Tuesday, April 25, 2017

Tuesday reads

Happy DNA Day! Has been a little while since the last post but I have been busy with a lot of different tasks and working myself into new projects. Here is hope that things will go back to normal slowly. Lots of interesting reads today starting with two of my own.

The School Malaise Trap Program (SMTP) provides a technologically sophisticated and scientifically relevant educational experience that exposes students to the diversity of life, enhancing their understanding of biodiversity while promoting environmental stewardship. Since 2013, the SMTP has allowed 15,000 students at 350 primary and secondary schools to explore insect diversity in Canadian schoolyards. Students at each school collected hundreds of insects for an analysis of DNA sequence variation that enabled their rapid identification to a species. Through this hands-on approach, they participated in a learning exercise that conveys a real sense of scientific discovery. As well, the students contributed valuable data to the largest biodiversity genomics initiative ever undertaken: the International Barcode of Life project. To date, the SMTP has sequenced over 80,000 insect specimens, which includes representatives of 7,990 different species, nearly a tenth of the Canadian fauna. Both surprisingly and importantly, the collections generated the first DNA barcode records for 1,288 Canadian species.

To date the global initiative to barcode all fishes, FISH-BOL, has delivered barcodes for approximately 14,400 of the 30,000 fish species; there is still much to do to attain its ultimate goal of barcoding all the world’s fishes. One strategy to overcome local gaps is to initiate short but intensive efforts to collect and barcode as many species as possible from a small region – a barcode ‘blitz’. This study highlights one such event, for the marine waters around Lizard island in the Great Barrier Reef (Queensland, Australia). Barcode records were obtained from 983 fishes collected over a two-week period. The resulting dataset comprised 358 named species and another 13 species that presently can only be reliably identified to genus level. Overall, this short expedition provided DNA barcodes for 13% of all marine fish species known to occur in Queensland.

The difficulty of censusing marine animal populations hampers effective ocean management. Analyzing water for DNA traces shed by organisms may aid assessment. Here we tested aquatic environmental DNA (eDNA) as an indicator of fish presence in the lower Hudson River estuary. A checklist of local marine fish and their relative abundance was prepared by compiling 12 traditional surveys conducted between 1988–2015. To improve eDNA identification success, 31 specimens representing 18 marine fish species were sequenced for two mitochondrial gene regions, boosting coverage of the 12S eDNA target sequence to 80% of local taxa. We collected 76 one-liter shoreline surface water samples at two contrasting estuary locations over six months beginning in January 2016. eDNA was amplified with vertebrate-specific 12S primers. Bioinformatic analysis of amplified DNA, using a reference library of GenBank and our newly generated 12S sequences, detected most (81%) locally abundant or common species and relatively few (23%) uncommon taxa, and corresponded to seasonal presence and habitat preference as determined by traditional surveys. Approximately 2% of fish reads were commonly consumed species that are rare or absent in local waters, consistent with wastewater input. Freshwater species were rarely detected despite Hudson River inflow. These results support further exploration and suggest eDNA will facilitate fine-scale geographic and temporal mapping of marine fish populations at relatively low cost.

We report the discovery of a non-native gammarid, Gammarus fossarum (Koch, 1836) (Crustacea, Amphipoda), in UK rivers. Gammarus fossarum is a common freshwater gammarid in many parts of mainland Europe, but was previously considered absent from the UK. Gammarus fossarum was detected in a number of UK rivers following DNA metabarcoding of a mini-barcode region of the COI gene in macroinvertebrate kick samples, and environmental DNA (eDNA) from water and sediment samples. Subsequent morphological analysis and standard DNA barcoding showed that the species could be reliably identified and separated from Gammarus pulex (Linnaeus, 1758), the most dominant and widespread native freshwater gammarid in the UK. Our data demonstrate extensive geographical coverage of G. fossarum in the UK, spanning distant river catchments. At present there is no data to confirm the likely origin of G. fossarum’s introduction. Subsequent re-examination of historic archive material shows the species to have been present in the UK since at least 1964. This study is among the first to demonstrate the potential of eDNA metabarcoding for detection of new non-native species. 

Consumption of frog legs is increasing worldwide, with potentially dramatic effects for ecosystems. More and more functioning frog farms are reported to exist. However, due to the lack of reliable methods to distinguish farmed from wild-caught individuals, the origin of frogs in the international trade is often uncertain. Here, we present a new methodological approach to this problem. We investigated the isotopic composition of legally traded frog legs from suppliers in Vietnam and Indonesia. Muscle and bone tissue samples were examined for δ15N, δ13C, and δ18O stable isotope compositions, to elucidate the conditions under which the frogs grew up. We used DNA barcoding (16S rRNA) to verify species identities. We identified three traded species (Hoplobatrachus rugulosus, Fejervarya cancrivora and Limnonectes macrodon); species identities were partly deviating from package labeling. Isotopic values of δ15N and δ18O showed significant differences between species and country of origin. Based on low δ15N composition and generally little variation in stable isotope values, our results imply that frogs from Vietnam were indeed farmed. In contrast, the frogs from the Indonesian supplier likely grew up under natural conditions, indicated by higher δ15N values and stronger variability in the stable isotope composition. Our results indicate that stable isotope analyses seem to be a useful tool to distinguish between naturally growing and intensively farmed frogs. We believe that this method can be used to improve the control in the international trade of frog legs, as well as for other biological products, thus supporting farming activities and decreasing pressure on wild populations. However, we examined different species from different countries and had no access to samples of individuals with confirmed origin and living conditions. Therefore, we suggest improving this method further with individuals of known origin and history, preferably including samples of the respective nutritive bases.

Biological diversity is depleting at an alarming rate. Additionally, a vast amount of biodiversity still remains undiscovered. Taxonomy has been serving the purpose of describing, naming, and classifying species for more than 250 years. DNA taxonomy and barcoding have accelerated the rate of this process, thereby providing a tool for conservation practice. DNA barcoding and traditional taxonomy have their own inherent merits and demerits. The synergistic use of both methods, in the form of integrative taxonomy, has the potential to contribute to biodiversity conservation in a pragmatic timeframe and overcome their individual drawbacks. In this review, we discuss the basics of both these methods of biological identification- traditional taxonomy and DNA barcoding, the technical advances in integrative taxonomy, and future trends. We also present a comprehensive compilation of published examples of integrative taxonomy that refer to nine topics within biodiversity conservation. Morphological and molecular species limits were observed to be congruent in ~41% of the 58 source studies. The majority of the studies highlighted the description of cryptic diversity through the use of molecular data, whereas research areas like endemism, biological invasion, and threatened species were less discussed in the literature.

Effective vector and arbovirus surveillance requires timely and accurate screening techniques that can be easily upscaled. Next-generation sequencing (NGS) is a high-throughput technology that has the potential to modernise vector surveillance. When combined with DNA barcoding, it is termed 'metabarcoding'. The aim of our study was to establish a metabarcoding protocol to characterise pools of mosquitoes and screen them for virus. Pools contained 100 morphologically identified individuals, including one Ross River virus (RRV) infected mosquito, with three species present at different proportions: 1, 5, 94%. Nucleic acid extracted from both crude homogenate and supernatant was used to amplify a 269 bp section of the mitochondrial cytochrome c oxidase subunit I (COI) locus. Additionally, a 67 bp region of the RRV E2 gene was amplified from synthesised cDNA to screen for RRV. Amplicon sequencing was performed using an Illumina MiSeq, and bioinformatic analysis was performed using a DNA barcode database of Victorian mosquitoes. Metabarcoding successfully detected all mosquito species and RRV in every positive sample tested. The limits of species detection were also examined by screening a pool of 1000 individuals, successfully identifying the species and RRV from a single mosquito. The primers used for amplification, number of PCR cycles, and total number of individuals present all have effects on the quantification of species in mixed bulk samples. Based on the results, a number of recommendations for future metabarcoding studies are presented. Overall, metabarcoding shows great promise for providing a new alternative approach to screening large insect surveillance trap catches. 

