Abstract
Insects of different orders produce elaborate structures to protect their eggs from the many threats they may face from the environment and natural enemies. In the weevil genus Gonipterus, their dark, hardened egg capsule is possibly generated by a mixture of the insects' excrement and glandular substances. To test this hypothesis, this study focused on the elucidation of protein components present in the egg capsule cover and interrogated them through comparative analysis and gene expression to help infer potential functions. First, female Gonipterus sp. n. 2 reproductive and alimentary tissues were isolated to establish a reference transcriptome-derived protein database. Then, proteins from weevil frass (excrement) and egg capsule cover were identified through mass spectrometry proteomics. We found that certain egg capsule cover proteins were both exclusive and shared between frass and egg capsule cover, including those of plant origin (e.g. photosystem II protein) and others secreted by the weevil, primarily from reproductive tissue. Among them, a mucin/spidroin-like protein and novel proteins with repetitive units that likely play a structural role were identified. We have confirmed the dual origin of the egg capsule cover substance as a blend of the insects' frass and secretions. Novel proteins secreted by the weevils are key candidates for holding the egg case cover together.