Last spring we made a call out for willing pond owners across Cornwall to be included in our county wide screening of ponds for disease. The aim of this was to determine whether Ranavirus could be detected in these ponds, and if we did find it, its prevalence and distribution. We tried to spread the message far and wide and we were met with a huge response – it seemed that many pond owners were keen to find out if their pond was being affected by this sometimes devastating disease! By mid March the amphibian breeding season was in full swing and we were driving across the county to try and catch frogs, toads and palmate newts whilst they were in their high breeding densities. This was a fun activity that pond owners were often keen to be involved in – some ponds were literally bubbling with amphibian activity!
A common frog about to be swabbed at one of our study sites!
Upon arriving at a pond, amphibians were caught and each was placed in a separate plastic container where they happily sat until it was their turn to be swabbed and released back into the pond. Swabbing is a technique that can be used to pick up pathogen DNA from the amphibians skin, which can later be detected when prepared in the lab.
Palmate newts at one of our sites - amphibians can show great variation in colour and size
Bio-control is really important when conducting field work to ensure the research activities are not contributing to the spread of disease (and/or invasive species). For this reason each amphibian was held in its own box, new disposable gloves were used for handling each amphibian and all equipment and footwear washed and disinfected between sites.
Bio-control is really important when conducting field work to ensure the research activities are not contributing to the spread of disease (and/or invasive species). For this reason each amphibian was held in its own box, new disposable gloves were used for handling each amphibian and all equipment and footwear washed and disinfected between sites.
Once all our field samples were collected it was time to head into the lab to start the screening. DNA was extracted from the swabs and a specific region amplified through a polymerase chain reaction (PCR). PCR is a thermocycling process that uses a mix of primers (short DNA strands that match the area you want to amplify), enzymes and spare nucleotides needed to build new DNA strands, to enable exponential amplification of the target DNA. This meant that if ranavirus was present, there would now be enough genetic material for it to be detected. Each sample was then run against a known positive sample via gel electrophoresis to determine whether the swab was positive or negative for ranavirus. Gel electrophoresis separates DNA according to its molecular size as it moves from one charged end of the gel to the other. All samples are run against a 'ladder' which has a known length and effectively acts as a ruler. By comparing our known positive sample - of which we know the expected length - to our own samples, we can confirm whether other samples are positive for the virus. We didn't find any evidence of ranavirus in the samples tested; whilst this is promising, further screening is needed in warmer months to confirm low levels of the virus in the county!