Quantitative monitoring of diverse fish communities on a large scale combining eDNA metabarcoding and qPCR

eDNA metabarcoding is an effective method for studying fish communities but allows only an estimation of relative species abundance (density / biomass). Here, we combine metabarcoding with an estimation of the total abundance of eDNA amplified by our universal marker (teleo) using a qPCR approach to infer the absolute abundance of fish species. We carried out a 2,850 km eDNA survey within the Danube catchment using a spatial integrative sampling protocol coupled with traditional electrofishing for fish biomass and density estimation. Total fish eDNA concentrations and total fish abundance were highly correlated. The correlation between eDNA concentrations per taxon and absolute specific abundance was of comparable strength when all sites were pooled and remained significant when the sites were considered separately. Furthermore, a non-linear mixed model showed that species richness was underestimated when the amount of teleo-DNA extracted from a sample was below a threshold of 0.65.106 copies of eDNA. This result, combined with the decrease in teleo-DNA concentration by several orders of magnitude with river size, highlights the need to increase sampling effort in large rivers. Our results show a comprehensive description of longitudinal changes in fish communities and underline our combined metabarcoding/qPCR approach for biomonitoring and bioassessment surveys when a rough estimate of absolute species abundance is sufficient.