Molecular taxonomic tools provide more accurate estimates of species richness at less cost than traditional morphology-based taxonomic practices in a vegetation survey

Vegetation surveys are conducted to obtain a catalogue of the plant species that occupy an area of interest, and are used to inform the decisions of policymakers about conservation, development, and remediation efforts. Currently, vegetation surveys rely on traditional morphology-based taxonomic practices to identify collected specimens. By implementing recent advances in molecular taxonomy, it may be possible to improve upon these methods and reduce the associated costs. In this study, we used both morphological and molecular taxonomic methods to sample 337 forest vegetation plots in northeastern Ontario, Canada. DNA barcoding—a molecular taxonomic tool—was used to identify specimens collected in the molecular taxonomic survey. The molecular taxonomic survey identified a mean of 12.4 species per plot and 202 species in total, whereas the morphological taxonomic survey identified a mean of 9.8 species per plot and 142 species in total; both surveys provided identical estimates of community similarity. The morphological taxonomic survey was 37 % more expensive than molecular taxonomic survey, owing largely to the increased time required in the field to collect specimens that flowered at different times. Our results indicate that molecular taxonomic tools are more cost-effective than traditional morphology-based taxonomic practices for species identification in vegetation surveys. Taxonomy underpins all conservation, and the implementation of molecular taxonomic tools for vegetation surveys has promise to lessen the consequences of the taxonomic impediment and increase the effectiveness of conservation efforts.     

Taxonomic Notes on Nasutitermes and Bulbitermes (Termitidae, Nasutitermitinae) from the Sunda region of Southeast Asia based on morphological and molecular characters

The Sunda region of Southeastern Asia is rich in termite fauna, but termites from this region have been poorly described. In this study, we described eight species from two diverse genera from this region, and from the family Termitidae. We describe Bulbitermes 4 spp. and Nasutitermes 4 spp. from new field collections. Where possible we examine original holotype specimens, and describe the essential morphological characters for soldier and worker castes. We devise two new bifurcating keys to guide the field identification of each species. In addition, we develop a nucleotide sequence profile for the COI gene. From this molecular character matrix, we use Neighbour-Joining analysis to test the monophyly of each morphospecies and genus. We find that the morphological and molecular characters are highly concordant, whereby all taxa appear to represent distinct molecular clades. For termites, there is therefore agreement between the morphological taxonomic characters used to sort species from a bifurcating key and the molecular taxonomic characters used to sort species on a bifurcating tree. This joint analysis suggests that DNA barcoding holds considerable promise for termite taxonomy, especially for diverse clades like Bulbitermes and Nasutitermes for which a global morphological key would be intractable.     

An Integrative Taxonomy of Amphibians of Nepal: An Updated Status and Distribution

Amphibians are declining worldwide.At the same time,each year many species of amphibians are described,indicating that biological diversity on Earth remains poorly understood.This is especially true for mountainous regions,such as the Himalayas,which are also regarded as a biodiversity hotspot.However,it is most likely that the current diversity of amphibians in Nepal is highly underestimated.Amphibian inventories by the past researchers primarily used phenotypic characters as the basis for species identification.However,the application of molecular systematics methods to taxonomic studies has successfully uncovered the taxonomic status of several cryptic taxa.This study used both molecular(16 S rDNA sequences)and morphological comparisons and estimated the diversity of amphibians from Nepal.This study reports the occurrence of 38 species of amphibians based on both molecular and morphological comparisons.Among them,three species were recorded for the first time in Nepal.This study also invalids the taxonomic status of Sphaerotheca paschima.This study highlights the importa nce of integrative taxonomic approaches that help to resolve the current taxonomic ambiguities of Nepalese amphibians.     

