Temperature and substrate availability are important variables controlling marine heterotrophic bacterial activity. However, particularly in tropical regions it remains to be determined how these variables jointly affect bacterial activity. In this study we show how bacterial carbon cycling in two tropical coastal ecosystems (the Great Barrier Reef [GBR, Australia] and the Red Sea [Saudi Arabia]) are influenced by changing temperature (using a 6°C gradient) and the addition of dissolved organic carbon (DOC) from different sources (addition of mangroves and seagrass leachates, plus natural seawater). Our study demonstrates that elevated temperatures in the GBR increased bacterial organic carbon processing, while in the Red Sea no clear effects were found. More of the added DOC was degraded in the Red Sea but this additional carbon did not increase the biomass production, due to low bacterial growth efficiencies in all treatments. In addition, increasing temperatures in the GBR resulted in lower bacterial growth efficiencies, while no clear impact were found in the Red Sea. In conclusion, this study suggests that site-specific ecosystem differences (e.g., different microbial and macrophyte community composition) may override general responses to temperature and substrate in tropical coastal waters.