Environmental DNA sequencing will allow scientists to explore the diversity of Amazonian pools without catching fish

A view of the river where environmental DNA was collected for comparison with captured specimens. The researchers aim to deposit in public databases sequences of genetic material representing all orders and families and most genera of Amazonian fish. Credit: David de Santana

A scientific expedition to the Javari River basin on the border between Brazil, Colombia and Peru has shown that the use of environmental DNA sequencing is feasible to study the diversity of fish in the Amazon. The eDNA method consists of extracting DNA molecules present in water samples and identifying the species to which they belong by means of genetic markers.

A research article published in the journal Scientific reports also discusses the limits of the technique for the study of environments as diverse as the Amazon.

“We must continue to capture and identify animals by traditional methods in order to create libraries of genetic material. They will serve as a benchmark to compare everything in the water samples. As the technique progresses, in a few years we may be able to know all the fish species present in one place without catching them, ”said Carlos David de Santana, associate researcher at the National Museum of Natural History. from the Smithsonian Institution in the United States and first author of the article.

The study was part of the “Diversity and Evolution of Gymnotiformes” project, supported by FAPESP and led by Naércio Menezes, professor at the Museum of Zoology of the University of São Paulo (MZ-USP) in Brazil. “Extraction of DNA from water samples creates very favorable expectations for environmental protection, as the usual methods of collecting samples from aquatic animals include the use of nets and other fishing gear having a negative impact, ”said Menezes, co-author of the article.

The group of researchers spent 18 days on the Javari River, collecting water samples at three of the 46 locations where they collected fish specimens. The number of species represented reached a surprising total of 443, and 60 were new to science.

At the sites where water was collected for eDNA analysis, 201 species were captured using traditional methods. However, only 58, or 26% of the total, were accurately identified to the species level from eDNA analysis.

“One possible explanation is the lack of reference genetic material in databases that can be used for comparison. Additionally, many species in these locations are entirely new and have never been previously identified at the. using conventional techniques, ”said Gislene Torrente-Vilara, a professor at the Institute of Marine Sciences at the Federal University of São Paulo (IMAR-UNIFESP) in Santos and co-author of the article.

Torrente-Vilara led the expedition as part of the Amazon Fish Project, an international collaboration supported by FAPESP that has provided insight into the distribution of fish species in the region.

DNA in 100 milliliters of water

To sequence eDNA, the researchers first collected 100 milliliter surface water samples from three predetermined locations. The samples were filtered and mixed with a solution to prevent degradation of eDNA.

The 12S mitochondrial RNA gene is the most widely used marker in the world to identify fish species from eDNA. To find this small piece of genetic code in the water samples, the researchers used DNA extraction kits designed for blood and tissue analysis. The droppings and animal parts present in the water can be “captured” by the technique.

However, 12S is a slowly evolving piece of genetic code and might not be sufficient to identify all fish specimens to the species level, as many species in the Amazon have diverged millions of times ago. years (recently in terms of evolution).

For the same reason, eDNA sequencing only provided an accurate portrait of the orders represented in the samples. He also differentiated the communities that live in large rivers from those that inhabit small streams deep in the forest (igarapés).

The Amazon basin is home to the greatest diversity of freshwater fish in the world, with more than 2,700 scientifically described species belonging to 18 orders, 60 families and more than 500 genera.

“Even with an adequate library, it would be very difficult to identify everything to the species level with just this marker,” Santana said. “Two electric eels which have diverged recently, for example Electrophorus voltai and E. electricus, might appear to be a single species.”

The researchers expect the technique to improve enough in the coming years that more than one DNA sample will be sequenced simultaneously, so that species can be precisely defined. Until then, it will be necessary to create genetic reference libraries, and Santana plans to catalog the genetic material of at least all fish families and most fish genera in the Amazon.

In this context, the authors emphasize that natural history museums are ideal institutions for creating genetic reference libraries and storing environmental samples. As technologies advance, deposited material can be sequenced with increasing precision.

“Museums keep samples of biodiversity for a very long time and make them available for study by future generations. To keep genetic material viable for such long periods of time, however, they must set up cryogenic facilities, or significantly expand those at their disposal, by reorganizing their physical layout and acquiring massive amounts of equipment, such as freezers. at ultra-low temperature and liquid nitrogen tanks, ”said Aléssio Datovo, another co-author of the article and curator of fish at MZ-USP, the first institution in Brazil to collect eDNA samples.

The technique can also be used for environmental monitoring and even to involve schools and riparian communities in conservation initiatives through citizen science programs.


Using DNA to research fish species


More information:
C. David de Santana et al, The critical role of natural history museums in advancing eDNA for biodiversity studies: a case study with Amazonian fish, Scientific reports (2021). DOI: 10.1038 / s41598-021-97128-3

Quote: Environmental DNA sequencing will allow scientists to explore the diversity of Amazonian pools without catching fish (2021, 23 November) Retrieved 23 November 2021 from https://phys.org/news/2021-11 -environmental-dna-sequencing-enable-scientists. html

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