Seasonal and Diel Signature of Eastern Hellbender Environmental DNA

Examination of environmental DNA (eDNA) is a non-invasive conservation tool that has been used for the detection of aquatic organisms. When coupled with quantitative PCR (qPCR), eDNA sampling may be utilized to infer seasonal or diel activities of target species. In order to survey the status of eastern hellbender (Cryptobranchus a. alleganiensis), a fully-aquatic cryptic salamander of conservation concern, through eDNA analyses, we collected water samples monthly from 13 sites across eight tributaries of the Susquehanna River in Pennsylvania, USA, from June through October 2014. We also examined the effects of the breeding season, diel nocturnal activity, and stream environmental variables on eDNA concentration estimates. We repeatedly detected hellbender eDNA from all four tributaries with previously known records, as well as from downstream sites of two of the four tributaries without known records. In the known tributaries, we observed notable increases in eDNA concentrations during the September breeding season, suggesting possible reproductive events. However, such seasonal eDNA signature was lacking from the eDNA positive sites of the unknown tributaries. In contrast to our prediction, there was no difference in eDNA estimates between day and night samples, indicating that diel activity was inconsequential to eDNA estimates. Our findings concur with recent studies on the importance of temporal sampling in interpreting eDNA signature in relation to life histories of target species. Further studies are needed to characterize the core habitats of the newly found populations for the future management of the declining hellbender populations