Deep Lineages Research Articles

Correction to: Genome-Scale Data Reveal Deep Lineage Divergence and a Complex Demographic History in the Texas Horned Lizard (Phrynosoma cornutum) throughout the Southwestern and Central United States

Correction to: Genome-Scale Data Reveal Deep Lineage Divergence and a Complex Demographic History in the Texas Horned Lizard (<i>Phrynosoma cornutum</i>) throughout the Southwestern and Central United States

Phylogenomic Coalescent Analyses of Avian Retroelements Infer Zero-Length Branches at the Base of Neoaves, Emergent Support for Controversial Clades, and Ancient Introgressive Hybridization in Afroaves.

Retroelement insertions (RIs) are low-homoplasy characters that are ideal data for addressing deep evolutionary radiations, where gene tree reconstruction errors can severely hinder phylogenetic inference with DNA and protein sequence data. Phylogenomic studies of Neoaves, a large clade of birds (>9000 species) that first diversified near the Cretaceous–Paleogene boundary, have yielded an array of robustly supported, contradictory relationships among deep lineages. Here, we reanalyzed a large RI matrix for birds using recently proposed quartet-based coalescent methods that enable inference of large species trees including branch lengths in coalescent units, clade-support, statistical tests for gene flow, and combined analysis with DNA-sequence-based gene trees. Genome-scale coalescent analyses revealed extremely short branches at the base of Neoaves, meager branch support, and limited congruence with previous work at the most challenging nodes. Despite widespread topological conflicts with DNA-sequence-based trees, combined analyses of RIs with thousands of gene trees show emergent support for multiple higher-level clades (Columbea, Passerea, Columbimorphae, Otidimorphae, Phaethoquornithes). RIs express asymmetrical support for deep relationships within the subclade Afroaves that hints at ancient gene flow involving the owl lineage (Strigiformes). Because DNA-sequence data are challenged by gene tree-reconstruction error, analysis of RIs represents one approach for improving gene tree-based methods when divergences are deep, internodes are short, terminal branches are long, and introgressive hybridization further confounds species–tree inference.

An integrative taxonomic revision and redefinition of Gephyromantis (Laurentomantis) malagasius based on archival DNA analysis reveals four new mantellid frog species from Madagascar

The subgenus Laurentomantis in the genusGephyromantiscontains some of the least known amphibian species of Madagascar. The six currently valid nominal species are rainforest frogs known from few individuals, hampering a full understanding of the species diversity of the clade. We assembled data on specimens collected during field surveys over the past 30 years and integrated analysis of mitochondrial and nuclear-encoded genes of 88 individuals, a comprehensive bioacoustic analysis, and morphological comparisons to delimit a minimum of nine species-level lineages in the subgenus. To clarify the identity of the speciesGephyromantis malagasius, we applied a target-enrichment approach to a sample of the 110 year-old holotype ofMicrophryne malagasiaMethuen and Hewitt, 1913 to assign this specimen to a lineage based on a mitochondrial DNA barcode. The holotype clustered unambiguously with specimens previously namedG. ventrimaculatus. Consequently we propose to considerTrachymantis malagasia ventrimaculatusAngel, 1935 as a junior synonym ofGephyromantis malagasius. Due to this redefinition ofG. malagasius, no scientific name is available for any of the four deep lineages of frogs previously subsumed under this name, all characterized by red color ventrally on the hindlimbs. These are here formally named asGephyromantis fiharimpesp. nov.,G. matsilosp. nov.,G. oelkrugisp. nov., andG. portonaesp. nov.The new species are distinguishable from each other by genetic divergences of &amp;gt;4% uncorrected pairwise distance in a fragment of the 16S rRNA marker and a combination of morphological and bioacoustic characters.Gephyromantis fiharimpeandG. matsilooccur, respectively, at mid-elevations and lower elevations along a wide stretch of Madagascar’s eastern rainforest band, whileG. oelkrugiandG. portonaeappear to be more range-restricted in parts of Madagascar’s North East and Northern Central East regions. Open taxonomic questions surroundG. horridus, to which we here assign specimens from Montagne d’Ambre and the type locality Nosy Be; andG. ranjomavo, which contains genetically divergent populations from Marojejy, Tsaratanana, and Ampotsidy.

