Laboratory-based assessment of quantitative PCR to describe Arctic fish assemblages in surface water affected by acid rock drainage

Permafrost thaw has been identified as a cause of metal mobilization from acid rock drainage and transport to Arctic rivers. Elevated metal concentrations and precipitation of Fe(III) minerals (i.e., rusting) in surface waters deteriorates water quality and raises questions about ecosystem effects, especially to the health of fish species. Environmental DNA (eDNA) analyses could be used for assessing impacts of acid rock drainage to fish assemblages but may be prone to biases like polymerase chain reaction (PCR) inhibition. This study tested whether water from Arctic rivers affected by acid rock drainage influenced the efficacy of quantitative PCR (qPCR) for detecting fish. Water samples from rivers influenced by acid rock drainage were collected, characterized, and sterilized. Fish tissue was then added and mixed, and aliquots were analyzed using qPCR to assess species-specific detection sensitivity, specificity, and inhibition. In experiments, qPCR exhibited perfect species-specific assay sensitivity. There was little evidence for qPCR inhibition, whereas amplification efficiency was higher in approximately half of the samples. In mixed species assemblages, the proportion of species-specific target genes detected was significantly correlated to the proportion of tissue present for most targets. Results indicate that qPCR-based approaches, such as those often used in eDNA analyses, may help describe Arctic fish species assemblages within surface waters affected by acid rock drainage. Evaluation of qPCR methods described in this laboratory-based study using field-collected samples could help determine the effectiveness of these approaches in natural systems.