High mountain lakes (HMLs) are considered unique and comparable ecosystems for monitoring global climate change. The food web structure can indicate the response of these ecosystems to ecological threats, such as fish introduction, by analyzing the trophic dynamics. Nonetheless, the food webs of tropical HMLs are less well-studied than temperate HMLs. The present study assessed the food webs of two neighboring (600 m apart) tropical HMLs, El Sol and La Luna, inside the crater of the Nevado de Toluca volcano, Mexico. It used stable isotopes (δ13C and δ15N) and Bayesian mixing models with different trophic discrimination factors and priors to assess the impacts of introduced rainbow trout, persisting only in the larger lake, El Sol. The food web in Lake El Sol was more complex than in Lake La Luna, mainly due to its larger size, extensive vegetated littoral zone, and being fueled by autochthonous primary production. In contrast, the smaller and fishless Lake La Luna has a reduced and bare littoral zone that harbored a simple food web substantially sustained by allochthonous carbon inputs. The persistence of introduced rainbow trout in Lake El Sol but not in Lake La Luna accentuated the differences between the lakes. The models suggested that rainbow trout fed on key consumers of littoral macroinvertebrates (70–80%) and pelagic zooplankton (20–30%), increasing the linkage between sub-networks. In both tropical HMLs, the species richness and herbivorous fraction were elevated compared with temperate HMLs, while the linkage density and omnivorous fraction were lower. Basal nodes dominated these tropical HMLs, and the vegetated littoral zone of Lake El Sol had more intermediate (omnivore) nodes. Our results showed the convenience of food web analysis to compare the effects of introduced fish in originally fishless lakes in different latitudes.