Inconsistent estimates of diversity between traditional and DNA taxonomy in bdelloid rotifers

Microscopic animals offer great potential in the analysis of spatial patterns of diversity, as they may provide different scenarios for biogeography and macroecology, but understanding diversity of microscopic animals is hampered by lack of comprehensive data on species distribution and by unreliable taxonomy. DNA taxonomy may prove useful in obtaining reliable data in the future, but we still do not know to what extent traditional and DNA taxonomy can be comparable for microscopic organisms. In this paper, we compare analyses and estimates of diversity at the level of species assemblage between traditional and DNA taxonomy for a group of moss-dwelling microscopic animals, bdelloid rotifers. The results are straightforward: Traditional species identification underestimates diversity by factors of 2 at the local and 2.5 at the regional scale. We discuss the results in the framework of current hypotheses on the distribution of microscopic animals.     

Global diversity of rotifers (Rotifera) in freshwater

Rotifera is a Phylum of primary freshwater Metazoa containing two major groups: the heterogonic Monogononta and the exclusively parthenogenetic Bdelloidea. Monogononta contains 1,570 species-level taxa, of which a majority (1,488) are free-living fresh or inland water taxa. Bdelloidea contains 461 “species,” only one of which is marine, but with many limnoterrestrial representatives or animals of unknown ecology. Actual numbers may be much higher, considering the occurrence of cryptic speciation in Monogononta and the unsatisfactory nature of taxonomic knowledge. Rotifers, mostly monogononts, occur in all types of water bodies, worldwide. They are particularly diverse in the littoral zone of stagnant waterbodies with soft, slightly acidic water and under oligo- to mesotrophic conditions. The rotifer record is highest in the Northern hemisphere, which may be due to the concentration of studies in those regions. Diversity is highest in the (sub)tropics; hotspots are northeast North America, tropical South America, Southeast Asia, Australia, and Lake Baikal, endemicity is low in Africa (including Madagascar), Europe, the Indian subcontinent, and Antarctica. Although the lack of fossil evidence and of molecular phylogenetic studies are major hindrances, contrasting hypotheses on the origin and evolutionary history of Brachionus, Macrochaetus, and Trichocerca are presented. Guest editors: E. V. Balian, C. Lévêque, H. Segers & K. Martens Freshwater Animal Diversity Assessment     

DNA条形码与动植物分类学的研究

DNA条形码在动植物分类学中的研究最近几年非常火热。这项技术在国外研究的比较多,国内的许多方面的研究还没达到国际水平。我国的动植物种类比较繁多,地区差异也较大,怎样能将这些物种准确,快速的进行分类,是众多动植物学家一直在研究的难题。针对DNA条形码研究的现状和他在动植物学中应用以及它存在的争议来进一步认识DNA条形码。     

DNA barcoding should accompany taxonomy - the case of Cerebratulus spp (Nemertea)

Many issues in DNA barcoding need to be solved before it can reach its goal to become a general database for species identification. While species delimitations are more or less well established in several taxa, there are still many groups where this is not the case. Without the proper taxonomic background/knowledge and corroboration with other kinds of data, the DNA barcoding approach may fail to identify species accurately. The classification and taxonomy of phylum Nemertea (nemerteans, ribbon worms) are traditionally based on morphology, but are not corroborated by an increasing amount of genetic data when it comes to classification either into species or into higher taxa. The taxonomy of the phylum needs to be improved before the full potential of DNA barcoding can be utilized to make sure that valid Linnean names accompany the barcode sequences. We illustrate the problematic situation in the phylum Nemertea by a case study from the genus Cerebratulus.     

Utility of DNA taxonomy and barcoding for the inference of larval community structure in morphologically cryptic Chironomus (Diptera) species

Biodiversity studies require species level analyses for the accurate assessment of community structures. However, while specialized taxonomic knowledge is only rarely available for routine identifications, DNA taxonomy and DNA barcoding could provide the taxonomic basis for ecological inferences. In this study, we assessed the community structure of sediment dwelling, morphologically cryptic Chironomus larvae in the Rhine-valley plain/Germany, comparing larval type classification, cytotaxonomy, DNA taxonomy and barcoding. While larval type classification performed poorly, cytotaxonomy and DNA-based methods yielded comparable results: detrended correspondence analysis and permutation analyses indicated that the assemblages are not randomly but competitively structured. However, DNA taxonomy identified an additional species that could not be resolved by the traditional method. We argue that DNA-based identification methods such as DNA barcoding can be a valuable tool to increase accuracy, objectivity and comparability of the taxonomic assessment in biodiversity and community ecology studies.     

