Hyporheic exchange (HE) contributes to the biogeochemical turnover of macro- and micro-pollutants in rivers. However, the spatiotemporal complexity and variability of HE hinder understanding of its role in the overall functioning of riverine ecosystems. The present study focuses on investigating the role of bacterial diversity and sediment morphology on HE using a multi-flume experiment. A fully coupled surface–subsurface numerical model was used to highlight complex exchange patterns between surface water and the underlying flow field in the sediments. Under the experimental conditions, the surface water flow induced by bedforms has a prominent effect on both local trajectories and residence time distributions of hyporheic flow paths, whereas mean hyporheic retention times are mainly modulated by average surface flowrates. In case of complex bedform morphologies, the numerical model successfully reproduces the HE estimated by means of salt dilution tests. However, the 2D numerical representation of the system falls short in predicting HE in absence of bedforms, highlighting the intrinsic complexity of water circulation patterns in real scenarios. Finally, results show that higher bacterial diversities in the stream sediments can significantly reduce hyporheic fluxes. This work provides a framework to interpret micropollutants turnover in light of the underlying physical transport processes in the hyporheic zone. The study emphasizes the importance of better understanding the tradeoff between physically driven transport processes and bacterial dynamics in the hyporheic zone to quantify the fate of pollutants in streams and rivers.