PS 13-145 - Snail slime in real time: qPCR detection of environmental DNA from apple snails

Monday, August 7, 2017
Exhibit Hall, Oregon Convention Center
Madison E. S. Granier, Southwestern University, Georgetown, TX, Matthew A. Barnes, Natural Resources Management, Texas Tech University, Lubbock, TX and Romi L. Burks, Biology, Southwestern University, Georgetown, TX

Environmental DNA (eDNA) represents extra-organismal DNA that individuals release into their environment such as sloughed cells or bodily fluids. Conservation efforts have recently documented more sensitive, cost effective results from eDNA rather than traditional survey methods. Improved understanding of the origin, state, transport, and fate of eDNA provides insight into its utility and limitations, thereby improving confidence in conservation studies. We focused on the freshwater apple snail (Pomacea maculata) eDNA production and investigated how abiotic factors influenced eDNA degradation.

We placed adult snails in a two-by-two mesocosm design (N=5) with warm and cool temperature treatments crossed with freshwater and salt treatments (6 ppt). DNA accumulated over 72 hours, at which point snails were removed and DNA degradation occurred over the next 72 hours. We collected water samples (250 mL) at 12 time points and then ran material through 1.2 μm Isopore membrane filters to retain eDNA (feces, slime, tissue, etc). We extracted the material from the filters with a cetrimonium bromide-chloroform method to obtain total genomic DNA for use in quantitative PCR (qPCR). We created a standard curve using total genomic DNA from an individual P. maculata to determine the number of eDNA copies per filtered water sample.


Using a newly developed specific primer, we successfully amplified eDNA from all samples at peak eDNA accumulation. At this time point, we saw an increase in eDNA production at higher salt and warmer temperature, possibly due to the active nature of the snails or stabilization of eDNA by salt. Similar amounts of eDNA accumulated in the control (no salt, no heat = 0.77 ± 0.56 as 1 SE) and the single treatments (salt or heat only; 0.60 ± 0.98, 0.69 ± 1.1, respectively). Only combination of salt and heat resulted in an observable difference in eDNA production.

Conservation work increasingly employs eDNA, but we know relatively little about the ‘ecology of eDNA,’ prompting the need for basic science about this applied approach. Aquatic eDNA studies commonly use amphibians and fishes. As an alternative model for developing eDNA approaches, freshwater snails represent a highly diverse invertebrate group with broad size ranges. Overall, our research contributes to the eDNA literature by providing the possibility of comparison between similarly sized invertebrates and vertebrates and examining both trends in accumulation and degradation. Future analyses will include our full time series to add more insight into the persistence of eDNA.