No filters, no fridges: a method for preservation of water samples for eDNA analysis.
BACKGROUND
Advancements in the detection of environmental DNA (eDNA) for detecting species of interest will likely allow for expanded use of these techniques in the field. One obstacle that continues to hinder applications in the field is the requirement of a cold chain of storage for water samples containing eDNA. While eDNA has been successfully preserved using Longmire's lysis buffer applied to filters, it has yet to be tried with freshwater samples collected for eDNA detection of an invasive species. We tested the utility of Longmire's solution (100 mM Tris, 100 mM EDTA, 10 mM NaCl, 0.5 % SDS, 0.2 % sodium azide) as an additive to freshwater samples for preservation of eDNA.


RESULTS
Environmental DNA was effectively preserved in 15 mL water samples with Longmire's solution added; eDNA positive detection was comparable to freezing the samples at -80 °C and occurred out to 56 days at the highest concentration (5 mL Longmire's solution: 15 mL sample water). Medium and low concentrations of Longmire's solution added to 15 mL of sample water generally preserved eDNA out to 56 days but not as well as did freezing or application of the highest concentration of Longmire's lysis buffer. Treatment and degradation time had a significant effect on average DNA concentration of samples, although not the interaction of treatment and time. Perfect detection occurred out to 56 days with the high Longmire's treatment group but DNA concentration was significantly lower at this time point compared to 28 days.


CONCLUSIONS
We conclude that Longmire's lysis buffer is a viable alternative to cold chain storage that can simplify the collection of eDNA by eliminating the need for filtering and allow more time for sample collection when added at our highest concentration (1 part Longmire's:3 parts water sample), which could translate to an increase in the chances of detecting a rare or elusive species.
Department of Fish, Wildlife, and Conservation Biology, Colorado State University, Fort Collins, CO, 80523, USA. kellwill@rams.colostate.edu.|Wildlife Services, National Wildlife Research Center, Wildlife Genetics Lab, USDA, 4101 LaPorte Avenue, Fort Collins, CO, 80521, USA. kellwill@rams.colostate.edu.
Kelly E
KE
Williams
Department of Fish, Wildlife, and Conservation Biology, Colorado State University, Fort Collins, CO, 80523, USA.
Kathryn P
KP
Huyvaert
Wildlife Services, National Wildlife Research Center, Wildlife Genetics Lab, USDA, 4101 LaPorte Avenue, Fort Collins, CO, 80521, USA.
Antoinette J
AJ
Piaggio
D000818:Animals; D059001:Aquatic Organisms / Q000235:genetics* / Q000254:growth & development; D004247:DNA / Q000032:analysis / Q000235:genetics*; D017753:Ecosystem; D005618:Fresh Water / Q000737:chemistry*; D058865:Introduced Species; D016133:Polymerase Chain Reaction; D011309:Preservation, Biological / Q000379:methods*; D015203:Reproducibility of Results; D013997:Time Factors
D004247:DNA
Environmental DNA; Freshwater; Longmire’s lysis buffer; Noncryogenic; Wild pigs