The potential of DNA barcoding approaches to identify single species and characterize species compositions strongly depends on the marker choice. The prominent "Folmer region", a 648 basepair fragment at the 5' end of the mitochondrial CO1 gene, has been traditionally applied as a universal DNA barcoding region for metazoans. In order to find a suitable marker for biomonitoring odonates (dragonflies and damselflies), we here explore a new region of the CO1 gene (CO1B) for DNA barcoding in 51 populations of 23 dragonfly and damselfly species. We compare the "Folmer region", the mitochondrial ND1 gene (NADH dehydrogenase 1) and the new CO1 region with regard to (i) speed and reproducibility of sequence generation, (ii) levels of homoplasy and (iii) numbers of diagnostic characters for discriminating closely related sister taxa and populations. The performances of the gene regions regarding these criteria were quite different. Both, the amplification of CO1B and ND1 was highly reproducible and CO1B showed the highest potential for discriminating sister taxa at different taxonomic levels. In contrast, the amplification of the "Folmer region" using the universal primers was difficult and the third codon positions of this fragment have experienced nucleotide substitution saturation. Most important, exploring this new barcode region of the CO1 gene identified a higher discriminating power between closely related sister taxa. Together with the design of layered barcode approaches adapted to the specific taxonomic "environment", this new marker will further enhance the discrimination power at the species level.

Monday, April 24, 2017

From the inbox: Postdoc @ Kartzinel Lab at Brown

Position Title: Postdoctoral Research Associate in Environment and Society: Molecular Ecology/Conservation Biology

Location: Department of Ecology and Evolutionary Biology; Institute at Brown for Environment and Society, Brown University, Providence, Rhode Island, USA

Position Description: The Kartzinel Lab at Brown University is seeking a postdoctoral research associate in molecular ecology and conservation biology to collaborate on studies of the ecology and evolution of animal diets, especially in African savannas (http://www.kartzinellab.com/). The postdoctoral project will emphasize conceptual and analytical advances in our understanding of plant-herbivore, predator-prey, and/or host-microbe interaction networks using DNA- based analyses. These results will be placed in the context of manipulative field experiments, broad geographic gradients, and different land use histories in order to illuminate the biological processes that determine population and community dynamics in a changing world. The postdoc will have ample opportunities to integrate both field and lab-based research on wildlife in Kenya, and other potentially relevant field sites. The Department of Ecology and Evolutionary Biology and Institute at Brown for Environment and Society are home to a diverse community of scholars and world-class scientific resources.

Qualifications: The successful candidate will have a recent Ph.D. and relevant experience in molecular ecology, such as environmental DNA analysis, microbiome analyses, DNA (meta) barcoding, metagenomics, or related approaches. A strong interest in community ecology and conservation is required, and prior field experience is a plus. Responsibilities include: helping develop and coordinate laboratory and field research activities; data management; analysis of ecological and molecular data; publication of manuscripts and dissemination of results. Candidates should demonstrate strong communication skills and an ability to work both independently and collaboratively with groups from diverse backgrounds.

Applicants should submit: (1) a cover letter describing research interests, qualifications, and motivations, (2) a CV, and (3) contact information for three references. Applications will be reviewed starting May 12, 2017, and accepted until the position is filled. The ideal start date is September 2017, but flexible. The initial appointment will be for one year with an opportunity for extension based on satisfactory performance. Please contact Tyler Kartzinel directly with any questions.

Equal Employment Opportunity Statement: Brown University is committed to fostering a diverse and inclusive academic global community; as an EEO/AA employer, Brown considers applicants for employment without regard to, and does not discriminate on the basis of, gender, race, protected veteran status, disability, or any other legally protected status.

Tuesday, April 18, 2017

Metabarcoding and Metagenomics journal

Not too long ago I posted a call for participation at a questionnaire which solicited feedback and suggestions for a new open access journal focusing on metabarcoding and metagenomics. Today, just a few months later the journal out in the open and ready for submission:

Metabarcoding and Metagenomics (MBMG) is an innovative open access journal which facilitates the publication of articles on metabarcoding and metagenomics both in basic and applied context. The journal welcomes submissions representing all stages of the research cycle: data, models, methods, workflows, software, perspectives, opinions, and conventional research articles. The journal will consider manuscripts for publication related (but not limited) to the following topics: Environmental MBMG, Microbial MBMG, Applied MBMG (biomonitoring, quarantine, environmental assessment, nature conservation, eDNA, species invasions and others), and other emerging fields related to MBMG. Submissions of bioinformatic approaches to MBMG (algorithms, software) are also encouraged.

So, if you are currently working on a metabarcoding or metagenomics study and looking for a home for your publication you might want to consider this new option.

Thursday, April 13, 2017


7th International Barcode of Life Conference, 20 – 24 November 2017 at Kruger National Park


The African Centre for DNA Barcoding (ACDB), the University of  Johannesburg (UJ), International Barcode of Life Project (iBOL), and the Department of Environmental Affairs (DEA) are proud to announce and welcome delegates to our hosting of the 7th International Barcode of Life Conference, 20 – 24 November 2017. This is the first time that this event will be held on the African continent. The venue for the hosting of this prestigious event will be the Nombolo Mdhluli Conference Centre, 

The major theme of the conference is exploring mega-diverse biotas with DNA barcodes. A series of presentations and workshops will focus on the use of DNA to understand diversity patterns and ecological processes in species-rich and complicated ecosystems. The conference also provides a 
general forum for presentations, posters, and discussion on the wider field of DNA barcoding.

Delegates are encouraged to register as soon as possible as space is limited.

Abstracts should be submitted here before or on 28th April 2017.

More information at:

Website: http://dnabarcodes2017.org
Facebook: @DNABarcodes2017
Twitter: @DNABarcodes

Wednesday, April 12, 2017

Wednesday reads

Two days delay and a lot to go through but I think I came up with a list of quite interesting reads:

Phlebotomine sand flies are haematophagous dipterans of primary medical importance. They represent the only proven vectors of leishmaniasis worldwide and are involved in the transmission of various other pathogens. Studying the ecology of sand flies is crucial to understand the epidemiology of leishmaniasis and further control this disease. A major limitation in this regard is that traditional morphological-based methods for sand fly species identifications are time-consuming and require taxonomic expertise. DNA metabarcoding holds great promise in overcoming this issue by allowing the identification of multiple species from a single bulk sample. Here, we assessed the reliability of a short insect metabarcode located in the mitochondrial 16S rRNA for the identification of Neotropical sand flies, and constructed a reference database for 40 species found in French Guiana. Then, we conducted a metabarcoding experiment on sand flies mixtures of known content and showed that the method allows an accurate identification of specimens in pools. Finally, we applied metabarcoding to field samples caught in a 1-ha forest plot in French Guiana. Besides providing reliable molecular data for species-level assignations of phlebotomine sand flies, our study proves the efficiency of metabarcoding based on the mitochondrial 16S rRNA for studying sand fly diversity from bulk samples. The application of this high-throughput identification procedure to field samples can provide great opportunities for vector monitoring and eco-epidemiological studies.

Translating the vast amounts of molecular barcodes from global surveys of microbial eukaryotes into ecological insight depends critically on a well-curated reference database with adequate taxonomic coverage. In this respect, the choanoflagellates resemble other eukaryotic lineages: reasonable coverage at higher taxonomic levels, but missing diversity at the species level. The acanthoecid (loricate) choanoflagellates are well-characterized morphologically, with over 115 species described, but less than 10% with any sequence data. Because lorica shape is species-specific, the acanthoecids represent an opportunity to link morphological with molecular data within a lineage of eukaryotes. To match morphospecies to sequences, we sampled the Kattegat and the Isefjord in Denmark in September 2014 and February 2015. We identified 45 morphospecies and sequenced ribosomal DNA of nine previously unsequenced species, roughly doubling the number of acanthoecid species with sequence data, including the first data representing five genera: Bicosta, Calliacantha, Cosmoeca, Crinolina and Pleurasiga. Our phylogenetic analysis is mainly congruent with morphology-based systematics. Five of the newly sequenced species match a previously unidentified barcode from Tara Oceans, providing access to the global distribution of species isolated from Danish waters. One species, Calliacantha natans, is the second most globally abundant choanoflagellate present in Tara Oceans. Our project translating new ribosomal DNA sequences to distributions of described species on a global scale supports the approach linking morphology to molecular barcodes for microbial eukaryote ecology.

