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Metazoa-level universal single-copy orthologs (mzl-USCOs) are universally applicable markers for DNA taxonomy in animals that can replace or supplement single-gene barcodes. Previously, mzl-USCOs from target enrichment data were shown to reliably distinguish species. Here, we tested whether USCOs are an evenly distributed, representative sample of a given metazoan genome and therefore able to cope with past hybridization events and incomplete lineage sorting. This is relevant for coalescent-based species delimitation approaches, which critically depend on the assumption that the investigated loci do not exhibit autocorrelation due to physical linkage. Based on 239 chromosome-level assembled genomes, we confirmed that mzl-USCOs are genetically unlinked for practical purposes and a representative sample of a genome in terms of reciprocal distances between USCOs on a chromosome and of distribution across chromosomes. We tested the suitability of mzl-USCOs extracted from genomes for species delimitation and phylogeny in four case studies: Anopheles mosquitos, Drosophila fruit flies, Heliconius butterflies and Darwin's finches. In almost all instances, USCOs allowed delineating species and yielded phylogenies that corresponded to those generated from whole genome data. Our phylogenetic analyses demonstrate that USCOs may complement single-gene DNA barcodes and provide more accurate taxonomic inferences. Combining USCOs from sources that used different versions of ortholog reference libraries to infer marker orthology may be challenging and, at times, impact taxonomic conclusions. However, we expect this problem to become less severe as the rapidly growing number of reference genomes provides a better representation of the number and diversity of organismal lineages.

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Between 1951-1958, most of the Hula Lake and its surrounding swamps in the Upper Jordan River (Rift) Valley of Israel were drained with the supposed purposes to eliminate malaria and to reclaim land for agriculture; both reasons later proved to be unnecessary decisions. With the paucity of biological knowledge of the Hula region, especially its aquatic invertebrates, accurate assessment of the environmental damage from this drainage is still being realized. Based on natural history museum collection specimen records, the pre-drainage presence of some aquatic insect species has been verified. Among these was Donaciabicolora, a member of a semi-aquatic subfamily (Donaciinae) of Leaf Beetles (Chrysomelidae) and whose Israeli populations were thought to have gone extinct because of the drainage of the Hula and other locations. Recently this species was rediscovered in two populations. However, the molecular identification of two of these recently collected specimens from one population revealed that the identity of this species is actually Donaciasimplex. In this work, the re-discovery of this species is detailed, and its conservation importance discussed.

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BACKGROUND: The infraorder Rhigonematomorpha comprises a group of obligate parasitic nematodes of millipedes (Arthropoda: Diplopoda). The current species identification of Rhigonematomorpha nematodes remains mainly based on morphological features, with molecular-based identification still in its infancy. Also, current knowledge of the phylogeny of Rhigonematomorpha is far from comprehensive. METHODS: The morphology of Rhigonematomorpha nematodes belonging to the genus Rhigonema, collected from the millipede Spirobolus bungii Brandt (Diplopoda: Spirobolida) in China, was studied in detail using light and scanning electron microscopy. Five different genetic markers, including the nuclear small ribosomal subunit (18S), internal transcribed spacer (ITS) and large ribosomal subunit (28S) regions and the mitochondrial cox1 and cox2 genes of these Rhigonematomorpha nematodes collected from China and Rhigonema naylae collected from Japan were sequenced and analyzed using Bayesian inference (BI) and Assemble Species by Automatic Partitioning (ASAP) methods. Phylogenetic analyses that included the most comprehensive taxa sampling of Rhigonematomorpha to date were also performed based on the 18S + 28S genes using maximum likelihood (ML) and BI methods. RESULTS: The specimens of Rhigonema collected from S. bungii in China were identified as a new species, Rhigonema sinense n. sp. Striking variability in tail morphology was observed among individuals of R. sinense n. sp. ASAP analyses based on the 28S, ITS, cox1 and cox2 sequences supported the species partition of R. sinense n. sp. and R. naylae, but showed no evidence that the different morphotypes of R. sinense n. sp. represent distinct genetic lineages. BI analyses also indicated that R. sinense n. sp. represents a separated species from R. naylae based on the cox1 and cox2 genes, but showed that R. naylae nested in samples of R. sinense n. sp. based on the ITS and 28S data. Phylogenetic results showed that the representatives of Rhigonematomorpha formed two large clades. The monophyly of the families Carnoyidae and Ichthyocephalidae and the genus Rhigonema was rejected. The representatives of the family Ransomnematidae clustered together with the family Hethidae with strong support. CONCLUSIONS: A new species of Rhigonematomorpha, R. sinense n. sp. is described based on morphological and molecular evidence. ASAP analyses using 28S, ITS, cox1 and cox2 data indicate the striking variability in tail morphology of R. sinense n. sp. as intraspecific variation, and also suggest that partial 28S, ITS, cox1 and cox2 markers are effective for molecular identification of Rhigonematomorpha nematodes. The phylogenetic results support the traditional classification of Rhigonematomorpha into the two superfamilies Rhigonematoidea and Ransomnematoidea, and indicate that the families Carnoyidae and Ichthyocephalidae and the genus Rhigonema are non-monophyletic. The present phylogeny strongly supports resurrection of the family Brumptaemiliidae, and also indicates that the family Ransomnematidae is sister to the family Hethidae.

