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.