Depth as a driver of evolution and diversification of ancient squat lobsters (Decapoda, Galatheoidea, Phylladiorhynchus)

The exceptional hidden diversity included in the squat lobster genus Phylladiorhynchus and its wide bathymetric and geographic range make it an interesting group to thoroughly study its evolutionary history. Here we have analyzed the entire currently known species diversity of Phylladiorhynchus using an integrative approach that includes morphological and molecular characters. The aim was to establish whether depth range (bathymetry) has played a role in their morphological and molecular evolution and in their diversification pathways. Phylogenetic analyses recovered the genus as monophyletic and as the sister group of Coralliogalathea, conforming with current systematic hypotheses, although their placement in a monophyletic Galatheidae is doubted. All the analyzed species represent well-supported lineages, structured in ten clades, correlated in most part with the morphological phylogeny. The reconstruction of ancestral habitat showed that the most recent common ancestor of Phylladiorhynchus most likely lived in shallow water environments. The divergence time estimation analyses dated the origin of the genus back to the Upper Jurassic, preceding the origin of all the other galatheoid lineages. Morphological analyses suggested that species from deeper waters exhibit greater morphological divergences and lower genetic divergences in comparison to species from shallower waters. In Phylladiorhynchus, the colonization of deeper waters has taken place independently multiple times since the Lower-Cretaceous. Our reconstruction of ancestral habitat suggests that shallow water ancestors might show an acceleration in the molecular rate of evolution and a slowdown in the rates of morphological evolution in comparison to deep sea lineages. However, although lineages from shallow and deep sea habitats show slight differences in diversification trends, bathymetry does not significantly affect the diversification rate in Phylladiorhynchus according to our diversification analyses.

Accessory Genome Dynamics of Local and Global Staphylococcus pseudintermedius Populations.

Staphylococcus pseudintermedius is a major bacterial colonizer and opportunistic pathogen in dogs. Methicillin-resistant S. pseudintermedius (MRSP) continues to emerge as a significant challenge to maintaining canine health. We sought to determine the phylogenetic relationships of S. pseudintermedius across five states in the New England region of the United States and place them in a global context. The New England dataset consisted of 125 previously published S. pseudintermedius genomes supplemented with 45 newly sequenced isolates. The core genome phylogenetic tree revealed many deep branching lineages consisting of 142 multi-locus sequence types (STs). In silico detection of the mecA gene revealed 40 MRSP and 130 methicillin-susceptible S. pseudintermedius (MSSP) isolates. MRSP were derived from five structural types of SCCmec, the mobile genetic element that carries the mecA gene conferring methicillin resistance. Although many genomes were MSSP, they nevertheless harbored genes conferring resistance to many other antibiotic classes, including aminoglycosides, macrolides, tetracyclines and penams. We compared the New England genomes to 297 previously published genomes sampled from five other states in the United States and 13 other countries. Despite the prevalence of the clonally expanding ST71 found worldwide and in other parts of the United States, we did not detect it in New England. We next sought to interrogate the combined New England and global datasets for the presence of coincident gene pairs linked to antibiotic resistance. Analysis revealed a large co-circulating accessory gene cluster, which included mecA as well as eight other resistance genes [aac (6′)-Ie-aph (2″)-Ia, aad (6), aph (3′)-IIIa, sat4, ermB, cat, blaZ, and tetM]. Furthermore, MRSP isolates carried significantly more accessory genes than their MSSP counterparts. Our results provide important insights to the evolution and geographic spread of high-risk clones that can threaten the health of our canine companions.

Culture-Independent Survey of Thermophilic Microbial Communities of the North Caucasus.

