Comprehensive DNA barcode coverage of North American birds

DNA barcoding seeks to assemble a standardized reference library for DNA-based identification of eukaryotic species. The utility and limitations of this approach need to be tested on well-characterized taxonomic assemblages. Here we provide a comprehensive DNA barcode analysis for North American birds including 643 species representing 93% of the breeding and pelagic avifauna of the USA and Canada. Most (94%) species possess distinct barcode clusters, with average neighbour-joining bootstrap support of 98%. In the remaining 6%, barcode clusters correspond to small sets of closely related species, most of which hybridize regularly. Fifteen (2%) currently recognized species are comprised of two distinct barcode clusters, many of which may represent cryptic species. Intraspecific variation is weakly related to census population size and species age. This study confirms that DNA barcoding can be effectively applied across the geographical and taxonomic expanse of North American birds. The consistent finding of constrained intraspecific mitochondrial variation in this large assemblage of species supports the emerging view that selective sweeps limit mitochondrial diversity.     

Establishment and dominance of an introduced herbivore has limited impact on native host-parasitoid food webs

The gypsy moth is considered one of the most harmful invasive forest insects in North America. It has been suggested that gypsy moth may indirectly impact native caterpillar communities via shared parasitoids. However, the impact of gypsy moth on forest insect food webs in general remains unstudied. Here we assess such potential impacts by surveying forest insect food webs in Ontario, Canada. We systematically collected caterpillars using burlap bands at sites with and without histories of gypsy moth outbreak, and then reared these caterpillars until potential parasitoid emergence. This procedure allowed us to generate quantitative food webs describing caterpillar-parasitoid interactions. We estimated the degree of parasitoid sharing between gypsy moth and native caterpillars. We also statistically modeled the effect of gypsy moth outbreak history and current gypsy moth abundance on standard indices of quantitative food web structure and the diversity of parasitoid communities. Rates of gypsy moth parasitism were very low and gypsy moth shared very few parasitoids with native caterpillars, suggesting limited potential for indirect interactions. We did not detect any significant effects of gypsy moth on either food web structure or parasitoid diversity, and the small amount of parasitoid sharing strongly implies that this lack of significance is not merely due to low statistical power. Our study suggests that gypsy moth has limited impact on native host-parasitoid food webs, at least for species that use burlap bands. Our results emphasize that extrapolations of theoretical and experimental conclusions on the impacts of invasive species should be tested in natural settings.     

Challenges to barcoding an entire flora

DNA barcodes are species‐specific genetic markers that allow taxonomic identification of biological samples. The promise of DNA barcoding as a rapid molecular tool for conducting biodiversity inventories has catalysed renewed efforts to document and catalogue the diversity of life, parallel to the large‐scale sampling conducted by Victorian naturalists. The unique contribution of DNA barcode data is in its ability to identify biotic material that would be impossible to classify using traditional taxonomic keys. However, the utility of DNA barcoding relies upon the construction of accurate barcode libraries that provide a reference database to match to unidentified samples. Whilst there has been much debate in the literature over the choice and efficacy of barcode markers, there has been little consideration of the practicalities of generating comprehensive barcode reference libraries for species‐rich floras. Here, we discuss several challenges to the generation of such libraries and present a case study from a regional biodiversity hotspot in southern Quebec. We suggest that the key challenges include (i) collection of specimens for rare or ephemeral species, (ii) limited access to taxonomic expertise necessary for reliable identification of reference specimens and (iii) molecular challenges in amplifying and matching barcode data. To be most effective, we recommend that sampling must be both flexible and opportunistic and conducted across the entire growing season by expert taxonomists. We emphasize that the success of the global barcoding initiative will depend upon the close collaboration of taxonomists, plant collectors, and molecular biologists.     

