Sampling and Extraction Method for Environmental DNA (eDNA) in Freshwater Ecosystems

Abstract Potamodromous fish are considered important indicators of habitat connectivity in freshwater ecosystems, but they are globally threatened by anthropogenic impacts. Hence, non-invasive techniques are necessary for monitoring during spawning migrations. The use of environmental DNA (eDNA) potentially facilitates these efforts, albeit quantitative examinations of spawning migrations remain so far mostly uncharted. Here, we investigated spawning migrations of Danube bleak, Alburnus mento, and Vimba bream, Vimba vimba, and found a strong correlation between daily visual fish counts and downstream eDNA signals obtained from filtered water samples analysed with digital PCR and end-point PCR coupled with capillary electrophoresis. By accounting for daily discharge fluctuations, it was possible to predict eDNA signal strength from the number of migrating fish: first, the whole spawning reach was taken into account. Second, the model was validated using eDNA signals and fish counts obtained from the upper half of the examined river stretch. Consequently, fish counts and their day-to-day changes could be described via an eDNA-based time series model for the whole migration period. Our findings highlight the capability of eDNA beyond delivering simple presence/absence data towards efficient and informative monitoring of highly dynamic aquatic processes such as spawning migrations of potamodromous fish species.

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FIRST ENVIRONMENTAL DNA (EDNA) RECORD OF CENTRAL AMAZON IN A FLOODPLAIN LAKE: EXTRACTION METHOD SELECTION AND VALIDATION / PRIMEIRO REGISTRO DE DNA AMBIENTAL (EDNA) DA AMAZÔNIA CENTRAL EM UM LAGO DE VÁRZEA: SELEÇÃO E VALIDAÇÃO DE MÉTODO DE EXTRAÇÃO

Brazilian Journal of Development ◽

10.34117/bjdv6n11-254 ◽

2020 ◽

Vol 6 (11) ◽

pp. 87606-87621

Author(s):

Danniel Rocha Bevilaqua ◽

Sabrina Araújo de Melo ◽

Carlos Edwar de Carvalho Freitas ◽

Ana Caroline Viana da Silva ◽

Jacqueline da Silva Batista

Keyword(s):

Extraction Method ◽

Environmental Dna ◽

Floodplain Lake ◽

Central Amazon ◽

Method Selection

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Environmental DNA allows upscaling spatial patterns of biodiversity in freshwater ecosystems

Nature Communications ◽

10.1038/s41467-020-17337-8 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 3

Author(s):

Luca Carraro ◽

Elvira Mächler ◽

Remo Wüthrich ◽

Florian Altermatt

Keyword(s):

Spatial Patterns ◽

Environmental Dna ◽

Freshwater Ecosystems

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Effective removal of the American bullfrog (Lithobates catesbeianus) on a landscape level: long term monitoring and removal efforts in Yosemite Valley, Yosemite National Park

Biological Invasions ◽

10.1007/s10530-019-02116-4 ◽

2019 ◽

Vol 22 (2) ◽

pp. 617-626 ◽

Cited By ~ 1

Author(s):

Colleen Kamoroff ◽

Ninette Daniele ◽

Robert L. Grasso ◽

Rebecca Rising ◽

Travis Espinoza ◽

...

Keyword(s):

Invasive Species ◽

National Parks ◽

National Park ◽

Environmental Dna ◽

Freshwater Ecosystems ◽

Yosemite National Park ◽

American Bullfrog ◽

Yosemite Valley ◽

Park Service ◽

Landscape Level

Abstract Invasive alien species are a major threat to freshwater ecosystems, and American bullfrogs are among the world’s 100 most prominent aquatic invasive species causing negative direct and indirect effect on native aquatic fauna worldwide. Bullfrogs were intentionally introduced into Yosemite Valley, Yosemite National Park in the 1950s where they became well established in the subsequent years. Starting in 2005, the National Park Service (NPS) began bullfrog removal, targeting various life stages using hand, net, and spear techniques. Starting in 2015, the NPS conducted environmental DNA (eDNA) surveys and deployed audio recordings devices to ensure adequate detection of bullfrogs. During the first year of cencerted effort in the Valley in 2005, the NPS removed 86% of all recorded bullfrog. The subsequent decade was spent searching for individuals with lower return on effort. In 2012, the NPS removed the last observed signs of bullfrog breeding, and the last observed bullfrog in 2019. Following removal of the breeding bullfrog population, the NPS began restoration projects for species of special concern. The NPS introduced the federally threatened California red-legged frogs (Rana draytonii) into Yosemite Valley beginning in 2016. This is the first published successful eradication of bullfrogs on a landscape level. National Parks and Monuments often provide refuges for imperiled wildlife and should be managed to remove invasive species. Our work highlights effective bullfrog removal is obtainable and can lead to local recovery of endangered species.