Phylogenetic analyses of marine sponges within the order Verongida: a comparison of morphological and molecular data

Abstract. Because the taxonomy of marine sponges is based primarily on morphological characters that can display a high degree of phenotypic plasticity, current classifications may not always reflect evolutionary relationships. To assess phylogenetic relationships among sponges in the order Verongida, we examined 11 verongid species, representing six genera and four families. We compared the utility of morphological and molecular data in verongid sponge systematics by comparing a phylogeny constructed from a morphological character matrix with a phylogeny based on nuclear ribosomal DNA sequences. The morphological phylogeny was not well resolved below the ordinal level, likely hindered by the paucity of characters available for analysis, and the potential plasticity of these characters. The molecular phylogeny was well resolved and robust from the ordinal to the species level. We also examined the morphology of spongin fibers to assess their reliability in verongid sponge taxonomy. Fiber diameter and pith content were highly variable within and among species. Despite this variability, spongin fiber comparisons were useful at lower taxonomic levels (i.e., among congeneric species); however, these characters are potentially homoplasic at higher taxonomic levels (i.e., between families). Our molecular data provide good support for the current classification of verongid sponges, but suggest a re-examination and potential reclassification of the genera Aiolochroia and Pseudoceratina . The placements of these genera highlight two current issues in morphology-based sponge taxonomy: intermediate character states and undetermined character polarity.     

Evolutionary trends in Tetrastigma (Vitaceae): Morphological diversity and taxonomic implications

Tetrastigma (Miq.) Planch. (Vitaceae) is a genus with ca. 100 species showing great morphological diversity. Previous molecular phylogenetic studies suggested that traditional classification systems are not consistent with the molecular phylogeny, and Tetrastigma is undergoing further systematic investigation. We traced the evolutionary trends of 20 morphological characters within a robust phylogenetic framework. Our results revealed that many morphological characters show either multiple transitions or few state changes, however, some characters show distinct variation. The two subgenera in Tetrastigma (subgen. Tetrastigma and subgen. Palmicirrata) based on unbranched/bifurcate versus digitately branched tendrils are not supported because subgen. Tetrastigma is paraphyletic. However, the unbranched versus bifurcate/digitately branched tendril is of taxonomic utility to characterize some of the major clades. Inflorescences in Tetrastigma appear axillary, but are leaf‐opposed on a compressed axillary shoot. We found most of the species in Tetrastigma retained the ancestral compound dichasial inflorescence, except those of clade IV that have derived pseudo‐umbellate inflorescences. Other characters including habit, leaf organization, and berry shape provide additional morphological support for the major clades. Our morphological analysis and recent molecular study suggest each of the five major clades within Tetrastigma be treated as distinct taxonomic sections (five sections in the genus).     

The application of DNA sequence data for the identification of benthic nematodes from the North Sea

Nematodes or roundworms represent one of the most diverse and dominant taxon in marine benthic habitats. Whereas a morphological identification of many species is challenging, the application of molecular markers represents a promising approach for species discrimination and identification. In this study, we used an integrative taxonomic approach, combining both molecular and morphological methods, to characterize nematodes of distinct sex and ontogenetic stages from three sampling sites of the North Sea. Morphospecies were discriminated after first visual determination, followed by a molecular analysis of the nuclear 28S rDNA: D2–D3 marker. By linking each sequence to a morphological voucher, discordant morphological identification was subjected to a so-called reverse taxonomic approach. Molecular operational taxonomic units (MOTUs) and morphospecies were compared for all of the three sampling sites to assess concordance of methodology. In total, 32 MOTUs and 26 morphospecies were assigned, of which 12 taxa were identified as described species. Both approaches showed high concordance in taxon assignment (84.4 %) except for a cluster comprising various Sabatieria species. Our study revealed the high potential of the analyzed fragment as a useful molecular marker for the identification of the North Sea nematodes and highlighted the applicability of this combined taxonomic approach in general.     