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There is a growing interest in the exploitation of deep-sea mineral deposits, particularly on the abyssal seafloor of the central Pacific Clarion-Clipperton Zone (CCZ), which is rich in polymetallic nodules. In order to effectively manage potential exploitation activities, a thorough understanding of the biodiversity, community structure, species ranges, connectivity, and ecosystem functions across a range of scales is needed. The benthic megafauna plays an important role in the functioning of deep-sea ecosystems and represents an important component of the biodiversity. While megafaunal surveys using video and still images have provided insight into CCZ biodiversity, the collection of faunal samples is needed to confirm species identifications to accurately estimate species richness and species ranges, but faunal collections are very rarely carried out. Using a Remotely Operated Vehicle, 55 specimens of benthic megafauna were collected from seamounts and abyssal plains in three Areas of Particular Environmental Interest (APEI 1, APEI 4, and APEI 7) at 3100-5100 m depth in the western CCZ. Using both morphological and molecular evidence, 48 different morphotypes belonging to five phyla were found, only nine referrable to known species, and 39 species potentially new to science. This work highlights the need for detailed taxonomic studies incorporating genetic data, not only within the CCZ, but in other bathyal, abyssal, and hadal regions, as representative genetic reference libraries that could facilitate the generation of species inventories.

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The pygmy marmoset, the smallest of the anthropoid primates, has a broad distribution in Western Amazonia. Recent studies using molecular and morphological data have identified two distinct species separated by the Napo and Solimões-Amazonas rivers. However, reconciling this new biological evidence with current taxonomy, i.e., two subspecies, Cebuella pygmaea pygmaea (Spix, 1823) and Cebuella pygmaea niveiventris (Lönnberg, 1940), was problematic given the uncertainty as to whether Spix's pygmy marmoset ( Cebuella pygmaea pygmaea) was collected north or south of the Napo and Solimões-Amazonas rivers, making it unclear to which of the two newly revealed species the name pygmaea would apply. Here, we present the first molecular data from Spix's type specimen of Cebuella pygmaea, as well as novel mitochondrial genomes from modern pygmy marmosets sampled near the type locality (Tabatinga) on both sides of the river. With these data, we can confirm the correct names of the two species identified, i.e., C. pygmaea for animals north of the Napo and Solimões-Amazonas rivers and C. niveiventris for animals south of these two rivers. Phylogenetic analyses of the novel genetic data placed into the context of cytochrome b gene sequences from across the range of pygmy marmosets further led us to re-evaluate the geographical distribution for the two Cebuella species. We dated the split of these two species to 2.54 million years ago. We discuss additional, more recent, subdivisions within each lineage, as well as potential contact zones between the two species in the headwaters of these rivers.