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Ethanol eDNA Reveals Unique Community Composition of Aquatic Macroinvertebrates Compared to Bulk Tissue Metabarcoding in a Biomonitoring Sampling Scheme

Diversity ◽

10.3390/d13010034 ◽

2021 ◽

Vol 13 (1) ◽

pp. 34

Author(s):

Sadhna Fiona Persaud ◽

Karl Cottenie ◽

Jennifer Erin Gleason

Keyword(s):

Land Use ◽

Community Composition ◽

Agricultural Land ◽

Environmental Dna ◽

Ecological Condition ◽

Freshwater Ecosystems ◽

Aquatic Macroinvertebrates ◽

Biological Communities ◽

Non Destructive ◽

Bulk Tissue

Freshwater ecosystems provide essential ecosystem services and support biodiversity; however, their water quality and biological communities are influenced by adjacent agricultural land use. Aquatic macroinvertebrates are commonly used as bioindicators of stream conditions in freshwater biomonitoring programs. Sorting benthic samples for molecular identification is a time-consuming process, and this study investigates the potential of ethanol-collected environmental DNA (eDNA) for metabarcoding macroinvertebrates, especially for common bioindicator groups. The objective of this study was to compare macroinvertebrate composition between paired bulk tissue and ethanol eDNA samples, as eDNA could provide a less time-consuming and non-destructive method of sampling macroinvertebrates. We collected benthic samples from streams in Ontario, Canada, and found that community composition varied greatly between sampling methods and that few taxa were shared between paired tissue and ethanol samples, suggesting that ethanol eDNA is not an acceptable substitute. It is unclear why we did not detect all the organisms that were preserved in the ethanol, or the origin of the DNA we did detect. Furthermore, we also detected no difference in community composition for bioindicator taxa due to surrounding land use or water chemistry, suggesting sites were similar in ecological condition.

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Filtration extraction method using microfluidic channel for measuring environmental DNA

10.22541/au.163854179.99663512/v1 ◽

2021 ◽

Author(s):

Takashi Fukuzawa ◽

Yuichi Kameda ◽

Hisao Nagata ◽

Naofumi Nishizawa ◽

Hideyuki Doi

Keyword(s):

Water Sample ◽

River Water ◽

Dna Extraction ◽

Water Samples ◽

Extraction Method ◽

Microfluidic Channel ◽

Pcr Amplification ◽

Environmental Dna ◽

Laboratory Analysis ◽

River Water Samples

The environmental DNA (eDNA) method, which is widely applied for biomonitoring, is limited to laboratory analysis and processing. In this study, we developed a filtration/extraction component using a microfluidic channel, Biryu-Chip (BC), and a filtration/extraction method, BC method, to minimize the volume of the sample necessary for DNA extraction and subsequent PCR amplification. We tested the performance of the BC method and compared it with the Sterivex filtration/extraction method using aquarium and river water samples. We observed that using the BC method, the same concentration of the extracted DNA was obtained with 1/20–1/40 of the filtration volume of the Sterivex method, suggesting that the BC method can be widely used for eDNA measurement. In addition, we could perform on-site measurements of eDNA within 30 min using a mobile PCR device. Using the BC method, filtration and extraction could be performed easily and quickly. The PCR results obtained by the BC method were similar to those obtained by the Sterivex method. The BC method required fewer steps and therefore, the risk of DNA contamination could be reduced. When combined with a mobile PCR, the BC method can be applied to easily detect eDNA within 30 min from a few 10 mL of the water sample, even on-site.

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Using environmental DNA to monitor the reintroduction success of the Rhine sculpin (Cottus rhenanus) in a restored stream

10.7287/peerj.preprints.27574v1 ◽

2019 ◽

Author(s):

Christopher A Hempel ◽

Bianca Peinert ◽

Arne J Beermann ◽

Vasco Elbrecht ◽

Jan-Niklas Macher ◽

...

