The evolutionary process of invasion in the fall armyworm (Spodoptera frugiperda).

The fall armyworm (FAW; Spodoptera frugiperda) is one of the major agricultural pest insects. FAW is native to the Americas, and its invasion was first reported in West Africa in 2016. Then it quickly spread through Africa, Asia, and Oceania, becoming one of the main threats to corn production. We analyzed whole genome sequences of 177 FAW individuals from 12 locations on four continents to infer evolutionary processes of invasion. Principal component analysis from the TPI gene and whole genome sequences shows that invasive FAW populations originated from the corn strain. Ancestry coefficient and phylogenetic analyses from the nuclear genome indicate that invasive populations are derived from a single ancestry, distinct from native populations, while the mitochondrial phylogenetic tree supports the hypothesis of multiple introductions. Adaptive evolution specific to invasive populations was observed in detoxification, chemosensory, and digestion genes. We concluded that extant invasive FAW populations originated from the corn strain with potential contributions of adaptive evolution.

A novel reference dated phylogeny for the genus Spodoptera Guenée (Lepidoptera: Noctuidae: Noctuinae): new insights into the evolution of a pest-rich genus.

The noctuid genus Spodoptera currently consists of 31 species with varied host plant breadths, ranging from monophagous and oligophagous non-pest species to polyphagous pests of economic importance. Several of these pest species have become major invaders, colonizing multiple continents outside their native range. Such is the case of the infamous fall armyworm, Spodoptera frugiperda (J.E. Smith), which includes two recognized host strains that have not been treated as separate species. Following its accidental introduction to Africa in 2016, it quickly spread through Africa and Asia to Australia. Given that half the described Spodoptera species cause major crop losses, comparative genomics studies of several Spodoptera species have highlighted major adaptive changes in genetic architecture, possibly relating to their pest status. Several recent population genomics studies conducted on two species enable a more refined understanding of their population structures, migration patterns and invasion processes. Despite growing interest in the genus, the taxonomic status of several Spodoptera species remains unstable and evolutionary studies suffer from the absence of a robust and comprehensive dated phylogenetic framework. We generated mitogenomic data for 14 Spodoptera taxa, which are combined with data from 15 noctuoid outgroups to generate a resolved mitogenomic backbone phylogeny using both concatenation and multi-species coalescent approaches. We combine this backbone with additional mitochondrial and nuclear data to improve our understanding of the evolutionary history of the genus. We also carry out comprehensive dating analyses, which implement three distinct calibration strategies based on either primary or secondary fossil calibrations. Our results provide an updated phylogenetic framework for 28 Spodoptera species, identifying two well-supported ecologically diverse clades that are recovered for the first time. Well-studied larvae in each of these clades are characterized by differences in mandibular shape, with one clade's being more specialized on silica-rich C4 grasses. Interestingly, the inferred timeframe for the genus suggests an earlier origin than previously thought for the genus: about 17-18 million years ago.

Genome-wide macroevolutionary signatures of key innovations in butterflies colonizing new host plants.

The mega-diversity of herbivorous insects is attributed to their co-evolutionary associations with plants. Despite abundant studies on insect-plant interactions, we do not know whether host-plant shifts have impacted both genomic adaptation and species diversification over geological times. We show that the antagonistic insect-plant interaction between swallowtail butterflies and the highly toxic birthworts began 55 million years ago in Beringia, followed by several major ancient host-plant shifts. This evolutionary framework provides a valuable opportunity for repeated tests of genomic signatures of macroevolutionary changes and estimation of diversification rates across their phylogeny. We find that host-plant shifts in butterflies are associated with both genome-wide adaptive molecular evolution (more genes under positive selection) and repeated bursts of speciation rates, contributing to an increase in global diversification through time. Our study links ecological changes, genome-wide adaptations and macroevolutionary consequences, lending support to the importance of ecological interactions as evolutionary drivers over long time periods.

Whole Genome Shotgun Phylogenomics Resolves the Pattern and Timing of Swallowtail Butterfly Evolution.