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Due to superficial morphological similarities, there is often confusion in the identification of some species of the genus Septaria. A combined analysis of the genital anatomy, morphometric and DNA, based on a portion of the COI gene, applied to three nominal species of this genus, confirmed the validity of Septaria tahitiana Eichhorst, 2016, and suggested that Septaria borbonica (Bory de Saint Vincent, 1804) is a subspecies of Septaria porcellana (Linnaeus, 1758), both taxa with disjunct distribution areas: Septaria borbonica in the western Indian Ocean and Septaria porcellana the western Pacific and eastern Indian Oceans. The possible presence of Septaria tesselata (Lamarck, 1816) in Mayotte (Comoros Archipelago) needs to be confirmed.

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Here, we describe the taxon hypothesis (TH) paradigm, which covers the construction, identification, and communication of taxa as datasets. Defining taxa as datasets of individuals and their traits will make taxon identification and most importantly communication of taxa precise and reproducible. This will allow datasets with standardized and atomized traits to be used digitally in identification pipelines and communicated through persistent identifiers. Such datasets are particularly useful in the context of formally undescribed or even physically undiscovered species if data such as sequences from samples of environmental DNA (eDNA) are available. Implementing the TH paradigm will to some extent remove the impediment to hastily discover and formally describe all extant species in that the TH paradigm allows discovery and communication of new species and other taxa also in the absence of formal descriptions. The TH datasets can be connected to a taxonomic backbone providing access to the vast information associated with the tree of life. In parallel to the description of the TH paradigm, we demonstrate how it is implemented in the UNITE digital taxon communication system. UNITE TH datasets include rich data on individuals and their rDNA ITS sequences. These datasets are equipped with digital object identifiers (DOI) that serve to fix their identity in our communication. All datasets are also connected to a GBIF taxonomic backbone. Researchers processing their eDNA samples using UNITE datasets will, thus, be able to publish their findings as taxon occurrences in the GBIF data portal. UNITE species hypothesis (species level THs) datasets are increasingly utilized in taxon identification pipelines and even formally undescribed species can be identified and communicated by using UNITE. The TH paradigm seeks to achieve unambiguous, unique, and traceable communication of taxa and their properties at any level of the tree of life. It offers a rapid way to discover and communicate undescribed species in identification pipelines and data portals before they are lost to the sixth mass extinction.

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Caecidae is a species-rich family of microsnails with a worldwide distribution. Typical for many groups of gastropods, caecid taxonomy is largely based on overt shell characters. However, identification of species using shell characteristics is problematic due to their rather uniform, tubular shells, the presence of different growth stages, and a high degree of intraspecific variability. In the present study, a first integrative approach to caecid taxonomy is provided using light-microscopic investigation with microsculptural analyses and multi-marker barcoding, in conjunction with molecular species delineation analyses (ABGD, haplotype networks, GMYC, and bPTP). In total 132 specimens of Caecum and Meioceras collected during several sampling trips to Central America were analyzed and delineated into a minimum of 19 species to discuss putative synonyms, and supplement the original descriptions. Molecular phylogenetic analyses suggest Meioceras nitidum and M. cubitatum should be reclassified as Caecum, and the genus Meioceras might present a junior synonym of Caecum. Meiofaunal caecids morphologically resembling C. glabrum from the Northeast Atlantic are a complex of cryptic species with independent evolutionary origins, likely associated with multiple habitat shifts to the mesopsammic environment. Caecum invisibile Egger & Jörger, sp. nov. is formally described based on molecular diagnostic characters. This first integrative approach towards the taxonomy of Caecidae increases the known diversity, reveals the need for a reclassification of the genus Caecum and serves as a starting point for a barcoding library of the family, thereby enabling further reliable identifications of these taxonomically challenging microsnails in future studies.