DNA Barcoding of Japanese Click Beetles (Coleoptera,Elateridae)

Click beetles (Coleoptera: Elateridae) represent one of the largest groups of beetle insects. Some click beetles in larval form, known as wireworms, are destructive agricultural pests. Morphological identification of click beetles is generally difficult and requires taxonomic expertise. This study reports on the DNA barcoding of Japanese click beetles to enable their rapid and accurate identification. We collected and assembled 762 cytochrome oxidase subunit I barcode sequences from 275 species, which cover approximately 75% of the common species found on the Japanese main island, Honshu. This barcode library also contains 20 out of the 21 potential pest species recorded in Japan. Our analysis shows that most morphologically identified species form distinct phylogenetic clusters separated from each other by large molecular distances. This supports the general usefulness of the DNA barcoding approach for quick and reliable identification of Japanese elaterid species for environmental impact assessment, agricultural pest control, and biodiversity analysis. On the other hand, the taxonomic boundary in dozens of species did not agree with the boundary of barcode index numbers (a criterion for sequence-based species delimitation). These findings urge taxonomic reinvestigation of these mismatched taxa.     

Wedding biodiversity inventory of a large and complex Lepidoptera fauna with DNA barcoding

By facilitating bioliteracy, DNA barcoding has the potential to improve the way the world relates to wild biodiversity. Here we describe the early stages of the use of cox1 barcoding to supplement and strengthen the taxonomic platform underpinning the inventory of thousands of sympatric species of caterpillars in tropical dry forest, cloud forest and rain forest in northwestern Costa Rica. The results show that barcoding a biologically complex biota unambiguously distinguishes among 97% of more than 1000 species of reared Lepidoptera. Those few species whose barcodes overlap are closely related and not confused with other species. Barcoding also has revealed a substantial number of cryptic species among morphologically defined species, associated sexes, and reinforced identification of species that are difficult to distinguish morphologically. For barcoding to achieve its full potential, (i) ability to rapidly and cheaply barcode older museum specimens is urgent, (ii) museums need to address the opportunity and responsibility for housing large numbers of barcode voucher specimens, (iii) substantial resources need be mustered to support the taxonomic side of the partnership with barcoding, and (iv) hand-held field-friendly barcorder must emerge as a mutualism with the taxasphere and the barcoding initiative, in a manner such that its use generates a resource base for the taxonomic process as well as a tool for the user.     

DNA barcoding of Pedicularis L.(Orobanchaceae): Evaluating four universal barcode loci in a large and hemiparasitic genus

One application ofDNA barcoding is species identification based on sequences of a short and standardized DNA region.In plants,various DNA regions,alone or in combination,have been proposed and investigated,but consensus on a universal plant barcode remains elusive.In this study,we tested the utility of four candidate barcoding regions (rbcL,matK,trnH-psbA,and internal transcribed spacer (ITS)) as DNA barcodes for discriminating species in a large and hemiparasitic genus Pedicularis (Orobanchaceae).Amplification and sequencing was successful using single primer pairs for rbcL,trnH-psbA,and ITS,whereas two primer pairs were required for matK.Patterns of sequence divergence commonly showed a “barcoding gap”,that is,a bimodal frequency distribution of pairwise distances representing genetic diversity within and between species,respectively Considering primer universality,ease of amplification and sequencing,and performance in discriminating species,we found the most effective single-region barcode for Pedicularis to be ITS,and the most effective two-region barcode to be rbcL +ITS.Both discriminated at least 78％ of the 88 species and correctly identified at least 89％ of the sequences in our sample,and were effective in placing unidentified samples in known species groups.Our results suggest that DNA barcoding has the potential to aid taxonomic research in Pedicularis,a species-rich cosmopolitan clade much in need of revision,as well as ecological studies in its center of diversity,the Hengduan Mountains region of China.     

Reading the complex skipper butterfly fauna of one tropical place

Background

An intense, 30-year, ongoing biodiversity inventory of Lepidoptera, together with their food plants and parasitoids, is centered on the rearing of wild-caught caterpillars in the 120,000 terrestrial hectares of dry, rain, and cloud forest of Area de Conservacion Guanacaste (ACG) in northwestern Costa Rica. Since 2003, DNA barcoding of all species has aided their identification and discovery. We summarize the process and results for a large set of the species of two speciose subfamilies of ACG skipper butterflies (Hesperiidae) and emphasize the effectiveness of barcoding these species (which are often difficult and time-consuming to identify).

