Cost-effective DNA extraction method for detection of amphibian fungal pathogen in invertebrate samples

The amphibian fungal pathogen Batrachochytrium dendrobatidis (Bd) has devastated global amphibian biodiversity. Non-amphibian hosts might facilitate Bd spread across the landscape, but our understanding of their role remains severely limited. Several species of invertebrates have been implicated to varying degrees as potential carriers of Bd, but research is needed to understand the impact these invertebrates have on the spread and maintenance of this pathogen in the wild. Detecting Bd presence in field-collected invertebrates is a necessary first step for identifying potential vectors, but methodological comparisons have not been conducted. Detection thresholds of DNA extraction methods for Bd detection from amphibian skin swabs may not be applicable for invertebrate samples, because they may contain lower Bd loads and higher amounts of PCR-inhibitory substances. This study aimed to identify the most cost-effective and reliable method of DNA extraction for detecting Bd DNA in invertebrate samples using qPCR. We compared the effectiveness of 5 commonly used DNA extraction kits (QIAGEN, Zymo, SPINeasy, PrepMan Ultra, and Chelex resin) for Bd detection in homogenised cricket samples spiked with known concentrations of Bd. We found PrepMan Ultra to be the optimal extraction kit for a broad screening of field-collected invertebrate samples, due to its relatively low cost and the ability to detect Bd presence in homogenised cricket samples containing at least 100 zoospore equivalents. However, researchers need to conduct their own cost-benefit analysis when choosing an extraction method to ensure that the method suits their needs.