Insects don’t talk, but new DNA-based technologies are helping to tell their stories

Insects don’t talk, but new DNA-based technologies are helping to tell their stories

Guilherme Oliveira

Researchers unlock clues about vertebrate diversity by examining arthropod samples
Malaise trap deployed in Carajás National Forest in Brazil. PHOTO CREDIT: Christina Lynggaard

The study of arthropod diversity often uses primers solely targeting arthropod DNA, ignoring those originating from non-arthropod sources. Yet, some arthropods (e.g. sand flies, blowflies, mosquitoes) feed on vertebrates and vertebrate remains. By specifically targeting these vertebrate-feeding arthropods, invertebrate derived DNA (iDNA) can be used to understand vertebrate diversity as well. Our study has found that by using samples collected for arthropod diversity research, we can tap into a rich information source of vertebrate diversity.

Arthropods are highly abundant and provide vital ecosystem services such as pollination, decomposition of organic matter, and they serve as a food source for other animals. Because of this, hundreds to thousands of arthropod samples are collected globally every year for monitoring programs, conservation efforts, and ecosystem assessments.

However, identifying arthropod species based on physical characteristics is difficult, requiring specific expertise and a significant amount of time. Due to this, molecular analyses, such as DNA metabarcoding, are being used to identify the arthropods present in bulk samples, speeding-up the otherwise time-demanding identification process by screening the contents of hundreds of samples simultaneously.

Bulk arthropod sample

PHOTO CREDIT: Christina Lynggaard

In this study, bulk arthropod samples were collected using pitfall and Malaise traps deployed at the Carajás National Forest in Brazil and in the Udzungwa Mountains in Tanzania. Our aim was to study the arthropod diversity in these areas by using metabarcoding primers targeting arthropod DNA. Nevertheless, we conducted an additional analysis targeting vertebrate DNA remains in the bulk samples by using metabarcoding vertebrate primers.

We were able to detect vertebrate DNA in 19 per cent of our 265 analyzed samples, identifying more than 30 vertebrate taxa including mammals, amphibians and birds. For example, we found South American tapir (Tapirus terrestris), chirinda screeching frog (Arthroleptis xenodactyloides), velvety free-tailed bat (Molossus molossus), Zanzibar bushbaby (Paragalago zanzibaricus) and honeyguide greenbul (Baeopogon indicator). The presence of some species was further confirmed through visual observations during the sample collections.

Some cases represent the first time those vertebrate species have been detected in the area. This could indicate that we are detecting vertebrates that live in different areas to where the arthropods were collected or that with this method it is possible to detect local animals that are difficult to observe.

Tapir footprint found in one of the Brazilian study sites during bulk arthropod sample collection confirmed the presence of some of the detected vertebrate taxa PHOTO CREDIT: Christina Lynggaard

Bushbaby (Paragalago zanzibaricus) found in the Udzungwa Mountains in the Tanzanian study site during bulk arthropod sample collection confirmed the presence of some of the detected vertebrate taxa

PHOTO CREDIT: mnielsenphotography

The sensitivity of new DNA-based technologies is changing the way we explore and understand the relationships between species. Approaches like DNA metabarcoding can not only tell us about the different types of insects in our environment, but also what they eat and where they have been. Our study is the first to not target a specific invertebrate group to detect vertebrate DNA but instead to use bulk arthropod samples. We encourage others to consider bulk samples during environmental monitoring not only as a source for arthropod diversity information, but one for vertebrate diversity as well.

This study is published in the eDNA Journal.

Written by

Christina Lynggaard

Christina Lynggaard

The GLOBE Institute, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark

Martin Nielsen

Martin Nielsen

The GLOBE Institute, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark

Luisa Santos-Bay

Luisa Santos-Bay

The GLOBE Institute, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark

NTNU University Museum, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
Markus Gastauer

Markus Gastauer

Instituto Tecnológico Vale, Belém, Pará, Brazil

Guilherme Oliveira

Guilherme Oliveira

Instituto Tecnológico Vale, Belém, Pará, Brazil

Kristine Bohmann

Kristine Bohmann

The GLOBE Institute, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark

University of East Anglia, Norwich Research Park, Norwich, UK

October 17, 2019
https://doi.org/10.21083/ibol.v9i1.5727

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DNA Barcoding and Genomics in the Megadiverse Amazon Altitude Fields

DNA Barcoding and Genomics in the Megadiverse Amazon Altitude Fields

DNA BARCODING AND GENOMICS IN THE MEGADIVERSE AMAZON ALTITUDE FIELDS

Scientists are contributing to the most profound molecular representation of biodiversity in any Brazilian environment.

The entrance of a ferruginous cave at the Bocaina mountain in the Carajás region, Pará State, Brazil.

PHOTO CREDIT: João Marcos Rosa.

Industrial activities in the Brazilian Amazon are highly regulated by governmental agencies. However, the lack of knowledge about megadiverse areas is a problem for the establishment of best conservation practices; this is the case for mining operations in the eastern Amazonian Carajás, a region comprised of a mosaic of national parks, indigenous peoples’ conservation areas, and nature reserves. Of particular interest are the ferruginous altitude fields known as the Canga. Our floristic survey described the presence of 1,094 species from just over 200 previously known1. The lack of biodiversity data is even more significant for the ferruginous caves where only around 10 invertebrate species are identified to the species level. Unfortunately, this is typical for the Amazon basin.

To provide reliable scientific data that contributes to the implementation of best conservation practices, Instituto Tecnológico Vale is developing DNA barcode reference libraries for the flora, cave invertebrates, and bats of the region, and providing deeper genomic references for species that are endangered or difficult to identify. To achieve this goal, we established the necessary infrastructure to conduct DNA sequencing using Sanger, Illumina, and PacBio technologies, coupled with high-performance computing, artificial intelligence algorithms, and highly trained personnel.

