Using carbon and nitrogen stable isotope analysis, we characterised food web properties of the deep subtropical Fei-Tsui Reservoir (FTR), which was recently altered from a lotic to a lentic system after dam construction. In the littoral zone, zoobenthos showed strong reliance (83.9%) on benthic algal production. Zoobenthos were never found in the profundal zone because of anoxia. Zooplankton depleted 13 C more than that of particulate organic matter as their putative food source, suggesting a contribution of methane-derived carbon to pelagic food webs. Excluding juveniles, non-native and fluvial species, adult fish showed strong reliance (on average 80.9%) on benthic production, weakly coupled with pelagic food webs. These results contrast low benthic production reliance (on average 27.4%) for a fish community in Lake Biwa, which is also classified as a subtropical lake. Both lakes are characterised by deep pelagic waters but quite different in their geological ages, suggesting that the aquatic communities in the FTR have fluvial origins, and their lacustrine history was too short for them to adapt to newly emerged deep pelagic habitat. Our isotope data are useful as a reference of newly established lentic food webs to monitor ongoing ecological and evolutionary dynamics as a result of anthropogenic disturbances.