are still successful at colonizing new environments. It is possible that the evolutionary processes during the post-colonization period may increase the genetic diversity and gene flow among invasive populations over time, facilitating their long-term success. Therefore, genetic diversity and population structure would be expected to show greater temporal variation for successful introduced populations than for native populations. We studied the population genetics of the walnut husk fly, Rhagoletis completa, which was introduced into California from the Midwestern US in the early 1900s. We used microsatellites and allozymes to genotype current and historic fly populations, providing a rare perspective on temporal variability in population genetic parameters. We found that introduced populations showed greater temporal fluctuations in allele frequencies than native populations. Some introduced populations also showed an increase in genetic diversity over time, indicating multiple introductions had occurred. Population genetic structure decreased in both native and introduced populations over time. Our study demonstrates that introduced species are not at equilibrium and post-colonization processes may be important in ameliorating the loss of genetic diversity associated with biological invasions.

How to better predict long-term benefits and risks in weed biocontrol: an evolutionary perspective.

Classical biological control (also called importation biological control) of weeds has a remarkable track record for efficiency and safety, but further improvement is still needed, particularly to account for potential evolutionary changes after release. Here, we discuss the increasing yet limited evidence of post-introduction evolution and describe approaches to predict evolutionary change. Recent advances include using experimental evolution studies over several generations that combine -omics tools with behavioral bioassays. This novel approach in weed biocontrol is well suited to explore the potential for rapid evolutionary change in real-time and thus can be used to estimate more accurately potential benefits and risks of agents before their importation. We outline this approach with a chrysomelid beetle used to control invasive common ragweed.

The Global Museum: natural history collections and the future of evolutionary science and public education.

Natural history museums are unique spaces for interdisciplinary research and educational innovation. Through extensive exhibits and public programming and by hosting rich communities of amateurs, students, and researchers at all stages of their careers, they can provide a place-based window to focus on integration of science and discovery, as well as a locus for community engagement. At the same time, like a synthesis radio telescope, when joined together through emerging digital resources, the global community of museums (the 'Global Museum') is more than the sum of its parts, allowing insights and answers to diverse biological, environmental, and societal questions at the global scale, across eons of time, and spanning vast diversity across the Tree of Life. We argue that, whereas natural history collections and museums began with a focus on describing the diversity and peculiarities of species on Earth, they are now increasingly leveraged in new ways that significantly expand their impact and relevance. These new directions include the possibility to ask new, often interdisciplinary questions in basic and applied science, such as in biomimetic design, and by contributing to solutions to climate change, global health and food security challenges. As institutions, they have long been incubators for cutting-edge research in biology while simultaneously providing core infrastructure for research on present and future societal needs. Here we explore how the intersection between pressing issues in environmental and human health and rapid technological innovation have reinforced the relevance of museum collections. We do this by providing examples as food for thought for both the broader academic community and museum scientists on the evolving role of museums. We also identify challenges to the realization of the full potential of natural history collections and the Global Museum to science and society and discuss the critical need to grow these collections. We then focus on mapping and modelling of museum data (including place-based approaches and discovery), and explore the main projects, platforms and databases enabling this growth. Finally, we aim to improve relevant protocols for the long-term storage of specimens and tissues, ensuring proper connection with tomorrow's technologies and hence further increasing the relevance of natural history museums.

The joint evolutionary histories of Wolbachia and mitochondria in Hypolimnas bolina.

BACKGROUND: The interaction between the Blue Moon butterfly, Hypolimnas bolina, and Wolbachia has attracted interest because of the high prevalence of male-killing achieved within the species, the ecological consequences of this high prevalence, the intensity of selection on the host to suppress the infection, and the presence of multiple Wolbachia infections inducing different phenotypes. We examined diversity in the co-inherited marker, mtDNA, and the partitioning of this between individuals of different infection status, as a means to investigate the population biology and evolutionary history of the Wolbachia infections. RESULTS: Part of the mitochondrial COI gene was sequenced from 298 individuals of known infection status revealing ten different haplotypes. Despite very strong biological evidence that the sample represents a single species, the ten haplotypes did not fall within a monophyletic clade within the Hypolimnas genus, with one haplotype differing by 5% from the other nine. There were strong associations between infection status and mtDNA haplotype. The presence of wBol1 infection in association with strongly divergent haplotypes prompted closer examination of wBol1 genetic variation. This revealed the existence of two cryptic subtypes, wBol1a and wBol1b. The wBol1a infection, by far the most common, was in strict association with the single divergent mtDNA haplotype. The wBol1b infection was found with two haplotypes that were also observed in uninfected specimens. Finally, the wBol2 infection was associated with a large diversity of mtDNA haplotypes, most often shared with uninfected sympatric butterflies. CONCLUSION: This data overall supports the hypothesis that high prevalence of male-killing Wolbachia (wBol1) in H. bolina is associated with very high transmission efficiency rather than regular horizontal transmission. It also suggests this infection has undergone a recent selective sweep and was introduced in this species through introgression. In contrast, the sharing of haplotypes between wBol2-infected and uninfected individuals indicates that this strain is not perfectly transmitted and/or shows a significant level of horizontal transmission.