Evolutionary relationships have remained unresolved in many well-studied groups, even though advances in next-generation sequencing and analysis, using approaches such as transcriptomics, anchored hybrid enrichment, or ultraconserved elements, have brought systematics to the brink of whole genome phylogenomics. Recently, it has become possible to sequence the entire genomes of numerous nonbiological models in parallel at reasonable cost, particularly with shotgun sequencing. Here, we identify orthologous coding sequences from whole-genome shotgun sequences, which we then use to investigate the relevance and power of phylogenomic relationship inference and time-calibrated tree estimation. We study an iconic group of butterflies-swallowtails of the family Papilionidae-that has remained phylogenetically unresolved, with continued debate about the timing of their diversification. Low-coverage whole genomes were obtained using Illumina shotgun sequencing for all genera. Genome assembly coupled to BLAST-based orthology searches allowed extraction of 6621 orthologous protein-coding genes for 45 Papilionidae species and 16 outgroup species (with 32% missing data after cleaning phases). Supermatrix phylogenomic analyses were performed with both maximum-likelihood (IQ-TREE) and Bayesian mixture models (PhyloBayes) for amino acid sequences, which produced a fully resolved phylogeny providing new insights into controversial relationships. Species tree reconstruction from gene trees was performed with ASTRAL and SuperTriplets and recovered the same phylogeny. We estimated gene site concordant factors to complement traditional node-support measures, which strengthens the robustness of inferred phylogenies. Bayesian estimates of divergence times based on a reduced data set (760 orthologs and 12% missing data) indicate a mid-Cretaceous origin of Papilionoidea around 99.2 Ma (95% credibility interval: 68.6-142.7 Ma) and Papilionidae around 71.4 Ma (49.8-103.6 Ma), with subsequent diversification of modern lineages well after the Cretaceous-Paleogene event. These results show that shotgun sequencing of whole genomes, even when highly fragmented, represents a powerful approach to phylogenomics and molecular dating in a group that has previously been refractory to resolution.

Serial horizontal transfer of vitamin-biosynthetic genes enables the establishment of new nutritional symbionts in aphids' di-symbiotic systems.

Many insects depend on obligate mutualistic bacteria to provide essential nutrients lacking from their diet. Most aphids, whose diet consists of phloem, rely on the bacterial endosymbiont Buchnera aphidicola to supply essential amino acids and B vitamins. However, in some aphid species, provision of these nutrients is partitioned between Buchnera and a younger bacterial partner, whose identity varies across aphid lineages. Little is known about the origin and the evolutionary stability of these di-symbiotic systems. It is also unclear whether the novel symbionts merely compensate for losses in Buchnera or carry new nutritional functions. Using whole-genome endosymbiont sequences of nine Cinara aphids that harbour an Erwinia-related symbiont to complement Buchnera, we show that the Erwinia association arose from a single event of symbiont lifestyle shift, from a free-living to an obligate intracellular one. This event resulted in drastic genome reduction, long-term genome stasis, and co-divergence with aphids. Fluorescence in situ hybridisation reveals that Erwinia inhabits its own bacteriocytes near Buchnera's. Altogether these results depict a scenario for the establishment of Erwinia as an obligate symbiont that mirrors Buchnera's. Additionally, we found that the Erwinia vitamin-biosynthetic genes not only compensate for Buchnera's deficiencies, but also provide a new nutritional function; whose genes have been horizontally acquired from a Sodalis-related bacterium. A subset of these genes have been subsequently transferred to a new Hamiltonella co-obligate symbiont in one specific Cinara lineage. These results show that the establishment and dynamics of multi-partner endosymbioses can be mediated by lateral gene transfers between co-ocurring symbionts.

Opposite macroevolutionary responses to environmental changes in grasses and insects during the Neogene grassland expansion.

The rise of Neogene C4 grasslands is one of the most drastic changes recently experienced by the biosphere. A central - and widely debated - hypothesis posits that Neogene grasslands acted as a major adaptive zone for herbivore lineages. We test this hypothesis with a novel model system, the Sesamiina stemborer moths and their associated host-grasses. Using a comparative phylogenetic framework integrating paleoenvironmental proxies we recover a negative correlation between the evolutionary trajectories of insects and plants. Our results show that paleoenvironmental changes generated opposing macroevolutionary dynamics in this insect-plant system and call into question the role of grasslands as a universal adaptive cradle. This study illustrates the importance of implementing environmental proxies in diversification analyses to disentangle the relative impacts of biotic and abiotic drivers of macroevolutionary dynamics.