Simple SummaryThe Republic of North Ossetia-Alania, located in the southern part of the North Caucasus, possess a number of hydrothermal habitats, including both subterranean thermal reservoirs and terrestrial hot springs. At the same time, reports on microbiology of numerous geothermal sites are rather scarce for the whole North Caucasus region. In this paper, we report on the first culture-independent metabarcoding study of thermal habitats in the North Caucasus, coupled with a chemical analysis of the elemental composition of water. The results of this work include the conclusions regarding key metabolic characteristics of these habitats as well as detection of few but abundant deep lineages of uncultivated microorganisms which could be regarded as endemic. This study may represent a first step in closing the knowledge gap in extremophilic microbial communities of the North Caucasus.The Greater Caucasus is a part of seismically active Alpine–Himalayan orogenic belt and has been a center of significant volcanic activity during the Quaternary period. That led to the formation of the number of hydrothermal habitats, including subterranean thermal aquifers and surface hot springs. However, there are only a limited number of scientific works reporting on the microbial communities of these habitats. Moreover, all these reports concern only studies of specific microbial taxa, carried out using classical cultivation approaches. In this work, we present first culture-independent study of hydrotherms in the Republic of North Ossetia-Alania, located in the southern part of the North Caucasus. Using 16S metabarcoding, we analyzed the composition of the microbial communities of two subterranean thermal aquifers and terrestrial hot springs of the Karmadon valley. Analysis of correlations between the chemical composition of water and the representation of key taxa allowed us to identify the key factors determining the formation of microbial communities. In addition, we were able to identify a significant number of highly abundant deep phylogenetic lineages. Our study represents a first glance on the thermophilic microbial communities of the North Caucasus and may serve as a basis for further microbiological studies of the extreme habitats of this region.

Mesophotic Gorgonian Corals Evolved Multiple Times and Faster Than Deep and Shallow Lineages

Mesophotic Coral Ecosystems (MCEs) develop on a unique environment, where abrupt environmental changes take place. Using a time-calibrated molecular phylogeny (mtDNA: mtMutS), we examined the lineage membership of mesophotic gorgonian corals (Octocorallia: Cnidaria) in comparison to shallow and deep-sea lineages of the wider Caribbean-Gulf of Mexico and the Tropical Eastern Pacific. Our results show mesophotic gorgonians originating multiple times from old deep-sea octocoral lineages, whereas shallow-water species comprise younger lineages. The mesophotic gorgonian fauna in the studied areas is related to their zooxanthellate shallow-water counterparts in only two clades (Gorgoniidae and Plexauridae), where the bathymetrical gradient could serve as a driver of diversification. Interestingly, mesophotic clades have diversified faster than either shallow or deep clades. One of this groups with fast diversification is the family Ellisellidae, a major component of the mesophotic gorgonian coral assemblage worldwide.

Genome-Scale Data Reveal Deep Lineage Divergence and a Complex Demographic History in the Texas Horned Lizard (Phrynosoma cornutum) throughout the Southwestern and Central United States.

The southwestern and central United States serve as an ideal region to test alternative hypotheses regarding biotic diversification. Genomic data can now be combined with sophisticated computational models to quantify the impacts of paleoclimate change, geographic features, and habitat heterogeneity on spatial patterns of genetic diversity. In this study, we combine thousands of genotyping-by-sequencing (GBS) loci with mtDNA sequences (ND1) from the Texas horned lizard (Phrynosoma cornutum) to quantify relative support for different catalysts of diversification. Phylogenetic and clustering analyses of the GBS data indicate support for at least three primary populations. The spatial distribution of populations appears concordant with habitat type, with desert populations in AZ and NM showing the largest genetic divergence from the remaining populations. The mtDNA data also support a divergent desert population, but other relationships differ and suggest mtDNA introgression. Genotype–environment association with bioclimatic variables supports divergence along precipitation gradients more than along temperature gradients. Demographic analyses support a complex history, with introgression and gene flow playing an important role during diversification. Bayesian multispecies coalescent analyses with introgression (MSci) analyses also suggest that gene flow occurred between populations. Paleo-species distribution models support two southern refugia that geographically correspond to contemporary lineages. We find that divergence times are underestimated and population sizes are overestimated when introgression occurred and is ignored in coalescent analyses, and furthermore, inference of ancient introgression events and demographic history is sensitive to inclusion of a single recently admixed sample. Our analyses cannot refute the riverine barrier or glacial refugia hypotheses. Results also suggest that populations are continuing to diverge along habitat gradients. Finally, the strong evidence of admixture, gene flow, and mtDNA introgression among populations suggests that P. cornutum should be considered a single widespread species under the General Lineage Species Concept.