Methodology/Principal Findings

Adults are DNA barcoded by the Biodiversity Institute of Ontario, Guelph, Canada; and they are identified by correlating the resulting COI barcode information with more traditional information such as food plant, facies, genitalia, microlocation within ACG, caterpillar traits, etc. This process has found about 303 morphologically defined species of eudamine and pyrgine Hesperiidae breeding in ACG (about 25% of the ACG butterfly fauna) and another 44 units indicated by distinct barcodes (n = 9,094), which may be additional species and therefore may represent as much as a 13% increase. All but the members of one complex can be identified by their DNA barcodes.

Conclusions/Significance

Addition of DNA barcoding to the methodology greatly improved the inventory, both through faster (hence cheaper) accurate identification of the species that are distinguishable without barcoding, as well as those that require it, and through the revelation of species “hidden” within what have long been viewed as single species. Barcoding increased the recognition of species-level specialization. It would be no more appropriate to ignore barcode data in a species inventory than it would be to ignore adult genitalia variation or caterpillar ecology.     

Parasitism rate, parasitoid community composition and host specificity on exposed and semi-concealed caterpillars from a tropical rainforest

The processes maintaining the enormous diversity of herbivore—parasitoid food webs depend on parasitism rate and parasitoid host specificity. The two parameters have to be evaluated in concert to make conclusions about the importance of parasitoids as natural enemies and guide biological control. We document parasitism rate and host specificity in a highly diverse caterpillar-parasitoid food web encompassing 266 species of lepidopteran hosts and 172 species of hymenopteran or dipteran parasitoids from a lowland tropical forest in Papua New Guinea. We found that semi-concealed hosts (leaf rollers and leaf tiers) represented 84 % of all caterpillars, suffered a higher parasitism rate than exposed caterpillars (12 vs. 5 %) and their parasitoids were also more host specific. Semi-concealed hosts may therefore be generally more amenable to biological control by parasitoids than exposed ones. Parasitoid host specificity was highest in Braconidae, lower in Diptera: Tachinidae, and, unexpectedly, the lowest in Ichneumonidae. This result challenges the long-standing view of low host specificity in caterpillar-attacking Tachinidae and suggests higher suitability of Braconidae and lower suitability of Ichneumonidae for biological control of caterpillars. Semi-concealed hosts and their parasitoids are the largest, yet understudied component of caterpillar—parasitoid food webs. However, they still remain much closer in parasitism patterns to exposed hosts than to what literature reports on fully concealed leaf miners. Specifically, semi-concealed hosts keep an equally low share of idiobionts (2 %) as exposed caterpillars.     

Food web structure changes with elevation but not rainforest stratum

Changes in species richness along elevational gradients are well documented. However, little is known about how trophic interactions between species and, in particular, the food webs that these interactions comprise, change with elevation. Here we present results for the first comparison of quantitative food webs in forest understorey and canopy along an elevational gradient. Replicate quantitative food webs were constructed for assemblages involving 23 species of cavity‐nesting Hymenoptera and 12 species of their parasitoids and kleptoparasites in subtropical rainforest in Australia. A total of 1589 insects were collected using trap nests across 20 plots distributed at sites ranging from 300 to 1100 m a.s.l. Insect abundance, insect diversity and parasitism rate generally decreased with increasing elevation. Food web structure significantly changed with elevation. In particular, weighted quantitative measures of linkage density, interaction evenness, nestedness (weighted NODF) and potential for enemy mediated interactions (PAC) decreased with increasing elevation, and network specialisation (H₂′) increased with increasing elevation, even after controlling for matrix size; but there was no change in weighted connectance. Changes in forest type and temperature along the elevational gradient are likely to be, at least partly, responsible for the patterns observed. We found no significant differences in insect abundance, insect diversity or parasitism rate between canopy and understorey. Furthermore, there were no differences in food web structure between strata. These results contribute further evidence to studies revealing changes in food web structure along natural environmental gradients and provide information that can potentially be used for predicting how communities may respond to climate change.     