基于模式产地的分子证据澄清南平鳞毛蕨的分类学地位

地球上有多少物种一直以来都是人们极度关注的问题, 而存疑物种是造成该问题难以解决的重要阻碍之一。目前, 模式标本是存疑物种得以确定的唯一凭证标准, 然而, 因其特殊的分类学地位而难以进行有效的观察和研究。因此, 前往模式产地寻找存疑物种的原始信息成为最有效的替代途径。随着分子生物学, 特别是DNA条形码技术在物种分类与鉴定中的广泛应用, 准确而快速鉴定存疑物种成为可能。以存疑物种——南平鳞毛蕨(Dryopteris yenpingensis)为例, 利用来自模式产地的分子数据进行DNA条形码分析, 并结合形态学特征澄清了南平鳞毛蕨的分类学地位, 确定南平鳞毛蕨与荔波鳞毛蕨(D. liboensis)为同一物种, 同意将荔波鳞毛蕨作为其异名。研究结果和方法可为今后澄清更多存疑物种的分类学地位提供参考。     

Fast Census of Moth Diversity in the Neotropics: A Comparison of Field-Assigned Morphospecies and DNA Barcoding in Tiger Moths

The morphological species delimitations (i.e. morphospecies) have long been the best way to avoid the taxonomic impediment and compare insect taxa biodiversity in highly diverse tropical and subtropical regions. The development of DNA barcoding, however, has shown great potential to replace (or at least complement) the morphospecies approach, with the advantage of relying on automated methods implemented in computer programs or even online rather than in often subjective morphological features. We sampled moths extensively for two years using light traps in a patch of the highly endangered Atlantic Forest of Brazil to produce a nearly complete census of arctiines (Noctuoidea: Erebidae), whose species richness was compared using different morphological and molecular approaches (DNA barcoding). A total of 1,075 barcode sequences of 286 morphospecies were analyzed. Based on the clustering method Barcode Index Number (BIN) we found a taxonomic bias of approximately 30% in our initial morphological assessment. However, a morphological reassessment revealed that the correspondence between morphospecies and molecular operational taxonomic units (MOTUs) can be up to 94% if differences in genitalia morphology are evaluated in individuals of different MOTUs originated from the same morphospecies (putative cases of cryptic species), and by recording if individuals of different genders in different morphospecies merge together in the same MOTU (putative cases of sexual dimorphism). The results of two other clustering methods (i.e. Automatic Barcode Gap Discovery and 2% threshold) were very similar to those of the BIN approach. Using empirical data we have shown that DNA barcoding performed substantially better than the morphospecies approach, based on superficial morphology, to delimit species of a highly diverse moth taxon, and thus should be used in species inventories.     

Hiding deep in the trees: discovery of divergent mitochondrial lineages in Malagasy chameleons of the Calumma nasutum group

WE CONDUCTED A COMPREHENSIVE MOLECULAR PHYLOGENETIC STUDY FOR A GROUP OF CHAMELEONS FROM MADAGASCAR (CHAMAELEONIDAE: Calumma nasutum group, comprising seven nominal species) to examine the genetic and species diversity in this widespread genus. Based on DNA sequences of the mitochondrial gene (ND2) from 215 specimens, we reconstructed the phylogeny using a Bayesian approach. Our results show deep divergences among several unnamed mitochondrial lineages that are difficult to identify morphologically. We evaluated lineage diversification using a number of statistical phylogenetic methods (general mixed Yule-coalescent model; SpeciesIdentifier; net p-distances) to objectively delimit lineages that we here consider as operational taxonomic units (OTUs), and for which the taxonomic status remains largely unknown. In addition, we compared molecular and morphological differentiation in detail for one particularly diverse clade (the C. boettgeri complex) from northern Madagascar. To assess the species boundaries within this group we used an integrative taxonomic approach, combining evidence from two independent molecular markers (ND2 and CMOS), together with genital and other external morphological characters, and conclude that some of the newly discovered OTUs are separate species (confirmed candidate species, CCS), while others should best be considered as deep conspecific lineages (DCLs). Our analysis supports a total of 33 OTUs, of which seven correspond to described species, suggesting that the taxonomy of the C. nasutum group is in need of revision.     

