Mitochondrial DNA diversity and taxa delineation in the land snails of the Iberus gualtieranus (Pulmonata,Helicidae) complex

Iberus gualtieranus is a species complex of land snails that is endemic to the Iberian Peninsula. The species taxonomy of the group is based merely on the basis of shell morphology, but validity of the existing taxonomy is uncertain. Using mitochondrial DNA (mtDNA) data (cytochrome oxidase I and 16S rRNA sequences) we were able to validate the observed phylogenetic taxa within the I. gualtieranus s.l. complex by means of the analysis of specimens of the different morphospecies, together with the study of topotypes. Strong incongruences were obtained between morphology and molecular data. The Iberus alonensis morphospecies comprised several genetically divergent but morphologically cryptic lineages. Considering (1) the allopatric distribution of the operational taxonomic units (OTUs), (2) the morphological differentiation, (3) the possible occurrence of hybridization among the different lineages, and (4) the strong differentiation of the mtDNA phylogroups, we suggest the main lineages obtained, for the time being, may be treated as evolutionary species. The robust phylogenetic reconstruction obtained allows us to consider I. alonensis s.s., Iberus campesinus, Iberus carthaginiensis, and Iberus gualtieranus s.s. as valid species. Two additional unnominated taxa of the alonensis shell type have also been identified. Further subdivisions are also considered, including Iberus gualtieranus mariae and Iberus gualtieranus ornatissimus. © 2008 The Linnean Society of London, Zoological Journal of the Linnean Society, 2008, 154 , 722–737.     

AFLP analysis shows high incongruence between genetic differentiation and morphology‐based taxonomy in a widely distributed tortoise

Morphology has traditionally been used to diagnose the taxa of various taxonomic ranks. However, there is growing evidence that morphology is not always able to reveal cryptic taxa, and that pronounced morphological variation could reflect phenotypic plasticity rather than evolutionary divergence. Spur‐thighed tortoises (the Testudo graeca complex), distributed in the western Palaearctic region, are characterized by high morphological variability and complicated taxonomy, which are under debate. Previous molecular studies using mainly mitochondrial DNA (mtDNA) sequences have revealed incongruence between genetic differentiation and morphology‐based taxonomy, suggesting that morphological variability is the result of phenotypic plasticity and stabilizing selection, which masks the true genealogies. In the present study, we used a range‐wide sampling and nuclear Amplified fragment length polymorphism (AFLP) markers to investigate genetic differentiation within the T. graeca complex. We found that spur‐thighed tortoises are differentiated into four geographically well‐defined AFLP groups: Balkans–Middle Eastern, western Mediterranean, Caucasian and central‐eastern Iranian. Compared with the distribution of mtDNA lineages, the groups are largely concordant, although the AFLP markers are less sensitive and distinguish fewer groups than do mtDNA sequences. The AFLP groups show an allopatric or parapatric distribution. The AFLP differentiation conflicts with the previously proposed morphology‐based taxonomy of the complex, suggesting that local adaptation to different environmental conditions may have led to the great extent of morphological variation within the same lineages. We propose a re‐evaluation of the taxa that were confirmed genetically using a thorough morphological analysis corrected for phenotypic plasticity. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ●● , ●●–●●.     

Molecular diversity of species of the Elymus trachycaulus species complex and their relationships to non-North American taxa

Species of the E. trachycaulus complex species are known for their morphological variability, but little is known about their genetic basis. The delimitation of taxa within the complex has been controversial and difficult. E. trachycaulus is predominantly self-pollinating, and lacks clear morphological boundaries between it and E. alaskanus. Another controversial taxonomic issue of E. trachycaulus is the relationships of this complex species to non-North American E. caninus. The objectives of this study were to examine genetic diversity and the systematic relationships among the species of the E. trachycaulus complex and their relationships with E. caninus, E. alaskanus and E. mutabilis. Random amplified polymorphic DNA method was used to study 35 accessions of E. trachycaulus complex and other Elymus species. Higher genetic variation was detected within species of E. trachycaulus complex. Eurasian accessions are as variable as the North American ones. Both UPGMA and NJ analyses did not show clearly separation among species of the E. trachycaulus complex. No clear association between geographic origin and genetic grouping among these species was found. Eurasian E. trachycaulus probably originated from multiple North American populations.     

