Sammendrag
In 2016 the North-American signal crayfish Pacifastacus leniusculus was included in the list of Invasive Alien Species of concern to the European Union. In Norway the species is black-listed and rated as a high-risk species and eradication has been carried out whenever feasible. This is because it is a chronic carrier of the oomycete Aphanomyces astaci, which is lethal to all European freshwater crayfish species and listed among the 100 worst invasive species. Environmental DNA (eDNA) methodology is now being used on a wide variety of target species and on many different platforms including targeted PCR and broad spectred sequencing methods. Detection and monitoring of invasive, endangered and elusive species is commonly performed using species specific quantitative real-time PCR (qPCR) or droplet digital PCR (ddPCR), and has proved useful for targeting species of specific concern. Recently eDNA monitoring of A. astaci has been implemented in the national crayfish plague surveillance in Norway. For the carrier of the crayfish plague, the signal crayfish, a qPCR assay for eDNA detection has also been published recently.
In this study we present simultaneous eDNA monitoring of the host-pathogen complex P. leniusculus - A.astaci under varying environmental conditions both in aquarium and field experiments. We used the published qPCR assays for both species and redesigned them so they can be run as a duplex ddPCR assay. We compared water samples by means of qPCR and ddPCR from two lakes in Sweden and Norway with different signal crayfish population densities and A. astaci prevalence in the population. We also studied eDNA emission from the host-pathogen couple in aquarium-experiments with A. astaci positive signal crayfish held at different densities, temperatures and feeding regimes. Samples were obtained by filtering water (1 & 5 L) on-site through glass fibre filters. These were subsequently analysed using the species-specific qPCR and ddPCR assays for the respective targets. The concentrations of detectable eDNA copies of the two targets are influenced differently and by several factors, including population density of the crayfish, pathogen prevalence, temperature, and turbidity including microbiological activity in the water. They are therefore subject to significant fluctuation. Thus, there seems to be no straightforward correlation between eDNA copy-number and crayfish density and the probability of detecting one target rather than the other varies according to different conditions. Therefore, for a reliable monitoring of crayfish plague (A. astaci) and signal crayfish alone or together, the simultaneous monitoring concept for both targets is recommended.
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