Some ‘ant’swers: Application of a layered barcode approach to problems in ant taxonomy

DNA barcoding has emerged as a routine tool in modern taxonomy. Although straightforward, this approach faces new challenges, when applied to difficult situation such as defining cryptic biodiversity. Ants are prime examples for high degrees of cryptic biodiversity due to complex population differentiation, hybridization and speciation processes. Here, we test the DNA barcoding region, cytochrome c oxidase 1 and two supplementary markers, 28S ribosomal DNA and long‐wavelength rhodopsin, commonly used in ant taxonomy, for their potential in a layered, character‐based barcoding approach across different taxonomic levels. Furthermore, we assess performance of the character‐based barcoding approach to determine cryptic species diversity in ants. We found (i) that the barcode potential of a specific genetic marker varied widely among taxonomic levels in ants; (ii) that application of a layered, character‐based barcode for identification of specimens can be a solution to taxonomical challenging groups; (iii) that the character‐based barcoding approach allows us to differentiate specimens even within locations based on pure characters. In summary, (layered) character‐based barcoding offers a reliable alternative for problematic species identification in ants and can be used as a fast and cost‐efficient approach to estimate presence, absence or frequency of cryptic species.     

藻类DNA条形码研究进展

DNA barcode,又称为DNA条形码,是指利用短的标准DNA序列的核苷酸多样性进行物种的鉴定和快速识别.目前该方法在动物分类研究中应用广泛,其中线粒体的细胞色素c氧化酶亚基1(cytochrome c oxidase subunit 1,COI或cox 1)基因中的约700bp长度的一段被用来作为标准DNA片段.在陆地植物条形码研究中,生命-植物条形码联盟会(Consortium for the Barcode of Life-Plant Working Group,CBOL-Plant Working Group)近期推荐将植物叶绿体中的两个基因片段rbcL+ matK作为初步的陆生植物条形码,此组合能在70％的程度上进行植物物种的鉴别.在海藻的分类研究中,DNA条形码的应用较少,已有的研究主要集中在硅藻、红藻和褐藻,尚没有学者明确提出适合藻类的DNA条形码.总结了能够作为藻类DNA条形码的序列特点、应用流程及分析方法,综述了DNA条形码在藻类中的研究现状和存在的问题,展望了藻类DNA条形码的应用前景.     

Integrative taxonomy of Leptonetela spiders (Araneae,Leptonetidae), with descriptions of 46 new species

Extreme environments,such as subterranean habitats,are suspected to be responsible for morphologically inseparable cryptic or sibling species and can bias biodiversity assessment.A DNA barcode is a short,standardized DNA sequence used for taxonomic purposes and has the potential to lessen the challenges presented by a biotic inventory.Here,we investigate the diversity of the genus Leptonetela Kratochvíl,1978 that is endemic to karst systems in Eurasia using DNA barcoding.We analyzed 624 specimens using one mitochondrial gene fragment (COI).The results show that DNA barcoding is an efficient and rapid species identification method in this genus.DNA barcoding gap and automatic barcode gap discovery (ABGD) analyses indicated the existence of 90 species,a result consistent with previous taxonomic hypotheses,and supported the existence of extreme male pedipalpal tibial spine and median apophysis polymorphism in Leptonetela species,with direct implications for the taxonomy of the group and its diversity.Based on the molecular and morphological evidence,we delimit and diagnose 90 Leptonetela species,including the type species Leptonetela kanellisi (Deeleman-Reinhold,1971).Forty-six of them are previously undescribed.The female of Leptonetela zhai Wang & Li,2011 is reported for the first time.Leptonetela tianxinensis (Tong & Li,2008) comb.nov.is transferred from the genus Leptoneta Simon,1872;the genus Guineta Lin & Li,2010 syn.nov.is a junior synonym of Leptonetela;Leptonetela gigachela (Lin & Li,2010)comb.nov.is transferred from Guineta.The genus Sinoneta Lin & Li,2010 syn.nov.is a junior synonym of Leptonetela;Leptonetela notabilis (Lin & Li,2010) comb.nov.and Leptonetela sexdigiti (Lin & Li,2010) comb.nov.are transferred from Sinoneta;Leptonetela sanchahe Wang & Li nom.nov.is proposed as a replacement name for Sinoneta palmata (Chen et al.,2010) because Leptonetela palmata is preoccupied.     