Automated Deep Lineage Tree Analysis Using a Bayesian Single Cell Tracking Approach

Single-cell methods are beginning to reveal the intrinsic heterogeneity in cell populations, arising from the interplay of deterministic and stochastic processes. However, it remains challenging to quantify single-cell behaviour from time-lapse microscopy data, owing to the difficulty of extracting reliable cell trajectories and lineage information over long time-scales and across several generations. Therefore, we developed a hybrid deep learning and Bayesian cell tracking approach to reconstruct lineage trees from live-cell microscopy data. We implemented a residual U-Net model coupled with a classification CNN to allow accurate instance segmentation of the cell nuclei. To track the cells over time and through cell divisions, we developed a Bayesian cell tracking methodology that uses input features from the images to enable the retrieval of multi-generational lineage information from a corpus of thousands of hours of live-cell imaging data. Using our approach, we extracted 20,000 + fully annotated single-cell trajectories from over 3,500 h of video footage, organised into multi-generational lineage trees spanning up to eight generations and fourth cousin distances. Benchmarking tests, including lineage tree reconstruction assessments, demonstrate that our approach yields high-fidelity results with our data, with minimal requirement for manual curation. To demonstrate the robustness of our minimally supervised cell tracking methodology, we retrieve cell cycle durations and their extended inter- and intra-generational family relationships in 5,000 + fully annotated cell lineages. We observe vanishing cycle duration correlations across ancestral relatives, yet reveal correlated cyclings between cells sharing the same generation in extended lineages. These findings expand the depth and breadth of investigated cell lineage relationships in approximately two orders of magnitude more data than in previous studies of cell cycle heritability, which were reliant on semi-manual lineage data analysis.

Characterizing chloroplast genomes and inferring maternal divergence of the Triticum–Aegilops complex

The Triticum (wheat)–Aegilops (goatgrass) complex has been extensively studied, but the evolutionary history of polyploid wheats has not been fully elucidated. The chloroplast (cp) with maternal inheritance and homoplasy can simplify the sequence-based evolutionary inferences, but informative inferences would require a complete and accurate cp genome sequence. In this study, 16 cp genomes representing five Aegilops and 11 Triticum species and subspecies were sequenced, assembled and annotated, yielding five novel circular cp genome sequences. Analyzing the assembled cp genomes revealed no marked differences in genome structure and gene arrangement across the assayed species. A polymorphism analysis of 72 published cp genome sequences representing 10 Aegilops and 15 Triticum species and subspecies detected 1183 SNPs and 1881 SSRs. More than 80% SNPs detected resided on the downstream and upstream gene regions and only 2.78% or less SNPs were predicted to be deleterious. The largest nucleotide diversity was observed in the short single-copy genomic region. Relatively weak selection pressure on cp coding genes was detected. Different phylogenetic analyses confirmed that the maternal divergence of the Triticum–Aegilops complex had three deep lineages each representing a diploid species with nuclear A, B, or D genome. Dating the maternal divergence yielded age estimates of divergence that matched well with those reported previously. The divergence between emmer and bread wheats occurred at 8200–11,200 years ago. These findings are useful for further genomic studies, provide insight into cp genome evolvability and allow for better understanding of the maternal divergence of the Triticum–Aegilops complex.

Diversity and Distribution of Uncultured and Cultured Gaiellales and Rubrobacterales in South China Sea Sediments.