Species delimitation in the Aloysia gratissima complex (Verbenaceae) following the phylogenetic species concept

The Aloysia gratissima complex is a group of 22 accepted, specific and infraspecific taxa, the boundaries of which are controversial. Various different authors have chosen different characters as criteria for delimiting taxa in this complex, which has been reflected in the variable number of accepted species. To resolve the taxonomy of this group, a modified population aggregation analysis, based on the phylogenetic species concept, was employed. As a result, the Aloysia gratissima complex is here circumscribed as compromising only four species, A. decipiens, A. gratissima s.s., A. oblanceolata and A. schulziana. These species are sustained by a different combination of four characters, of the 16 qualitative attributes analysed. A new approach based on cluster analysis is proposed here delimiting varieties. As a consequence two varieties, supported by significant differences in quantitative characters, are recognized here: A. gratissima var. gratissima and A. gratissima var. sellowii. A complete taxonomic revision of the accepted taxa is presented and six new synonyms are proposed. A key, illustrations and geographical distribution maps are included. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 180 , 193–212.     

Revision of the Euthalia phemius complex (Lepidoptera: Nymphalidae) based on morphology and molecular analyses

Adults of the Euthalia phemius complex, which is composed of three South‐East Asian nymphalid species, Euthalia phemius, Euthalia ipona, and Euthalia euphemia, were genetically analysed by examining mitochondrial and nuclear genes. The E. phemius complex was also examined morphologically, with particular emphasis on wing markings and male genitalia. No significant differences amongst the three species in the complex were detected with respect to either genetic distance or genital morphology. We therefore conclude that the three currently recognized Euthalia species belong to a single species. Accordingly, E. ipona is synonymized with E. phemius. Euthalia euphemia is treated as a subspecies of E. phemius. Type specimens of all taxa and a synonymic list for the E. phemius complex are also given. In addition, we briefly discuss the evolution and biogeography of the species complex. © 2011 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 164 , 304–327.     

Character analysis and taxonomic revision of the Microtoena insuavis complex (Lamiaceae)

The taxonomy of the Microtoena insuavis complex (Lamiaceae) is controversial. No agreement on the taxonomic treatments of M. esquirolii, M. insuavis, M. mollis and M. patchoulii has been reached. In addition, some species and varieties described on the basis of a single specimen or a limited number of specimens are doubtful. Based on the examination of a large number of herbarium specimens, extensive field observations, population sampling, scanning electron microscopy observation and statistical analysis of some characters, M. esquirolii is separated from the M. insuavis complex, and M. subspicata and M. subspicata var. intermedia are treated as synonyms of M. esquirolii, M. pauciflora is treated as a synonym of M. patchouli, and M. siamica is treated as a synonym of M. insuavis. Only three species (M. insuavis, M. mollis and M. patchouli) are recognized in this complex. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 165 , 315–327.     

An integrative taxonomic revision of the Tarentola geckos (Squamata,Phyllodactylidae) of the Cape Verde Islands

Recent phylogeographical analyses using mitochondrial DNA (mtDNA) sequences indicate that the Tarentola geckos from the Cape Verde archipelago originated from a propagule that dispersed from the Canary Islands approximately 7.7 Mya and that underwent a fast evolutionary radiation. Molecular analyses carried out to date clearly show some incongruences with the current taxonomy of Tarentola from the Cape Verde Islands, with some species being paraphyletic or polyphyletic, and several independently evolving lineages needing formal taxonomic recognition. The aim of this study was to clarify the systematics of this group to unravel its taxonomy by applying an integrative approach based on information from three independent sources: mtDNA, nuclear genes, and morphology. As a result of this taxonomic revision, two novel species for the islands of S. Nicolau and Fogo are described and eight subspecies are upgraded to species level. Moreover, an identification key for the genus Tarentola from the Cape Verde archipelago is presented. This study reconciles taxonomy and phylogeny in this group, provides a better understanding of diversity patterns, new insights on evolutionary hypotheses, and supports the basic framework for the future management and conservation of this unique reptile radiation. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 164 , 328–360.     

