Major ongoing declines in global biodiversity necessitate biomonitoring strategies that enable precise estimates of community diversity on a fine spatial and temporal scale. While environmental DNA (eDNA) has been established as a powerful tool for biodiversity assessment, studies investigating the comparative performance of environmental RNA (eRNA) are limited. Here, we performed eDNA/eRNA metabarcoding of zooplankton communities in outdoor freshwater mesocosms subject to a dynamic range of pH conditions. We comparatively assessed i) the sensitivity of eRNA metabarcoding relative to eDNA and traditional survey methods in capturing zooplankton diversity, ii) the influence of pH on eDNA/eRNA detectability, and iii) the propensity of eRNA to capture contemporary biological assemblages (i.e., rapid species turnover) with high spatial and temporal acuity. Zooplankton richness was similar amongst eDNA/eRNA metabarcoding and traditional survey methods; however, the composition of zooplankton communities detected was more analogous between eDNA and eRNA metabarcoding than with traditional methods. Both eDNA and eRNA captured similar ZOTU richness and frequency of false negative detections (irrespective of site-specific pH); however, eRNA captured species turnover more rapidly than eDNA. Collectively, our findings suggest that i) relative to traditional methods, eDNA and eRNA metabarcoding may provide users with complementary rather than congruent estimates of biodiversity, ii) eDNA and eRNA provide comparable estimates of species richness irrespective of site-specific pH conditions, and iii) eRNA is able to capture short-term community responses with higher spatial and temporal acuity than eDNA. Overall, our findings support the use of eRNA for characterizing contemporary biodiversity in complex and dynamic aquatic environments.