Actinobacteria are ubiquitous in marine ecosystems, and they are regarded as an important, underexplored, potential pharmaceutical resource. The orders Gaiellales and Rubrobacterales are deep taxonomic lineages of the phylum Actinobacteria, both are represented by a single genus and contain only a few species. Although they have been detected frequently by high-throughput sequencing, their functions and characteristics in marine habitats remain unknown due to the lack of indigenous phenotypes. Here, we investigated the status of the orders in South China Sea (SCS) sediments using culture-independent and culture-dependent methods. Gaiellales is the second-most abundant order of Actinobacteria and was widely distributed in SCS sediments at water depths of 42–4,280 m, and four novel marine representatives in this group were successfully cultured. Rubrobacterales was present at low abundance in energy-limited marine habitats. An isolation strategy for Rubrobacterales from marine samples was proposed, and a total of 138 mesophilic Rubrobacterales strains were isolated under conditions of light and culture time combined with high-salinity or low-nutrient media. Marine representatives recovered in this study formed branches with a complex evolutionary history in the phylogenetic tree. Overall, the data indicate that both Gaiellales and Rubrobacterales can adapt to and survive in extreme deep-sea environments. This study lays the groundwork for further analysis of the distribution and diversity of the orders Gaiellales and Rubrobacterales in the ocean and provides a specific culture strategy for each group. The results open a window for further research on the ecological roles of the two orders in marine ecosystems.

On the utility of taxonomy to reflect biodiversity: the example of Lasiurini (Chiroptera: Vespertilionidae)

The taxonomic history of bats of the tribe Lasiurini (Chiroptera: Vespertilionidae) has undergone significant changes over time. Authors at different times have recognized various numbers of genera and subgenera within the tribe. The most recent proposed change to generic level taxonomy (that there should be three genera recognized instead of a single genus) has been debated in the literature. We reviewed papers that commented on the recent changes to lasiurine generic taxonomy, as well as those that have adopted the new taxonomy and the ones that have not. We also reviewed the relevant taxonomic literature from 1942 to the present that shows the fluid taxonomic history of these bats. The literature review shows that the recently proposed taxonomic change recognizing the three groups of lasiurine bats as distinct genera is the only taxonomy that differentiates the tribe from the genera. Examination of times to most recent common ancestor (TMRCA) of 24 vespertilionid genera shows Lasiurus, if it comprises all Lasiurini, to be an outlier. Here, we support the recognition of three genera and explain how this arrangement best reflects the evolutionary history and biodiversity of the tribe by bringing the three distinct lineages in line with other vespertilionid genera with respect to divergence times and genetic distances. Considering the Lasiurini to comprise a single genus, Lasiurus, that genus has the greatest TMRCA of all vespertilionid genera analyzed, comparable only to the genus Kerivoula of the monotypic subfamily Kerivoulinae. However, recognizing the three deeply diverged lasiurine lineages (red bats, yellow bats, and hoary bats) as genera brings their TMRCAs in line with other genera and approximates the mean TMRCA of the 24 genera analyzed. Opponents of Baird et al.’s taxonomy argued that these three lineages should be considered as subgenera to avoid changing scientific names for purpose of nomenclatural stability and ease of conducting a literature search and because the three deep lineages are all monophyletic. That argument ignores the biological reality that these lineages are morphologically distinct, and that they are genetically as distinct from one another as other genera of vespertilionid bats; there is ample precedent in the mammalian literature to use values of TMRCA as a metric to maintain consistency of higher taxonomic categories such as genus. We encourage other mammalogists to utilize taxonomy to its maximum descriptive potential, while taking into account phylogenetic relationships of the taxa of interest.

Endozoicomonadaceae symbiont in gills of Acesta clam encodes genes for essential nutrients and polysaccharide degradation.

ABSTRACTGammaproteobacteria from the family Endozoicomonadaceae have emerged as widespread associates of dense marine animal communities. Their abundance in coral reefs involves symbiotic relationships and possibly host nutrition. We explored functions encoded in the genome of an uncultured Endozoicomonadaceae ‘Candidatus Acestibacter aggregatus’ that lives inside gill cells of large Acesta excavata clams in deep-water coral reefs off mid-Norway. The dominance and deep branching lineage of this symbiont was confirmed using 16S rRNA gene sequencing and phylogenomic analysis from shotgun sequencing data. The 4.5 Mb genome binned in this study has a low GC content of 35% and is enriched in transposon and chaperone gene annotations indicating ongoing adaptation. Genes encoding functions potentially involved with the symbiosis include ankyrins, repeat in toxins, secretion and nutritional systems. Complete pathways were identified for the synthesis of eleven amino acids and six B-vitamins. A minimal chitinolytic machinery was indicated from a glycosyl hydrolase GH18 and a lytic polysaccharide monooxygenase LPMO10. Expression of the latter was confirmed using proteomics. Signal peptides for secretion were identified for six polysaccharide degrading enzymes, ten proteases and three lipases. Our results suggest a nutritional symbiosis fuelled by enzymatic products from extracellular degradation processes.