Almost all plants in nature harbour fungi in their roots but the knowledge on distribution and the underlying principles of assemblage is still poorly developed for the root-associated fungi. In this study we analysed the root endophytic fungal communities associated with switchgrass, rosette grass, and pitch pine in the acidic, oligotrophic pine barrens ecosystem. A total of 434 fungal isolates were obtained from 600 root segments of 60 plant samples. DNA barcoding and morphological analyses identified 92 fungal species, which belong to 39 genera in six classes. Compared to other ecosystems, the pine barrens has a higher proportion of Leotiomycetes. The fungal community associated with pitch pine was significantly different from those associated with the grasses, while less difference was found between those associated with the two grasses. Our results suggest that edaphic factors and host specificity play a role in shaping root endophytic fungal community. This study also corroborates our previous finding that plant roots in the pine barrens are a rich reservoir of novel fungi.

The introduction of exotic species can have serious consequences for marine ecosystems. On the shores of the Cantabrian Sea (North of Spain) there are no routine examinations of seaweeds that combine molecular and morphological methods for early detection of exotic species making it difficult to assess in the early stages their establishment and expansion processes as a result of anthropogenic activities (e.g., shipping and/or aquaculture).
In this work we used both morphological identification and molecular barcoding (COI-5P and rbcL genes) of red algae collected in Asturias, Bay of Biscay (Gijón and Candás harbours) and from the University of Oviedo's herbarium samples.
The results confirmed the presence of exotic Asian seaweeds Pachymeniopsis gargiuli and Grateloupia turuturu Yamada on Cantabrian Sea shores. Several individuals of these species were fertile and developing cystocarps when collected, underlining the risk of possible expansion or continued establishment. This study constitutes the first report of the Asian P. gargiuli in this area of the Bay of Biscay.
Here the presence of the exotic species of the Halymeniales P. gargiuli is confirmed. We hypothesize that this species may have been established some time ago as a cryptic introduction with G. turuturu in Galician shores. The detection of these species on the shores of the Cantabrian Sea is relevant since introductions of Pachymeniopsis species could have been overlooked on other European coasts, probably mixed with G. turuturu and P. lanceolata. Our results confirm one new alien seaweed species that has been detected using molecular methods (COI-5P region and rbcL genes barcoding) on North Atlantic shores: the Asian native P. gargiuli. This demonstrates that routine screening for early detection of exotic algae in the Cantabrian Sea can be used for risk assessment. Genetic barcoding should be done using both rbcL gene and COI-5P regions since, although COI-databases are still poorer in sequences and this inhibits successful outcomes in Grateloupia-related species identifications, it is nonetheless a useful marker for species-level identifications in seaweeds.

The difficulty of censusing marine animal populations hampers effective ocean management. Analyzing water for DNA traces shed by organisms may aid assessment. Here we tested aquatic environmental DNA (eDNA) as an indicator of fish presence in the lower Hudson River estuary. A checklist of local marine fish and their relative abundance was prepared by compiling 12 traditional surveys conducted between 1988–2015. To improve eDNA identification success, 31 specimens representing 18 marine fish species were sequenced for two mitochondrial gene regions, boosting coverage of the 12S eDNA target sequence to 80% of local taxa. We collected 76 one-liter shoreline surface water samples at two contrasting estuary locations over six months beginning in January 2016. eDNA was amplified with vertebrate-specific 12S primers. Bioinformatic analysis of amplified DNA, using a reference library of GenBank and our newly generated 12S sequences, detected most (81%) locally abundant or common species and relatively few (23%) uncommon taxa, and corresponded to seasonal presence and habitat preference as determined by traditional surveys. Approximately 2% of fish reads were commonly consumed species that are rare or absent in local waters, consistent with wastewater input. Freshwater species were rarely detected despite Hudson River inflow. These results support further exploration and suggest eDNA will facilitate fine-scale geographic and temporal mapping of marine fish populations at relatively low cost.

Monday, April 10, 2017

Student Travel Bursaries for African and South African Students

Happy to pass on some important conference related information:

Dear Delegates, 

We are pleased to announce that there are Travel Bursaries available for African and South African students to attend the upcoming 7th International Barcode of Life Conference in November 2017.

To apply and for more information, please go to to this link.

We encourage all African students to apply - please note the application deadline is 01 May 2017.

Good luck!

With best wishes, 

7th iBOL Local Organizing Committee

Monday, April 3, 2017

Monday reads

Happy April! Another week of light posting passed and I am afraid there will be one more of the same. A number of things got in the way despite the fact that there is no shortage of things to blog about. In short - I am too busy. Here is hope that this will change in the near future. And now to this weeks selection:

DNA-barcoding is a rapidly developing method for efficiently identifying samples to species level by means of short standard DNA sequences. However, reliable species assignment requires the availability of a comprehensive DNA barcode reference library, and hence numerous initiatives aim at generating such barcode databases for particular taxa or geographic regions. Historical museum collections represent a potentially invaluable source for the DNA-barcoding of many taxa. This is particularly true for birds and mammals, for which collecting fresh (voucher) material is often very difficult to (nearly) impossible due to the special animal welfare and conservation regulations that apply to vertebrates in general, and birds and mammals in particular. Moreover, even great efforts might not guarantee sufficiently complete sampling of fresh material in a short period of time. DNA extracted from historical samples is usually degraded, such that only short fragments can be amplified, rendering the recovery of the barcoding region as a single fragment impossible. Here, we present a new set of primers that allows the efficient amplification and sequencing of the entire barcoding region in most higher taxa of Central European birds and mammals in six overlapping fragments, thus greatly increasing the value of historical museum collections for generating DNA barcode reference libraries. Applying our new primer set in recently established NGS protocols promises to further increase the efficiency of barcoding old bird and mammal specimens.

We foresee a new global-scale, ecological approach to biomonitoring emerging within the next decade that can detect ecosystem change accurately, cheaply, and generically. Next-generation sequencing of DNA sampled from the Earth’s environments would provide data for the relative abundance of operational taxonomic units or ecological functions. Machine-learning methods would then be used to reconstruct the ecological networks of interactions implicit in the raw NGS data. Ultimately, we envision the development of autonomous samplers that would sample nucleic acids and upload NGS sequence data to the cloud for network reconstruction. Large numbers of these samplers, in a global array, would allow sensitive automated biomonitoring of the Earth’s major ecosystems at high spatial and temporal resolution, revolutionising our understanding of ecosystem change.

Anthropogenic activities are having devastating impacts on marine systems with numerous knock-on effects on trophic functioning, species interactions and an accelerated loss of biodiversity. Establishing conservation areas can not only protect biodiversity, but also confer resilience against changes to coral reefs and their inhabitants. Planning for protection and conservation in marine systems is complex, but usually focuses on maintaining levels of biodiversity and protecting special and unique landscape features while avoiding negative impacts to socio-economic benefits. Conversely, the integration of evolutionary processes that have shaped extant species assemblages is rarely taken into account. However, it is as important to protect processes as it is to protect patterns for maintaining the evolutionary trajectories of populations and species. This review focuses on different approaches for integrating genetic analyses, such as phylogenetic diversity, phylogeography and the delineation of management units, temporal and spatial monitoring of genetic diversity and quantification of adaptive variation for protecting evolutionary resilience, into marine spatial planning, specifically for coral reef fishes. Many of these concepts are not yet readily applied to coral reef fish studies, but this synthesis highlights their potential and the importance of including historical processes into systematic biodiversity planning for conserving not only extant, but also future, biodiversity and its evolutionary potential.

