The idea that atmospheric oxygen has dictated the maximum body size of insects across their evolutionary history is ingrained in popular and scientific literature1–3. In Nature 30 years ago, the hypothesis was put forward that a limitation on oxygen diffusion at the level of the tracheoles constrains the maximum body size of insects and that increased atmospheric oxygen concentration in the late Palaeozoic permitted insect gigantism4. Here we contest this hypothesis by showing that the relative space occupied by tracheoles in the flight muscle of insects (1) increases by only 1.8-fold over a 10,000-fold body mass range (1,320 micrographs, 44 species, 10 orders), (2) is typically 1% or less in most species, and (3) that this observation holds when we extend our relationship to the long-extinct gigantic dragonfly-like Meganeuropsis permiana (approximately 100 g). The small space requirement and the lack of a strong increase in tracheolar investment with body size, despite clear evolutionary potential to do so, provide convincing evidence that diffusive oxygen transport through the tracheolar–muscle system does not constrain the maximum body size of extant or gigantic prehistoric insects.