Global phylogeography of sailfish: deep evolutionary lineages with implications for fisheries management

Since the Miocene profound climatic changes have influenced the biology and ecology of species worldwide, such as their connectivity, genetic population structure, and biogeography. The goal herein is to evaluate the phylogeography of sailfish Istiophorus platypterus between the Atlantic, Indian, and Pacific oceans. Our results evidenced a high genetic diversity and three distinct populations among the ocean basins with limited gene flow among them. In addition, the species is characterized by two deep evolutionary lineages that diverged during the Miocene/Pliocene transition, one of them is circumtropical while the other is restricted to the Atlantic Ocean. These lineages evolved along the successive glacial-interglacial cycles from the Pleistocene and remained isolated from each other in glacial refugium until deglaciation. Assessments of sailfish suggest it may be subject to overfishing and the results herein imply the need to re-evaluate the current stock delimitations and management measures adopted by the Regional Fisheries Management Organizations, especially in the Atlantic and the Indo-Western Pacific oceans to effectively manage the species. In addition, this work highlights that both lineages should at least be treated as two distinct management units in the Atlantic Ocean until their taxonomic status is fully resolved, given their high genetic divergence.

Evidence of historical isolation and genetic structuring among broadnose sevengill sharks (Notorynchus cepedianus) from the world’s major oceanic regions

Cosmopolitan marine pelagic species display variable patterns of population connectivity among the world’s major oceans. While this information is crucial for informing management, information is lacking for many ecologically important species, including apex predators. In this study we examine patterns of genetic structure in the broadnose sevengill shark, Notorynchus cepedianus across its global distribution. We estimate patterns of connectivity among broadnose sevengill shark populations from three major oceanic regions (South Atlantic, Oceania and Eastern Pacific) by contrasting mitochondrial and nuclear DNA haplotype frequencies. We also produced time calibrated Bayesian Inference phylogenetic reconstructions to analyses global phylogeographic patterns and estimate divergence times among distinctive shark lineages. Our results demonstrate significant genetic differentiation among oceanic regions (ΦST = 0.9789, P < 0.0001) and a lack of genetic structuring within regions (ΦST = − 0.007; P = 0.479). Time calibrated Bayesian Inference phylogenetic reconstructions indicate that the observed patterns of genetic structure among oceanic regions are historical, with regional populations estimated to have diverged from a common ancestor during the early to mid-Pleistocene. Our results indicate significant genetic structuring and a lack of gene flow among broadnose sevengill shark populations from the South Atlantic, Oceania and Eastern Pacific regions. Evidence of deep lineage divergences coinciding with the early to mid-Pleistocene suggests historical glacial cycling has contributed to the vicariant divergence of broadnose sevengill shark populations from different ocean basins. These finding will help inform global management of broadnose sevengill shark populations, and provides new insights into historical and contemporary evolutionary processes shaping populations of this ecologically important apex predator.

Interrogating Phylogenetic Discordance Resolves Deep Splits in the Rapid Radiation of Old World Fruit Bats (Chiroptera: Pteropodidae).

The family Pteropodidae (Old World fruit bats) comprises n}{}>200 species distributed across the Old World tropics and subtropics. Most pteropodids feed on fruit, suggesting an early origin of frugivory, although several lineages have shifted to nectar-based diets. Pteropodids are of exceptional conservation concern with n}{}>50% of species considered threatened, yet the systematics of this group has long been debated, with uncertainty surrounding early splits attributed to an ancient rapid diversification. Resolving the relationships among the main pteropodid lineages is essential if we are to fully understand their evolutionary distinctiveness, and the extent to which these bats have transitioned to nectar-feeding. Here we generated orthologous sequences for n}{}>1400 nuclear protein-coding genes (2.8 million base pairs) across 114 species from 43 genera of Old World fruit bats (57% and 96% of extant species- and genus-level diversity, respectively), and combined phylogenomic inference with filtering by information content to resolve systematic relationships among the major lineages. Concatenation and coalescent-based methods recovered three distinct backbone topologies that were not able to be reconciled by filtering via phylogenetic information content. Concordance analysis and gene genealogy interrogation show that one topology is consistently the best supported, and that observed phylogenetic conflicts arise from both gene tree error and deep incomplete lineage sorting. In addition to resolving long-standing inconsistencies in the reported relationships among major lineages, we show that Old World fruit bats have likely undergone at least seven independent dietary transitions from frugivory to nectarivory. Finally, we use this phylogeny to identify and describe one new genus. [Chiroptera; coalescence; concordance; incomplete lineage sorting; nectar feeder; species tree; target enrichment.]

