The generation of ASVs has several advantages over OTUs including finer resolution, accurate measures of diversity and easy comparison between independently processed data sets (Callahan, McMurdie, & Holmes, 2017). Studies in marine systems have also proposed a disconnect between eDNA concentrations in surface and deep sampling points, although the overall effect of stratification was less clear because of the release of extraneous eDNA into the bay due to waste from a local fish market (Yamamoto et al., 2016). Stratification is a desirable strategy to provide efficient room air conditioning with much less effort than using the piston strategy. This analysis was performed for each of the three temporal data sets of telemetry data collected (one day, one week and one month before the point of sampling), to test whether differences in the temporal range of habitat selection better explained the distribution of eDNA, as it is known to persist in the water column for several days to weeks. We used nonmetric multidimensional scaling to visualize fish communities, by specifying either two or three dimensions (to minimize stress and achieve convergence) and 200 random starts. However, there have been few studies that look at within‐habitat eDNA distribution particularly with respect to habitat niche specialization or behavioural preferences (although see Macher & Leese, 2017; Nichols, Königsson, Danell, & Spong, 2012), and fewer still have examined how this might change seasonally. Thus, habitat use by obligate cold‐water species can be greatly reduced and constrained to deeper depths during summer stratification, especially in small temperate lakes where habitat volume reductions of >60% are common due to lack of preferred temperature and dissolved oxygen conditions (Paterson, Podemski, Wesson, & Dupuis, 2011; Plumb & Blanchfield, 2009). Typically, this point coincided with the 15°C isotherm, which is the threshold for lake trout thermal preference. All lakes have overlapping community compositions, including S. namaycush, a cold‐water top predator, in every lake. By contrast, Li et al., (2019) found eDNA of deep‐water species in shoreline samples during winter sampling, but as it is not clear to what degree (if any) the study lakes were stratified during winter months, this may have been the result of thorough mixing during autumn turnover. Performance of one high size real thermocline thermal energy storage system is analysed. Thank you for your interest in spreading the word about bioRxiv. Students can meet the goals for this lesson by completing a directed study or an inquiry lesson. Briefly, S. namaycush were captured by angling and surgically implanted with coded, acoustic, pressure‐sensing telemetry tags (model V13P‐1L; Vemco, Innovasea, Bedford, NS). There are 14 species of fish across all the study lakes (mean 8, range 6–10 species per lake, Table S2). Please check your email for instructions on resetting your password. We hypothesized that (1) lake thermal stratification (i.e. By contrast, rates of vertical diffusion of tracer across the thermocline of stratified lakes are much slower (Quay, 1980). A forest community is a typical example of terrestrial stratification because here a number of strata both above and below the soil can be recognised. Studies at the habitat scale have already suggested possible generalities linking eDNA to biological activity; for example, that peaks of eDNA can indicate the onset of reproduction (Bylemans et al., 2018; Spear et al., 2015) or relative abundance of species (Li et al., 2019). Chrosomus neogaeus (finescale dace) and Chrosomus eos (redbelly dace) were both assigned at genus level, possibly because pure C. eos does not exist in this region but instead forms both cytoplasmic and nuclear hybrids with C. neogaeus (Mee & Taylor, 2012). We used a variance stabilizing transformation on our sample x ASV matrix to account for uneven library size across our samples. This approach allows us to control for library size while retaining interpretable response data (e.g. At the end of thermal stratification the surface waters of the epilimnion gradually cool as a result of conduction, evaporation and convection. We recovered 94,013 ± 6,389 sequences per demultiplexed sample with an initial quality score of 33.0 ± 0.23. The receivers logged acoustic signals emitted by the tags through an omnidirectional hydrophone. Equally, the ecological significance of these factors cannot be tested when examined in isolation (Carpenter, Chisholm, Krebs, Schindler, & Wright, 1995). If you do not receive an email within 10 minutes, your email address may not be registered, There was a significant interaction between lake depth and lake state affecting fish community assemblages detected by eDNA (PERMANOVA, F1,335 = 4.35, p = .0002). DNA was extracted from individual fish samples using the Qiagen Blood and Tissue kit following the manufacturer's instructions, equimolarized to 6.5 ng/μl and combined to create the mock community. For example, S. namaycush displays clear shifts away from littoral habitats when epilimnetic temperatures rise above 15°C, suggesting that water temperature phenology is a strong determinant of seasonal habitat use (Guzzo, Blanchfield, & Rennie, 2017). By contrast, during lake turnover (b) there is an isothermal water column with mixing between deep and shallow waters. All filtration, extraction and PCR negative controls were amplified in triplicate. SUMMARY. Habitat use by S. namaycush was validated by acoustic telemetry and was significantly related to eDNA distribution during stratification. For example, Klobucar, Rodgers, and Budy, (2017) found that surface sampling points had lower Salvelinus alpinus (arctic char) eDNA concentrations than the deeper sampling point during summer stratification in North Alaskan lakes, probably due to limited thermal habitat for the deep‐water species. 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