Improved methods for capture, extraction, and quantitative assay of environmental DNA from Asian bigheaded carp (Hypophthalmichthys spp.)

PLoS One. 2014 Dec 4;9(12):e114329. doi: 10.1371/journal.pone.0114329. eCollection 2014.

Abstract

Indirect, non-invasive detection of rare aquatic macrofauna using aqueous environmental DNA (eDNA) is a relatively new approach to population and biodiversity monitoring. As such, the sensitivity of monitoring results to different methods of eDNA capture, extraction, and detection is being investigated in many ecosystems and species. One of the first and largest conservation programs with eDNA-based monitoring as a central instrument focuses on Asian bigheaded carp (Hypophthalmichthys spp.), an invasive fish spreading toward the Laurentian Great Lakes. However, the standard eDNA methods of this program have not advanced since their development in 2010. We developed new, quantitative, and more cost-effective methods and tested them against the standard protocols. In laboratory testing, our new quantitative PCR (qPCR) assay for bigheaded carp eDNA was one to two orders of magnitude more sensitive than the existing endpoint PCR assays. When applied to eDNA samples from an experimental pond containing bigheaded carp, the qPCR assay produced a detection probability of 94.8% compared to 4.2% for the endpoint PCR assays. Also, the eDNA capture and extraction method we adapted from aquatic microbiology yielded five times more bigheaded carp eDNA from the experimental pond than the standard method, at a per sample cost over forty times lower. Our new, more sensitive assay provides a quantitative tool for eDNA-based monitoring of bigheaded carp, and the higher-yielding eDNA capture and extraction method we describe can be used for eDNA-based monitoring of any aquatic species.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Biodiversity
  • Carps / genetics*
  • DNA / genetics
  • DNA / isolation & purification*
  • Ecosystem*
  • Metagenomics*

Substances

  • DNA

Grants and funding

Funding was provided by the Great Lakes Restoration Initiative (FY10-S-T0240-O169-2) and the Great Lakes Fishery Trust (2011.1197). CRT was supported by National Science Foundation (NSF) IGERT grant award #0504495 to the GLOBES graduate training program at the University of Notre Dame. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.