ABOL BioBlitz: DNA barcoding safeguards taxonomic knowledge

ABOL BioBlitz: DNA barcoding safeguards taxonomic knowledge

ABOL BioBlitz: DNA barcoding safeguards taxonomic knowledge

The Austrian Barcode of Life (ABOL) initiative uses DNA barcoding to safeguard and make publicly accessible rare knowledge on biodiversity generated in the course of local BioBlitz events
Local fisher presents catch of the day from lake Millstatt PHOTO CREDIT: Susanne Glatz-Jorde, Biosphere Reserve Nockberge

A BioBlitz is an event organized to survey the biodiversity of a designated area as fast and as comprehensively as possible1. Along with the general public, taxonomic experts from various fields, whether laymen or academics, swarm a location to record every identifiable species, usually within 24 hours.

The primary goal of these events is to increase bioliteracy—awareness of biodiversity and its importance for healthy, functioning ecosystems. We believe increased bioliteracy could transform societal perceptions of nature and, ultimately, humanity’s relationship with other species. Consequently, we deem it a prerequisite in our arsenal to abate the dramatic loss of biodiversity2. Additionally, the BioBlitz approach is becoming increasingly important for conservation efforts as it generates high-quality biodiversity data while simultaneously enhancing research capacity3.

In 2019, the Austrian Barcode of Life Initiative (ABOL) successfully introduced a new extended BioBlitz format to Austria—the ABOL BioBlitz—which combines existing collecting events with DNA barcoding. The organisms acquired and identified by experts during BioBlitz events are subsequently DNA barcoded. Importantly, specimens are photographed and stored in a scientific collection along with their associated metadata to be fully compliant with DNA barcoding standards for reference sequences.

The Great Lakes

Locations of the ‘Days of Biodiversity’ 2019 in Austria. The logos represent the organizing institutions.

Base map from d-maps.com

Although running for the first time, the ABOL BioBlitz efforts were very well received and highly successful. From 2000 individuals comprising 1400 taxa, approximately 1500 DNA barcodes were obtained and these data are now available on BOLD.

The number of individuals per higher taxon provided for DNA barcode analysis in the course of the ABOL BioBlitzes 2019. Sites indicated by colours.

Images from ClipArt ETC

In 2019, ABOL joined six ‘Days of Biodiversity’ with ABOL BioBlitzes in five different federal states of Austria. These events covered very different habitat types, from an organic farmstead in Upper Austria, a valley in the mountainous area in Tyrol, to annually organized events in protected areas like National Park Hohe Tauern or the Biosphere Reserves of Nockberge and Wienerwald. It is important to note that these events are only possible with the generosity and cooperation of the institutional hosts to whom we wish to extend our deepest thanks.

In total 54 taxonomic experts joined our efforts at these events. They not only provided us with samples from their collected material, along with the respective metadata and photos, but they also prepared reference individuals and assured their appropriate storage in public collections. After receiving samples from the experts, the ABOL team transferred tissue into microplates and shipped them to the Canadian Centre for DNA Barcoding (CCDB) in Canada for barcode analysis. We thank all the experts for their incredible efforts as well as staff at the CCDB for their obligingness and cooperation.

Day of Nature in the Biosphere Reserve Nockberge

The 4th Day of Nature named ‘Shores and mountain slopes of Lake Millstatt in Carinthia’ in the Biosphere Reserve Nockberge contributed substantially to the species inventory of the Carinthian part of the Biosphere Reserve. 45 taxonomic experts recorded 1166 species, some rare while others new records for the Biosphere Reserve Nockberge and for Carinthia. The event was organized by the team at the Biosphere Reserve, together with two ecological agencies, Ökoteam and E.C.O.

IMAGE: Local fisher presents catch of the day from lake Millstatt
CREDIT: Susanne Glatz-Jorde, Biosphere Reserve Nockberge

Day of Biodiversity in Upper Austria

The Day of Biodiversity in Upper Austria, organized by the Naturschutzbund Oberösterreich and the Biologiezentrum of the Upper Austrian State museums, took place around the Mühlbergerhof, an organic farmstead covering around 20 ha of species-rich grassland, pastures, and deciduous forest. In addition to the species inventory, excursions with different topics (e.g. moths and bats, plants, fungi and lichens, insects) were offered to the public.

