Conservation Genetics Resources

, Volume 10, Issue 2, pp 247–257 | Cite as

Evaluation of eDNA for groundwater invertebrate detection and monitoring: a case study with endangered Stygobromus (Amphipoda: Crangonyctidae)

  • Matthew L. NiemillerEmail author
  • Megan L. Porter
  • Jenna Keany
  • Heather Gilbert
  • Daniel W. Fong
  • David C. Culver
  • Christopher S. Hobson
  • K. Denise Kendall
  • Mark A. Davis
  • Steven J. Taylor
Methods and Resources Article


Effective conservation and management of biodiversity is limited by a lack of critical knowledge on species’ distributions and abundances. This problem is particularly exacerbated for species living in habitats that are exceptionally difficult to access or survey, such as groundwater habitats. Environmental DNA (eDNA) represents a rapid, noninvasive, and potentially cost-effective new tool for detection and monitoring of biodiversity that occur in such habitats. In this study, we investigated the utility of eDNA in detecting the federally endangered Hay’s Spring Amphipod Stygobromus hayi and a co-occurring common congener S. tenuis potomacus from unique groundwater-associated habitats—hypotelminorheic seepage springs—in the Washington, DC metro area. We developed taxon-specific primers and probes for each species to amplify Stygobromus DNA using qPCR. In silico and in vitro validation demonstrated specificity of each designed assay. Assays were then used to screen water samples collected from ten seepage springs. Stygobromus hayi was detected at four seepage springs, including one potential new locality, while S. t. potomacus was detected at four springs, two of which were new localities. This study is the first to our knowledge to successfully employ an eDNA approach to detect rare or threatened invertebrates from subterranean ecosystems. Our study highlights challenges of employing an eDNA approach for the detection and monitoring of invertebrates in groundwater habitats that are difficult to study, including accounting for PCR inhibition and the potential for cryptic genetic diversity.


District of Columbia Hypotelminorheic seep, real-time PCR Short-range endemic Stygobromus hayi Stygobromus tenuis potomacus Subterranean 



This study was supported by a grant from the Friends of the Capital Crescent Trail and the town of Chevy Chase, Maryland, and the Illinois Natural History Survey. Sampling of S. hayi was authorized by a subpermit under U.S. Fish and Wildlife Service Regional Endangered Species Recovery Permit TE-697823. This research was also authorized under National Park Service permit no. ROCR-2016-SCI-0014. We thank Jonathan Cybulski and Emily Petersen for assistance with field collections of water samples. We thank Michael Slay for providing specimens of non-target species for DNA analyses.

Supplementary material

12686_2017_785_MOESM1_ESM.docx (21 kb)
Supplementary material 1 (DOCX 20 KB)


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Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  • Matthew L. Niemiller
    • 1
    Email author
  • Megan L. Porter
    • 2
  • Jenna Keany
    • 3
  • Heather Gilbert
    • 3
  • Daniel W. Fong
    • 4
  • David C. Culver
    • 3
  • Christopher S. Hobson
    • 5
  • K. Denise Kendall
    • 6
  • Mark A. Davis
    • 1
  • Steven J. Taylor
    • 1
  1. 1.Illinois Natural History Survey, Prairie Research InstituteUniversity of Illinois at Urbana-ChampaignChampaignUSA
  2. 2.Department of BiologyUniversity of Hawai’i at MānoaHonoluluUSA
  3. 3.Department of Environmental ScienceAmerican UniversityWashingtonUSA
  4. 4.Department of BiologyAmerican UniversityWashingtonUSA
  5. 5.Virginia Natural Heritage ProgramVirginia Department of Conservation and RecreationRichmondUSA
  6. 6.School of Integrative BiologyUniversity of Illinois Urbana-ChampaignUrbanaUSA

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