DNA barcoding gap: reliable species identification over morphological and geographical scales

The philosophical basis and utility of DNA barcoding have been a subject of numerous debates. While most literature embraces it, some studies continue to question its use in dipterans, butterflies and marine gastropods. Here, we explore the utility of DNA barcoding in identifying spider species that vary in taxonomic affiliation, morphological diagnosibility and geographic distribution. Our first test searched for a ‘barcoding gap’ by comparing intra‐ and interspecific means, medians and overlap in more than 75 000 computed Kimura 2‐parameter (K2P) genetic distances in three families. Our second test compared K2P distances of congeneric species with high vs. low morphological distinctness in 20 genera of 11 families. Our third test explored the effect of enlarging geographical sampling area at a continental scale on genetic variability in DNA barcodes within 20 species of nine families. Our results generally point towards a high utility of DNA barcodes in identifying spider species. However, the size of the barcoding gap strongly depends on taxonomic groups and practices. It is becoming critical to define the barcoding gap statistically more consistently and to document its variation over taxonomic scales. Our results support models of independent patterns of morphological and molecular evolution by showing that DNA barcodes are effective in species identification regardless of their morphological diagnosibility. We also show that DNA barcodes represent an effective tool for identifying spider species over geographic scales, yet their variation contains useful biogeographic information.     

Taxonomy and species delimitation in Cryptosporidium

Amphibians, reptiles, birds and mammals serve as hosts for 19 species of Cryptosporidium. All 19 species have been confirmed by morphological, biological, and molecular data. Fish serve as hosts for three additional species, all of which lack supporting molecular data. In addition to the named species, gene sequence data from more than 40 isolates from various vertebrate hosts are reported in the scientific literature or are listed in GenBank. These isolates lack taxonomic status and are referred to as genotypes based on the host of origin. Undoubtedly, some will eventually be recognized as species. For them to receive taxonomic status sufficient morphological, biological, and molecular data are required and names must comply with the rules of the International Code for Zoological Nomenclature (ICZN). Because the ICZN rules may be interpreted differently by persons proposing names, original names might be improperly assigned, original literature might be overlooked, or new scientific methods might be applicable to determining taxonomic status, the names of species and higher taxa are not immutable. The rapidly evolving taxonomic status of Cryptosporidium sp. reflects these considerations.     

Rolling into the deep of the land planarian genus <Emphasis Type="Italic">Choeradoplana</Emphasis> (Tricladida,Continenticola, Geoplanidae) taxonomy

The land planarian genus Choeradoplana (Plathelminthes, Tricladida) is currently integrated by 13 species. In previous works, morphological variation in its type species, Choeradoplana iheringi, was reported, but no attempt to test whether it is just a single species has been made yet. In order to disentangle the taxonomy of this species and further members of the genus, we sampled new specimens and combined morphological and molecular data and also have evaluated the performance of diverse methods of molecular species delimitation. Our data point to the presence of two cryptic species named C. iheringi, plus two new species, all hidden under the same general appearance. An in-depth morphological study of the specimens allowed detection of diagnostic morphological traits in each species, for which we also propose a molecular diagnosis. This integrative taxonomic study demonstrates once again the usefulness of molecular tools to weigh minor morphological characteristics and thus reveal the existence of species that would otherwise remain cryptic. However, under certain parameters, the molecular methods may over-split species with a high genetic structure, maybe pointing to incipient speciation. This makes critical the use of these methods combined with a comprehensive morphological approach. We also present a comprehensive phylogenetic tree including most Choeradoplana species. The tree, well supported, allows making some preliminary inferences on the evolution of the group and its historical biogeography.     