IMAGE: Members of the ABOL coordination team examine collected insects from Mühlbergerhof
CREDIT: Heidi Kurz, Naturschutzbund OÖ

Day of Biodiversity in the Biosphere Reserve Wienerwald

The annual Day of Biodiversity in the Biosphere Reserve Wienerwald took place in 2019 in Pressbaum, Lower Austria. Within 24 hours, 1151 species were recorded. This data forms a valuable basis for scientific research and nature conservation activities in the Biosphere Reserve. Excursions led by experts explored water insects, plants, and birds. More than 30 stands provided information on various topics for adults and children. The festival of biodiversity was completed with regional food, kid’s programs and an open-air concert.

IMAGE: ABOL information stand at the festival of biodiversity in Pressbaum
CREDIT: Michaela Sonnleitner

Day of Biodiversity in Vienna
The Day of Biodiversity in Vienna was the 1st transnational event of its kind within the Interreg project CITY NATURE jointly organized by the Vienna municipality (MA22), the University of Natural Resources & Life Sciences (BOKU), and Bratislava, Slovakia. The collecting event around the BOKU area was complemented by excursions exploring birds, plants, insects & mammals.

IMAGE: Group photo from the closing event with project partner from Bratislava at the Day of Biodiversity in Vienna CREDIT: Barbara Reinwein, MA22

Day of Biodiversity in Tyrol

During the Day of Biodiversity in Tyrol, the Brandenberg Valley attracted many experts and visitors with its diverse natural habitat. A highlight of the event was the demonstration of different types of light traps catching night-active insects. Experts held lectures on butterflies, which fascinated children and adults. Arctia matronula, a rare and locally distributed Noctuidae was among the observed species. The days of biodiversity in Tyrol are organized by the Tyrolean State Museums, the University of Innsbruck and the State of Tyrol.

IMAGE: Expert demonstrates light traps and explains nocturnal butterflies
CREDIT: Michaela Sonnleitner

Day of Biodiversity in the National Park Hohe Tauern

The motto of the 13th Day of Biodiversity in the National Park Hohe Tauern was Summit of biodiversity at the foot of the Großglockner (the highest mountain in Austria). More than 60 experts investigated the species inventory of the Gössnitztal, an elongated alpine valley up to more than 2000 m and the surroundings of Heiligenblut in Carinthia. In addition to plants, insects, birds etc., bats were observed in different altitudes and their call sequences recorded. The event was perfectly organized by the Team of the National Park.

IMAGE: Group photo from the Day of Biodiversity in the National Park Hohe Tauern in Heiligenblut with Großglockner in the background.
CREDIT: Manfred Schmucker

Due to the current Coronavirus pandemic, some of the 2020 ‘Days of Biodiversity’ were cancelled or postponed to 2021, while some will take place but with great care. For the future, we hope to be able to increase the number of participants and, ultimately, the data collected.

The concerted actions of the ABOL team at the ‘Days of Biodiversity’ significantly enhance the value and reach of these local BioBlitz events. For example, our efforts contribute to the completion of DNA barcode reference databases, important resources for society in the long term. They also support taxonomic research by providing the genetic resources important to the morphological determination of species. Additionally, we significantly promote these events amongst the public, raise awareness of the importance of biodiversity as well as build and strengthen bonds within the biodiversity community by enabling a process of mutual learning between cross-disciplinary experts.

As events this past year have shown, combining a BioBlitz with DNA barcoding is a very successful way to obtain both taxonomic and barcode data as well as integrate experts, especially for national DNA barcoding initiatives, which are not always fully funded, as is the case in Austria.

These events also foster a greater appreciation of biodiversity and ecosystems amongst the public and, therefore, are a very successful approach to increase bioliteracy. The currently ongoing biodiversity crisis implies rapidly growing importance of biodiversity data. As taxonomic expertise is increasingly rare in the academic environment, it is becoming more dependent on nature enthusiasts. This knowledge of biodiversity has always received too little attention, although a substantial part of it is held outside of academic institutions. Therefore, we wish to emphasize, that increased appreciation of nature should go hand in hand with that of private taxonomic expertise. ABOL BioBlitzes take all this into account.