A Freeloader? The Highly Eroded Yet Large Genome of the Serratia symbiotica Symbiont of Cinara strobi.

Genome reduction is pervasive among maternally inherited bacterial endosymbionts. This genome reduction can eventually lead to serious deterioration of essential metabolic pathways, thus rendering an obligate endosymbiont unable to provide essential nutrients to its host. This loss of essential pathways can lead to either symbiont complementation (sharing of the nutrient production with a novel co-obligate symbiont) or symbiont replacement (complete takeover of nutrient production by the novel symbiont). However, the process by which these two evolutionary events happen remains somewhat enigmatic by the lack of examples of intermediate stages of this process. Cinara aphids (Hemiptera: Aphididae) typically harbor two obligate bacterial symbionts: Buchnera and Serratia symbiotica. However, the latter has been replaced by different bacterial taxa in specific lineages, and thus species within this aphid lineage could provide important clues into the process of symbiont replacement. In the present study, using 16S rRNA high-throughput amplicon sequencing, we determined that the aphid Cinara strobi harbors not two, but three fixed bacterial symbionts: Buchnera aphidicola, a Sodalis sp., and S. symbiotica. Through genome assembly and genome-based metabolic inference, we have found that only the first two symbionts (Buchnera and Sodalis) actually contribute to the hosts' supply of essential nutrients while S. symbiotica has become unable to contribute towards this task. We found that S. symbiotica has a rather large and highly eroded genome which codes only for a few proteins and displays extensive pseudogenization. Thus, we propose an ongoing symbiont replacement within C. strobi, in which a once "competent" S. symbiotica does no longer contribute towards the beneficial association. These results suggest that in dual symbiotic systems, when a substitute cosymbiont is available, genome deterioration can precede genome reduction and a symbiont can be maintained despite the apparent lack of benefit to its host.

Convergent herbivory on conifers by Choristoneura moths after boreal forest formation.

Mitogenomes are useful markers for phylogenetic studies across a range of taxonomic levels. Here, we focus on mitogenome variation across the tortricid moth genus Choristoneura and particularly the spruce budworm (Choristoneura fumiferana) species complex, a notorious pest group of North American conifer forests. Phylogenetic relationships of Tortricidae, representing two subfamilies, four tribes and nine genera, were analyzed using 21 mitogenomes. These included six newly-sequenced mitogenomes for species in the spruce budworm complex plus three additional Choristoneura species and 12 previously published mitogenomes from other tortricids and one from the Cossidae. We evaluated the phylogenetic informativeness of the mitogenomes and reconstructed a time-calibrated tree with fossil and secondary calibrations. We found that tortricid mitogenomes had conserved protein and ribosomal regions, and analysis of all protein-coding plus ribosomal genes together provided an efficient marker at any taxonomic rank. The time-calibrated phylogeny showed evolutionary convergence of conifer feeding within Choristoneura, with two independent lineages, the Nearctic spruce budworm complex and the Palearctic species Choristoneura murinana, both shifting onto conifers about 11 million years ago from angiosperms. These two host-plant shifts both occurred after the formation of boreal forest in the late Miocene. Haplotype diversification within the spruce budworm complex occurred in the last 4 million years, and is probably linked to the initial cooling cycles of the Northern Hemisphere in the Pliocene.

A first higher-level time-calibrated phylogeny of antlions (Neuroptera: Myrmeleontidae).