An integrative approach to reveal speciation and species richness in the genus Diasporus (Amphibia: Anura: Eleutherodactylidae) in eastern Panama

We have applied an integrative taxonomic approach, including bioacoustics, ecology, morphology, and molecular genetics (barcoding and phylogeography), to explore species richness in the genus Diasporus in eastern Panama, from where only Diasporus quidditus (Lynch, 2001) was previously known. During fieldwork in eastern Panama in 2011 and 2012 we found six additional species, four of which we are describing here as new to science, plus two species that are new for this region. We have evaluated the presence of Diasporus diastema (Cope, 1875) in eastern Panama by comparing morphological, genetic, and bioacoustic characters of specimens from near the type locality in central Panama with specimens from eastern Panama. We further describe and compare male advertisement calls of most Diasporus species. The phylogeographic analysis suggests the allopatric speciation of Diasporus species in eastern Panama following the completion of the Panamanian isthmus in the middle Miocene. Subsequent geological events concur with the vicariant evolution of different lineages in situ, suggesting eastern Panama to be a centre of endemism for this group of frogs. We present an integrative analysis of the species from eastern Panama and include an identification key for all species of the genus.     

Proposal of Monorhizochytrium globosum gen. nov., comb. nov. (Stramenopiles,Labyrinthulomycetes) for former Thraustochytrium globosum based on morphological features and phylogenetic relationships

Thraustochytrium is the type genus of the family Thraustochytriaceae in the class Labyrinthulomycetes. This genus is characterized by zoospore formation, namely, shape of the cell wall of sporangia and presence or absence of a proliferous body. However, there are several issues associated with the taxonomy of this genus, and these include polyphyletic taxa and overlapping of taxonomic features among species. In particular, the first and second species, T. proliferum and T. globosum, were described based on observations of the morphological features of natural samples in the absence of culture conditions. Before addressing the taxonomic issues associated with this genus, it is important to consider the taxonomic features of each species, i.e., the life history under culture conditions and the phylogenetic position. The objective of the present study was to isolate T. globosum, the second described species in the genus Thraustochytrium, from the type locality. We successfully isolated strain NBRC 112723, which exhibited characteristic features of T. globosum. Under culture conditions, strain NBRC 112723 exhibited taxonomic features observed in other thraustochytrid species. Our molecular phylogeny indicated that this strain isolated from the type locality was located in an unidentified thraustochytrid group; moreover, some strains located in this group exhibited characteristic features of strain NBRC 112723. We clearly distinguished T. globosum based on the taxonomic criteria used to classify the T. proliferum type species. Therefore, we propose the establishment of a new genus, Monorhizochytrium, for the species T. globosum in the family Thraustochytriaceae.     

Tragelaphus nakuae: evolutionary change,biochronology, and turnover in the African Plio‐Pleistocene

Fossil Bovidae constitute one of the most significant proxy records for evolutionary and palaeoecological change in Africa. Tragelaphus nakuae is a regularly encountered antelope in the East African Plio‐Pleistocene, and is a common component of hominin faunas. As previously understood, this species ranged for almost 2 million years, encompassed a large range of morphological variation, exhibited relative stasis in the face of environmental perturbations, and left no known living descendants. I here review and revise the fossil record of this tragelaphin bovid, finding that specimens older than ～2.8 Mya and previously attributed to T. nakuae or a close form are in fact referable to a distinct, but ancestral, species. This new interpretation adds these fossil tragelaphins to the body of evidence supporting major faunal turnover occurring around 2.8 Mya in concert with global climatic change. I also document morphological changes that occur through the duration of T. nakuae, particularly after 2.3 Mya. These taxonomic revisions allow for refined biochronological estimates for several East African Plio‐Pleistocene sites and specimen assemblages of uncertain age. A phylogenetic analysis suggests that the T. nakuae lineage is related to the extant bongo (Tragelaphus eurycerus), relating this living but enigmatic forest antelope to the fossil record. One resulting palaeoecological hypothesis is that the bongo's modern fragmented range represents the relicts of a much more widely distributed late Pliocene African forest belt. This study highlights the importance of specimen‐based approaches for elucidating the pattern and timing of major evolutionary events. © 2011 The Linnean Society of London, Zoological Journal of the Linnean Society, 2011, 162 , 699–711.     

