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Designing monitoring programs for amphibians and their pathogens using environmental DNA Caren Goldberg, Katherine Strickler, Alexander Fremier.

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Presentation on theme: "Designing monitoring programs for amphibians and their pathogens using environmental DNA Caren Goldberg, Katherine Strickler, Alexander Fremier."— Presentation transcript:

1 Designing monitoring programs for amphibians and their pathogens using environmental DNA Caren Goldberg, Katherine Strickler, Alexander Fremier

2 Environmental DNA (eDNA)

3 Di Rosa et al Bd Grayfer et al Ranavirus Environmental DNA sampling

4 eDNA project focus  Under what circumstances is eDNA sampling more efficient than standard field surveys?  How do we maximize detection probabilities? Effort Detection High density populations Low density populations eDNA sampling Field sampling more cost- effective eDNA sampling more cost effective

5 Processes affecting eDNA detection Production Diffusion/Transport Degradation Detection

6 Field sites Fort Huachuca, AZ Eglin AFB, FL North Idaho

7 Water sampling

8 Lab analysis DNA extraction (Qiashredder/DNeasy) Quantitative PCR (qPCR)

9  Potential covariates of detection UV exposure Conductivity Water temperature pH Area Volume Modeling amphibian eDNA detection

10  Collect replicate water filter samples in coordination with field surveys  Model factors influencing eDNA detection of species to inform eDNA monitoring design Modeling amphibian eDNA detection

11 Fort Huachuca, AZ American bullfrog (invasive) Sonoran tiger salamander (endangered) Chiricahua leopard frog (threatened) Di Rosa et al Bd Grayfer et al Iridoviruses Arizona treefrog (candidate)

12 Diffusion: Low Degradation: Moderate High temperatures High UV High pH (low degradation) YesNo Yes249 No539 Year 1 Field detection eDNA Detection YesNo Yes1920 No117 Year 2 Field detection eDNA Detection Fort Huachuca, AZ

13 Sonora tiger salamander detection Year 1  4 replicates  ≤250 mL each  0.45 µm CN filter

14 Sonora tiger salamander detection probability Year 1 (per sample = 0.73) AIC weight = 1.0

15 Sonora tiger salamander detection probability Year 2 (per sample = 0.77) YesNo Yes103 No06 Field Detection eDNA Detection  4 replicates  250 mL each  6 µm cellulose filter

16 Simultaneous ATV (iridovirus) detection Species# Sites Positive sites Detection probability per sample Tiger salamander (year 1) 239%0.88 Tiger salamander (year 2) 1718%0.68

17 Take samples at 2 locations Take samples at 3 locations Adaptive sampling design - spatial AIC weight =

18 Chiricahua leopard frog (per sample = 0.63)  3 replicates per location, adaptive  250 mL each  0.45 µm CN filter  Still missed 1 site ->

19 Simultaneous Bd detection Species# Sites Bd positive sites Prob detection Chiricahua leopard frog 1625%0.71 Bullfrog 119%0.75

20 Eglin Air Force Base, FL Acidic wetlands in long-leaf pine system Reticulated flatwoods salamander (endangered) Ornate chorus frog (at-risk) Di Rosa et al Bd

21 Diffusion: Very low Degradation: Very high High temperatures High UV Low pH (high degradation) YesNo Yes91 No722 Field detection eDNA Detection mL/sample Pooled samples from 4 locations/wetland Eglin Air Force Base, FL (year 1)

22 Flatwoods salamander detection probability AIC weight = Ornate chorus frogs had perfect detection pH > 5, 0 below Year 2 (now): Adaptive spatially distributed sampling (pH) Doubled sample volume

23 Simultaneous Bd detection # Sites Bd positive sites Prob detection 1926% frogs and 1 salamander found dead March 2014 COD may have been Bd for frogs COD uncertain for salamander (too degraded)

24 Bd field testing North Idaho  September 2013, April 2014  4 ponds of paired sampling (swabs, filters)  0.45 µm filters collected in quadruplicate

25 Bd field testing (in progress)

26 Ranavirus field testing P< Hall, Crespi, Goldberg, Brunner, in review Detection probability per sample: 0.90

27 Ranavirus field testing Hall, Crespi, Goldberg, Brunner, in review

28  Diffusion, acidity, and filter clogging are limiting factors in eDNA detection. Recommendations:  Pilot study  Adaptive sampling design  Distributed sampling in larger wetlands (≥1200 m 2 in this study)  Increased filter pore size in turbid systems (5-6 µ m) May reduce detection probability of pathogens Sampling conclusions

29 eDNA monitoring  How does eDNA complement or replace existing protocols? 29

30 Resources Website (this summer or fall) Field protocol (draft available) Protocol for choosing a lab (available soon) Guidelines for eDNA sampling programs (available next fall)

31 Thank you


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