Assessing anatomical changes in male reproductive organs in response to larval crowding using micro-computed tomography imaging

Ecological conditions shape (adaptive) responses at the molecular, anatomical, and behavioural levels. Understanding these responses is key to predict the outcomes of intra- and inter-specific competitions and the evolutionary trajectory of populations. Recent technological advances have enabled large-scale molecular (e.g., RNAseq) and behavioural (e.g., computer vision) studies, but the study of anatomical responses to ecological conditions has lagged behind. Here, we highlight the role of X-Ray micro-computed tomography (micro-CT) in generating in vivo and ex vivo 3D imaging of anatomical structures, which can enable insights into adaptive anatomical responses to ecological environments. To demonstrate the application of this method, we manipulated the larval density of Drosophila melanogaster flies and applied micro-CT to investigate the anatomical responses of the male reproductive organs to varying intra-specific competition levels during development. Our data is suggestive of two classes of anatomical responses which broadly agree with sexual selection theory: increasing larval density led to testes and ejaculatory duct to be overall larger (in volume), while the volume of accessory glands and, to a lesser extent, ejaculatory duct decreased. These two distinct classes of anatomical responses might reflect shared developmental regulation of the structures of the male reproductive system. Overall, we show that micro-CT can be an important tool to advance the study of anatomical (adaptive) responses to ecological environments.