Taxonomic revision of the dolphin genus Lagenorhynchus

Evolutionary relationships among cetaceans within the family Delphinidae have been difficult to resolve due to the high number of species and their relatively rapid radiation. This is the case for the dolphin species currently placed in the genus Lagenorhynchus, and their relations to Cephalorhynchus and Lissodelphis species. Phylogenetic relationships among these species have been investigated using multiple lines of evidence, and that evidence consistently suggests that the six species currently assigned to Lagenorhynchus do not form a monophyletic group. Here, we summarize findings from studies of morphology, genetics, historical biogeography, and acoustics that offer insight into the phylogenetics of these taxa. We present the taxonomic basis for revision of Lagenorhynchus, propose retention of Lagenorhynchus albirostris and reassignment of the remaining five species into other existing generic names, namely Leucopleurus acutus, Sagmatias australis, Sagmatias cruciger, Sagmatias obliquidens, and Sagmatias obscurus. Making these revisions now so that the taxonomy better reflects evolutionary relationships among these species will ensure that future studies include the most appropriate taxa for investigating the complex phylogenetic and systematic relationships among cetaceans. However, comprehensive analyses using multiple lines of evidence are still needed to clarify the phylogenetic relationships within and among the Lissodelphininae genera Cephalorhynchus, Lissodelphis, and Sagmatias.     

Revision and phylogeny of the bee genus Paratetrapedia Moure,with description of a new genus from the Andean Cordillera (Hymenoptera,Apidae, Tapinotaspidini)

The bee genus Paratetrapedia represents a commonly collected group of bees and is especially diverse in forested areas of the Neotropics. Its taxonomy has remained poorly understood because of a lack of modern revisionary work and numerous species described as Tetrapedia whose type specimens have not been re‐examined in recent times. Here, a comprehensive study was carried out to review the taxonomy of the genus Paratetrapedia and to investigate cladistically the relationships amongst its species. Eighteen new species of Paratetrapedia are described, giving a total of 32 species in the genus. A phylogenetic analysis of the species of Paratetrapedia was carried out using 61 morphological characters for 41 terminal taxa. The phylogenetic results confirmed the monophyly of Paratetrapedia and allowed the recognition of five species groups: the lugubris, moesta, bicolor, lineata, and flavipennis groups. Nasutopedia gen. nov. , recognized as the sister group of Paratetrapedia and with its distribution restricted to the western forested portions of the Andean highlands, is proposed based on distinct morphology, its placement in the phylogenetic tree, and biogeographical patterns. Species of Paratetrapedia are especially diverse in the Amazon Forest; the eastern Brazilian Atlantic Forest contains four endemic species, and one species is endemic to the Cerrado of central Brazil. Paratetrapedia shows a biogeographical pattern similar to other Neotropical groups of bees and birds, with wide distribution and high diversity in lowland forests and whose sister taxon occurs on highlands of north‐western portions of the Andean cordillera. Identification keys for males and females of all species are provided, as well as distribution maps and illustrations of general external morphology and genitalia. © 2011 The Linnean Society of London, Zoological Journal of the Linnean Society, 2011, 162 , 351–442.     

Taxonomic implications of foliar epidermis in Impatiens (Balsaminaceae): Investigating 12 Pakistani taxa as an example

Impatiens L. is taxonomically a complicated genus that necessitates fresh characterization to resolve its taxonomy. This study therefore aimed at ascertaining micro-morphological characters in the foliar epidermis of Impatiens. For this purpose, we gathered twelve taxa (10 species and 2 subspecies) mostly from the northern regions of Pakistan and studied qualitative and quantitative characters depicting variation in epidermal cells, stomata, and trichomes on both surfaces. Multicellular, uniseriate trichomes were observed only in I. balsamina L., and raphides in I. balfourii Hook. f. and I. thomsonii Hook. f. while stomata were exclusively anomocytic in all species. Morphological variation in the foliar epidermal characters and their diagnostic value has been depicted by constructing a taxonomic key to all taxa. The study revealed that although foliar epidermal characters in Impatiens did not desirably assort taxa on the basis of structural topographies, yet demonstrated adequate variation distinguishing the Pakistani taxa.     