In this study, we reconstruct the first time-calibrated phylogeny of the iconic antlion family, the Myrmeleontidae (Neuroptera: Myrmeleontiformia). We use maximum likelihood and Bayesian inference to analyse a molecular dataset based on seven mitochondrial and nuclear gene markers. The dataset encompasses 106 species of Neuroptera, including 94 antlion species. The resulting phylogenetic framework provides support for a myrmeleontid classification distinguishing four subfamilies: Acanthaclisinae, Myrmeleontinae, Palparinae, and Stilbopteryginae. Within Myrmeleontinae, Myrmecaelurini and Nemoleontini are recovered as monophyletic clades; Gepini also appears as a valid tribe, distinct from Myrmecaelurini whereas Myrmecaelurini and Nesoleontini on one hand and Brachynemurini and Dendroleontini on the other hand, appear closely related. Some preliminary information related to generic and specific levels are also implied from our results, such as the paraphyly of several genera. Dating analyses based on thoroughly evaluated fossil calibrations indicate that the antlion family likely originated in the Cretaceous, between 135 and 138 million years ago (depending on the set of fossil calibrations), and that all higher-level lineages appeared during the Early Cretaceous. This first phylogenetic hypothesis will provide a valuable basis to further expand the taxonomic coverage and molecular sampling, and to lay the foundations of future systematic revisions.

Deciphering the evolution of birdwing butterflies 150 years after Alfred Russel Wallace.

One hundred and fifty years after Alfred Wallace studied the geographical variation and species diversity of butterflies in the Indomalayan-Australasian Archipelago, the processes responsible for their biogeographical pattern remain equivocal. We analysed the macroevolutionary mechanisms accounting for the temporal and geographical diversification of the charismatic birdwing butterflies (Papilionidae), a major focus of Wallace's pioneering work. Bayesian phylogenetics and dating analyses of the birdwings were conducted using mitochondrial and nuclear genes. The combination of maximum likelihood analyses to estimate biogeographical history and diversification rates reveals that diversity-dependence processes drove the radiation of birdwings, and that speciation was often associated with founder-events colonizing new islands, especially in Wallacea. Palaeo-environment diversification models also suggest that high extinction rates occurred during periods of elevated sea level and global warming. We demonstrated a pattern of spatio-temporal habitat dynamics that continuously created or erased habitats suitable for birdwing biodiversity. Since birdwings were extinction-prone during the Miocene (warmer temperatures and elevated sea levels), the cooling period after the mid-Miocene climatic optimum fostered birdwing diversification due to the release of extinction. This also suggests that current global changes may represent a serious conservation threat to this flagship group.

Phylogenetic molecular species delimitations unravel potential new species in the pest genus Spodoptera Guenée, 1852 (Lepidoptera, Noctuidae).

Nowadays molecular species delimitation methods promote the identification of species boundaries within complex taxonomic groups by adopting innovative species concepts and theories (e.g. branching patterns, coalescence). As some of them can efficiently deal with large single-locus datasets, they could speed up the process of species discovery compared to more time consuming molecular methods, and benefit from the existence of large public datasets; these methods can also particularly favour scientific research and actions dealing with threatened or economically important taxa. In this study we aim to investigate and clarify the status of economically important moths species belonging to the genus Spodoptera (Lepidoptera, Noctuidae), a complex group in which previous phylogenetic analyses and integrative approaches already suggested the possible occurrence of cryptic species and taxonomic ambiguities. In this work, the effectiveness of innovative (and faster) species delimitation approaches to infer putative species boundaries has been successfully tested in Spodoptera, by processing the most comprehensive dataset (in terms of number of species and specimens) ever achieved; results are congruent and reliable, irrespective of the set of parameters and phylogenetic models applied. Our analyses confirm the existence of three potential new species clusters (for S. exigua (Hübner, 1808), S. frugiperda (J.E. Smith, 1797) and S. mauritia (Boisduval, 1833)) and support the synonymy of S. marima (Schaus, 1904) with S. ornithogalli (Guenée, 1852). They also highlight the ambiguity of the status of S. cosmiodes (Walker, 1858) and S. descoinsi Lalanne-Cassou & Silvain, 1994. This case study highlights the interest of molecular species delimitation methods as valuable tools for species discovery and to emphasize taxonomic ambiguities.

Spodoptera frugiperda (Lepidoptera: Noctuidae) host-plant variants: two host strains or two distinct species?

