Understanding litter and plant decomposition rates is vital to ecosystem restoration efforts. Further, these processes are impacted by natural and engineered hydrological fluctuations. Decomposition of leaf litter in lotic systems has been explored to increase understanding of nutrient cycling and food web ecology in moving bodies of water, yet little has been done to understand these mechanisms in lentic systems despite their capacity as biological hotspots. Further, the effects of ephemerality on lentic systems and the role of the microbial community in the decomposition process in these systems are poorly understood. To assess these processes, we designed a study to examine the effects of hydrological fluctuation on known macrophyte species and their associated microbial communities in experimental restoration ponds. Specifically, we assessed how these fluctuations impacted these ponds to characterize and quantify microbial assemblages on isotopically labeled leaf litter to better understand the decomposition process and the fate of carbon and nitrogen in lentic systems. Leaf packs with 13C- and 15N-labeled leaf litter were deployed inside herbivore protection cages with Sagittaria and Vallisneria species within replicated 0.33-acre freshwater ponds (n = 3 ambient and n = 3 ephemeral) to examine microbial assemblages and C and N fluxes from leaf litter to microbes in situ. Hydrological fluctuations were induced to emulate natural drying events over a two-week period to create ecologically relevant impacts affecting decomposition rates, community assemblages, and nutrient assimilation of the microbial species in their natural environment in the recovery phase after the simulated drought. By placing the leaf packs inside cages with known macrophyte species, we were able to examine the interactive effects of ephemerality and macrophyte species on leaf litter decomposition and microbial communities associated with this leaf litter. Results from this study will inform restoration efforts on how detrital food webs function in lentic system, as well as provide applied information to land managers about how to manage these systems in the context of increased drying from human activities and changing climate.