Lakes and rivers are important sources of potable water in many countries, but taste-and-odour compounds (T&O) such as geosmin (GSM) and 2-methylisoborneol (2-MIB) may affect the aesthetic water quality and require high-cost treatment. In this study, we compared changes in T&O concentrations (GSM and 2-MIB) and the microbial community at seven sites along the subtropical Brisbane River, which is the main source of potable water to the city of Brisbane. GSM and 2-MIB had distinctly different spatial patterns. Concentrations of 2-MIB entering the river appeared to be sourced from the upstream Lake Wivenhoe reservoir. Several Oscillatoriales species (filamentous cyanobacteria) were identified as a possible source of 2-MIB in the lake. However, concentrations of both compounds were also observed to increase along the upper reaches of the river at different times when there were no inputs from tributaries, implying in-river production. 2-MIB and GSM concentrations in the river were correlated with the abundance of several actinobacteria species in the water column. Concentrations of 2-MIB and GSM decreased downstream, at times, with this decrease more likely to occur in the lower reaches of the river. Concentrations were typically close to analytical detection levels (2 ng L−1) at the drinking water treatment plant (DWTP) 60 km downstream of the reservoir. These findings suggest that both cyanobacteria and non-photosynthetic bacteria are important in controlling 2-MIB and GSM concentrations in the river, but water depth and release from sediments may also affect riverine concentrations of T&Os. Different strategies may be required for predicting and managing each compound in rivers.