Using environmental DNA methods to improve detectability in a hellbender (Cryptobranchus alleganiensis) monitoring program

Spear, Stephen F.; Groves, John D.; Williams, Lori A.; Waits, Lisette P.

doi:10.1016/j.biocon.2014.11.016

Cited by 225 publications

(270 citation statements)

References 24 publications

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“…In contrast, DNeasy Blood and Tissue kits (Qiagen) were commonly used to isolate and extract DNA from both marine and freshwater (e.g. Thomsen et al 2012a;Goldberg et al 2013;Spear et al 2015;Takahara et al 2015). DNeasy kits are available in a 96 well extraction format, making them more attractive for the analysis of a large number of samples.…”

Section: Summary Of Dna Extraction Methodsmentioning

confidence: 99%

“…Studies targeting macro-organisms, including fish and amphibians, most commonly extracted DNA using DNeasy Blood and Tissue kits (e.g. Takahara & Minimoto 2013;Laramie et al 2015;Spear et al 2015). In contrast, DNeasy PowerSoil ® and PowerMax ® kits were most commonly used in studies targeting the DNA of both prokaryotic and eukaryotic micro-organisms (e.g.…”

Section: Summary Of Dna Extraction Methodsmentioning

confidence: 99%

“…Most of these studies investigated the presence of fish, including Chinook salmon (Oncorhynchus tshawytscha; Laramie et al 2015), brook and bull trout (Salvelinus fontinalis and S. confluentus; Wilcox et al 2013), and slackwater darter (Etheostoma boschungi; Janosik & Johnston 2015), as well as amphibians such as the great crested newt (Triturus cristatus; Rees et al 2014;Biggs et al 2015), Idaho salamander (Dicamptodon aterrimus), Rocky Mountain tailed frog (Ascaphus montanus; Goldberg et al 2011;Pilliod et al 2013) and eastern hellbender (Cryptobranchus alleganiensis alleganiensis; Olson et al 2012;Spear et al 2015). With the exception of the bull trout and slackwater darter, which are respectively classified as vulnerable and endangered (IUCN Red List version 2.3), these organisms may be locally rare, but are otherwise classified as low risk in terms of their conservation status.…”

Section: Studies Conducted On Rare Organismsmentioning

confidence: 99%

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Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

Lear¹,

Dickie²,

Banks³

et al. 2018

107

112

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Advances in the sequencing of DNA extracted from media such as soil and water offer huge opportunities for biodiversity monitoring and assessment, particularly where the collection or identification of whole organisms is impractical. However, there are myriad methods for the extraction, storage, amplification and sequencing of DNA from environmental samples. To help overcome potential biases that may impede the effective comparison of biodiversity data collected by different researchers, we propose a standardised set of procedures for use on different taxa and sample media, largely based on recent trends in their use. Our recommendations describe important steps for sample pre-processing and include the use of (a) Qiagen DNeasy PowerSoil ® and PowerMax ® kits for extraction of DNA from soil, sediment, faeces and leaf litter; (b) DNeasy PowerSoil ® for extraction of DNA from plant tissue; (c) DNeasy Blood and Tissue kits for extraction of DNA from animal tissue; (d) DNeasy Blood and Tissue kits for extraction of DNA from macroorganisms in water and ice; and (e) DNeasy PowerWater ® kits for extraction of DNA from microorganisms in water and ice. Based on key parameters, including the specificity and inclusivity of the primers for the target sequence, we recommend the use of the following primer pairs to amplify DNA for analysis by Illumina MiSeq DNA sequencing: (a) 515f and 806RB to target bacterial 16S rRNA genes (including regions V3 and V4); (b) #3 and #5RC to target eukaryote 18S rRNA genes (including regions V7 and V8); (c) #3 and #5RC are also recommended for the routine analysis of protist community DNA; (d) ITS6F and ITS7R to target the chromistan ITS1 internal transcribed spacer region; (e) S2F and S3R to target the ITS2 internal transcribed spacer in terrestrial plants; (f) fITS7 or gITS7, and ITS4 to target the fungal ITS2 region; (g) NS31 and AML2 to target glomeromycota 18S rRNA genes; and (h) mICOIintF and jgHCO2198 to target cytochrome c oxidase subunit I (COI) genes in animals. More research is currently required to confirm primers suitable for the selective amplification of DNA from specific vertebrate taxa such as fish. Combined, these recommendations represent a framework for efficient, comprehensive and robust DNA-based investigations of biodiversity, applicable to most taxa and ecosystems. The adoption of standardised protocols for biodiversity assessment and monitoring using DNA extracted from environmental samples will enable more informative comparisons among datasets, generating significant benefits for ecological science and biosecurity applications.

