Correlated evolution of elaborate intromission mechanics during copulation between the sexes in leaf beetles

Diverse female and male genitalia have evolved. Although the evolutionary mechanisms behind this phenomenon have been studied extensively, the mechanical interactions between female and male genitalia are much less understood. Here, we provide an in-depth biomechanical study on the intromission of elongated tube-like female and male genitalia, termed the spermathecal duct and flagellum, in tortoise leaf beetles. Our findings reveal that this seemingly straightforward penetration mechanism is underpinned by structural and material specializations of the genitalia. We employed synchrotron-based micro-computed tomography and confocal laser scanning microscopy (CLSM) to visualize the precise mechanical interactions between female and male genitalia. Mechanical tests revealed a stiffness gradient within the flagellum and correlated stiffness variations between the sexes. Combining CLSM and cryo-microtome techniques, we characterized a heterogeneous material distribution at the flagellum tip; this specialization was more remarkable in the species with a more sclerotized spermathecal duct. Finite element analyses incorporating the observed material properties demonstrated that the observed material distribution reduced the penetration force required tremendously and dispersed the stress at the flagellum tip. This study unveils that structural and material adaptations in female and male genitalia have probably coevolved and highlights the importance of biomechanics in genital studies.