The Rotifera of Hummock-Hollow Formations in a Poor (Mesotrophic) Fen in Newfoundland

Rotifers encountered from a poor (mesotrophic) fen, Avalon Peninsula of Newfoundland, are listed. Sixty-four species and varieties, including 25 Bdelloidea and 39 Monogononta, were recorded. All but 4 of the reported species and varieties are new records for the Province of Newfoundland, while 27 are new records for Canada; of these, 13 are new for North America. A seasonal quantitative study of rotifer communities in the hummock-hollow formations on the fen was carried out from August 1973 to October 1974. An average of 354 rotifers/cm² (17 species) was found per formation. Monogononta almost totally disappeared during winter, while the Bdelloidea decreased somewhat in numbers. The tops of hummocks contained mainly bdelloid species. The number of monogononts increased both in species and individuals on the slopes of hummocks and was greatest in the hollows, although the total number of rotifers was less in these areas than in the hummock tops. Some physical and biological factors affecting the rotifer communities were examined. Rotifers in these sites are not subjected to dessication. Predation was not an important factor. A quantitative sampling method was devised for procuring relatively accurate samples from living Sphagnum, and for extracting rotifers from these samples.     

First description of the serotonergic nervous system in a bdelloid rotifer: Macrotrachela quadricornifera Milne 1886 (Philodinidae)

Class Bdelloidea of phylum Rotifera comprises aquatic microinvertebrates that are known for both obligate parthenogenesis and for resisting desiccation through a dormant reversible state. In the frame of an investigation about the role of the nervous system in controlling life cycle, reproduction and dormancy, we describe the serotonergic system of a bdelloid, Macrotrachela quadricornifera, using serotonin immunohistochemistry and confocal laser scanning microscopy. Serotonin immunoreactivity is present in the cerebral ganglion, lateral nerve cords and peripheral neurites. The cerebral ganglion consists of perikarya that send neurites cephalically to the rostrum and corona. A pair of neurites exits the cerebral ganglion as lateral nerve cords, and proceeds caudally to the pedal ganglion where additional neurites enter the foot. Based on the location of serotonergic immunoreactivity, we hypothesize that the neurotransmitter is involved in both motor activity (e.g., ciliary beating, inchworm-like locomotion) and sensory activity. A comparison between the serotonergic nervous systems of M. quadricornifera and species of Monogononta reveals differences in the numbers and patterns of cerebral perikarya, peripheral perikarya, and periperhal neurites. These differences may have functional significance for understanding adaptations to specific environments and/or systematic significance for reconstructing the rotiferan ground pattern.     

Spatial patterns of plant diversity below-ground as revealed by DNA barcoding

Our understanding of the spatial organization of root diversity in plant communities and of the mechanisms of community assembly has been limited by our ability to identify plants based on root tissue, especially in diverse communities. Here, we test the effectiveness of the plastid gene rbcL, a core plant DNA barcoding marker, for investigating spatial patterns of root diversity, and relate observed patterns to above-ground community structure. We collected 3800 root fragments from four randomly positioned, 1-m-deep soil profiles (two vertical transects per plot), located in an old-field community in southern Ontario, Canada, and extracted and sequenced DNA from 1531 subsampled fragments. We identified species by comparing sequences with a DNA barcode reference library developed previously for the local flora. Nearly 85% of sampled root fragments were successfully sequenced and identified as belonging to 29 plant species or species groups. Root abundance and species richness varied in horizontal space and were negatively correlated with soil depth. The relative abundance of taxa below-ground was correlated with their frequency above-ground (r = 0.73, P = 0.0001), but several species detected in root tissue were not observed in above-ground quadrats. Multivariate analyses indicated that diversity was highly structured below-ground, and associated with depth, root morphology, soil chemistry and soil texture, whereas little structure was evident above-ground. Furthermore, analyses of species co-occurrence indicates strong species segregation overall but random co-occurrence among confamilials. Our results provide insights into the role of environmental filtering and competitive interactions in the organization of plant diversity below-ground, and also demonstrate the utility of barcoding for the identification of plant roots.     