References:

1. Baker G.M., Duncan N., Gostomski T., Horner M.A., Manski D. (2014). The bioblitz: Good science, good outreach, good fun. Park Science 31(1): 39–45.
2. IPBES (2019). Global assessment report on biodiversity and ecosystem services of the Intergovernmental Science-Policy Platform on Biodiversity and Ecosystem Services. E. S. Brondizio, J. Settele, S. Díaz and H. T. Ngo (eds.) Bonn, IPBES secretariat.
3. Parker S.S., Pauly G.B., Moore J., Fraga N.S., Knapp J.J., Principe Z., Brown B.V., Randall J.M., Cohen B.S. Wake T.A. (2018). Adapting the bioblitz to meet conservation needs. Conservation Biology 32(5): 1007–1019. https://doi.org/10.1111/cobi.13103

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The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

Forest dynamics, spatial distribution patterns, and sampling scale are associated with mitochondrial DNA variation in Argentinian butterflies

Andean forests in northwestern Argentina.
PHOTO CREDIT: Ezequiel Núñez Bustos

Argentina harbours more than 1,200 species of butterflies, most of them found in two biodiversity hotspots and priority areas for conservation: the Atlantic Forest and the Andean forests1.

Figure 1: Sampling localities in northwestern Argentina (NWA, black squares) and northeastern Argentina (NEA, white triangles)2. The Atlantic Forest (dark blue) extends along the Brazilian coast and reaches its southernmost portion in NEA, while the Central Andean forests (red) descend from southern Peru and reach NWA. The distribution of eight other ecoregions indicated.

Despite their current isolation (Figure 1), these two areas have been cyclically and transiently connected in the past, promoting the interchange of flora and fauna, and resulting in a pattern of disjunctly co-distributed taxa. While the historical relationship between these allopatric forests and its evolutionary effects on shared fauna has been the subject of recent (and ongoing) research, studies have been concentrated mostly on vertebrates.

This study explores the butterflies of the Atlantic Forest and the Andean forests providing new insights into both the diversification patterns in southern South America and the impact of increasing the geographic and taxonomic scale of sampling on DNA barcoding performance in the region.

Atlantic Forest in northeastern Argentina. PHOTO CREDIT: Ezequiel Núñez Bustos

In 2017, we assembled and analyzed a DNA barcode reference library for 417 species from northeastern Argentina (NEA)2, focusing on the Atlantic Forest and covering around one-third of the butterfly fauna of the country. To expand the geographic and taxonomic distribution of this library, we generated DNA barcodes for 213 butterfly species from northwestern Argentina (NWA) with a focus on the Andean forests.

We then used these libraries to examine three themes, outlined below.

1.The effectiveness of DNA barcodes for species discrimination and identification

 

The mean intraspecific distance for the butterflies of NWA was 0.29%, while mean interspecific distance among congeneric species was 7.24% (Figure 2). More importantly, mean distance to the nearest neighbour (7.56%) was nearly 13 times larger than the mean distance to the furthest conspecific (0.60%), resulting in a distinct barcode gap for all but two species represented by two or more individuals (Figure 2).

Genetic distance or sequence variation in the COI sequences within and between species was estimated using the Kimura-2-parameter (K2P) model of nucleotide substitution

Substitution models describe the process of genetic variation through fixed mutations, constituting the foundation of evolutionary analysis at the molecular level.

Arenas M (2015) Trends in substitution models of molecular evolution. Frontiers in Genetics 6(319). 

Figure 2: Frequency histogram of COI sequence distances within species (orange) and among congeneric species (blue) of butterflies in NWA. The inset graph shows the barcode gap analysis for species represented by two or more COI sequences, where each dot represents a specimen. Red dots correspond to individuals with a maximum intraspecific distance higher than the distance to the nearest heterospecific. The vertical dashed line shows the 95th percentile of all intraspecific distances (2.02%), while the horizontal line corresponds to the lower 5% of all congeneric distances (3.36%).