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The genus Copelatus in Australia is revised and nine species are recognised. One new species, Copelatus martinbaehri sp. nov., is described from Papua New Guinea (Central Province) and Cape York Peninsula (Iron Range NP and Mt Tozer). Copelatus divisus Watts, 1978, syn. nov., is considered a junior synonym of C. portior Guignot, 1956, described from New Guinea. Species delimitation is based on the morphological characters and Cox1 data. All species are (re)described, and their important species characters (median lobes, parameres, habitus and colour patterns) are illustrated. A key to all nine species is provided. The known distribution and habitat preferences of each species are outlined briefly. In Australia, all nine species are distributed in the northern half of the continent. Four species are also reported from New Guinea: in addition to C. martinbaehri sp. nov., we record C. clarki Sharp, 1882 for the first time from southern New Guinea, and consider literature records of C. irregularis W.J. Macleay, 1871 and C. marginatus Sharp, 1882 from New Guinea as doubtful. Copelatus portior is widely distributed in Australasia, while C. tenebrosus is widely distributed in the Indomalayan and Australasian realms. All Australian Copelatus are confirmed to be lentic, found in a large variety of stagnant water, mainly in lowland areas up to 250 m.

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Land leeches in the family Haemadipsidae are mostly from the humid tropical rainforest habitats and habitually take blood from the body of human and other animals. In the present study, we report a new species, Sinospelaeobdella wulingensis sp. n., from caves in the northern subtropical Wuling Mountains of central-south China that feeds blood exclusively on cave-dwelling bats. Based on morphological characteristics, COI gene sequence divergence, and phylogenetic analysis, a new genus Sinospelaeobdella gen. n. is established for the new species, to which a previously described species Haemadipsa cavatuses Yang et al., 2009 is transferred as S. cavatuses comb. n. We also provided extended discussion on phylogenetic relationship within the "Tritetrabdellinae" clade uncovered in a previous study, DNA taxonomy, morphological and behavioral adaptions, biogeography, and possible involvement of Sinospelaeobdella gen. n. in bat transmitted diseases of public concerns.

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Eriophyoids affect crops around the globe directly or indirectly as virus vectors. Eriophyoid systematics initiated over a century ago, yet more than 90% of their fauna remain undescribed. Morphological identification is challenging because of a limited number of traits, cryptic speciation and complex life cycle reported for many species in the group. Nucleic acids extraction for mite identification is challenging due to their microscopic size with researchers using pooled samples leading to polymorphisms and inconclusive results. Identification of mite virus vectors is a tiresome task that could be simplified with a protocol that allows for the detection of viruses in the individual specimen. This communication describes an innovative, highly efficient extraction and detection pipeline. Direct Reverse Transcriptase - Polymerase Chain Reaction (Drt-PCR) assays were implemented in the molecular identification of eriophyoids and detection of viruses present in their bodies. The reverse transcription step allows for amplification from a single mite or egg, as in addition to the genomic DNA, it incorporates the abundant transcripts of targeted genes, whereas it also allows for the amplification of viruses. This communication provides an efficient, sensitive and cost-effective alternative that can be implemented in pest identification and detection as well as biological and ecological studies.

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Ostracods (Crustacea, Ostracoda) are small bivalved crustaceans, contributing over 200 described species to the marine zooplankton community. They are widely distributed and are relatively abundant components of the mesozooplankton, playing an important role in the transport of organic matter to deep layers. However, identification of ostracods based on micro-morphological characters is extremely difficult and time-consuming. Previous fragmentary taxonomic studies of ostracods in the South China Sea (SCA), were based solely on morphology. Here, by analysing the mitochondrial COI gene, we explore the taxa across the SCA using molecular tools for the first time. Our results show that sequence divergence among species varies within a large range, from 12.93% to 35.82%. Sixteen of the taxonomic units recovered by DNA taxonomy agree well with morphology, but Paraconchoecia oblonga, Conchoecia magna and Halocypris brevirostris split into two clades each, each of which contains cryptic species.

