Detection of Limnoperna fortunei DNA from plankton samples: a new protocol

Abstract

Limnoperna fortunei (golden mussel) is an invasive mollusk, native to Southeast Asia. The detection of L. fortunei, during early stages of the invasion, is an interesting strategy for its control and eradication. For these aims, traditional methods are used based on identifying the species in samples of zooplankton or benthos community. In addition, molecular methods based on the detection of DNA in different environmental samples (eDNA) have been proposed. We describe and discuss a protocol developed for the detection of L. fortunei based on mitochondrial eDNA in plankton samples from freshwater systems. Water samples were obtained during 2018-2019, from cooling systems of the hydroelectric power plant in Río Negro, Uruguay. eDNA was detected by end-time PCR and PCR products were resolved on polyacrylamide gels and silver staining. Primers specificity was determined in related limnic and brackish native species. Sensitivity was analyzed as genomic DNA concentration and number of larvae. Different methodologies were tested to remove inhibitors from samples. Finally, the eDNA PCR products were sequenced to confirm the specificity of the reaction. The results obtained by molecular techniques were more sensitive than traditional methods of species detection.