Intensive agriculture reduces soil biodiversity across Europe

Soil biodiversity plays a key role in regulating the processes that underpin the delivery of ecosystem goods and services in terrestrial ecosystems. Agricultural intensification is known to change the diversity of individual groups of soil biota, but less is known about how intensification affects biodiversity of the soil food web as a whole, and whether or not these effects may be generalized across regions. We examined biodiversity in soil food webs from grasslands, extensive, and intensive rotations in four agricultural regions across Europe: in Sweden, the UK, the Czech Republic and Greece. Effects of land‐use intensity were quantified based on structure and diversity among functional groups in the soil food web, as well as on community‐weighted mean body mass of soil fauna. We also elucidate land‐use intensity effects on diversity of taxonomic units within taxonomic groups of soil fauna. We found that between regions soil food web diversity measures were variable, but that increasing land‐use intensity caused highly consistent responses. In particular, land‐use intensification reduced the complexity in the soil food webs, as well as the community‐weighted mean body mass of soil fauna. In all regions across Europe, species richness of earthworms, Collembolans, and oribatid mites was negatively affected by increased land‐use intensity. The taxonomic distinctness, which is a measure of taxonomic relatedness of species in a community that is independent of species richness, was also reduced by land‐use intensification. We conclude that intensive agriculture reduces soil biodiversity, making soil food webs less diverse and composed of smaller bodied organisms. Land‐use intensification results in fewer functional groups of soil biota with fewer and taxonomically more closely related species. We discuss how these changes in soil biodiversity due to land‐use intensification may threaten the functioning of soil in agricultural production systems.     

Ecology in the age of DNA barcoding: the resource,the promise and the challenges ahead

Ten years after DNA barcoding was initially suggested as a tool to identify species, millions of barcode sequences from more than 1100 species are available in public databases. While several studies have reviewed the methods and potential applications of DNA barcoding, most have focused on species identification and discovery, and relatively few have addressed applications of DNA barcoding data to ecology. These data, and the associated information on the evolutionary histories of taxa that they can provide, offer great opportunities for ecologists to investigate questions that were previously difficult or impossible to address. We present an overview of potential uses of DNA barcoding relevant in the age of ecoinformatics, including applications in community ecology, species invasion, macroevolution, trait evolution, food webs and trophic interactions, metacommunities, and spatial ecology. We also outline some of the challenges and potential advances in DNA barcoding that lie ahead.     

The potential for indirect effects between a weed, one of its biocontrol agents and native herbivores: A food web approach

The safety of biological control is a contentious issue. We suggest that constructing and analyzing food webs may be a valuable addition to standard biological control research techniques, as they offer a means of assessing the post-release safety of control agents. Using preliminary data to demonstrate the value of food webs in biocontrol programs, we quantified the extent to which a key agent has infiltrated natural communities in Australia and, potentially, impacted on non-target species. Using these data, we also demonstrate how food webs can be used to generate testable hypotheses regarding indirect interactions between introduced agents and non-target species. We developed food webs in communities invaded to varying degrees by an exotic weed, bitou bush, Chrysanthemoides monilifera ssp. rotundata, and a key biocontrol agent for this weed in Australia, the tephritid fly, Mesoclanis polana. Three food webs were constructed during springtime showing the interactions between plants, seed-feeding insects and their parasitoids. One food web was constructed in a plot of native Australian vegetation that was free of bitou bush (‘bitou-free’), another in a plot of Australian vegetation surrounded by an invasion of bitou bush (‘bitou-threatened’) and a third from a plot infested with a monoculture of bitou bush (‘bitou-infested’). The bitou-free web contained 36 species, the bitou-threatened plot 9 species and the bitou-infested web contained 6 species. One native Australian herbivore attacked the seeds of bitou bush. M. polana, a seed-feeding fly, was heavily attacked by native parasitoids, these being more abundant than the parasitoids feeding on the native seed feeders. A surprising result is that none of the three species of native parasitoids reared from M. polana were reared from any of the native herbivores. The food webs revealed how a highly host-specific biocontrol agent, such as M. polana has the potential to change community structure by increasing the abundance of native parasitoids. The webs also suggest that indirect interactions between M. polana and native non-target species are possible, these been mediated by shared parasitoids. The experiments necessary to determine the presence of these interactions are outlined.     