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The acanthocephalans are characterized by a retractible proboscis, armed with rows of recurved hooks, which serves as the primary organ for attachment of the adult worm to the intestinal wall of the vertebrate definitive host. Whilst there is a considerable variation in the size, shape and armature of the proboscis across the phylum, intraspecific variation is generally regarded to be minimal. Consequently, subtle differences in proboscis morphology are often used to delimit congeneric species. In this study, striking variability in proboscis morphology was observed among individuals of Neorhadinorhynchus nudus (Harada, 1938) collected from the frigate tuna Auxis thazard Lacépède (Perciformes: Scombridae) in the South China Sea. Based on the length of the proboscis, and number of hooks per longitudinal row, these specimens of N. nudus were readily grouped into three distinct morphotypes, which might be considered separate taxa under the morphospecies concept. However, analysis of nuclear and mitochondrial DNA sequences revealed a level of nucleotide divergence typical of an intraspecific comparison. Moreover, the three morphotypes do not represent three separate genetic lineages. The surprising, and previously undocumented level of intraspecific variation in proboscis morphology found in the present study, underscores the need to use molecular markers for delimiting acanthocephalan species.

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Daphnia on the Tibetan Plateau has been little studied, and information on species diversity and biogeography is lacking. Here, we conducted a 4-year survey using the barcoding fragment of the mitochondrial COI gene to determine the distribution and diversity of Daphnia species found across the Plateau. Our results show that species richness is higher than previously thought, with total described and provisional species number doubling from 5 to 10. Six of the taxonomic units recovered by DNA taxonomy agreed well with morphology, but DNA barcoding distinguished three clades each for the D. longispina (D. galeata, D. dentifera, and D. longispina) and D. pulex (D. pulex, D. cf. tenebrosa, and D. pulicaria) complexes. The sequence divergence between congeneric species varied within a large range, from 9.25% to 30.71%. The endemic D. tibetana was the most common and widespread species, occurring in 12 hyposaline to mesosaline lakes. The lineage of D. longispina is the first confirmed occurrence in west Tibet.

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Body size is a multi-functional trait related to various fitness components, but the relative importance of different selection pressures is seldom resolved. In Carabus japonicus beetles, of which the larvae exclusively prey on earthworms, adult body size is related to the presence/absence of a larger congener and habitat temperature. In sympatry, C. japonicus consistently exhibits smaller body size which is effective for avoiding interspecific mating, but in allopatry, it shows size variation unrelated to temperature. Here, we show that this predator-size variation is attributed to prey-size variation, associated with high phylogenetic diversity in earthworm communities. In allopatry, the predator size was larger where larger prey occurred. Larger adult size may have been selected because larger females produce larger larvae, which can subdue larger prey. Thus, in the absence of a larger congener, variation in prey body size had a pronounced effect on geographic body size divergence in C. japonicus.

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Cryptosporidium spp. (Apicomplexa) causing cryptosporidiosis are of medical and veterinary significance. The genus Cryptosporidium has benefited from the application of what is considered a DNA-barcoding approach, even before the term 'DNA barcoding' was formally coined. Here, the objective to define the DNA barcode diversity of Cryptosporidium infecting mammals is reviewed and considered to be accomplished. Within the Cryptosporidium literature, the distinction between DNA barcoding and DNA taxonomy is indistinct. DNA barcoding and DNA taxonomy are examined using the latest additions to the growing spectrum of named Cryptosporidium species and within-species and between-species identity is revisited. Ease and availability of whole-genome DNA sequencing of the relatively small Cryptosporidium genome offer an initial perspective on the intra-host diversity. The opportunity emerges to apply a metagenomic approach to purified field/clinical Cryptosporidum isolates. The outstanding question remains a reliable definition of Cryptosporidium phenotype. The complementary experimental infections and metagenome approach will need to be applied simultaneously to address Cryptosporidium phenotype with carefully chosen clinical evaluations enabling identification of virulence factors.

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A preliminary phylogeny of 36 species of Carmenta (Lepidoptera: Sesiidae) was reconstructed based on mitochondrial cytochrome oxidase I DNA sequences using parsimony and Bayesian inference in order to assess the placement of a suspected new species. Although the phylogeny was not completely resolved, there were well-supported species groups associated with geography. Based on these results and diagnostic morphological characters, Carmenta wildishorum, n. sp., is described and illustrated from the Cimarron Mountain Range in northeastern New Mexico. The new species is sister to C. texana with a 5.2% nucleotide difference between the two, which is similar to the distance between other Carmenta species and exceeds the intraspecific difference observed within C. texana (0.3%). The phylogeny also suggests additional hidden species diversity among Carmenta species that have large geographic distributions.