Assessing the diversity of New Zealand freshwater harpacticoid copepods (Crustacea: Copepoda) using mitochondrial DNA (COI) barcodes

Taxonomic and ecological studies of freshwater harpacticoid copepods are limited globally by the ability to easily and accurately identify specimens. Here, we test the use of the mitochondrial cytochrome c oxidase subunit I (COI) gene locus as a tool for assessing the diversity of freshwater Harpacticoida. We obtained sequences from New Zealand harpacticoid copepods, representing two families, five genera and nine species, including the non-indigenous Elaphoidella sewelli. All species were delineated by the COI gene. However, high intraspecific diversity was evident among populations of Elaphoidella bidens (>12%), and between North and South Island populations of Bryocamptus pygmaeus (>18%), potentially indicating the presence of morphologically cryptic taxa. We suggest that mitochondrial DNA (COI) sequences can provide a useful tool for the routine identification of freshwater harpacticoid copepods. Applications of these data will include assessing species diversity and biogeography as well as assisting with the detection of non-indigenous species.     

Chloroplast DNA diversity in the genusRubus (Rosaceae) revealed by Southern hybridization

The variability in chloroplast DNA type of 20Rubus genotypes was examined by Southern hybridization. DNA extracted from theRubus accessions was digested with two restriction enzymes (EcoRI and EcoRV) and heterologous chloroplast DNA sequences from barley and pea were used as probes to detectRubus chloroplast DNA sequences on Southern blots ofRubus total DNA. Restriction fragment length polymorphism was detected and a total of 92 restriction fragments were generated by the probe/enzyme combinations examined. Cladistic principles based on the parsimony assumption were used to assemble a phylogenetic tree based on chloroplast restriction fragment length data. The phylogenetic tree grouped the taxonomically defined species and is in general agreement with information based on morphological criteria. However, the Japanese red raspberryR. illecebrosus was shown to have diverged considerably in terms of evolutionary time from other species in subg.Idaeobatus. Furthermore, the molecular approach provides a quantitative estimate of the relationship between species that is difficult to obtain from morphological data. In order to complement the chloroplast DNA information a ribosomal DNA probe was also included in the analysis and provided further information on the phylogenetic relationships withinRubus.     

Cryptic clonal lineages and genetic diversity in the loach Misgurnus anguillicaudatus (Teleostei: Cobitidae) inferred from nuclear and mitochondrial DNA analyses

In the loach Misgurnus anguillicaudatue, the asexual lineage, which produces unreduced clonal diploid eggs, has been identified. Among 833 specimens collected from 54 localities in Japan and two localities in China, 82 candidates of other lineage(s) of cryptic clones were screened by examining RFLP (restriction fragment length polymorphism)-PCR haplotypes in the control region of mtDNA. This analysis was performed because triploid loaches arise from the accidental incorporation of the sperm nucleus into unreduced diploid eggs of a clone. The categorization of members belonging to three newly identified lineages (clones 2–4) and the previously identified clonal lineage (clone 1) was verified by evaluating the genetic identity between two or more individuals from each clonal lineage based on RAPD (random amplified polymorphic DNA)-PCR and multilocus DNA fingerprints. We detected 75 haplotypes by observing the nucleotide status at variable sites from the control region of mtDNA. Phylogenic trees constructed from such sequences showed two highly diversified clades, A and B, that were beyond the level common for interspecific genetic differentiation. That result suggests that M. anguillicaudatus in Japan is not a single species entity. Two clone-specific mtDNA sequences were included in clade A, and the loaches with such sequences may be the maternal origin of the clones. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Distribution of Brachionidae (Rotifera,Monogononta) in Inner Mongolian Waters

Two-hundred and fourty-six freshwater and saline-alkaline waters in Inner Mongolia, P.R. China were surveyed for Brachionidae. Twenty-nine brachionid morphospecies (33 taxa) are recorded, four are new to China. Some data on their distribution, and comments on selected taxa are provided. Brachionus asplanchnoides (stat.nov.) is reinstated at the morphospecies level. We suggest that this taxon may be used in aquaculture, considering that, by its large size, it extends the size range of brackish-water Rotifera potentially available as food for fish larvae.     