The moth Spodoptera frugiperda is a well-known pest of crops throughout the Americas, which consists of two strains adapted to different host-plants: the first feeds preferentially on corn, cotton and sorghum whereas the second is more associated with rice and several pasture grasses. Though morphologically indistinguishable, they exhibit differences in their mating behavior, pheromone compositions, and show development variability according to the host-plant. Though the latter suggest that both strains are different species, this issue is still highly controversial because hybrids naturally occur in the wild, not to mention the discrepancies among published results concerning mating success between the two strains. In order to clarify the status of the two host-plant strains of S. frugiperda, we analyze features that possibly reflect the level of post-zygotic isolation: (1) first generation (F1) hybrid lethality and sterility; (2) patterns of meiotic segregation of hybrids in reciprocal second generation (F2), as compared to the meiosis of the two parental strains. We found a significant reduction of mating success in F1 in one direction of the cross and a high level of microsatellite markers showing transmission ratio distortion in the F2 progeny. Our results support the existence of post-zygotic reproductive isolation between the two laboratory strains and are in accordance with the marked level of genetic differentiation that was recovered between individuals of the two strains collected from the field. Altogether these results provide additional evidence in favor of a sibling species status for the two strains.

Cretaceous environmental changes led to high extinction rates in a hyperdiverse beetle family.

BACKGROUND: As attested by the fossil record, Cretaceous environmental changes have significantly impacted the diversification dynamics of several groups of organisms. A major biome turnover that occurred during this period was the rise of angiosperms starting ca. 125 million years ago. Though there is evidence that the latter promoted the diversification of phytophagous insects, the response of other insect groups to Cretaceous environmental changes is still largely unknown. To gain novel insights on this issue, we assess the diversification dynamics of a hyperdiverse family of detritivorous beetles (Tenebrionidae) using molecular dating and diversification analyses. RESULTS: Age estimates reveal an origin after the Triassic-Jurassic mass extinction (older than previously thought), followed by the diversification of major lineages during Pangaean and Gondwanan breakups. Dating analyses indicate that arid-adapted species diversified early, while most of the lineages that are adapted to more humid conditions diversified much later. Contrary to other insect groups, we found no support for a positive shift in diversification rates during the Cretaceous; instead there is evidence for an 8.5-fold increase in extinction rates that was not compensated by a joint increase in speciation rates. CONCLUSIONS: We hypothesize that this pattern is better explained by the concomitant reduction of arid environments starting in the mid-Cretaceous, which likely negatively impacted the diversification of arid-adapted species that were predominant at that time.

Integrative taxonomy of New Caledonian beetles: species delimitation and definition of the Uloma isoceroides species group (Coleoptera, Tenebrionidae, Ulomini), with the description of four new species.

New Caledonia is an important biodiversity hotspot with much undocumented biodiversity, especially in many insect groups. Here we used an integrative approach to explore species diversity in the tenebrionid genus Uloma (Coleoptera, Tenebrionidae, Ulomini), which encompasses about 150 species, of which 22 are known from New Caledonia. To do so, we focused on a morphologically homogeneous group by comparing museum specimens with material collected during several recent field trips. We also conducted molecular phylogenetic analyses based on a concatenated matrix of four mitochondrial and three nuclear genes for 46 specimens. The morphological study allowed us to discover and describe four new species that belong to the group of interest, the Uloma isoceroides group. Molecular analyses confirmed the species boundaries of several of the previously described species and established the validity of the four new species. The phylogenetic analyses also provided additional information on the evolutionary history of the group, highlighting that a species that was thought to be unrelated to the group was in fact a member of the same evolutionary lineage. Molecular species delimitation confirmed the status of the sampled species of the group and also suggested some hidden (cryptic) biodiversity for at least two species of the group. Altogether this integrative taxonomic approach has allowed us to better define the boundaries of the Uloma isoceroides species group, which comprises at least 10 species: Uloma isoceroides (Fauvel, 1904), Uloma opacipennis (Fauvel, 1904), Uloma caledonica Kaszab, 1982, Uloma paniei Kaszab, 1982, Uloma monteithi Kaszab, 1986, Uloma robusta Kaszab, 1986, Uloma clamensae sp. n., Uloma condaminei sp. n., Uloma jourdani sp. n., and Uloma kergoati sp. n. We advocate more studies on other New Caledonian groups, as we expect that much undocumented biodiversity can be unveiled through the use of similar approaches.