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A new species, Gracixalus jinggangensis sp. nov., is described based on a series of specimens collected from Mount Jinggang, Jiangxi Province, southeastern China. The new species is distinguished from all other known congeners by the following combination of morphological characters: relatively small body size, SVL 27.9-33.8 mm in nine males and 31.6 mm in a single female; upper eyelid and dorsum lacking spines; skin of dorsal and lateral surface of head, body and limbs rough with sparsely scattered with tubercles; ventral skin granular; tibiotarsal projection absent; finger webbing rudimentary; toes with moderately developed webbing; brown to beige above in life, with an inverse Y-shaped dark brown marking extending from the interorbital region to the middle of dorsum; males with a single, subgular vocal sac, barely visible nuptial pads with minute granules on the dorsal surface of the bases of first and second fingers. The new species is also genetically divergent from all other Gracixalus for which comparable 16S rRNA gene sequences are available. The discovery of Gracixalus jinggangensis sp. nov. represents the twelfth known species in the genus.

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Land-locked anchialine blue holes are karstic sinkholes and caves with tidally influenced, vertically stratified water bodies that harbor endemic fauna exhibiting variable troglomorphic features. These habitats represent island-like systems, which can serve to elucidate evolutionary and biogeographic processes at local scales. We investigated whether the 'continuous spelean corridor' hypothesis may elucidate the biogeographical distributions of the stygobitic annelid Pelagomacellicephala iliffei (Polynoidae) collected from the Great Bahama and Caicos Banks of the Bahamas Archipelago. Phylogenetic reconstructions were performed using Bayesian Inference on individual and combined datasets of three molecular markers (16S rDNA, COI, 18S rDNA) and species delimitation employed three widely accepted methods in DNA taxonomy, namely GMYC, bPTP, and ABGD. Mantel tests were used to test the effect of geography on genetic structure. Using these analyses, we recovered five independently evolving entities of the focal species across four islands of the Great Bahama Bank including Cat, Eleuthera, Exumas, and Long. Genetic data yielded strong correlations between islands and phylogenetic entities, signifying independent evolutionary histories within anchialine caves across the platform. The island of Eleuthera showed intra-island gene flow and dispersal capabilities between blue holes separated by 115km, providing evidence of a crevicular spelean corridor within the island. However, no evidence of inter-island dispersal is present in the analyzed system. Consistent with previous biogeographic studies of cave crustaceans, the major barriers shaping the cave biota of the Bahamas Archipelago appears to be the deep trenches and channels separating the Bahamian banks.

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We describe a novel species Cheiracanthium ilicis sp. n. (Araneae, Eutichuridae) collected in the province of Toledo (Central Spain). It was found during a systematic sampling campaign carried out in an agricultural landscape with isolated Holm oaks Quercus ilex and small forest patches. Its morphology and affinities with other species of the genus are discussed. Furthermore, one mitochondrial gene was sequenced to confirm species membership and its differentiation from other Cheiracanthium species. The molecular phylogenies based on mitochondrial and nuclear genes showed a close relationship of Cheiracanthium ilicis sp. n. with Cheiracanthium inclusum and Cheiracanthium mildei, with which it also shares morphological similarities. Nonetheless, the sparse sampling of the phylogeny, due to the low number of sequences available, impedes drawing any definitive conclusion about these relationships; it is first necessary to perform an extensive review of the genus worldwide and more thorough phylogenies. Cheiracanthium ilicis sp. n. also shares certain ecological and phenological characteristics with Cheiracanthium inclusum and Cheiracanthium mildei. Like them, Cheiracanthium ilicis sp. n. is an obligate tree dweller that prefers a tree canopy habitat and reproduces primarily in late spring and summer. From a conservation perspective, the present study suggests the need to preserve isolated trees in agricultural landscapes. They are not only the refuge of common forest organisms but also of novel species yet to be discovered.