Deep genetic divergences in Aoraki denticulata (Arachnida, Opiliones, Cyphophthalmi): a widespread 'mite harvestman' defies DNA taxonomy

Aoraki denticulata (Arachnida, Opiliones, Cyphophthalmi, Pettalidae), a widespread 'mite harvestman' endemic to the South Island of New Zealand, is found in leaf littler habitats throughout Nelson and Marlborough, and as far south as Arthur's Pass. We investigated the phylogeography and demographic history of A. denticulata in the first genetic population-level study within Opiliones. A total of 119 individuals from 17 localities were sequenced for 785 bp of the gene cytochrome c oxidase subunit I; 102 of these individuals were from the Aoraki subspecies A. denticulata denticulata and the remaining 17 were from the subspecies A. denticulata major. An extraordinarily high degree of genetic diversity was discovered in A. denticulata denticulata, with average uncorrected p-distances between populations as high as 19.2%. AMOVA, average numbers of pairwise differences, and pairwise F(ST) values demonstrated a significant amount of genetic diversity both within and between populations of this subspecies. Phylogenetic analysis of the data set revealed many well-supported groups within A. denticulata denticulata, generally corresponding to clusters of specimens from single populations with short internal branches, but separated by long branches from individuals from other populations. No haplotypes were shared between populations of the widespread small subspecies, A. denticulata denticulata. These results indicate a subspecies within which very little genetic exchange occurs between populations, a result consistent with the idea that Cyphophthalmi are poor dispersers. The highly structured populations and deep genetic divergences observed in A. denticulata denticulata may indicate the presence of cryptic species. However, we find a highly conserved morphology across sampling localities and large genetic divergences within populations from certain localities, equivalent to those typically found between populations from different localities. Past geological events may have contributed to the deep genetic divergences observed between sampling localities; additionally, the high divergence within populations of A. denticulata denticulata suggests that the rate of COI evolution may be accelerated in this taxon. In contrast, the larger subspecies A. denticulata major shows much less differentiation between and within sampling localities, suggesting that it may disperse more easily than its smaller counterpart. The fact that the remarkable genetic divergences within populations of A. denticulata denticulata from certain localities are equivalent to divergences between localities poses a challenge to the rapidly spreading practice of DNA taxonomy.     

Study of COI sequences from endemic New Zealand aphids highlights high mitochondrial DNA diversity in Rhopalosiphina (Hemiptera: Aphididae)

Focussed searches were made across New Zealand between 2013 and 2016, for endemic aphids from the Schizaphis (Rhopalosiphina) genus, which is currently represented by two putative, undescribed species from the endemic host plants Aciphylla and Dracophyllum. Cytochrome c oxidase I (COI) gene sequences (48 in total) from the Schizaphis were analysed together with those from a broader collection of New Zealand endemic aphids that has been assembled since the year 2000. The bulk of the Schizaphis belonged to two clusters corresponding to the host plant genera. Two aphids from central North Island Dracophyllum represented a much diverged lineage without clear affiliations to other New Zealand Schizaphis. Inter-population variation in the New Zealand Schizaphis was high compared with that seen in international studies of Aphidinae and among populations of other endemic New Zealand Aphidina. Within Schizaphis from Dracophyllum, geography played an apparent role in genetic structuring, with populations from Taranaki (North Island) and especially Mt Lyford (South Island) being divergent from those on the South Island main divide. Two distinct lineages of Schizaphis, which co-occurred at some sites, were found on Aciphylla. Our sequence comparisons, including GMYC analyses, indicated up to five New Zealand Schizaphis lineages, and two newly discovered endemic Aphis species from the host plants Clematis and Hebe.     

The hidden diversity of Coleodactylus amazonicus (Sphaerodactylinae,Gekkota) revealed by molecular data

Coleodactylus amazonicus, a small leaf-litter diurnal gecko widely distributed in Amazon Basin, has been considered a single species with no significant morphological differences between populations along its range. A recent molecular study, however, detected large genetic differences between populations of central Amazonia and those in the easternmost part of the Amazon Basin, suggesting the presence of taxonomically unrecognised diversity. In this study, DNA sequences of three mitochondrial (16S, cytb, and ND4) and two nuclear genes (RAG-1, c-mos) were used to investigate whether the species currently identified as C.amazonicus contains morphologically cryptic species lineages. The present phylogenetic analysis reveals further genetic subdivision including at least five potential species lineages, restricted to northeastern (lineage A), southeastern (lineage B), central-northern (lineage E) and central-southern (lineages C and D) parts of Amazon Basin. All clades are characterized by exclusive groups of alleles for both nuclear genes and highly divergent mitochondrial haplotype clades, with corrected pairwise net sequence divergence between sister lineages ranging from 9.1% to 20.7% for the entire mtDNA dataset. Results of this study suggest that the real diversity of “C. amazonicus” has been underestimated due to its apparent cryptic diversification.     