Determining the ecosystem function of high-order predators is critical for evaluation of food web interactions. Insectivorous birds are abundant predators in many ecosystems yet because they forage upon small taxa, it remains largely unknown whether birds are providing ecosystem services in the form of pest control or disservices by preying upon predaceous arthropod species. We extracted DNA from noninvasive fecal samples of adult and nestling Western Bluebirds (Sialia mexicana) in California vineyards. Using universal arthropod-specific primers, we sequenced prey items via massively parallel sequencing on the Illumina MiSeq platform. Bluebirds consumed a broad diet comprising 66 unique arthropod species from 6 orders and 28 families. Aedes sp. (mosquitoes: Culicidae), a previously unknown prey, was the most common item recovered, occurring in 49.5% of the fecal samples. Ectoparasitic bird blowfly (Protocalliphora) DNA was found in 7% of adult and 11% of nestling samples, presenting clear evidence of active feeding by the avian hosts on adult or larval ectoparasites. Herbivorous insects, primarily from the orders Hemiptera and Lepidoptera, represented over half (56%) of the prey items in bluebird diets. Intraguild predation (consumption of predator or parasitoid arthropods) represented only 3% of adult and nestling dietary items. Diets of adults were significantly different from nestlings as were diets from birds sampled in different vineyard blocks. Sex, date, number of young, and individual bird (based on resampled individuals) were all insignificant factors that did not explain diet variability. Nestling age was a significant factor in explaining a small amount of the variability in dietary components. In addition, our analysis of subsampling larger fecal samples and processing them independently revealed highly dissimilar results in all 10 trials and we recommend avoiding this common methodology. Molecular scatology offers powerfully informative techniques that can reveal the ecosystem function and services provided by abundant yet cryptic avian foragers.

Monday, March 27, 2017

Monday reads

New reads, hot of the press. Very diverse spread of application, news and research.

The African Centre for DNA Barcoding (ACDB) was established in 2005 as part of a global initiative to accurately and rapidly survey biodiversity using short DNA sequences. The mitochondrial cytochrome c oxidase 1 gene (CO1) was rapidly adopted as the de facto barcode for animals. Following the evaluation of several candidate loci for plants, the Plant Working Group of the Consortium for the Barcoding of Life in 2009 recommended that the two plastid genes, rbcLa and matK, be adopted as core DNA barcodes for terrestrial plants. To date, numerous studies continue to test the discriminatory power of these markers across various plant lineages. Over the past decade, we at the African Centre for DNA Barcoding, have used these core DNA barcodes to generate a barcode library for southern Africa. To date, the ACDB has contributed more than 21 000 plant barcodes and over 3 000 CO1 barcodes for animals to the Barcode of Life Database (BOLD). Building upon this effort, we at the ACDB have addressed questions related to community assembly, biogeography, phylogenetic diversification, and invasion biology. Collectively, our work demonstrates the diverse applications of DNA barcoding in ecology, systematics, evolutionary biology, and conservation.

Sequences from the DNA barcode region of the mitochondrial COI gene are an effective tool for specimen identification and for the discovery of new species. The Barcode of Life Data Systems (BOLD) (www.boldsystems.org) currently hosts 4.5 million records from animals which have been assigned to more than 490,000 different Barcode Index Numbers (BINs), which serve as a proxy for species. Because a fourth of these BINs derive from Lepidoptera, BOLD has a strong capability to both identify specimens in this order and to support studies of faunal overlap. DNA barcode sequences were obtained from 4503 moths from 329 sites across Pakistan, specimens that represented 981 BINs from 52 families. Among 379 species with a Linnaean name assignment, all were represented by a single BIN excepting five species that showed a BIN split. Less than half (44%) of the 981 BINs had counterparts in other countries; the remaining BINs were unique to Pakistan. Another 218 BINs of Lepidoptera from Pakistan were coupled with the 981 from this study before being compared with all 116,768 BINs for this order. As expected, faunal overlap was highest with India (21%), Sri Lanka (21%), United Arab Emirates (20%) and with other Asian nations (2.1%), but it was very low with other continents including Africa (0.6%), Europe (1.3%), Australia (0.6%), Oceania (1.0%), North America (0.1%), and South America (0.1%). This study indicates the way in which DNA barcoding facilitates measures of faunal overlap even when taxa have not been assigned to a Linnean species.

Polystomes are monogenean parasites that infest mainly semi aquatic vertebrates, such as amphibians and chelonians. Owing to the lack of discriminative morphological characters and because polystomes are considered to be strictly host- and site-specific, host identity is often used as an additional character for parasite identification. Recent genetic studies, however, show that polystomes infecting freshwater turtles in outdoor turtle enclosures and natural environments, are not strictly host-specific. Therefore, we proposed a new procedure for turtle polystome taxonomy based on the combination of Cytochrome c Oxydase I sequences and two discriminant morphological characters, namely the number of genital spines and the testis shape. We tested the validity of this procedure with Polystomoides oris, which was collected from the pharyngeal cavity of the American painted turtle Chrysemys picta and two undescribed species, both collected from the pharyngeal cavity of the American slider Trachemys scripta and two other European turtles, namely the European pond turtle Emys orbicularis and the Mediterranean turtle Mauremys leprosa. A Principal Component Analysis based on both morphological characters allowed the separation of all specimens in three morphological groups, which matched well with the molecular data. As a result, we describe two new polystome species, i.e., Polystomoides soredensis n. sp. and Polystomoides scriptanus n. sp.

Fungus gnats (Sciaroidea) are a globally species rich group of lower Diptera. In Europe, Fennoscandian peninsula in particular holds a notable diversity, ca. 1000 species, of which 10 % are still unnamed. Fungus gnats are predominantly terrestrial insects, but some species dwell in wetland habitats.
Eight new fungus gnat species, belonging to the families Keroplatidae (Orfelia boreoalpina Salmela sp.n.) and Mycetophilidae (Sciophila holopaineni Salmela sp.n., S. curvata Salmela sp.n., Boletina sasakawai Salmela & Kolcsár sp.n., B. norokorpii Salmela & Kolcsár sp.n., Phronia sompio Salmela sp.n., P. reducta Salmela sp.n., P. prolongata Salmela sp.n.), are described. Four of the species are known from Fennoscandia only whilst two are supposed to have boreo-alpine disjunct ranges, i.e. having populations in Fennoscandia and the Central European Alps. One of the species probably has a boreal range (Finnish Lapland and Central Siberia). Type material of Boletina curta Sasakawa & Kimura from Japan was found to consist of two species, and a further species close to these taxa is described from Finland. Phronia elegantula Hackman is redescribed and reported for the first time from Norway. DNA barcodes are provided for the first time for five species.

Several methods of DNA extraction, coupled with 'DNA barcoding' species identification, were compared using specimens from early developmental stages of forensically important flies from the Calliphoridae and Sarcophagidae families. DNA was extracted at three immature stages - eggs, the first instar larvae, and empty pupal cases (puparia) - using four different extraction methods, namely, one simple 'homemade' extraction buffer protocol and three commercial kits. The extraction conditions, including the amount of proteinase K and incubation times, were optimized. The simple extraction buffer method was successful for half of the eggs and for the first instar larval samples. The DNA Lego Kit and DEP-25 DNA Extraction Kit were useful for DNA extractions from the first instar larvae samples, and the DNA Lego Kit was also successful regarding the extraction from eggs. The QIAamp DNA mini kit was the most effective; the extraction was successful with regard to all sample types - eggs, larvae, and pupari.

