Effective Amphibian Conservation Monitoring and Habitat Assessment Using eDNA

The European green toad (Bufotes viridis) is currently in decline and considered endangered across the northern extent of its native range, with large investments in ongoing conservation and translocation efforts. To assist conservation efforts, survey methods must be established that are cost-effective, non-invasive, and rapidly deployable. Here we evaluated the effectiveness of eDNA metabarcoding for amphibian conservation across three objectives: (1) Test B. viridis probability of detection before and after translocation efforts in 3 ponds in Öland, Sweden. (2) Assess pond biodiversity and biotic interactions across Öland and Kalmar using eDNA metabarcoding. (3) Determine which surveyed sites are suitable for future translocation efforts. We found that the detection probability of B. viridis increased 100% 24 h after the translocation was initiated, whereby they were undetected prior to release. Additionally, we detected 11 fish species, 14 bird species, 9 mammal species, and 4 amphibian species across the translocated sites. The results from the 37 pond eDNA surveys resulted in the detection of 15 fish species, 38 bird species, 8 amphibian species, and 17 mammal species. Species richness of the surveyed ponds ranged from 1 to 24, with an average richness of 8. Co-occurrence analysis found significant associations between several species, including a significant negative association between amphibian occurrence and cattle and gray heron and positive associations with duck and common crane. Multi-Criteria Decision Analysis (MCDA) suggests 6 sites had consistent lower site rankings, indicating them as more favorable locations for future amphibian translocation efforts. Overall, these findings showcase eDNA high-throughput sequencing as a viable means to non-invasively assess European green toads and simultaneously assess wider community dynamics that may help evaluate the sustainability of reintroduced and endemic populations.