Delimiting species in the genus Otospermophilus (Rodentia: Sciuridae), using genetics,ecology, and morphology

We apply an integrative taxonomy approach to delimit species of ground squirrels in the genus Otospermophilus because the diverse evolutionary histories of organisms shape the existence of taxonomic characters. Previous studies of mitochondrial DNA from this group recovered three divergent lineages within Otospermophilus beecheyi separated into northern, central, and southern geographical populations, with Otospermophilus atricapillus nested within the southern lineage of O. beecheyi. To further evaluate species boundaries within this complex, we collected additional genetic data (one mitochondrial locus, 11 microsatellite markers, and 11 nuclear loci), environmental data (eight bioclimatic variables), and morphological data (23 skull measurements). We used the maximum number of possible taxa (O. atricapillus, Northern O. beecheyi, Central O. beecheyi, and Southern O. beecheyi) as our operational taxonomic units (OTUs) and examined patterns of divergence between these OTUs. Phenotypic measures (both environmental and morphological) showed little differentiation among OTUs. By contrast, all genetic datasets supported the evolutionary independence of Northern O. beecheyi, although they were less consistent in their support for other OTUs as distinct species. Based on these data, we support the conclusions from a previous study that synonymized O. atricapillus with O. beecheyi, and we elevate the northern lineage of O. beecheyi to a separate species. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 113 , 1136–1151.     

New generic proposal for the European Neogene large testudinids (Cryptodira) and the first phylogenetic hypothesis for the medium and large representatives of the European Cenozoic record

Fossil testudinids are known in Europe since the Eocene, with several taxa of medium size (from more than 0.3 m to less than 0.7 m) recognized in the Palaeogene record, most of them being poorly known. The size of several European Neogene taxa was larger (between 1 and 2 m). These large testudinids were relatively abundant and diverse, ranging from the early Miocene to the Pleistocene. However, there is a nomenclatural gap at the generic level for the Neogene forms, as their generally used assignment to the more primitive Eocene Cheirogaster cannot be sustained. This is because relatively little material has been assigned to the described species, and also because of the absence of a detailed study comparing all of the European taxa. Here, the European Cenozoic taxa are incorporated for the first time in a data matrix, so that a hypothesis on their phylogenetic relationships is justified. This study identified the large testudinids from the Neogene of Europe as belonging to a monophyletic clade, assigned to the new genus T itanochelon . The hitherto poorly understood ‘Testudo’ bolivari, proposed nearly a century ago but lacking diagnosis, is analysed in detail. It is recognized as the best‐represented large testudinid from the European record, and is identified as the type species of T itanochelon gen. nov. Its comparison with the other Neogene species allowed a detailed study of the new genus and an analysis of its phylogenetic relationships with the other European taxa. © 2014 The Linnean Society of London     

A new genus of cave‐dwelling microcrustaceans from the Dinaric Region (south‐east Europe): adaptations of true stygobitic Cladocera (Crustacea: Branchiopoda)

We revise ‘true’ stygobitic cladocerans and lift three species from Alona Baird, 1843 (Cladocera: Chydoridae). Species of Brancelia gen. nov. are inhabitants of saturated karst, collected in pools of residual water in the amphibious zones of a few caves in the Dinaric Region, Europe. All species are blind (regression of eye and ocellus), have elongated sensorial equipment (aesthetascs) and a short rostrum, reduced antennal spines, and a globular body. In contrast to earlier hypotheses, there is no epigean chydorid taxon from the Palaearctic that can be linked to Brancelia gen. nov. The new genus may be an offshoot of six‐limbed Aloninae, but a littoral‐benthic ancestor is not apparent and most likely extinct. Evolution of Brancelia gen. nov. parallels that of other subterranean Cladocera like Phreatalona Van Damme, Brancelj & Dumont, 2009. We discuss the functional morphology of Brancelia gen. nov. and compare its adaptations to a subterranean life mode with those of Phreatalona. © 2011 The Linnean Society of London, Zoological Journal of the Linnean Society, 2011, 161 , 31–52.     