DNA barcoding of economically important freshwater fish species from north‐central Nigeria uncovers cryptic diversity

This study examines the utility of morphology and DNA barcoding in species identification of freshwater fishes from north‐central Nigeria. We compared molecular data (mitochondrial cytochrome c oxidase subunit I (COI) sequences) of 136 de novo samples from 53 morphologically identified species alongside others in GenBank and BOLD databases. Using DNA sequence similarity‐based (≥97% cutoff) identification technique, 50 (94.30%) and 24 (45.30%) species were identified to species level using GenBank and BOLD databases, respectively. Furthermore, we identified cases of taxonomic problems in 26 (49.00%) morphologically identified species. There were also four (7.10%) cases of mismatch in DNA barcoding in which our query sequence in GenBank and BOLD showed a sequence match with different species names. Using DNA barcode reference data, we also identified four unknown fish samples collected from fishermen to species level. Our Neighbor‐joining (NJ) tree analysis recovers several intraspecific species clusters with strong bootstrap support (≥95%). Analysis uncovers two well‐supported lineages within Schilbe intermedius. The Bayesian phylogenetic analyses of Nigerian S. intermedius with others from GenBank recover four lineages. Evidence of genetic structuring is consistent with geographic regions of sub‐Saharan Africa. Thus, cryptic lineage diversity may illustrate species’ adaptive responses to local environmental conditions. Finally, our study underscores the importance of incorporating morphology and DNA barcoding in species identification. Although developing a complete DNA barcode reference library for Nigerian ichthyofauna will facilitate species identification and diversity studies, taxonomic revisions of DNA sequences submitted in databases alongside voucher specimens are necessary for a reliable taxonomic and diversity inventory.     

The campaign to DNA barcode all fishes, FISH-BOL

FISH-BOL, the Fish Barcode of Life campaign, is an international research collaboration that is assembling a standardized reference DNA sequence library for all fishes. Analysis is targeting a 648 base pair region of the mitochondrial cytochrome c oxidase I (COI) gene. More than 5000 species have already been DNA barcoded, with an average of five specimens per species, typically vouchers with authoritative identifications. The barcode sequence from any fish, fillet, fin, egg or larva can be matched against these reference sequences using BOLD; the Barcode of Life Data System ( http://www.barcodinglife.org ). The benefits of barcoding fishes include facilitating species identification, highlighting cases of range expansion for known species, flagging previously overlooked species and enabling identifications where traditional methods cannot be applied. Results thus far indicate that barcodes separate c. 98 and 93% of already described marine and freshwater fish species, respectively. Several specimens with divergent barcode sequences have been confirmed by integrative taxonomic analysis as new species. Past concerns in relation to the use of fish barcoding for species discrimination are discussed. These include hybridization, recent radiations, regional differentiation in barcode sequences and nuclear copies of the barcode region. However, current results indicate these issues are of little concern for the great majority of specimens.     

DNA Barcoding for the Identification of Sand Fly Species (Diptera,Psychodidae, Phlebotominae) in Colombia

Sand flies include a group of insects that are of medical importance and that vary in geographic distribution, ecology, and pathogen transmission. Approximately 163 species of sand flies have been reported in Colombia. Surveillance of the presence of sand fly species and the actualization of species distribution are important for predicting risks for and monitoring the expansion of diseases which sand flies can transmit. Currently, the identification of phlebotomine sand flies is based on morphological characters. However, morphological identification requires considerable skills and taxonomic expertise. In addition, significant morphological similarity between some species, especially among females, may cause difficulties during the identification process. DNA-based approaches have become increasingly useful and promising tools for estimating sand fly diversity and for ensuring the rapid and accurate identification of species. A partial sequence of the mitochondrial cytochrome oxidase gene subunit I (COI) is currently being used to differentiate species in different animal taxa, including insects, and it is referred as a barcoding sequence. The present study explored the utility of the DNA barcode approach for the identification of phlebotomine sand flies in Colombia. We sequenced 700 bp of the COI gene from 36 species collected from different geographic localities. The COI barcode sequence divergence within a single species was <2% in most cases, whereas this divergence ranged from 9% to 26.6% among different species. These results indicated that the barcoding gene correctly discriminated among the previously morphologically identified species with an efficacy of nearly 100%. Analyses of the generated sequences indicated that the observed species groupings were consistent with the morphological identifications. In conclusion, the barcoding gene was useful for species discrimination in sand flies from Colombia.