Consistently, sequence-based specimen identification simulations showed that this library is extremely effective in the identification of the butterflies of NWA, exceeding a 98% success rate regardless of the identification criteria implemented.

We then used different clustering algorithms to assess the presence of cryptic species. Overall, these methods generated between 1.4–9.9% more Molecular Operational Taxonomic Units (MOTUs) than the number of reference species, suggesting that the butterfly diversity of NWA might be higher than currently recognized.

Figure 3: Taxonomic coverage of the complete DNA barcode reference library for the butterflies of Argentina. Dark shading indicates the proportion of species covered within each family based on the total known for the country.

Merging the NWA and NEA databases resulted in a DNA barcode reference library for nearly 500 butterfly species, covering ~40% of the butterfly fauna of Argentina (Figure 3) and representing 549 barcode clusters (BINs) on BOLD (170 of which are new to the platform).

2.The impact of increasing the spatial and taxonomic coverage on DNA barcoding performance

When we compared the two reference libraries, we found that the barcode gap was significantly narrower in the NEA than in the NWA library (Figure 4). This is most likely associated with the higher geographic and taxonomic coverage of the former, since expanding the spatial scale of sampling is expected to not only increase intraspecific variation as a result of isolation by distance but also reduce interspecific divergences as more closely related species appear.

Figure 4: Maximum intraspecific distance (blue) and minimum interspecific distance (red) for the three datasets. Note the different scales.

When we tried to identify specimens from NWA by using the reference library of NEA, a considerably high proportion of individuals representing shared species between these regions could not be identified or resulted in an ambiguous identification, even when we allowed a maximum intraspecific distance of as high as 2% in the identification procedure. This was due to the existence of deep intraspecific divergences between conspecifics from northeastern and northwestern Argentina, two regions separated on average by more than 1,000 km.

At the same time, however, we observed that the effect of increasing the geographic (and taxonomic) scale was more profound on the minimum interspecific distances than on the maximum intraspecific distances. Therefore, it is possible that butterfly species in NEA are also naturally more variable than in NWA based on our current sampling. While specimens from NWA came almost exclusively from the montane forest on the east slope of the Andes, the sampling in NEA covered not only larger geographic distances but also a more heterogeneous landscape, characterized by the existence of different ecoregions (Figure 1) and physical barriers such as river, specifically the Paraná-Paraguay River axis. Regardless, our results show that both large geographic distances and increased taxonomic coverage can affect DNA barcoding identification performance, especially when using a local library to identify the fauna from another distant region.

As expected, the maximum intraspecific distance was significantly higher and minimum interspecific distance was significantly lower in the complete database (NEA + NWA) than within the NWA and NEA libraries alone (Figure 4). However, the logical and anticipated reduction in the barcode gap did not have, in this case, a significant impact on the identification performance of DNA barcodes, which were able to correctly identify ~99% of the individuals. This reflects the importance of increasing the spatial and taxonomic coverage of DNA barcode libraries to improve identification success, and of considering the use of a local database to identify regional fauna when a more comprehensive COI database is not available.

Doxocopa cyane burmeisteri
Doxocopa cyane burmeisteri
Parides erithalion erlaces

Parides erithalion erlaces

Pteronymia ozia tanampaya

Pteronymia ozia tanampaya

Butterfly species from the Andean forests. PHOTO CREDIT: Ezequiel Núñez Bustos

3.Geographic patterns of intraspecific variation across Argentina

A total of 135 butterfly species are shared between the databases of NEA and NWA. Mean intraspecific distance for these species was significantly higher between regions (1.02%) than within them (NEA, mean 0.35%; NWA, mean 0.33%), especially for a subset of 43 species that showed particularly deeper distance (mean 2.43%) between NEA and NWA.

We then focused only on the 85 species that are present in both the Atlantic Forest and the Andean forests (Figure 5), 27 of which have a disjunct distribution between forests, being absent from intermediate ecoregions, while the remaining 57 have a continuous range across northern Argentina.

Figure 5: Proportion of shared species between NEA and NWA that occur in both forests. The spatial distribution pattern (disjunct vs continuous) and the percentage of species with a deep intraspecific divergence between forest populations indicated.

