Background: The correlations between Phanerozoic atmospheric oxygen fluctuations and insect body size suggest that higher oxygen levels facilitate the evolution of larger size in insects. Methods and Principal Findings: Testing this hypothesis we selected Drosophila melanogaster for large size in three oxygen atmospheric partial pressures (aPO2). Fly body sizes increased by 15% during 11 generations of size selection in 21 and 40 kPa aPO2. However, in 10 kPa aPO2, sizes were strongly reduced. Beginning at the 12th generation, flies were returned to normoxia. All flies had similar, enlarged sizes relative to the starting populations, demonstrating that selection for large size had functionally equivalent genetic effects on size that were independent of aPO2. Significance: Hypoxia provided a physical constraint on body size even in a tiny insect strongly selected for larger mass, supporting the hypothesis that Triassic hypoxia may have contributed to a reduction in insect size.
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