Keyword(s):

False Negative ◽

Ecological Status ◽

Environmental Dna ◽

Freshwater Ecosystems ◽

Freshwater Ecosystem ◽

Ecosystem Monitoring ◽

Modern Approach ◽

Negative Results ◽

False Negative Results ◽

Good Ecological Status

As a consequence of the strong human impact on freshwater ecosystems, restoration measures are increasingly applied to restore and maintain their good ecological status. The ecological status of freshwaters can be inferred by assessing the presence of indicator species, such as the Rhine sculpin (Cottus rhenanus). However, traditional methods of monitoring fish, such as electrofishing, are often challenging and invasive. To augment or even replace the traditional fish monitoring approach, the analysis of environmental DNA (eDNA) has recently been proposed as an alternative, sensitive approach. The present study employed this modern approach to monitor the Rhine sculpin, a species that has been reintroduced into a recently restored stream within the Emscher catchment in Germany, in order to validate the success of the restorations. We monitored the dispersal of the Rhine sculpin using replicated 12S end-point PCR eDNA surveillance at a fine spatial and temporal scale to investigate the applicability of analyzing eDNA for freshwater ecosystem monitoring. We also performed traditional electrofishing in one instance to compare visual and eDNA-based assessments. We could track the dispersal of the Rhine sculpin and showed a higher dispersal potential of the species than we assumed. Furthermore, the eDNA analysis showed higher sensitivity for detecting the species than traditional electrofishing, although false negative results occurred at early reintroduction stages. Our results show that analyzing eDNA is capable of validating and tracking ecological reintroductions and contribute to the assessment and modelling of ecological status of streams.

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Filtration extraction method using microfluidic channel for measuring environmental DNA

10.1101/2021.11.18.468931 ◽

2021 ◽

Author(s):

Fukuzawa Takashi ◽

Yuichi Kameda ◽

Hisao Nagata ◽

Naofumi Nishizawa ◽

Hideyuki Doi

Keyword(s):

Water Sample ◽

River Water ◽

Dna Extraction ◽

Water Samples ◽

Extraction Method ◽

Microfluidic Channel ◽

Pcr Amplification ◽

Environmental Dna ◽

Laboratory Analysis ◽

River Water Samples

The environmental DNA (eDNA) method, which is widely applied for biomonitoring, is limited to laboratory analysis and processing. In this study, we developed a filtration/extraction component using a microfluidic channel, Biryu-Chip (BC), and a filtration/extraction method, BC method, to minimize the volume of the sample necessary for DNA extraction and subsequent PCR amplification. We tested the performance of the BC method and compared it with the Sterivex filtration/extraction method using aquarium and river water samples. We observed that using the BC method, the same concentration of the extracted DNA was obtained with 1/20-1/40 of the filtration volume of the Sterivex method, suggesting that the BC method can be widely used for eDNA measurement. In addition, we could perform on-site measurements of eDNA within 30 min using a mobile PCR device. Using the BC method, filtration and extraction could be performed easily and quickly. The PCR results obtained by the BC method were similar to those obtained by the Sterivex method. However, the BC method required fewer steps and therefore, the risk of DNA contamination could be reduced. When combined with a mobile PCR, the BC method can be applied to easily detect eDNA within 30 min from 10 mL of the water sample, even on-site.

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Effects of Microbial Activity and Environmental Parameters on the Degradation of Extracellular Environmental DNA from a Eutrophic Lake

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph16183339 ◽

2019 ◽

Vol 16 (18) ◽

pp. 3339 ◽

Cited By ~ 6

Author(s):

Nur Syahidah Zulkefli ◽

Keon-Hee Kim ◽

Soon-Jin Hwang

Keyword(s):

Microbial Activity ◽

Degradation Rate ◽

Abiotic Factors ◽

Environmental Parameters ◽

Environmental Dna ◽

Eutrophic Lake ◽

Freshwater Ecosystems ◽

Extracellular Dna ◽

Biotic And Abiotic Factors ◽

Positive Effects

Extracellular DNA (exDNA) pool in aquatic environments is a valuable source for biomonitoring and bioassessment. However, degradation under particular environmental conditions can hamper exDNA detectability over time. In this study, we analyzed how different biotic and abiotic factors affect the degradation rate of extracellular environmental DNA using 16S rDNA sequences extracted from the sediment of a eutrophic lake and Anabaena variabilis cultured in the laboratory. We exposed the extracted exDNA to different levels of temperature, light, pH, and bacterial activity, and quantitatively analyzed the concentration of exDNA during 4 days. The solution containing bacteria for microbial activity treatment was obtained from the lake sediment using four consecutive steps of filtration; two mesh filters (100 μm and 60 μm mesh) and two glass fiber filters (2.7 μm and 1.2 μm pore-sized). We found that temperature individually and in combination with bacterial abundance had significant positive effects on the degradation of exDNA. The highest degradation rate was observed in samples exposed to high microbial activity, where exDNA was completely degraded within 1 day at a rate of 3.27 day−1. Light intensity and pH had no significant effects on degradation rate of exDNA. Our results indicate that degradation of exDNA in freshwater ecosystems is driven by the combination of both biotic and abiotic factors and it may occur very fast under particular conditions.

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