We found that mean intraspecific distance between forest populations was significantly higher for the disjunctly distributed species (1.65%) than for species with continuous ranges (0.78%), showing that spatial distribution patterns have an influence on the level of intraspecific variation. Moreover, the proportion of species showing the deep divergence between populations from the Atlantic Forest and the Andean forests was notably higher among species with fragmented distributions (nearly 50%) than for species with continuous ranges (less than 30%) (Figure 5).

Lastly, based on standard molecular rates and COI sequence divergence, all diversification events between forest populations were dated to the last two million years, a time period when the currently isolated Atlantic Forest and Andean forests experienced multiple transient connections across the open vegetation corridor, a diagonal of more open and drier savanna-like environments (Caatinga, Cerrado and Chaco) that isolates the Atlantic Forest from the Andean forests (and the adjacent Amazonia) (Figure 1). These past connections were promoted mainly by Pleistocene climatic changes and habitat shifts.

Catonephele numilia neogermanica

Catonephele numilia neogermanica

Callicore hydaspes

Callicore hydaspes

Doxocopa agathina vacuna

Doxocopa agathina vacuna

Butterfly species from the Atlantic Forest.
PHOTO CREDIT: Ezequiel Núñez Bustos

Conclusions

Our study has not only expanded the DNA barcode reference library for the butterflies of Argentina, but it also constitutes, to our knowledge, the first multi-species assessment of the historical relationship between the currently isolated Atlantic Forest and Andean forests using butterfly species as model organisms.

Importantly, our research supports the fact that, even in the era of genomic data, large-scale analyses of mitochondrial DNA variation are still extremely useful for evolutionary studies, as they unveil spatial diversification patterns and highlight cases that deserve further investigation.

ACKNOWLEDGEMENTS:

We thank our colleagues from the Museo Argentino de Ciencias Naturales and the staff at the Centre for Biodiversity Genomics (CBG) for their help during different stages of this ongoing investigation. We also thank Michelle D’Souza for her helpful comments and suggestions that improved this contribution. This project is supported by Richard Lounsbery Foundation, the CBG, the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), the Agencia Nacional de Promoción de la Investigación, el Desarrollo Tecnológico y la Innovación, Fundación Williams, Fundación Bosques Nativos Argentinos and Fundación Temaiken. For granting the permits and transit guides, we thank the Offices of Fauna of the Argentinian provinces in which fieldwork was conducted, the Administración de Parques Nacionales, and the Ministerio de Ambiente y Desarrollo Sostenible from Argentina.

References:

1. Klimaitis J, Núñez Bustos E, Klimaitis C, Güller R (2018) Mariposas-Butterflies-Argentina. Guía de Identificación-Identification Guide. Vazquez Mazzini Editores. Buenos Aires. pp. 327.

2. Lavinia P, Núñez Bustos E, Kopuchian C, Lijtmaer D, García N, Hebert P, Tubaro P (2017) Barcoding the butterflies of southern South America: Species delimitation efficacy, cryptic diversity and geographic patterns of divergence. PLOS ONE 12(10), e0186845. https://dx.doi.org/10.1371/journal.pone.0186845

Written by

Natalí Attiná

Natalí Attiná

Ezequiel Núñez Bustos

Ezequiel Núñez Bustos

Darío A. Lijtmaer

Darío A. Lijtmaer

Pablo L. Tubaro

Pablo L. Tubaro

Pablo D. Lavinia

Pablo D. Lavinia

Museo Argentino de Ciencias Naturales “Bernardino Rivadavia” (MACN–CONICET)

July 31, 2020

doi: 10.21083/ibol.v10i1.6256  

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Hunting for a water mite neotype in southern Norway

Hunting for a water mite neotype in southern Norway

Hunting for a water mite neotype in southern Norway

Scientists rediscover lost specimens of water mite in Norway 120 years after they were first described

A stream near the church of Vanse at Lista in southern Norwaythe type locality of Lebertia porosa Thor, 1900.

PHOTO CREDIT: Torbjørn Ekrem

Did you know that scientists can assess natural water quality by monitoring the diversity of aquatic invertebrates? Freshwater insect and arachnid populations are often important indicators of environmental change. This is evident in particularly species-rich groups, such as water mites and biting or non-biting midges, which have great potential for monitoring water quality. The problem is only that they are too difficult and time consuming to identify in routine water quality assessments. This hurdle can be overcome with DNA metabarcoding, but only if a good reference barcode library is available.

