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Mixotrophs, which combine photosynthesis and heterotrophy to gain energy, play an important role global biogeochemical cycles. Metabolic theory predicts that mixotrophs will become more heterotrophic with rising temperatures, potentially creating a positive feedback loop that accelerates carbon dioxide accumulation in the atmosphere. Studies testing this theory have focused on phenotypically plastic (short-term) thermal responses of mixotrophs. However, as small organisms with short generation times and large population sizes, mixotrophs may rapidly evolve in response to climate change. Here we present data from a 3-year experiment quantifying the evolutionary response of two mixotrophic nanoflagellates to temperature. We found evidence for adaptive evolution (increasing growth rates) in the obligately mixotrophic strain, but not in the facultative mixotroph, though all lineages evolved greater carbon use efficiency. Generally, mixotrophs evolved reduced photosynthesis and higher grazing with increased temperatures, suggesting that evolution may act to exacerbate mixotrophs’ effects on global carbon cycling.