The impact of drying and temperature on the reduction of Escherichia coli, Enterococcus faecalis and Pseudomonas aeruginosa in stormwater bioretention systems with and without a submerged zone was assessed using 16 pilot-scale bioretention columns under controlled laboratory conditions. The experimental design enabled analysis of possible interactions between the factors. First outflow and event-based samples were collected. Outflow concentrations were independent of inflow concentrations and hence controlled by internal processes. Overall TSS removal was high but sensitive to bacterial synthesis. Event-based samples had significantly higher bacteria concentrations than first outflow samples, suggesting that remaining/surviving bacteria in the bioretention cells have little effect on initial peak outflow concentrations. The effect of temperature varied between bacterial species and sample types. Long dry periods seemed beneficial for bacteria reduction, but outflow bacteria concentrations peaked during the second watering after long dry periods. Submerged zones significantly reduced bacteria outflow concentrations. However, sudden temperature increases caused bioretention cells with a submerged zone to produce significantly higher bacteria outflow concentrations than before the temperature increase, which was not the case for standard cells. Bioretention cells with submerged zones may thus be poor choices for reducing bacterial concentrations in stormwater runoff in areas experiencing winter conditions. Finally, our results suggest that adsorption (e.g. further enhanced by biofilm formation) is the major mechanism governing bacteria reduction in bioretention systems.