Elisabeth Stur of the Norwegian University of Science and Technology (NTNU) University Museum, along with her team, have been doing summer fieldwork for the Water Mites and Midges in southern Norway (Water M&M) project. One of the many goals for this year’s fieldwork was not only to contribute to the reference barcode library, but also to sample the type locality of the water mite Lebertia porosa, described 120 years ago by Sig Thor, a Norwegian priest and acarologist.

The Great Lakes
Phaenopsectra flavipes (Diptera: Chironomidae) with water mite larvae attached. PHOTO CREDIT: Aina Mærk Aspaas, NTNU University Museum

Barcode data indicate that there are at least six cryptic genetic lineages within this species, but it is unknown which of these applies to the nominal species. Since the original type material is lost, re-sampling L. porosa from its type locality is important in designating a neotype that most likely belongs to the species described by Thor in 1900. This way, researchers can stabilize the definition of the L. porosa species name, such that potential new species could be described. This species delineation is part of a MSc. project by Valentina Tyukosova at NTNU: Integrative taxonomy and species delimitation in the Lebertia porosa species complex (Acari, Parasitengona: Hydrachnidia).

The type locality of L. porosa was vaguely described in Thor’s original publication as a “stream near the church of Vanse”. After studying maps of the surrounding area, researchers learned that this church still stands, and were able to locate two nearby streams.

Now they wondered, would these streams still be in good condition 120 years later? As the team of researchers approached what they thought might be the stream in June 2020, they were pleased to see running, clear water under the bridge. Next mystery: could the streams hold a population of L. porosa 120 years after first collection? They found that yes, the waters could, and the water mite populations were bountiful!

The Great Lakes DNA Barcoding Project team

Water mites from the type locality of Lebertia porosa Thor, 1900.
PHOTO CREDIT: Torbjørn Ekrem

Stur and her team are now looking forward to getting these critters under the compound microscope. Using DNA analysis, they hope to identify which barcode clusters they match with, potentially revealing the nominal species of L. porosa. We’re sure that Sig Thor would be thrilled to learn that his identified species is still thriving, 120 years later.

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Local wildlife enthusiasts drive DNA barcode library building in the UK

Local wildlife enthusiasts drive DNA barcode library building in the UK

Local wildlife enthusiasts drive DNA barcode library building in the UK

Researchers in the UK are spearheading a number of high-profile initiatives designed to populate and fill gaps in the national DNA barcode library

Distribution of DNA barcode records for the United Kingdom.

IMAGE: BOLD Sytems 2020-03-10

Despite some notable achievements, such as a complete DNA barcode library for the native plants of Wales, the UK has lagged behind other European countries when it comes to growing its DNA barcode library. On BOLD there are 24,555 DNA barcode records for specimens collected in the UK (from 5,484 species) which is very similar to Austria (24,513 records, from 5,375 species), a landlocked country with roughly one third the land area and one seventh the human population. Germany leads Europe with 167,458 records from 14,805 species.

    The UK is working to catch-up through a number of high-profile initiatives designed to populate and fill gaps in the UK’s DNA barcode library and, in particular, bring BIOSCAN to UK insects.

    Distribution of DNA barcodes records for the United Kingdom.
    IMAGE: BOLD Systems from 2020-03-10

    The Darwin Tree of Life project is being led by the Wellcome Sanger Institute and involves a consortium of institutes, universities, museums, and agencies, including the Natural History Museum and Royal Botanical Gardens Kew. The project aims to deliver public DNA barcodes for 10,000 species by 2023 and ultimately sequence the genomes of all 66,000 species of plants, fungi, protozoa, and animals that are found in the UK.

    DEFRA (the Department for Environment, Food & Rural Affairs) has established a Centre of Excellence for Environmental Genomic Applications. This virtual centre recognises the absolute necessity of DNA barcode libraries to meet its aims of “applying genomics methods (eDNA and metabarcoding) to detect rare and invasive species, evaluate the effectiveness of conservation interventions, monitor the status and trends for key assemblages and taxa, and assess ecosystem health, functioning, and resilience”1.

