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DNA testing for Chytrid fungus 

(Batrachochytrium spp.)

 

 

We now offer testing of water samples or swabs for the presence of pathogenic Chytrid fungi. This allows you to check that mitigation ponds marked for amphibian relocations are chytrid-free, or more generally to determine the prevalence of the fungus on your site and implement appropriate management plans.

 

The cost of the test is £200 for both species, or £150 per sample for Bd only, and results are returned within 10 working days.

 

 

 

 

Background

Chytridiomycosis is an amphibian disease caused by Batrachochytrium fungi (aka chytrid fungus). It is associated with global population declines and extinctions in anurans. There are many species of chytrid fungi, but only two - B. dendrobatidis (Bd) and B. salamandrivorans (Bs) - are known to cause chytridiomycosis. Bd has been known about for over a decade, while Bs has only recently been discovered. Given the significant impact these fungi species have on the global amphibian population, much effort has focused on the quick detection of these fungi in the hope that they can be managed.

 

Development of the qPCR protocol

Historically, diagnosis of chytridiomycosis has relied on examination of amphibian skin scrapes, toe clips or webbing clips. Apart from the hugely invasive nature of these methods, they require a high level of expertise to accurately stain and diagnose the chytrid, are very time consuming, and lack sensitivity when parasitic load is low. Given that early detection of Bd and Bs is vitally important for controlling the impact of their infection, molecular diagnostic techniques have been developed that are superior to their conventional counterparts. These are based on quantitative PCR (qPCR), which is the same approach used ike the GCN eDNA test).

These tests were initially applied to amphibian toe clippings, but have more recently been used on swabs. Swabbing is more sensitive than conventional histological tests and less harmful to amphibians, and this approach has since been used in large-scale surveys to determine the distribution of Bd in Great Britain.

 

Environmental Filtration

The disadvantage of swabbing is that amphibians have to be caught in relatively large numbers (30 – 60 per pond) and each one brushed at least 50 times with the swab. This is difficult, time consuming, and still to some degree invasive. Moreover, qPCR assays based on swab samples can perform poorly when the infection load is low, which is typical of some amphibian species and early on in the infection.

In 2007, researchers successfully developed a filtration-based method to extract Bd spores from sterile water, and subsequent refinement of this method means that it is now possible to assay chytrid spores collected from natural water bodies using on-site water filtration. This bypasses the ethical and logistical concerns associated with collecting large numbers of individuals, and research has shown that an appropriate sampling regime can give a 95% or greater chance of detecting Bd where it is present.

The NatureMetrics protocol follows the most up to date research in chytrid assays. Water is collected from around the perimeter of the pond, mixed together, and passed through a filter. A DNA preservation fluid is added to the filter for transportation to the laboratory, where DNA is extracted following scientific best practice and assayed using the established qPCR tests for Bd and Bs. From these analyses, we can confidently determine whether Bd or Bs are present in the sampled water body.