Mitochondrial introns intermit coding regions of genes and feature characteristic secondary structures and splicing mechanisms. In metazoans, mitochondrial introns have only been detected in sponges, cnidarians, placozoans and one annelid species. Within demosponges, group I and group II introns are present in six families. Based on different insertion sites within the cox1 gene and secondary structures, four types of group I and two types of group II introns are known, which can harbor up to three encoding homing endonuclease genes (HEG) of the LAGLIDADG family (group I) and/or reverse transcriptase (group II). However, only little is known about sponge intron mobility, transmission, and origin due to the lack of a comprehensive dataset. We analyzed the largest dataset on sponge mitochondrial group I introns to date: 95 specimens, from 11 different sponge genera which provided novel insights into the evolution of group I introns.
For the first time group I introns were detected in four genera of the sponge family Scleritodermidae (Scleritoderma, Microscleroderma, Aciculites, Setidium). We demonstrated that group I introns in sponges aggregate in the most conserved regions of cox1. We showed that co-occurrence of two introns in cox1 is unique among metazoans, but not uncommon in sponges. However, this combination always associates an active intron with a degenerating one. Earlier hypotheses of HGT were confirmed and for the first time VGT and secondary losses of introns conclusively demonstrated.
This study validates the subclass Spirophorina (Tetractinellida) as an intron hotspot in sponges. Our analyses confirm that most sponge group I introns probably originated from fungi. DNA barcoding is discussed and the application of alternative primers suggested.

Tuesday, March 21, 2017

Reminder: Abstract deadline ends in 10 days

The 7th International Barcode of  Life Conference will be held from November 20 - 24, 2017 at the Nombolo Mdhluli Conference Centre, Skukuza, located within the heart of African wildlife at Kruger National Park, South Africa. 

On March 31st the abstract submission will end and because I know that we researchers like to cut it close I like to remind you to send in your's quickly. How about setting aside a couple of minutes this week and write it down? Here you go.

Monday, March 20, 2017

Monday reads

Now back to Monday reads on a Monday. Some new studies for you to catch up.

Premise of the study: DNA metabarcoding has broad-ranging applications in ecology, aerobiology, biosecurity, and forensics. A bioinformatics pipeline has recently been published for identification using a comprehensive database of ITS2, one of the common plant DNA barcoding markers. There is, however, no corresponding database for rbcL, the other primary marker used in plants.
Methods: Using publicly available data, we compiled a reference library of rbcL sequences and trained databases for use with UTAX and RDP classifier algorithms. We used this reference library, along with the existing bioinformatics pipeline and ITS2 reference library, to identify species in an artificial mixture of nine species of pollen. We have made this database publicly available in multiple formats, to allow use with multiple bioinformatics pipelines, now and in the future.
Results: Using the rbcL database, in addition to the ITS2 database, we succeeded in making species-level identifications for eight species and a family-level identification of the ninth species. This is an improvement on ITS2 sequence alone.
Discussion: The reference library described here will assist with identification of plant species using rbcL. By making another gene region available for standard barcoding, this will increase the resolution and accuracy of identifications.

Leucopis argenticollis (Zetterstedt) and Leucopis piniperda (Malloch) are known to feed on the lineage of Adelges tsugae Annand that is native to western North America, but it is not known if they will survive on the lineage that was introduced from Japan to the eastern USA. In 2014, western Leucopis spp. larvae were brought to the laboratory and placed on A. tsugae collected in either Washington (North American A. tsugae lineage) or Connecticut (Japanese lineage). There were no significant differences in survival or developmental times between flies reared on the two different adelgid lineages. In 2015 and 2016, western Leucopis spp. adults were released at two different densities onto enclosed branches of A. tsugae infested eastern hemlock (Tsuga canadensis (L.) Carr.) in Tennessee and New York. Cages were recovered and their contents examined 4 weeks after release at each location. Leucopis spp. larvae and puparia of the F1 generation were recovered at both release locations and adults of the F1 generation were collected at the Tennessee location. The number of Leucopis spp. offspring collected increased with increasing adelgid density, but did not differ by the number of adult flies released. Flies recovered from cages and flies collected from the source colony were identified as L.argenticollis and L. piniperda using DNA barcoding. These results demonstrate that Leucopis spp. from the Pacific Northwest are capable of feeding and developing to the adult stage on A. tsugae in the eastern USA and they are able to tolerate environmental conditions during late spring and early summer at the southern and northern extent of the area invaded by A. tsugae in the eastern USA.

Despite the high value of decapod crustaceans, relatively little research has focused on assessing the transparency in the marketing of these species. This study represents the first comprehensive evaluation of the quality of labelling, and the extent of mislabelling, of decapod crustacean products on the South African market. Data collected through surveys of supermarkets and seafood shops in three provinces (KwaZulu-Natal [KZN], Western Cape [WC] and Gauteng [GP]), indicated that the large majority of domestically available crustacean products were imported, but that 18% of these failed to comply with locally applicable country of origin labelling regulations. Voluntary information relating to the scientific name, production method (wild caught or farmed), and capture method of the species was supplied more frequently in supermarkets than in seafood shops, more frequently in the WC and GP than in KZN, and more frequently on shrimp products than on crab and lobster products. DNA sequencing of 77 products collected from the surveyed outlets revealed that 24 (31%) were misrepresented in some way. Species misrepresentations were most pronounced for shrimps, with Litopenaeus vannamei and Pleoticus muelleri being confirmed as the most common substitute species. One shrimp product was found to contain at least three different species, none of which matched the declared species, whereas a product labelled as crab turned out to be a member of the phylum Mollusca rather than the subphylum Crustacea. Overall, these findings demonstrate that the misrepresentation of crustaceans is commonplace on the South African market, signalling the need for a revision of the current seafood labelling and traceability legislation, as well as monitoring and enforcement efforts.

The aim of this study was to compare the performance of a DNA-barcode assay with fatty acid profile analysis to authenticate the botanical origin of olive oil. To achieve this aim, we performed a PCR-capillary electrophoresis (PCR-CE) approach on olive oil: seed oil blends using the plastid trnL (UAA) intron barcode. In parallel to genomic analysis, we subjected the samples to gas chromatography analysis of fatty acid composition. While the PCR-CE assay proved equally efficient as gas chromatography analysis in detecting adulteration with soybean, palm, rapeseed, sunflower, sesame, cottonseed and peanut oils, it was superior to the widely utilized analytical chemistry approach in revealing the adulterant species and detecting small quantities of corn and safflower oils in olive oil. Moreover, the DNA-based test correctly identified all tested olive oil: hazelnut oil blends whereas it was not feasible to detect hazelnut oil adulteration through fatty acid profile analysis. Thus, the present research has shown the feasibility of a PCR-CE barcode assay to detect adulteration in olive oil.

Determining the ecosystem function of high-order predators is critical for evaluation of food web interactions. Insectivorous birds are abundant predators in many ecosystems yet because they forage upon small taxa, it remains largely unknown whether birds are providing ecosystem services in the form of pest control or disservices by preying upon predaceous arthropod species. We extracted DNA from noninvasive fecal samples of adult and nestling Western Bluebirds (Sialia mexicana) in California vineyards. Using universal arthropod-specific primers, we sequenced prey items via massively parallel sequencing on the Illumina MiSeq platform. Bluebirds consumed a broad diet comprising 66 unique arthropod species from 6 orders and 28 families. Aedes sp. (mosquitoes: Culicidae), a previously unknown prey, was the most common item recovered, occurring in 49.5% of the fecal samples. Ectoparasitic bird blowfly (Protocalliphora) DNA was found in 7% of adult and 11% of nestling samples, presenting clear evidence of active feeding by the avian hosts on adult or larval ectoparasites. Herbivorous insects, primarily from the orders Hemiptera and Lepidoptera, represented over half (56%) of the prey items in bluebird diets. Intraguild predation (consumption of predator or parasitoid arthropods) represented only 3% of adult and nestling dietary items. Diets of adults were significantly different from nestlings as were diets from birds sampled in different vineyard blocks. Sex, date, number of young, and individual bird (based on resampled individuals) were all insignificant factors that did not explain diet variability. Nestling age was a significant factor in explaining a small amount of the variability in dietary components. In addition, our analysis of subsampling larger fecal samples and processing them independently revealed highly dissimilar results in all 10 trials and we recommend avoiding this common methodology. Molecular scatology offers powerfully informative techniques that can reveal the ecosystem function and services provided by abundant yet cryptic avian foragers.

