This study examined the trophic ecology of three sympatric tropical tuna species (bigeye BET, skipjack SKJ, and yellowfin YFT) sampled in the Western Indian Ocean throughout 2013. Specifically we explored inter-specific resource partitioning and ontogenetic variability using neutral fatty acids and stable isotope analysis of liver and muscle from small (<100 cm fork length, FL) and large (>100 cm FL) tuna collected in mixed schools at the surface by purse-seine. Both biochemical tracers were used to calculate trophic niche indices that collectively revealed high potential for resource overlap, especially among small tuna. Resource overlap appeared strongest between BET and YFT, with SKJ tissues having high carbon isotope (delta C-13) values (-17 +/- 0.3 parts per thousand), lower nitrogen isotope (delta N-15) values (11.4 +/- 0.6 parts per thousand), and higher relative proportion of poly-unsaturated fatty acids (PUFA) than the two other species, indicating a different diet. Size was found to be a strong predictor for most biochemical tracers in the three species with delta C-13, delta N-15 and total lipid content in the liver. In the larger species (YFT and BET), proportions of mono-unsaturated fatty acids typically increased with size, while quantities of PUFA decreased. In addition to ontogenetic variability, trophic markers were shown to vary between sampling area and season: higher lipid reserves and delta N-15 values, and lower delta C-13 values occurred during monsoon periods around Seychelles than in the Mozambique Channel (parted from about 1500 km). Our multi-tracer approach reveals the magnitude of potential competitive interactions in mixed tropical tuna schools at both small and large sizes and demonstrates that ontogenetic niche differentiation acts as a major factor of coexistence in tropical tuna