DNA barcoding and the associated PhylAphidB@se website for the identification of European aphids (Insecta: Hemiptera: Aphididae).

UNLABELLED: Aphids constitute a diverse group of plant-feeding insects and are among the most important crop pests in temperate regions. Their morphological identification is time-consuming and requires specific knowledge, training and skills that may take years to acquire. We assessed the advantages and limits of DNA barcoding with the standard COI barcode fragment for the identification of European aphids. We constructed a large reference dataset of barcodes from 1020 specimens belonging to 274 species and 87 genera sampled throughout Europe and set up a database-driven website allowing species identification from query sequences. RESULTS: In this unbiased sampling of the taxonomic diversity of European aphids, intraspecific divergence ranged from 0.0% to 3.9%, with a mean value of 0.29%, whereas mean congeneric divergence was 6.4%, ranging from 0.0% to 15%. Neighbor-joining analysis generated a tree in which most species clustered in distinct genetic units. Most of the species with undifferentiated or overlapping barcodes belonged to the genus Aphis or, to a lesser extent, the genera Brachycaudus, Dysaphis and Macrosiphum. The taxa involved were always morphologically similar or closely related and belonged to species groups known to present taxonomic difficulties. CONCLUSIONS: These data confirm that COI barcoding is a useful identification tool for aphids. Barcode identification is straightforward and reliable for 80% of species, including some difficult to distinguish on the basis of morphological characters alone. Unsurprisingly, barcodes often failed to distinguish between species from groups for which classical taxonomy has also reached its limits, leading to endless revisions and discussions about species and subspecies definitions. In such cases, the development of an effective procedure for the accurate identification of aphid specimens continues to pose a difficult challenge.

Disentangling dispersal, vicariance and adaptive radiation patterns: a case study using armyworms in the pest genus Spodoptera (Lepidoptera: Noctuidae).

Thanks to the recent development of integrative approaches that combine dated phylogenies with models of biogeographic evolution, it is becoming more feasible to assess the roles of dispersal and vicariance in creating complex patterns of geographical distribution. However, the historical biogeography of taxa with good dispersal abilities, like birds or flying insects, still remains largely unknown because of the lack of complete phylogenies accompanied by robust estimates of divergence times. In this study, we investigate the evolution and historical biogeography of the globally distributed pest genus Spodoptera (Lepidoptera: Noctuidae) using complete taxon sampling and an extensive set of analyses. Through the analysis of a combined morphological and molecular dataset, we provide the first robust phylogenetic framework for this widespread and economically important group of moths. Historical biogeography approaches indicate that dispersal events have been the driving force in the biogeographic history of the group. One of the most interesting findings of this study is the probable occurrence of two symmetric long-distance dispersal events between the Afrotropical and the Neotropical region, which appear to have occurred in the late Miocene. Even more remarkably, our dated phylogenies reveal that the diversification of the clade that includes specialist grass feeders has followed closely the expansion of grasslands in the Miocene, similar to the adaptive radiation of specialist grazing mammals during the same period.

Permanent genetic resources added to Molecular Ecology Resources Database 1 December 2011-31 January 2012.

This article documents the addition of 473 microsatellite marker loci and 71 pairs of single-nucleotide polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Barteria fistulosa, Bombus morio, Galaxias platei, Hematodinium perezi, Macrocentrus cingulum Brischke (a.k.a. M. abdominalis Fab., M. grandii Goidanich or M. gifuensis Ashmead), Micropogonias furnieri, Nerita melanotragus, Nilaparvata lugens Stål, Sciaenops ocellatus, Scomber scombrus, Spodoptera frugiperda and Turdus lherminieri. These loci were cross-tested on the following species: Barteria dewevrei, Barteria nigritana, Barteria solida, Cynoscion acoupa, Cynoscion jamaicensis, Cynoscion leiarchus, Cynoscion nebulosus, Cynoscion striatus, Cynoscion virescens, Macrodon ancylodon, Menticirrhus americanus, Nilaparvata muiri and Umbrina canosai. This article also documents the addition of 116 sequencing primer pairs for Dicentrarchus labrax.