A wide variety of DNA based methods have been developed to identify fish species, including those that employ a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. One such method developed by Dooley, Sage, Clarke, Brown, and Garrett (2005) used amplification of a portion of the mitochondrial cytochrome B (cytB) gene, with a three enzyme digestion, visualized and identified using the Agilent 2100 Bioanalyzer System. More recently this method was modified by Agilent Technologies to target a section of the cytochrome c oxidase 1 (CO1) gene, within the 655 base pair (bp) "barcoding" fragment, using a two enzyme digestion to increase sample throughput and to exploit publically available CO1 data generated through the Barcode of Life initiative (Mueller et al. 2015). Here we evaluate this method on fifteen different commercial fish species with five replicate specimens of each. DNA barcoding of the CO1 gene was used as an orthogonal confirmatory method and also to further understand the results found using the modified Agilent PCR-RFLP method.

Wednesday, March 15, 2017

Monday reads

Monday reads on a Wednesday - just the result of some conference travel. For the same reason there are a few more articles than usual. Interesting reads as always.

Land-use intensification threatens freshwater biodiversity. Freshwater eukaryotic communities are affected by multiple chemical contaminants with a land-use specific manner. However, biodiversities of eukaryotes and their associations with multiple chemical contaminants are largely unknown. This study characterized in situ eukaryotic communities in sediments exposed to mixtures of chemical contaminants and assessed relationships between various environmental variables and eukaryotic communities in sediments from the Nanfei River. Eukaryotic communities in the sediment samples were dominated by Annelida, Arthropoda, Rotifera, Ochrophyta, Chlorophyta and Ciliophora. Alpha-diversities (Shannon entropy) and structures of eukaryotic communities were significantly different between land-use types. According to the results of multiple statistical tests (PCoA, distLM, Mantel and network analysis), dissimilarity of eukaryotic community structures revealed the key effects of pyrethroid insecticides, manganese, zinc, lead, chromium and polycyclic aromatic hydrocarbons (PAHs) on eukaryotic communities in the sediment samples from the Nanfei River. Furthermore, taxa associated with land-use types were identified and several sensitive eukaryotic taxa to some of the primary contaminants were identified as potential indicators to monitor effects of the primary chemical contaminants. Overall, environmental DNA metabarcoding on in situ eukaryotic communities provided a powerful tool for biomonitoring and identifying primary contaminants and their complex effects on benthic eukaryotic communities in freshwater sediments.

no abstract

DNA barcoding methods use a single locus (usually the mitochondrial COI gene) to assign unidentified specimens to known species in a library based on a genetic distance threshold that distinguishes between-species divergence from within-species diversity. Recently developed species delimitation methods based on the multispecies coalescent (MSC) model offer an alternative approach to individual assignment using either single-locus or multi-loci sequence data. Here we use simulations to demonstrate three features of an MSC method implemented in the program bpp. First, we show that with one locus, MSC can accurately assign individuals to species without the need for arbitrarily determined distance thresholds (as required for barcoding methods). We provide an example in which no single threshold or barcoding gap exists that can be used to assign all specimens without incurring high error rates. Second, we show that bpp can identify cryptic species that may be mis-identified as a single species within the library, potentially improving the accuracy of barcoding libraries. Third, we show that taxon rarity does not present any particular problems for species assignments using bpp, and that accurate assignments can be achieved even when only one or a few loci are available. Thus, concerns that have been raised that MSC methods may have problems analyzing rare taxa (singletons) are unfounded. Currently barcoding methods enjoy a huge computational advantage over MSC methods and may be the only approach feasible for massively large datasets, but MSC methods may offer a more stringent test for species that are tentatively assigned by barcoding.

Partitioning tissue metal concentration into subcellular compartments reflecting toxicologically available pools may provide good descriptors of the toxicological effects of metals on organisms. Here we investigated the relationships between internal compartmentalization of Cd, Pb and Zn and biomarker responses in a model soil organism: the earthworm. The aim of this study was to identify metal fractions reflecting the toxic pressure in an endogeic, naturally occurring earthworm species (Aporrectodea caliginosa) exposed to realistic field-contaminated soils. After a 21 days exposure experiment to 31 field-contaminated soils, Cd, Pb and Zn concentrations in earthworms and in three subcellular fractions (cytosol, debris and granules) were quantified. Different biomarkers were measured: the expression of a metallothionein gene (mt), the activity of catalase (CAT) and of glutathione-s-transferase (GST), and the protein, lipid and glycogen reserves. Biomarkers were further combined into an integrated biomarker index (IBR). The subcellular fractionation provided better predictors of biomarkers than the total internal contents hence supporting its use when assessing toxicological bioavailability of metals to earthworms. The most soluble internal pools of metals were not always the best predictors of biomarker responses. metallothionein expression responded to increasing concentrations of Cd in the insoluble fraction (debris + granules). Protein and glycogen contents were also mainly related to Cd and Pb in the insoluble fraction. On the other hand, GST activity was better explained by Pb in the cytosolic fraction. CAT activity and lipid contents variations were not related to metal subcellular distribution. The IBR was best explained by both soluble and insoluble fractions of Pb and Cd. This study further extends the scope of mt expression as a robust and specific biomarker in an ecologically representative earthworm species exposed to field-contaminated soils. The genetic lineage of the individuals, assessed by DNA barcoding with cytochrome c oxidase subunit I, did not influence mt expression.

DNA barcoding is a commonly used bio-technology in multiple disciplines including biology, environmental science, forensics and inspection, etc. Forest dynamic plots provide a unique opportunity to carry out large-scale, comparative, and multidisciplinary research for plant DNA barcoding. The paper concisely reviewed four previous progresses in China; specifically, species discrimination, community phylogenetic reconstruction, phylogenetic community structure exploration, and biodiversity index evaluation. Further, we demonstrated three major challenges; specifically, building the impetus to generate DNA barcodes using multiple plant DNA markers for all woody species at forest community levels, analyzing massive DNA barcoding sequence data, and promoting theoretical innovation. Lastly, we raised five possible directions; specifically, proposing a "purpose-driven barcode" fit for multi-level applications, developing new integrative sequencing strategies, pushing DNA barcoding beyond terrestrial ecosystem, constructing national-level DNA barcode sequence libraries for special plant groups, and establishing intelligent identification systems or online server platforms. These efforts will be potentially valuable to explore large-scale biodiversity patterns, the origin and evolution of life, and will also facilitate preservation and utilization of biodiversity resources.

Tuesday, February 28, 2017

Metabarcoding to track indoor fungi

Environmental microbes can have both beneficial and harmful effects on health, e.g. bacterial biodiversity is discussed as a factor influencing immuntolerance. This might explain the lower incidence of allergic diseases in children living in rural environments in contrast to children living in urban environments. Rural environments are usually more diverse and researchers are particularly looking at the effect of environmental microbiota on the commensal microbiota. The latter has been shown to have a strong influence on human immuncompetence.

People spend most of their time in indoor environments, which contain a variety of microbes. Serious problems may develop in buildings with long-lasting dampness, where the moisture supports the growth of bacteria and fungi (i.e., mould). Based on epidemiological studies, mould in buildings is positively associated with several allergic and respiratory effects, and certain moulds are toxigenic, meaning that they can produce mycotoxins. There are estimates that allergic diseases caused by plant, animal, and fungal allergens affect more than 30% of the population in industrialized countries, and there is increasing awareness and concern over exposure to moulds in indoor environments. The phenomenon has become known as Sick Building Syndrome (SBS), where the occupants describe a complex range of vague and often subjective health complaints. 

The most widely used methods to identify indoor fungi are culture dependent which means that they are inherently biased. We know that among the thousands of microbial species that populate the world, only a few have been characterized by more than just a DNA sequence due to our inability to grow them in the laboratory. A very good reason to switch to DNA based methods as two researchers of the University of Helsinki just did. They utilized metabarcoding to determine the fungal diversity in samples collected from five buildings: two at the university, two from nursery schools in the city, and an old inhabited farmhouse in a more rural setting.