    What is special about these initiatives is that they capitalize on the UK’s large community of local wildlife enthusiasts. A recent workshop organised by BugLife (the Invertebrate Conservation Trust) and Natural England (the UK government’s adviser for the natural environment in England) to examine “gaps” in BOLD for “key English invertebrates” brought together members of the Caddisfly Recording Scheme, Cranefly Recording Scheme, the British Dragonfly Society, the Amateur Entomologists’ Society, amongst others. The UK’s exceptional network of dedicated volunteer wildlife recorders already contribute thousands of records to taxon-focussed databases such as the UK Butterfly Monitoring Scheme (UKBMS), Local Record Centres, and through apps such as iSpot and iRecord which transmit data to the NBN (National Biodiversity Network) Atlas.

    Ainsdale Sand Dunes National Nature Reserve, one of the most important wildlife sites in England.
    PHOTO CREDIT: Gary Hedges

    The Darwin Tree of Life project kicked off last summer about 25 km north of Liverpool at Ainsdale Sand Dunes National Nature Reserve with a DNA Bioblitz attended by a team of local recorders including National Museums Liverpool entomologists. These local experts are passionate, driven and keen to contribute to DNA barcode libraries, but don’t necessarily have background knowledge in molecular biology or a basic skill set in “wet” lab techniques.

    Participants during the World Museum DNA Barcoding Workshop in February 2020.
    PHOTO CREDIT: Leanna Dixon

    To address this we recently ran a DNA barcoding workshop at World Museum Liverpool for eleven prominent local recorders connected with the Tanyptera Project. The Tanyptera Project is a seven-year initiative funded by the Tanyptera Trust to promote the study and conservation of insects and other invertebrates in the Lancashire and Cheshire region of Northwest England. To our knowledge this was one of the first DNA barcoding workshops run solely for non-professional scientists.

    Sphecodes ferruginatus female blood bee collected in Cheshire, England.
    PHOTO CREDIT: Chloe Aldridge

    The 1.5-day workshop covered the key steps in DNA barcoding from lab to BOLD2. Participants brought along their own invertebrates collected during recent local fieldwork and all successfully produced DNA barcodes for their specimens, which included springtails, bees, a cranefly, other flies, beetles, and spiders. The specimens have been vouchered into World Museum Liverpool’s National Entomology Collection which includes over 1 million specimens, and the sequences have been submitted to BOLD. One participant was able to confirm the first record of a Nationally Scarce blood bee in Cheshire – Sphecodes ferruginatus – raising interesting hypotheses about its potential host species.

    At National Museums Liverpool, together with the Tanyptera Project, we are committed to continue developing our DNA barcoding educational offering for local wildlife enthusiasts and supporting their work driving forward national initiatives to get more UK barcodes onto BOLD.

    References:

    1. Nelson M, Woodcock P, Maggs C (2018) Using eDNA and metabarcoding for nature conservation. Joint Nature Conservation Committee (JNCC 18 25). Available at http://data.jncc.gov.uk/data/99e1f69f-c438-439f-8401-dd8a6ce17320/JNCC18-25-Using-eDNA-and-Metabarcoding-for-Nature-Conservation.pdf

    2. Wilson JJ, Sing KW, Jaturas N (2019) DNA barcoding: Bioinformatics workflows for beginners. In Bioinformatics and Computational Biology. The A to Z of Bioinformatics. Ranganathan S, Nakai K, Gribskov M & Schönbach C, Eds. Elsevier Ltd., Oxford.