Competing selfish genetic elements in the butterfly Hypolimnas bolina.

Maternally inherited selfish genetic elements are common in animals . Whereas host genetics and ecology are recognized as factors that may limit the incidence of these parasites , theory suggests one further factor-interference with other selfish elements-that could affect their prevalence . In this paper, we show that spatial heterogeneity in the occurrence of the male-killing Wolbachia wBol1 in the tropical butterfly Hypolimnas bolina is caused by a second infection that can exclude the male-killer. We first provide evidence of a second Wolbachia strain, wBol2, present in most populations that do not carry the male-killer but rare or absent when the male-killer is present. Crossing data indicate that wBol2 in males induces cytoplasmic incompatibility to both uninfected and wBol1-infected females. The wBol2 infection can therefore not only spread through uninfected populations but also resist invasion by wBol1. Thus, we provide empirical support for the hypothesis that the incidence of particular selfish genetic elements can limit the presence of competing types.

Alpine biogeography of Parnassian butterflies during Quaternary climate cycles in North America.

Growth of alpine glaciers during the Pleistocene had profound effects on montane landscapes in North America and the organisms now inhabiting alpine ecosystems. Biogeography of this region has often been viewed as a system of sky islands despite the fact that species richness patterns deviate from a strict island biogeographic model. One explanation is that alpine species are not in equilibrium because of late Quaternary geographic range shifts. Genetic data can provide evidence of nonequilibrium dynamics and the distributional shifts that occur during glaciation events in alpine landscapes. Using mitochondrial and nuclear sequence data, we examine the evolutionary history of butterflies in the Parnassius phoebus complex. We test explicit, alternative models of the biogeographic history of Parnassius smintheus and Parnassius behrii, including an equilibrium island model, ancestral radiation and fragmentation, an expanding alpine archipelago and an alpine archipelago refuge model. Our results support the alpine archipelago refuge model, in which alpine butterflies undergo population contraction during glacial climates followed by population expansion during interglacial phases. While butterflies can disperse between distant mountain ranges during glacial periods, gene flow is rare. We find evidence of recent connectivity between California and Colorado, population expansion events following deglaciation approximately 20,000 years B.P., and small population sizes during the last glacial period. An analysis of lineage splitting suggests that morphological differences in P. smintheus and P. behrii are the result of late Pleistocene divergence (approximately 48,000 years B.P.) with limited gene flow. Our results demonstrate that spatially complex and nonequilibrium population dynamics influence alpine diversity patterns.

Evolution of male-killer suppression in a natural population.

Male-killing bacteria are widespread in arthropods, and can profoundly alter the reproductive biology of their host species. Here we detail the first case of complete suppression of a male killer. The nymphalid butterfly Hypolimnas bolina is infected with a strain of the bacterium Wolbachia, wBol1, which kills male host embryos in Polynesian populations, but does not do so in many areas of Southeast Asia, where both males and female adults are naturally infected, and wBol1-infected females produce a 1:1 sex ratio. We demonstrate that absence of male killing by wBol1 is associated with dominant zygotic suppression of the action of the male killer. Simulations demonstrate host suppressors of male-killer action can spread very rapidly, and historical data indicating the presence of male killing in Southeast Asia in the very recent past suggests suppressor spread has been a very recent occurrence. Thus, male killer/host interactions are much more dynamic than previously recognised, with rapid and dramatic loss of the phenotype. Our results also indicate that suppression can render male killers completely quiescent, leading to the conclusion that some species that do not currently express a male killer may have done so in the past, and thus that more species have had their biology affected by these parasites than previously believed.

You can't keep a good parasite down: evolution of a male-killer suppressor uncovers cytoplasmic incompatibility.

Maternally inherited parasites are known to impose a wide variety of reproductive manipulations upon their host. These often produce strong selection on the host to suppress the parasite, resulting in a reduction in the frequency of the parasite. However, in the butterfly Hypolimnas bolina, infected with a Wolbachia bacterium, field data demonstrate that suppression of the male-killing phenotype does not depress parasite frequency. Here we test and verify one hypothesis to explain this apparent paradox-Wolbachia induces a second phenotype, Cytoplasmic Incompatibility (CI), in populations where host suppression has evolved. We further demonstrate that the capacity to induce CI has not evolved de novo, but instead is instantaneously expressed upon the survival of infected males. The significance of these results is threefold: (1) multiple phenotypes can provide Wolbachia with the means to maintain itself in a host following suppression of a single manipulative phenotype; (2) the ability to induce CI can remain hidden in systems in which male-killing is observed, just as the ability to induce male-killing may be obscured in strains exhibiting CI; (3) the evolutionary maintenance of CI in a system in which it is not expressed suggests a functional link with male-killing or other traits under selection.