The study represents an important proof of concept as it shows quite promising depth of results but also the potential pitfalls. Having identified the latter will allow for reliable high quality applications. The total number of fungal genera found during the study was 585 but when comparing fungal diversities and taxonomic composition between different types of buildings, no obvious differences or patters were found.

Of the 78 fungal genera listed to have been shown to induce allergies in persons that are hypersensitive to allergens, 51 were found in this study, although mostly at very low frequencies. 

The authors make clear that given the experimental design of their study there are clear limitations to what can be concluded: We discovered that making explicit conclusions on the relationship between the indoor air quality and mycoflora is complicated by the lack of appropriate indicators for air quality and by the occurrence of wide spatial and temporal changes in diversity and compositions among samples.

Monday, February 27, 2017

Monday reads

Last Monday was a statutory holiday here so I skipped the post. Now we're back. A couple of interesting reads for your working week.

In this study we compared DNA barcode-suggested species boundaries with morphology-based species identifications in the amphipod fauna of the southern European Atlantic coast. DNA sequences of the cytochrome c oxidase subunit I barcode region (COI-5P) were generated for 43 morphospecies (178 specimens) collected along the Portuguese coast which, together with publicly available COI-5P sequences, produced a final dataset comprising 68 morphospecies and 295 sequences. Seventy-five BINs (Barcode Index Numbers) were assigned to these morphospecies, of which 48 were concordant (i.e., 1 BIN = 1 species), 8 were taxonomically discordant, and 19 were singletons. Twelve species had matching sequences (<2% distance) with conspecifics from distant locations (e.g., North Sea). Seven morphospecies were assigned to multiple, and highly divergent, BINs, including specimens of Corophium multisetosum (18% divergence) and Dexamine spiniventris (16% divergence), which originated from sampling locations on the west coast of Portugal (only about 36 and 250 km apart, respectively). We also found deep divergence (4%-22%) among specimens of seven species from Portugal compared to those from the North Sea and Italy. The detection of evolutionarily meaningful divergence among populations of several amphipod species from southern Europe reinforces the need for a comprehensive re-assessment of the diversity of this faunal group.

Unexpected contaminants uncovered during routine COI-5P DNA barcoding of British Columbia Kallymeniaceae indicated the presence of a novel lineage allied to the family Meiodiscaceae, Palmariales. Available rbcL data for species of this family were used to design specific primers to screen for the presence of the meiodiscacean species in 534 kallymeniacean specimens primarily from British Columbia, Canada. Ultimately, 43 positive PCR products representing six diverse genetic groups from nine host species were uncovered; three are described here in the new genus Kallymenicola gen. nov., viz., K. invisiblis sp. nov., K. penetrans sp. nov., and K. superficialis sp. nov. Although genetic groups loosely displayed evidence of host specificity and cospeciation, examples of host switching with interesting biogeographical patterns were also documented.

DNA barcoding relies on short and standardized gene regions to identify species. The agricultural and horticultural applications of barcoding such as for marketplace regulation and copyright protection remain poorly explored. This study examines the effectiveness of the standard plant barcode markers (matK and rbcL) for the identification of plant species in private and public nurseries in northern Egypt. These two markers were sequenced from 225 specimens of 161 species and 62 plant families of horticultural importance. The sequence recovery was similar for rbcL (96.4%) and matK (84%), but the number of specimens assigned correctly to the respective genera and species was lower for rbcL (75% and 29%) than matK (85% and 40%). The combination of rbcL and matK brought the number of correct generic and species assignments to 83.4% and 40%, respectively. Individually, the efficiency of both markers varied among different plant families; for example, all palm specimens (Arecaceae) were correctly assigned to species while only one individual of Asteraceae was correctly assigned to species. Further, barcodes reliably assigned ornamental horticultural and medicinal plants correctly to genus while they showed a lower or no success in assigning these plants to species and cultivars. For future, we recommend the combination of a complementary barcode (e.g. ITS or trnH-psbA) with rbcL + matK to increase the performance of taxa identification. By aiding species identification of horticultural crops and ornamental palms, the analysis of the barcode regions will have large impact on horticultural industry.

Fungi are among the most abundant and diverse organisms on Earth. However, a substantial amount of the species diversity, relationships, habitats, and life strategies of these microorganisms remain to be discovered and characterized. One important factor hindering progress is the difficulty in correctly identifying fungi. Morphological and molecular characteristics have been applied in such tasks. Later, DNA barcoding has emerged as a new method for the rapid and reliable identification of species. The nrITS region is considered the universal barcode of Fungi, and the ITS1 and ITS2 sub-regions have been applied as metabarcoding markers. In this study, we performed a large-scale analysis of all the available Basidiomycota sequences from GenBank. We carried out a rigorous trimming of the initial dataset based in methodological principals of DNA Barcoding. Two different approaches (PCI and barcode gap) were used to determine the performance of the complete ITS region and sub-regions.
For most of the Basidiomycota genera, the three genomic markers performed similarly, i.e., when one was considered a good marker for the identification of a genus, the others were also; the same results were observed when the performance was insufficient. However, based on barcode gap analyses, we identified genomic markers that had a superior identification performance than the others and genomic markers that were not indicated for the identification of some genera. Notably, neither the complete ITS nor the sub-regions were useful in identifying 11 of the 113 Basidiomycota genera. The complex phylogenetic relationships and the presence of cryptic species in some genera are possible explanations of this limitation and are discussed.
Knowledge regarding the efficiency and limitations of the barcode markers that are currently used for the identification of organisms is crucial because it benefits research in many areas. Our study provides information that may guide researchers in choosing the most suitable genomic markers for identifying Basidiomycota species.

Evolutionary history of the sequestrate genus Rossbeevera (Boletaceae) reveals a new genus Turmalinea and highlights the utility of ITS minisatellite-like insertions for molecular identification
The sequestrate (truffle-like) basidiomycete genera Rossbeevera, Chamonixia, and Octaviania are closely related to the epigeous mushroom genera Leccinum and Leccinellum. In order to elucidate the properties and placement of several undescribed sequestrate taxa in the group and to reveal the evolutionary history of Rossbeevera and its allies, we conducted phylogenetic analyses based on three nuclear (ITS, nLSU, EF-1α) and two mitochondrial DNA loci (ATP6 and mtSSU) as well as precise morphological observations. Phylogenetic analyses of three nuclear loci suggest a complex evolutionary history with sequestrate fruiting bodies present in several clades, including a previously unrecognized sister clade to Rossbeevera. Here we propose a new sequestrate genus, Turmalinea, with four new species and one new subspecies as well as two new species of Rossbeevera. The three-locus nuclear phylogeny resolves species-level divergence within the Rossbeevera-Turmalinea lineage, whereas a separate phylogeny based on two mitochondrial genes corresponds to geographic distance within each species-level lineage and suggests incomplete lineage sorting (ILS) and gene introgression within several intraspecific lineages of Rossbeevera. Furthermore, topological incongruence among the three nuclear single-locus phylogenies suggests that ancient speciation within Rossbeevera probably involved considerable ILS. We also found an unusually long, minisatellite-like insertion within the ITS2 in all Rossbeevera and Turmalinea species. A barcode gap analysis demonstrates that the insertion is more informative for discrimination at various taxonomic levels than the rest of the ITS region and could therefore serve as a unique molecular barcode for these genera.

valuating diffuse sediment contamination in the environment is a major concern with the aim of reaching a good chemical and ecological state of the littoral zone. In this study the risks of chronic chemical contamination and consequences in the bivalves Crassostrea gigas, Mytilus sp. and Mimachlamys varia were evaluated in coastal environments. The objective here was to understand the anthropological phenomena that affect the functioning of the marina of La Rochelle (semi-closed environment). Harbours seeking ecomanagement accreditations (such as the international reference ISO 14001) constitute zones of interest to implement biomonitoring studies. The biological effects of chemical pollution in the Marina of La Rochelle were studied to develop a multi-biomarker biomonitoring approach on specific marine species of this site. Moreover, a genetic (DNA barcoding) approach was applied to validate the species identity of collected bivalves. Of the three species tested the scallop, M. varia, was the most sensitive to metal exposure.