Into the Chamber of Horrors: A proposal for the resolution of nomenclatural chaos in the Scaphiophryne calcarata complex (Anura: Microhylidae), with a new species-level phylogenetic hypothesis for Scaphiophryninae.

The genus Scaphiophryne (Anura: Microhylidae) contains at least nine species that, together with their probable sister genus Paradoxophyla, form the Madagascar-endemic subfamily Scaphiophryninae. Scaphiophryne are robust burrowing frogs with explosive breeding behavior and are characterized by a unique larval morphology, being intermediate between that of filter-feeding and generalized tadpole types. Based on tadpole morphology, the nominal subgenus Scaphiophryne can be distinguished from the subgenus Pseudohemisus, a clade occurring in arid biomes with a largely unsolved taxonomy and nomenclature. Here we combine a comprehensive molecular phylogeny of all scaphiophrynines based on five mitochondrial and two nuclear genes with a re-examination of the name-bearing type specimens of the seven species-level nomina assignable to the subgenus Pseudohemisus. Our molecular phylogeny supports monophyly of the two subgenera and assigns the enigmatic S. matsoko to the subgenus Scaphiophryne. In this subgenus, our analysis confirms all currently accepted species as independent evolutionary lineages based on concordant differentiation in mitochondrial and nuclear genes, and provides evidence for a combination of low mitochondrial divergence, distinct nuclear divergence, and high nuclear heterozygosity. Yet, contrary to previous hypotheses, we do not find evidence of ongoing hybridization. In the subgenus Pseudohemisus we find support for three clearly delimited species-level lineages, one of which contains two deep conspecific lineages that warrant further study. To stabilize the taxonomy in this subgenus, and considering the very poor state of preservation of several type series and the heterogeneity of some of them, we here designate lectotypes for the nomina: Hemisus obscurus Grandidier, 1872; Calophrynus calcaratus Mocquard, 1895; Pseudohemisus verrucosus Angel, 1930; Pseudohemisus longimanus Angel, 1930; and Pseudohemisus longimanus var. melanopleura Angel, 1934. Taxonomically, we accept Scaphiophryne brevis (Boulenger, 1896) as a well-defined species; restrict S. calcarata (Mocquard, 1995) to the southern lineage of the S. calcarata complex; resurrect Hemisus obscurus Grandidier, 1872 (previously considered to be a dubious name or nomen inquirendum) as Scaphiophryne obscura to refer to the western and north-western lineage of the S. calcarata complex; consider Pseudohemisus verrucosus Angel, 1930 as junior synonym of Scaphiophryne brevis; and consider Pseudohemisus longimanus Angel, 1930 and Pseudohemisus longimanus var. melanopleura Angel, 1934 as junior synonyms of Scaphiophryne obscura. Finally, to accelerate taxonomic progress, we (i) outline a set of criteria by which such old and ambiguous nomina can be applied to known lineages, (ii) suggest that the International Code of Zoological Nomenclature should slightly broaden the conditions under which neotypes are to be designated, and (iii) propose to redouble discussions on the introduction of the concept of the epitype to zoology.

