Metamorphosis revealed: time-lapse three-dimensional imaging inside a living chrysalis.

Studies of model insects have greatly increased our understanding of animal development. Yet, they are limited in scope to this small pool of model species: a small number of representatives for a hyperdiverse group with highly varied developmental processes. One factor behind this narrow scope is the challenging nature of traditional methods of study, such as histology and dissection, which can preclude quantitative analysis and do not allow the development of a single individual to be followed. Here, we use high-resolution X-ray computed tomography (CT) to overcome these issues, and three-dimensionally image numerous lepidopteran pupae throughout their development. The resulting models are presented in the electronic supplementary material, as are figures and videos, documenting a single individual throughout development. They provide new insight and details of lepidopteran metamorphosis, and allow the measurement of tracheal and gut volume. Furthermore, this study demonstrates early and rapid development of the tracheae, which become visible in scans just 12 h after pupation. This suggests that there is less remodelling of the tracheal system than previously expected, and is methodologically important because the tracheal system is an often-understudied character system in development. In the future, this form of time-lapse CT-scanning could allow faster and more detailed developmental studies on a wider range of taxa than is presently possible.