Thursday, February 23, 2017

From the inbox

Two symposia are held back-to-back in Uppsala 11-15 Sept, 2017: the 3rd Symposium on Ecological Networks and the 3rd Symposium on Molecular Analysis of Trophic Interactions.

Please note that registration is now open. To aim for a program where all attendants can participate with a talk or poster, we have opted for a two-stage process of registration. In the first stage open now, we ask you to submit a notice of interest and the abstract of a talk or poster. All themes from within the broad remits of the respective symposia are warmly welcome. In submitting the abstract, you are asked are to choose from whether you want to submit an oral presentation or a poster -- and whether (should your oral presentation not fit into the oral program) you will still be happy to present it as a poster. Once the final acceptance of abstracts is completed (31 May), you will be invited to update this notice of interest to a final conference registration, at which stage we will also ask you to deposit the conference fee.

Friday, February 17, 2017

Invasions continue

For all groups of organisms on all continents, the number of alien species has increased continuously during the last 200 years. For most groups, even the rate of introduction is highest recently. Barring mammals and fishes, there are no signs of a slow-down and we have to expect more new invasions in the near future.

Quite a sobering statement. However, it summarizes the results of a new study in which a large international group of researchers analysed alien species accumulation during the last centuries. They used more than 45,000 first occurrence records for more than 16,000 alien species.

The colleagues found that 37% of all recorded alien species have been introduced between 1970-2014 and thus recently. At its peak 585 new species were recorded within one year. This corresponds to more than 1.5 new alien species per day globally. This is likely an underestimate as the date of first record is not available for most alien species.

The trends of increase vary among taxonomic groups, which can be attributed to human activities. We observe a distinct increase in first record rates of vascular plants in the 19th century, probably as a result of the intensification of horticulture. The rates of new introductions of other organisms such as algae, molluscs or insects increased steeply after 1950. This is most likely a consequence of the ongoing globalisation of trade.

Although it was known that the number of alien species increased during the last decades, it remained unclear whether or not the accumulation of alien species has already reached a point of slow-down. Well, it clearly hasn't.

Tuesday, February 14, 2017

Climate change is not a future threat anymore

The rate of warming over the past 50 years (0.13 °C ± 0.03 °C per decade) is nearly twice that for the previous 50 years, and the global temperature by 2100 is likely to be 5–12 standard deviations above the Holocene mean. The effects of climate change on some species are already being witnessed, with changes documented in spatial distribution, abundance, demography, phenology and morphology. However, to date, no quantification of the number of species for which at least one population has been currently impacted by climate change, and the extent of these impacts, has been conducted, even for the better-studied taxa such as birds and mammals. 

In an international study published yesterday a team of international researchers present evidence of observed responses to recent climate changes in some 700 bird and mammal species. The researchers reviewed the observed impacts of climate change on birds and mammals using a total of 130 studies, making it the most comprehensive assessment to date on how climate change has affected our most well studied species.

Only 7% of mammals and 4% of birds for which the colleagues found evidence of a negative response are actually coded on the IUCN Red List of Threatened Species as threatened by ‘climate change and severe weather'. This severe under-reporting is also very likely in less studied species groups which represent the vast majority of life. The authors strongly argue for big improvements of assessments of the impacts of climate change on all species right now:

We need to communicate the impacts of climate change to the wider public and we need to ensure key decision makers know significant change needs to happen now to stop species going extinct. Climate change is not a future threat anymore.

Monday, February 13, 2017

Monday reads

Another week that begins with some interesting reads. I find it fascinating that every week I am able to find a good number of academic publications which use DNA-based identifications and/or DNA barcoding in particular. The field is now large enough and gained sufficient momentum that new discoveries and findings appear on a more or less daily basis. As a result every week I am in the fortunate situation to pick a few of many publications to highlight in the Monday reads:

Until now, the potential of NGS for the construction of barcode libraries or integrative taxonomy has been seldom realised. Here, we amplified (two-step PCR) and simultaneously sequenced (MiSeq) multiple markers from hundreds of fig wasp specimens. We also developed a workflow for quality control of the data. Illumina and Sanger sequences accumulated in the past years were compared. Interestingly, primers and PCR conditions used for the Sanger approach did not require optimisation to construct the MiSeq library. After quality controls, 87% of the species (76% of the specimens) had a valid MiSeq sequence for each marker. Importantly, major clusters did not always correspond to the targeted loci. Nine specimens exhibited two divergent sequences (up to 10%). In 95% of the species, MiSeq and Sanger sequences obtained from the same sampling were similar. For the remaining 5%, species were paraphyletic or the sequences clustered into divergent groups on the Sanger + MiSeq trees (>7%). These problematic cases may represent coding NUMTS or heteroplasms. Our results illustrate that Illumina approaches are not artefact-free and confirm that Sanger databases can contain non-target genes. This highlights the importance of quality controls, working with taxonomists and using multiple markers for DNA-taxonomy or species diversity assessment.

We compare the diversity of Chilean bees as understood from traditional taxonomy-based catalogues with that currently known from DNA barcodes using the BIN system informed by ongoing morphology-based taxonomy. While DNA barcode surveys of the Chilean bee fauna remain incomplete; it is clear that new species can readily be distinguished using this method and that morphological differentiation of distinct barcode clusters is sometimes very easy. We assess the situation in two genera in detail. In Lonchopria Vachal one "species" is readily separable into two BINs that are easily differentiated based upon male mandibular and genitalic morphology (characters generally used in this group) as well female hair patterns. Consequently, we describe Lonchopria (Lonchopria) heberti Packer and Ruz, new species. For Liphanthus Reed, a large number of new species has been detected using DNA barcoding and considerable additional traditional morphological work will be required to describe them. When we add the number of BINs (whether identified to named species or not) to the number of Chilean bee species that we know have not been barcoded (both described and new species under study in our laboratories) we conclude that the bee fauna of Chile is substantially greater than the 436 species currently known.

The escalating growth in illegal wildlife trade and anthropogenic habitat changes threaten the survival of pangolin species worldwide. All eight extant species have experienced drastic population size reductions globally with a high extinction risk in Asia. Consequently, forensic services have become critical for law enforcement, with a need for standardised and validated genetic methods for reliable identifications. The seizure of three tonnes of pangolin scales, believed to have originated from Africa, by Hong Kong Customs Authorities provided an opportunity for the application of DNA barcoding in identifying scales. Three mitochondrial DNA gene regions (COI, Cyt b, and D-loop) were amplified for a subsample of the confiscated material and compared with taxonomically verified references. All four African species were recovered as monophyletic with high interspecific uncorrected p-distance estimates (0.048-0.188) among genes. However, only three of four African species (Phataginus tricuspis, Phataginus tetradactyla, and Smutsia gigantea, originating from West and Central Africa) and one of four Asian species (Manis javanica from Southeast Asia) were identified among scales. Although the assignment of unknown scales to specific species was reliable, additional genetic tools and representative reference material are required to determine geographic origins of confiscated pangolin specimens.

Kampo is the general designation for traditional Japanese herbal medicines, which are recognized as official medicines and listed in the Japanese pharmacopoeia (JP). In most cases, it is difficult to identify the crude drug materials to species level using only traditional identification methods. We report the first online DNA barcode identification system, which includes standard barcode sequences from approximately 95% of the species recorded in the JP (16th edition). This tool provides users with basic information on each crude drug recorded in the JP, DNA barcoding identification of herbal material, and the standard operating procedure (SOP) from sampling to data analysis. ITS2 sequences (psbA-trnH was an alternative when ITS2 could not be amplified) were generated from a total of 576 samples to establish the database. An additional 100 samples (from different medicinal parts, from both single origin and multiple origins and from both retailers and the planting base) were identified using the system. A total of 78% of the test samples were identified as the species listed on their label. This system establishes a model platform for other pharmacopeias from countries like China, Korea, the US and the European Union, for the safe and effective utilization of traditional herbal medicines.