A DNA barcode-based survey of wild urban bees in the Loire Valley, France

The current decline of wild bees puts important ecosystem services such as pollination at risk. Both inventory and monitoring programs are needed to understand the causes of wild bee decline. Effective insect monitoring relies on both mass-trapping methods coupled with rapid and accurate identifications. Identifying wild bees using only morphology can be challenging, in particular, specimens from mass-trapped samples which are often in poor condition. We generated DNA barcodes for 2931 specimens representing 157 species (156 named and one unnamed species) and 28 genera. Automated cluster delineation reveals 172 BINs (Barcodes Index Numbers). A total of 36 species (22.93%) were found in highly urbanized areas. The majority of specimens, representing 96.17% of the species barcoded form reciprocally exclusive groups, allowing their unambiguous identification. This includes several closely related species notoriously difficult to identify. A total of 137 species (87.26%) show a “one-to-one” match between a named species and the BIN assignment. Fourteen species (8.92%) show deep conspecific lineages with no apparent morphological differentiation. Only two species pairs shared the same BIN making their identification with DNA barcodes alone uncertain. Therefore, our DNA barcoding reference library allows reliable identification by non-experts for the vast majority of wild bee species in the Loire Valley.

Evaluating DNA Barcoding for Species Identification and Discovery in European Gracillariid Moths

Gracillariidae is the most species-rich leaf-mining moth family with over 2,000 described species worldwide. In Europe, there are 263 valid named species recognized, many of which are difficult to identify using morphology only. Here we explore the use of DNA barcodes as a tool for identification and species discovery in European gracillariids. We present a barcode library including 6,791 COI sequences representing 242 of the 263 (92%) resident species. Our results indicate high congruence between morphology and barcodes with 91.3% (221/242) of European species forming monophyletic clades that can be identified accurately using barcodes alone. The remaining 8.7% represent cases of non-monophyly making their identification uncertain using barcodes. Species discrimination based on the Barcode Index Number system (BIN) was successful for 93% of species with 7% of species sharing BINs. We discovered as many as 21 undescribed candidate species, of which six were confirmed from an integrative approach; the other 15 require additional material and study to confirm preliminary evidence. Most of these new candidate species are found in mountainous regions of Mediterranean countries, the South-Eastern Alps and the Balkans, with nine candidate species found only on islands. In addition, 13 species were classified as deep conspecific lineages, comprising a total of 27 BINs with no intraspecific morphological differences found, and no known ecological differentiation. Double-digest restriction-site associated DNA sequencing (ddRAD) analysis showed strong mitonuclear discrepancy in four out of five species studied. This discordance is not explained by Wolbachia-mediated genetic sweeps. Finally, 26 species were classified as “unassessed species splits” containing 71 BINs and some involving geographical isolation or ecological specialization that will require further study to test whether they represent new cryptic species.

Viromics of extant insect orders unveil the evolution of the flavi-like superfamily.

Insects are the most diversified and species-rich group of animals and harbor an immense diversity of viruses. Several taxa in the flavi-like superfamily, such as the genus Flavivirus, are associated with insects; however, systematic studies on insect virus genetic diversity are lacking, limiting our understanding of the evolution of the flavi-like superfamily. Here, we examined the diversity of flavi-like viruses within the most complete and up-to-date insect transcriptome collection comprising 1,243 insect species by employing a Flaviviridae RdRp profile hidden Markov model search. We identified seventy-six viral sequences in sixty-one species belonging to seventeen insect, one entognathan, and one arachnidan orders. Phylogenetic analyses revealed that twenty-seven sequences fell within the Flaviviridae phylogeny but did not group with established genera. Despite the large diversity of insect hosts studied, we only detected one virus in a blood-feeding insect, which branched within the genus Flavivirus, indicating that this genus likely diversified only in hematophagous arthropods. Nine new jingmenviruses with novel host associations were identified. One of the jingmenviruses established a deep rooting lineage additional to the insect- and tick-associated clades. Segment co-segregation phylogenies support the separation of tick- and insect-associated groups within jingmenviruses, with evidence for segment reassortment. In addition, fourteen viruses grouped with unclassified flaviviruses encompassing genome length of up to 20 kb. Species-specific clades for Hymenopteran- and Orthopteran-associated viruses were identified. Forty-nine viruses populated three highly diversified clades in distant relationship to Tombusviridae, a plant-infecting virus family, suggesting the detection of three previously unknown insect-associated families that contributed to tombusvirus evolution.