Lateral and longitudinal fish environmental DNA distribution in dynamic riverine habitats

Abstract

Assessing the status and distribution of fish populations in rivers is essential for management and conservation efforts in these dynamic habitats. Currently, methods detecting environmental DNA (eDNA) are being established as an alternative and/or complementary approach to the traditional monitoring of fish species. In lotic systems, a sound understanding of hydrological conditions and their influence on the local target detection probability and DNA quantity is key for the interpretation of eDNA‐based results. However, the effect of seasonal and diurnal changes in discharge and the comparability of semi‐quantitative results between species remain hardly addressed. We conducted a cage experiment with four fish species (three salmonid and one cyprinid species) in a glacier‐fed, fish‐free river in Tyrol (Austria) during summer, fall, and winter discharge situations (i.e., 25‐fold increase from winter to summer). Each season, water samples were obtained on three consecutive days at 13 locations downstream of the cages including lateral sampling every 1–2 m across the wetted width. Fish eDNA was quantified by species‐specific endpoint PCR followed by capillary electrophoresis. Close to the cages, lateral eDNA distribution was heterogenous and mirrored cage placement within the stream. In addition to the diluting effect of increased discharge, longitudinal signal changes within the first 20 m were weakest at high discharge. For downstream locations with laterally homogenous eDNA distribution, the signals decreased significantly with increasing distance and discharge. Generally, the eDNA of the larger‐bodied salmonid species was less frequently detected, and signal strengths were lower compared to the cyprinid species. This study exemplifies the importance of hydrological conditions for the interpretation of eDNA‐based data across seasons. To control for heterogenous eDNA distribution and enable comparisons over time, sampling schemes in lotic habitats need to incorporate hydrological conditions and species traits.

The Effect of Activity, Energy Use, and Species Identity on Environmental DNA Shedding of Freshwater Fish

“…All used primers (Table 1) have been previously published after extensive specificity and sensitivity testing (Thalinger et al, 2016(Thalinger et al, , 2021b and additional specificity tests were carried out on the digital PCR (dPCR) system (see below) confirming the specificity of the molecular assays under the following conditions: each 22 µl dPCR master mix for droplet generation on the QX200 AutoDG (Biorad) consisted of one-time EvaGreen Supermix (Biorad), 0.25 µM forward and reverse primer ( Table 1) and up to 10.5 µl DNA extract. Depending on the results of initial tests with capillary electrophoresis PCR (i.e., the Relative Fluorescence Units (RFU) of the resulting band; see Supplementary Material 3), extracts were diluted with molecular grade water for dPCR as follows: RFU < 0.2: undiluted; 0.2 ≤ RFU < 1.3: 1:1 dilution; 1.3 ≤ RFU < 2: 1:3 dilution; 2 ≤ RFU: 1:7 dilution.…”

“…Different processes influence the distribution of eDNA in space and time and the detection probabilities of species from environmental samples, namely the origin, degradation, suspension, resuspension, and transport of eDNA (Barnes and Turner, 2016;Harrison et al, 2019). The latter processes are directly linked to local hydrology [e.g., flow and substrate type (Shogren et al, 2017;Pont et al, 2018;Thalinger et al, 2021b)] and environmental conditions [e.g., water temperature, pH, UV-radiation (Strickler et al, 2015;Lacoursière-Roussel et al, 2016;Tsuji et al, 2017)]. The amount of eDNA in the water column is directly linked to fish biomass and originally, this was confirmed for common carp (Cyprinus carpio) in an aquarium trial and in experimental ponds (Takahara et al, 2012).…”

Proper environmental DNA metabarcoding data transformation reveals temporal stability of fish communities in a dendritic river system

“…However, despite recent studies investigating the effect of seasonality on eDNA (de Souza et al, 2016;Goldberg et al, 2011;Handley et al, 2019;Sevellec et al, 2020;Sigsgaard et al, 2017;Stoeckle et al, 2017;Thalinger et al, 2021;Wacker et al, 2019), sources of temporal variation in abundance in species eDNA need further investigation to improve the reliability of long-term surveys of species community in aquatic ecosystems.…”

Endpoint PCR coupled with capillary electrophoresis (celPCR) provides sensitive and quantitative measures of environmental DNA in singleplex and multiplex reactions

“…regions [18,23]. Nevertheless, other factors such as inhibition can potentially affect measurements derived with celPCR and (less likely) dPCR [60] from field-collected samples.…”

“…In the past, celPCR has been used to determine if the fluorescence of a target amplicon exceeds a predefined threshold and samples can thus be scored "positive" [19,20]. Although the basic concept of this approach is not novel [21,22], there have been only rudimentary attempts to assess the general quantification capabilities of celPCR for eDNA analyses [18,23]. This possibility for quantification is especially appealing for target eDNA detection in large sample sets, as there is a high potential for cost-reduction based on PCR-chemicals alone (Table 1).…”