Integrative taxonomy of the stunt nematodes of the genera Bitylenchus and Tylenchorhynchus (Nematoda,Telotylenchidae) with description of two new species and a molecular phylogeny

Stunt nematodes are characterized by phenotypic plasticity, with overlapping morphology and morphometry leading to potential misidentification. Consequently, the application of integrative taxonomic approaches is useful to species delimitation based on a combination of different perspectives, e.g. morphology and DNA sequences. We conducted nematode surveys in cultivated and natural environments in Spain and the USA, from which we identified 18 known species of the family Telotylenchidae and two new taxa within the studied samples. These species were morphologically, morphometrically, and molecularly characterized. The results of light and scanning electron microscopic observations, and molecular and phylogenetic analysis also allowed two new species to be distinguished, described herein as B itylenchus hispaniensis sp. nov. and T ylenchorhynchus mediterraneus sp. nov. The phylogenetic analysis was carried out using molecular data from nuclear ribosomal DNA genes [D2–D3 expansion segments of the large ribosomal subunit (28S), internal transcribed spacer (ITS), and partial small ribosomal subunit (18S)]. We also provide here a test of alternative hypotheses that confirms the monophyly of both Tylenchorhynchus and Bitylenchus sensu Siddiqi's classification but does not support Fortuner & Luc's conceptual view of Tylenchorhynchus as a large genus. Ancestral state reconstructions of several diagnostic morphological characters using a maximum parsimony approach showed congruence in morphological and molecular evolution for stylet knob inclination and tail tip annulation. Our analysis emphasizes some of the problems related to the taxonomy and phylogeny of nematodes of Telotylenchinae. © 2014 The Linnean Society of London     

Resurrection in Carapa (Meliaceae): a reassessment of morphological variation and species boundaries using multivariate methods in a phylogenetic context

The taxonomy of the amphi‐Atlantic tree genus Carapa (Meliaceae) has long been controversial. Of the three species currently recognized in the genus, two are known to present substantial morphological variation that has been used in the past to distinguish several taxa, most of which are currently placed in synonymy. Here, a combination of field observations, univariate analyses of leaf, floral and seed characters and principal coordinate analyses of floral characters in the context of a molecular phylogenetic analysis was used to investigate the patterns of variation and delimit morphological species anew in the genus. These results support the recognition of 27 species in Carapa, of which 16 are previously described and 11 are new. In general, phylogenetically related species occurred in the same geographical area, but were morphologically distinct. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 165 , 186–221.     

An integrative approach to characterize Malagasy bats of the subfamily Vespertilioninae Gray, 1821, with the description of a new species of Hypsugo

Although important advances have been made in recent years in the taxonomy of different families and subfamilies of Malagasy bats, those belonging to the Vespertilioninae remain partially unresolved. Herein using a mitochondrial marker (cytochrome b) as the point of departure for 76 specimens of Malagasy vespers and appropriate African taxa, we diagnose the six taxa of this subfamily on the island by overlaying different morphological and bioacoustic characters on the clade structure of sequenced animals. The species include: endemic Neoromicia matroka, which is sister to African N. capensis; endemics N. malagasyensis and N. robertsi, which form sister species; a member of the genus Hypsugo, which is sister to African H. anchietae and described herein as new to science; Pipistrellus hesperidus for which Madagascar animals are genetically close but distinct from African populations of the same species; and endemic P. raceyi, which shows segregation of eastern (mesic) and western (dry) populations and its sister species relationships are unresolved. While the external and craniodental measurements, as well as bioacoustic variables, allow only partial differentiation of these six species of Vespertilioninae, molecular characters provide definitive separation of the taxa, as do male bacular morphology. © 2015 The Linnean Society of London