Frequently Asked Questions:
Do water voles shed enough DNA to allow detection?
A:Yes! Field testing of the technique, at 10 separate sites, demonstrated that in all sites where the presence of water vole was known, a positive result for eDNA was obtained.
Can I be certain that the test is detecting water vole DNA and not DNA from other species?
A:Yes! The assay has been designed to specifically target water vole DNA alone. The assay was tested in the laboratory against water vole DNA extracts from populations across the UK, including Scotland. All water vole DNA extracts produced positive results. The assay was then tested against a range of non-target species’ DNA extracts (see list of species below) and all correctly resulted in negative results.
Non-target species tested:
|Common name||Latin name||qPCR test result|
|Bank vole||Myodes glareolus||Negative|
|Brown rat||Rattus norvegicus||Negative|
|Domestic cat||Felis catus||Negative|
|Domestic dog||Canis lupus familiaris||Negative|
|Domestic pig||Sus scrofa domesticus||Negative|
|Eurasian beaver||Castor fiber||Negative|
|European badger||Meles meles||Negative|
|European otter||Lutra lutra||Negative|
|Field vole||Microtus agrestis||Negative|
|House mouse||Mus musculus||Negative|
|Pine marten||Martes martes||Negative|
|Water shrew||Neomys fodiens||Negative|
|Wood mouse||Apodemus sylvaticus||Negative|
How is cross-contamination or an incorrect (“false”) result avoided?
A: A straightforward water sampling procedure can be provided for you to follow. This includes simple measures such as using a separate sampling kit for each site, so that no transfer of equipment between sites would be needed and avoiding entering the water body, to reduce the risk of the transferral of DNA from one site to the next.
Throughout the laboratory analysis process, Crestwood Environmental’s trained molecular biologists employ strict contamination prevention procedures. These include a uni-directional workflow and physical separation of pre-and post-analysis work. A range of “negative” and “no template” controls are used to detect the presence of contamination in the laboratory reagents and equipment.
Who can take the water samples?
A: A basic understanding of water vole ecology is required. This could be a suitably qualified ecologist or a trained citizen scientist. There is no licensing requirement for collecting water samples for water vole eDNA analysis. A straightforward sampling protocol is provided with every sampling kit, containing step-by-step instructions to ensure the correct procedure is followed.
Alternatively, Crestwood Environmental can collect the water samples if required – please contact us to request a bespoke quote for this aspect.
What does the sampling kit contain?
A: Each sampling kit contains all the required equipment for collecting a water sample, carrying out filtration and preserving the filter paper ready for transportation back to the laboratory for analysis. A full list of the equipment is provided with each sampling protocol.
How are the samples collected?
A: First, the sampling site selection is made, following a simple flow chart provided with each sampling protocol. 5 x 100 ml water samples are collected using a 100ml ladle and pooled in a whirl-pak bag. The pooled sample is then filtered through 2 connected filter units. Ethanol is then added to the second filter unit and the filter unit is sealed for transportation back to the laboratory for analysis. Please contact us to receive a full water sampling protocol as well as a quick guide to water sampling.
How long does collection and filtering of each sample take?
A: Water voles are known to inhabit a range of habitats, including flowing and still water. Crestwood Environmental has developed sampling methods for a range of habitat types. Sample collection time can therefore vary from 20-40 minutes per site. Please ask for further advice if needed to select the correct protocol.
How are the samples analysed?
A: The DNA is extracted from the filters by a specifically-developed DNA extraction method. The DNA samples are then analysed via real-time PCR. If water vole DNA is present above the limit of detection in the sample, it will be detected by the real-time PCR machine. The technique is very sensitive, allowing low concentrations of DNA to be detected.
How quickly can samples be analysed?
A: It usually takes 2-3 working days for received samples to be analysed and for the results to be returned. Analysis may take longer during peak times. Contact firstname.lastname@example.org for specific turnaround estimates or if you require results within a set timeframe. We will always endeavour to return your results as quickly as possible and will happily work with you to help return your results for when they are needed.
Can the method be used in flowing water?
A: Yes! The advantage of using filtration is that the DNA is collected from a larger volume of water and concentrated, allowing better detection. This means that DNA is collected in sufficient quantities, despite potentially low concentrations of eDNA present in the water, to allow detection from flowing water.
Could water vole eDNA be transported from upstream?
A: There is evidence to demonstrate that DNA can be transported downstream from a population source. The distance that the DNA travels is dependent upon many variables, including the flow rate of the water. A sampling strategy, to help understand the source of eDNA, can be formulated to test targeted stretches of a water course to determine whether water vole DNA is being transferred from upstream. Further sampling can be used to help identify stretches upstream where the DNA may be originating. This is particularly useful for identifying links to inhabited stretches and for monitoring the growth or contraction of the species’ range.
Please contact us for advice on how to formulate a design for your sampling strategy.
What is the advantage of using eDNA analysis techniques for water vole?
A: The potential advantages are numerous:
- The eDNA technique will provide a reliable ‘detected’ or ‘not-detected’ result for water vole eDNA at a site, even where habitats may not be optimal and where signs of presence are otherwise absent, or missed by inexperienced surveyors or non-professionals.
- eDNA analysis is a non-invasive, non-destructive technique. At no point is the water vole handled or disturbed during the protocol and there is no need to damage water vole habitat throughout the sampling process.
- The method requires low time and labour input, highly advantageous when many sites require surveys or time is limited
- The method can be used in conjunction with traditional survey methods to provide an additional level of evidence, so that your survey results are as accurate and reliable as possible
Can the results be used to indicate likely population sizes of water vole?
A: Not at this time, unfortunately. Relating eDNA concentration directly to population size poses a complex challenge. There are many variables that are known to affect eDNA concentration (e.g. eDNA production rate of the individual organism, the environmental conditions, the population density and how long the species was present at the location) therefore more research is needed before this application can be relied upon.
When can I use the eDNA analysis technique?
A: The optimum time to utilise this technique is during the main recognised water vole survey season (April – October inclusive), i.e. when eDNA concentrations are at their highest in the habitats.
When can’t I use eDNA for water vole?
A: eDNA sampling is unsuitable in water bodies with particularly high sediment levels, such as dried ponds, due to the sediment clogging the filters preventing the collection of a full water sample.
It is estimated that, due to reduced water vole activity, the eDNA concentrations will be lower over the winter months (November-March).
It is recommended that eDNA sampling is not be carried out directly following a flood event or sustained period of heavy rainfall, due to the increased water flow-rate associated with these times, thus reducing the concentration of eDNA in the water.
Water voles are known to be less active overnight therefore collecting samples during the daytime, when the species is likely to have been active for a period will maximise the likelihood of detecting the species’ eDNA.
Is the technique recognised by SNCO’s, planning authorities and in best practice guidance?
A: This is a novel technique that has only been recently developed. We are currently in conversation with SNCO’s and planning authorities to increase the awareness of the technique, leading to the technique being recognised in the future.
Can the evidence from this technique be used to support planning applications?
A: The technique can be used as part of a traditional survey to provide additional evidence for presence or absence, supporting planning applications.
Can the technique be used outside of the UK?
A: The technique has been validated against several non-target species present in the UK. There are, however, species that are present in other parts of Europe, such as the Southern Water Vole (Arvicola spaidus) for which further validation would be required before the technique can be reliably used outside of the UK and where this species may also be present.