Molecular and chemical characters to evaluate species status of two cuckoo bumblebees: Bombus barbutellus and Bombus maxillosus (Hymenoptera,Apidae, Bombini)

Many methods, based on morphological, molecular or chemical characters, have been used to address the question of species taxonomic status. Integrative taxonomy aims to define stronger supported taxonomic hypotheses by considering complementary datasets from different characters. By following an integrative approach, the present study includes molecular, chemical and morphological criteria to establish the taxonomic status of two rare and doubtful cuckoo bumblebee taxa: Bombus (Psithyrus) barbutellus and Bombus (Psithyrus) maxillosus. These two sympatric taxa are discriminated by few morphological criteria (mainly wing darkness and hair length). We used these morphological character diagnoses to establish an a priori status of our samples (23 specimens). We developed a combined molecular dataset from one nuclear gene, elongation factor 1α (EF‐1α), and one mitochondrial gene, cytochrome c oxidase subunit I (COI), spanning 1623 bp, and a chemical dataset of sexual marking pheromones (73 compounds). The molecular data were subjected to maximum‐likelihood and Bayesian phylogenetic inference under partitioned model and maximum parsimony. The chemical data were analysed by clustering and the two‐group k‐means method to test divergences between the two species. The resulting phylogenetic trees show no consistent divergence between the two taxa. Moreover, we found no divergence in the sexual marking pheromones in the clustering and two‐group k‐means analyses. These converging results support the conspecificity of both taxa. Nonetheless, our determinations using the traditional morphological criteria separated our samples into two taxa. We conclude that the morphological criteria seem to relate to intraspecific variations: B. maxillosus is regarded as a syn.n. of B. barbutellus.     

Integrating a Numerical Taxonomic Method and Molecular Phylogeny for Species Delimitation of Melampsora Species (Melampsoraceae,Pucciniales) on Willows in China

The species in genus Melampsora are the causal agents of leaf rust diseases on willows in natural habitats and plantations. However, the classification and recognition of species diversity are challenging because morphological characteristics are scant and morphological variation in Melampsora on willows has not been thoroughly evaluated. Thus, the taxonomy of Melampsora species on willows remains confused, especially in China where 31 species were reported based on either European or Japanese taxonomic systems. To clarify the species boundaries of Melampsora species on willows in China, we tested two approaches for species delimitation inferred from morphological and molecular variations. Morphological species boundaries were determined based on numerical taxonomic analyses of morphological characteristics in the uredinial and telial stages by cluster analysis and one-way analysis of variance. Phylogenetic species boundaries were delineated based on the generalized mixed Yule-coalescent (GMYC) model analysis of the sequences of the internal transcribed spacer (ITS1 and ITS2) regions including the 5.8S and D1/D2 regions of the large nuclear subunit of the ribosomal RNA gene. Numerical taxonomic analyses of 14 morphological characteristics recognized in the uredinial-telial stages revealed 22 morphological species, whereas the GMYC results recovered 29 phylogenetic species. In total, 17 morphological species were in concordance with the phylogenetic species and 5 morphological species were in concordance with 12 phylogenetic species. Both the morphological and molecular data supported 14 morphological characteristics, including 5 newly recognized characteristics and 9 traditionally emphasized characteristics, as effective for the differentiation of Melampsora species on willows in China. Based on the concordance and discordance of the two species delimitation approaches, we concluded that integrative taxonomy by using both morphological and molecular variations was an effective approach for delimitating Melampsora species on willows in China.     

Molecular phylogeny and systematics of flowering plants of the family Crassulaceae DC

Crassulaceae (orpine or stonecrop family) is the most species-rich (ca. 1400 spp) family in the order Saxifragales. Most members of the family are succulent plants. Phenotypic diversity and large number of species complicates systematics of the family and obscures reconstruction of relationship within it. Phylogenetic analyzes based on morphological and molecular markers placed Crassulaceae as one of the crown clades of Saxifragales. In this contribution a review of phylogenetic studies on the family Crassulaceae, based on DNA nucleotide sequence comparisons is presented; major clades established in the family are characterised; their structure and polyphylesis of some genera are discussed. It is shown that the traditional taxonomic structure of Crassulaceae contradicts pattern of phylogenetic relationships between its members. We critically analyzed recent taxonomic systems of the family and stress that homoplasy of morphological characters does not allow to use them to reconstruct relationships between crassulacean taxa even at the low taxonomic levels.     