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Section: Summary Of Dna Extraction Methodsmentioning

confidence: 99%

Section: Summary Of Dna Extraction Methodsmentioning

confidence: 99%

Section: Studies Conducted On Rare Organismsmentioning

confidence: 99%

See 1 more Smart Citation

Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

Lear¹,

Dickie²,

Banks³

et al. 2018

107

112

View full text Add to dashboard Cite

show abstract

“…For example, stream mtDNA concentration from a fully aquatic amphibian was significantly higher in breeding season than other months (Spear et al 2015). Increased eDNA production from gamete release and lower DNA degradation in intact gametes are likely explanations.…”

Section: Sample Collection and Preparationmentioning

confidence: 98%

The ecology of environmental DNA and implications for conservation genetics

2015

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Environmental DNA (eDNA) refers to the genetic material that can be extracted from bulk environmental samples such as soil, water, and even air. The rapidly expanding study of eDNA has generated unprecedented ability to detect species and conduct genetic analyses for conservation, management, and research, particularly in scenarios where collection of whole organisms is impractical or impossible. While the number of studies demonstrating successful eDNA detection has increased rapidly in recent years, less research has explored the ''ecology'' of eDNA-myriad interactions between extraorganismal genetic material and its environment-and its influence on eDNA detection, quantification, analysis, and application to conservation and research. Here, we outline a framework for understanding the ecology of eDNA, including the origin, state, transport, and fate of extraorganismal genetic material. Using this framework, we review and synthesize the findings of eDNA studies from diverse environments, taxa, and fields of study to highlight important concepts and knowledge gaps in eDNA study and application. Additionally, we identify frontiers of conservation-focused eDNA application where we see the most potential for growth, including the use of eDNA for estimating population size, population genetic and genomic analyses via eDNA, inclusion of other indicator biomolecules such as environmental RNA or proteins, automated sample collection and analysis, and consideration of an expanded array of creative environmental samples. We discuss how a more complete understanding of the ecology of eDNA is integral to advancing these frontiers and maximizing the potential of future eDNA applications in conservation and research.

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“…The test was performed in a series of six 1:10 dilutions from the initial 150 ng/μl of DNA template. Generally the concentrations from eDNA extractions are covered from the spectrum of these dilutions (Spear et al 2015). To ensure the results, the extractions and PCR were carried out in separate rooms.…”

Section: Primer Designmentioning

confidence: 99%

Development of a real-time PCR assay for the detection of the golden mussel ( Limnoperna fortunei , Mytilidae) in environmental samples

Pie

Ströher

Agostinis

et al. 2017

An. Acad. Bras. Ciênc.

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The golden mussel, Limnoperna fortunei, is among the most devastating invasive species in freshwater habitats worldwide, leading to severe environmental disturbances and economic losses. Therefore, management efforts would be greatly improved by methods that efficiently detect and quantify the abundance of the golden mussel in freshwater habitats, particularly in early stages of colonization. In this study, we describe a highly-sensitive real-time PCR assay targeting a 100-bp region of the COI mitochondrial gene of the golden mussel. The method was able to detect as little as 0.225 pg of target DNA. This assay represents an important contribution to surveillance methods, as well as to optimize field measures to contain and manage populations of the golden mussel in its introduced range.

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Using environmental DNA methods to improve detectability in a hellbender (Cryptobranchus alleganiensis) monitoring program

Cited by 225 publications

References 24 publications

Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

The ecology of environmental DNA and implications for conservation genetics

Development of a real-time PCR assay for the detection of the golden mussel ( Limnoperna fortunei , Mytilidae) in environmental samples

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