    Written by

    John-James Wilson

    John-James Wilson

    Vertebrate Zoology at World Museum, National Museums Liverpool, United Kingdom

    Leanna Dixon

    Leanna Dixon

    Tanyptera Project, National Museums Liverpool, United Kingdom

    Gary Hedges

    Gary Hedges

    Tanyptera Project, National Museums Liverpool, United Kingdom

    March 20, 2020

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    Assessing herbarium material with novel molecular techniques reveals a wealth of new data from old treasure troves

    Assessing herbarium material with novel molecular techniques reveals a wealth of new data from old treasure troves

    Assessing herbarium material with novel molecular techniques reveals a wealth of new data from old treasure troves

    Using large-scale genome skimming to build a resilient resource for the future
    Hamersley Range, Pilbara, Western Australia PHOTO CREDIT: Stephen van Leeuwen

    Herbaria are valuable sources of extensive curated plant material that are important reference specimens for plant identification. These plant materials are now also accessible to genetic studies because of advances in high-throughput, next-generation sequencing (NGS) methods.

    In our study, we conducted a large-scale applied assessment of one such NGS approach – genome skimming – and its ability to recover plastid and ribosomal genome sequences from a broad taxonomic sampling of herbarium material for the Western Australian flora. We sequenced 672 samples covering 21 families, 142 genera, and 530 named and proposed named species, and explored the impact of sample age, DNA concentration and quality, read depth and fragment length on plastid assembly error.

    We demonstrate that herbaria are a valuable source of plant material for building a comprehensive DNA sequence database which serves various applications from modernizing plant surveys to improving the resolution of plant phylogenies.

    Gastrolobium grandiflorum, Pilbara, Western Australia PHOTO CREDIT: Stephen van Leeuwen

    Genome skimming1 was effective at producing genomic information at large scale. Substantial sequence information on the chloroplast genome was obtained from 96.1% of samples, and complete or near-complete sequences of the nuclear ribosomal RNA gene repeat were obtained from 93.3% of samples.

    Eucalyptus kingsmillii, Pilbara, Western Australia PHOTO CREDIT: Stephen van Leeuwen
    Grevillea wickhamii, Pilbara, Western Australia PHOTO CREDIT: Stephen van Leeuwen

    We extracted sequences for plastid markers rbcL and matK – the core DNA barcode regions – from 96.4% and 93.3% of samples, respectively. Read quality and DNA fragment length had significant effects on sequencing outcomes and error correction of reads proved essential. Assembly problems were specific to certain taxa with low GC and high repeat content (e.g. Goodenia, Scaevola, Cyperus, Bulbostylis, Fimbristylis), suggesting the influence of biological rather than technical factors. The structure of related genomes was needed to guide the assembly of repeats that exceeded the read length. DNA-based matching proved highly effective and showed that the efficacy for species identification declined in the following order: total chloroplast DNA >> ribosomal DNA > matK >> rbcL.

    Ptilotus rotundifolius, Pilbara, Western Australia PHOTO CREDIT: Stephen van Leeuwen

    Our success is important as it demonstrates that herbaria can be used as a source of plant material for building a comprehensive DNA sequence database. These data form the basis of development of a molecular identification system for the Western Australian flora. This will enable identification of specimens throughout the year (e.g., non-flowering times) and for hard-to-identify species (e.g., those with constrained or reduced morphological characters) or for specimens where only fragments of non-diagnostic material are available. The availability of this technology will modernize plant surveys by reducing constraints on survey effort through moderating sampling timing restrictions and seasonal effects, as well as enabling rapid verifiable identification. It will also have practical applications in a wide range of ecological contexts using eDNA metabarcoding, such as gut and scat analysis of animals to determine dietary preferences of threatened species and livestock, and checking the integrity of seed collections for seed banking and use in land restoration/revegetation programs. Other potential uses of extensive plastid sequence data, beyond species identification, include improving the resolution of plant phylogenies and studies on the evolution of plastid genome function, including understanding adaptive changes.

    References:

    1. Straub S, Parks M, Weitemier K, Fishbein M, Cronn R, Liston A (2012) Navigating the tip of the genomic iceberg: Next-generation sequencing for plant systematics. American Journal of Botany 99(2), 349-364. https://dx.doi.org/10.3732/ajb.1100335

    For full details, please refer to the publication in BMC Plant Methods.

    Written by

    Paul Nevill

    Paul Nevill

    Curtin University, School of Molecular and Life Sciences, ARC Centre for Mine Site Restoration

    & Trace and Environmental DNA (TrEnD) Lab, Perth, Western Australia

    February 4, 2020
    https://doi.org/10.21083/ibol.v10i1.5934 

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