Pattern of shape variation in the eastern African gerbils of the genus Gerbilliscus (Rodentia,Muridae): Environmental correlations and implication for taxonomy and systematics

Gerbilliscus has been in recent years the subject of new molecular and karyological investigations that shed new light on the evolutionary processes of this taxon. However, the patterns of phenotypic diversification of Gerbilliscus remain poorly understood. Furthermore, the molecular phylogenetic analyses posed new questions concerning the systematics and taxonomy of the whole genus and revealed the possible occurrence of cryptic species and hence the need to carry out a taxonomic revision.We used geometric morphometrics to investigate the adaptive value of morphological changes and to elucidate the systematic relationships and taxonomic status of the Gerbilliscus species of east Africa.Our results show concordance between morphological and genetic species boundaries. However, the observed morphological differences appear not only related to hereditary characters. In fact, the correlation with the climatic variables suggests an adaptive value of shape changes related to different trophic resources availability.Finally, discriminant analysis of the eastern Africa Gerbilliscus species highlight the distinctiveness of G. vicinus, clearly separated from G. robustus and G. nigricaudus.     

Concepts in protistology: species definitions and boundaries

This paper summarises the Symposium 'Concepts in Protistology', during the VI European Congress of Protistology, Berlin, 25-29 July 2011. There is an increasing focus on cataloguing the number of species on earth, species barcoding initiatives, and the increasing need to reconcile molecular with morphological data in protists within a taxonomic framework. We identify several obstructions to defining species in protists, including the high incidence of asexuality, high levels of both morphological conservation and evolutionary convergence, high levels of genetic diversity that cannot so far be correlated with phenotypic characters, conflicting signals between both genetic and phenotypic taxonomic markers, and different requirements and challenges of species definition in different protist groups. We assert that there is no species 'category' for protists, and recommend that a working definition of species is clarified on a case-by-case basis. Thus, a consensus approach may emerge within protist groups, but any one approach is unlikely to encompass a wide phylogenetic range. However, as long as clarity of intent and method is maintained, the utility of the term 'species' in protists will also be maintained as a reproducible and convenient (if artificial) way of referring to particular lineages within a tightly defined context.     

Review: Taxonomy of Nitella (Charales, Charophyceae) based on comparative morphology of oospores and multiple DNA marker phylogeny using cultured material

The genus Nitella is the largest group in the Charales and has the highest diversity of vegetative and oospore morphologies. In his worldwide monograph on the Charales, R. D. Wood characterized the sections and species of Nitella mainly on the basis of vegetative morphology and treated oospore wall morphology as diagnostic at the infraspecific level. Therefore, many species of Nitella exhibiting distinct external morphology of the oospore wall (EMOW) were reduced to infraspecific rank and only 53 of 204 species previously described were recognized within Nitella . However, recent morphological and molecular phylogenetic studies have demonstrated the phylogenetic validity of using EMOW for diagnosing some species of Nitella . More recently, a scanning electron microscopy (SEM) study of internal morphology of the oospore wall (IMOW) and multiple DNA marker analyses using both nuclear and chloroplast gene sequences were conducted to improve our understanding of the taxonomy of Nitella at the species level, on the basis of cultured material of a large number of species. Multiple DNA marker analyses resolved detailed and robust phylogenetic relationships within the genus and demonstrated the taxonomic and phylogenetic significance of IMOW. In addition, they supported the taxonomic decisions based on differences in oospore morphology, suggesting that an integrated approach, involving both SEM studies of the EMOW and IMOW and multiple DNA marker analyses, is appropriate to address problems at lower taxonomic levels within the genus, as well as to construct a natural taxonomic system for the genus. In this paper, recent morphological and molecular phylogenetic studies are reviewed and recent improvements in taxonomy of Nitella are summarized. Moreover, the evolution of morphological features and phylogenetic relationships within Nitella are discussed, focusing especially on oospore morphology.