 

Botanists collecting samples in a temporary lake in ferruginous altitude fields in the Carajás region, Pará State, Brazil.
PHOTO CREDIT: João Marcos Rosa

To date, over 8,575 barcodes for 3,548 specimens of plants and invertebrates have been produced, while a large number of species remain to be identified by morphological attributes. Morphological specimen determination is conducted by in-house specialists, as well as by an extensive network of specialists in universities and museums across Brazil and abroad.

For this purpose, nuclear and chloroplast or mitochondrial markers as well as low coverage to whole genome sequencing or restriction site-associated DNA sequencing (RADSeq) are being employed to unravel the vast genetic diversity of the biota of Carajás2-4. For several endemic plants, such as species of Asteraceae, Melastomataceae5, Convolvulaceae6, and Isoetaceae2, diversity analyses, based on next-generation sequencing, aim to characterize the genetic variability among and within populations, as well as the identification of markers under selective pressure. These methods also contribute to the understanding of population structure and the process of gene flow between populations affected by natural factors and industrial operations. Models of environmental distribution, including parameters sensitive to climate change, were determined for several taxonomic groups, including plants and bats7.

Specimens of the pinheirinho-da-canga (Paepalanthus fasciculoides) highly adapted to inhabit the canga (ancient ferruginous rock outcrops) at the altitude fields in the Carajás region, Pará State, Brazil.
PHOTO CREDIT: João Marcos Rosa
We are also establishing eDNA methods, as well as metagenomics and metaproteomics data for environmental monitoring of ferruginous fields phytophysiognomies, areas under rehabilitation processes, and caves8. Together these data constitute the most profound molecular representation of any environment in Brazil. We have contributed a total of 3,072 specimens to the Barcode of Life Data System (BOLD) comprising 398 genera (291 new) in addition to the 408 different genera collected through the national effort for angiosperms in Brazil. We have also provided 571 cave fauna specimens.

It is important to highlight that all of these data generated are being provided to the public and its use will be critical to the conservation of such a unique collection of species.

References:

1. Brazil Flora Group (2018) Growing knowledge: an overview of Seed Plant diversity in Brazil. Rodriguésia 66(4): 1085–1113. http://dx.doi.org/10.1590/2175-7860201566411

2. Nunes GL, Oliveira RRM, Guimarães JTF, Giulietti AM, Caldeira C, Vasconcelos S, et al., (2018) Quillworts from the Amazon: A multidisciplinary populational study on Isoetes serracarajensis and Isoetes cangae. PLoS ONE 13(8): e0201417. https://doi.org/10.1371/journal.pone.0201417

3. Ramalho AJ, Zappi DC, Nunes GL, Watanabe MTC, Vasconcelos S, Dias MC, Jaffé R, Prous X, Giannini TC, Oliveira G and Giulietti AM (2018) Blind testing: DNA barcoding sheds light upon the identity of plant fragments as a subsidy for cave conservation. Frontiers in Plant Science 9:1052. https://doi.org/10.3389/fpls.2018.01052

4. Oliveira RRMO, Vasconcelos S, Pires ES, Pietrobon T, Prous X and Oliveira G (2019) Complete mitochondrial genomes of three troglophile cave spiders (Mesabolivar, pholcidae), Mitochondrial DNA Part B 4(1): 251–252. https://doi.org/10.1080/23802359.2018.1547139

5. Carvalho CdS, Lanes ECM, Silva AR, Caldeira CF, Carvalho-Filho N, Gastauer M, Imperatriz-Fonseca VL, Nascimento W, Oliveira G, Siqueira JO, Viana PL, Jaffe R (2019) Habitat loss does not always entail negative genetic consequences. bioRxiv 528430. https://doi.org/10.1101/528430

6. Lanes ÉC, Pope NS, Alves R, Carvalho Filho NM, Giannini TC, Giulietti AM, Imperatriz-Fonseca VL, Monteiro W, Oliveira G, Silva AR, Siqueira JO, Souza-Filho PW, Vasconcelos S and Jaffé R (2018) Landscape genomic conservation assessment of a narrow-endemic and a widespread morning glory from Amazonian Savannas. Frontiers in Plant Science 9:532. https://doi.org/10.3389/fpls.2018.00532

7. Costa WF, Ribeiro M, Saraiva AM, Imperatriz-Fonseca VL, Giannini TC (2018) Bat diversity in Carajás National Forest (Eastern Amazon) and potential impacts on ecosystem services under climate change. Biological Conservation 218: 200–210. https://doi.org/10.1016/j.biocon.2017.12.034

8. Gastauer M, Vero MPO, de Souza KP, Pires ES, Alves R, Caldeira CF, Ramos SJ, Oliveira G (2019) A metagenomic survey of soil microbial communities along a rehabilitation chronosequence after iron ore mining. Scientific Data 6:190008. https://doi.org/10.1038/sdata.2019.8

Written by

Guilherme Oliveira

Guilherme Oliveira

Environmental Genomics Group, Instituto Tecnológico Vale, Belém, Brazil

Gisele Nunes Lopes

Gisele Nunes Lopes

Environmental Genomics Group, Instituto Tecnológico Vale, Belém, Brazil

Rafael Valadares

Rafael Valadares

Environmental Genomics Group, Instituto Tecnológico Vale, Belém, Brazil

Ronnie Alves

Ronnie Alves

Environmental Genomics Group, Instituto Tecnológico Vale, Belém, Brazil

Santelmo Vasconcelos

Santelmo Vasconcelos

Environmental Genomics Group, Instituto Tecnológico Vale, Belém, Brazil

April 7, 2019
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https://doi.org/10.21083/ibol.v9i1.5498

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