Larvae of the three important Central American malaria vectors, Anopheles albimanus, An. vestitipennis, and An. darlingi, are found in distinctly different habitats broadly defined by hydrology and aquatic vegetation, but little is known about the actual food quality and quantity of these habitats. Polyunsaturated fatty acids (PUFA) are of special interest, because mosquitoes require 20:5ω;3 (EPA), 20:4ω;6 (ARA), and 22:6ω;3 (DHA) and without an adequate supply of these PUFAs they are not able to complete their life cycle. We collected samples of larvae and their corresponding habitats and analyzed their fatty acid (FA) composition to reveal if there are any species-specific and habitat-specific differences in FA composition, and if habitat FA differences can be linked to differences in the mosquito FA pattern and, ultimately, mosquito performance. We also assessed how FA of wild larvae compare to the laboratory-reared larvae. Habitats were generally low in essential PUFAs and there were no significant differences among the FA composition of habitat samples. There were significant differences in FA composition of larvae. An. darlingi contained significantly higher amounts of FA, specifically a higher content of ω;-6 PUFA, represented mainly by the linoleic acid (18:2ω;-6). Large differences were found between field-collected and laboratory-reared An. vestitipennis larvae, especially in the content of PUFAs. The laboratory-reared larvae contained significantly more of the total FA, ω;3 PUFA, and MUFA. The laboratory-reared larvae contained three to five times more essential PUFAs, EPA, and DHA. However, there were no differences in the total dry weight of the 4^th instar larvae between the wild vs laboratory-reared larvae. Total FA in both larvae and habitats of An. albimanus and An. darlingi were positively correlated with the concentration of particulate organic carbon and nitrogen (POC, PON) in their respective habitats, but no such correlation was found for An. vestitipennis. PUFA are a good indicator of nutritional quality, although factors controlling the success of anopheline development from larval habitats are likely to be more complex and would include the presence of predators, pathogens, and toxins as interacting factors.