Integrative taxonomy at work: DNA barcoding of taeniids harboured by wild and domestic cats

In modern taxonomy, DNA barcoding is particularly useful where biometric parameters are difficult to determine or useless owing to the poor quality of samples. These situations are frequent in parasitology. Here, we present an integrated study, based on both DNA barcoding and morphological analysis, on cestodes belonging to the genus Taenia, for which biodiversity is still largely underestimated. In particular, we characterized cestodes from Italian wildcats (Felis silvestris silvestris), free‐ranging domestic cats (Felis silvestris catus) and hybrids populations. Adult taeniids were collected by post‐mortem examinations of the hosts and morphologically identified as Taenia taeniaeformis. We produced cox1 barcode sequences for all the analysed specimens, and we compared them with reference sequences of individuals belonging to the genus Taenia retrieved from GenBank. In order to evaluate the performance of a DNA barcoding approach to discriminate these parasites, the strength of correlation between species identification based on classical morphology and the molecular divergence of cox1 sequences was measured. Our study provides clear evidence that DNA barcoding is highly efficient to reveal the presence of cryptic lineages within already‐described taeniid species. Indeed, we detected three well‐defined molecular lineages within the whole panel of specimens morphologically identified as T. taeniaeformis. Two of these molecular groups were already identified by other authors and should be ranked at species level. The third molecular group encompasses only samples collected in Italy during this study, and it represents a third candidate species, still morphologically undescribed.     

Exploring the species diversity of Trichoderma in Norwegian drinking water systems by DNA barcoding

A total of 123 Trichoderma strains were isolated from Norwegian surface-sourced drinking water. The water samples included raw water, treated water, and water from private homes and hospital installations. Trichoderma species are difficult to differentiate morphologically, but recent molecular identification tools, including DNA barcoding, successfully distinguish between closely related species. The diversity of Trichoderma spp. was explored by DNA sequencing of internal transcribed spacer (ITS) and translation elongation factor 1 alpha (TEF-1α). Sequence identification was performed in the TrichOKEY version 2.0 barcode program and in the multilocus similarity search database TrichoBLAST, combined with traditional blast searches in the EMBL/GenBank. A total of 11 known Trichoderma/Hypocrea species were identified. In addition, one group of unidentified Trichoderma strains was found to represent a separate, strongly supported subclade within the Pachybasium'A'/Hamatum clade, based on their TEF-1α haplotypes. Trichoderma viride comprised 49% of the identified strains, and was represented by four and eight slightly different ITS and TEF-1α haplotypes, respectively. Approximately 22% of the surface-derived water samples were positive for T. viride, and the species was frequently isolated throughout the surface-sourced drinking water distribution system. The results indicate that a broad range of Trichoderma species are present in Norwegian surface-sourced drinking. Water treatment has minor effect in removing Trichoderma from raw water, and active growth in the water distribution system is likely to occur.     

Molecular taxonomy and species delimitation in Andean Schistocerca (Orthoptera: Acrididae)

The acridian genus Schistocerca comprises about 50 species which are endemic to the New World, except the Old World locust S. gregaria. Their morphological identification is rendered difficult by phase polyphenism, geographical overlap due to migrations or swarming, the difficulty to easily differentiate genitalia and the occurrence of interspecific hybrids. The three species reported from Peru include the swarming species S. interrita, a pest that can be recognized only by taxonomists. We show that it can be unambiguously identified using a mitochondrial DNA fragment known to have barcoding properties in this genus. We used several methods to delimitate Peruvian species. While S. interrita and S. pallens were well characterized, S. piceifrons peruviana was split into several taxa by a phylogeny-based method, whereas a combination of population genetics methods led one to identify only the three nominal species. A tentative reconstruction of the species history shows that several populations of S. piceifrons peruviana have recently increased in number, while exchanging some migrants, whereas an isolated population at the northern margin of the species range is substantially differentiated while exchanging no migrants with the others. This complex history has resulted in an atypical lineage pattern that appears to have confounded the standard assumptions underlying available species delimitation methods. Because of its behavioral property which tends to keep it panmictic, the identification of the swarming S. interrita remained unaffected.