The role of the cuticle in the stress responses of aquatic beetles

Abstract

The colonization success of insects is largely due to a protective cuticle, a strong extracellular material that serves as skin and skeleton and functions as a first physical line of defence, providing the primary structural and biochemical barrier against environmental challenges, mechanical damage, and penetration by potentially infectious organisms. In aquatic beetles, cuticle composition could be an adaptation to colonisation of saline environments, where the presence of high levels of salinity and desiccation are frequent. However, the characteristics of the cuticle in aquatic organisms have been little studied compared to those of terrestrial ones, despite to its importance in the context of increasing aridity and salinization of inland waters, especially in arid and semiarid regions. In addition, the physiological adaptations to live in these stressful habitats, entail an energy cost, to the possible detriment of the immune response in saline species compared to those of freshwater ones, a response totally unexplored to date. The thesis focuses on the waterproofing role of cuticle in aquatic beetles in terms of composition of hydrocarbons and its plasticity as an adaptive response to cope with the multiple natural stressors such as salinity and desiccation as well as its relationship with immune capacity. Congeneric species of aquatic beetles of two families (Hydrophilidae and Dityscidae) along a salinity gradient were used as species model. Cuticular hydrocarbons profiles were identified and quantified by gas chromatography-mass spectrometry (GC-MS), the characterization was made by interpreting their EI mass spectra. To measure immune responses, phenoloxidase activity, encapsulation and antimicrobial peptide activity were studied. Results showed a more permeable cuticle of larvae than adults and the cuticle hydrocarbon profile of saline aquatic beetles seems to be more similar to that of some terrestrial beetles (e.g., desert Tenebrionidae) compared with other aquatic Coleoptera (freshwater Dytiscidae). Cuticle hydrocarbon profiles were highly species-specific and saline Enochrus species had higher waterproofing capacity showing longer chain compounds, higher relative abundance of branched alkanes, and lower unsaturated compounds than Nebrioporus ones in agreement with their higher resistance to salinity and desiccation. In addition, salinity and desiccation exposure induces plastic changes in cuticle hydrocarbons in both saline aquatic species (Nebrioporus baeticus and Enochrus jesusarribasi) and in the last, cuticular composition changes triggered the reduction of cuticular permeability to avoid water loss showing a beneficial acclimation response. Regarding immune responses, saline species of the two water beetle genera had generally lower basal immune responses than their freshwater relatives, measured at the typical salinity conditions of their natural habitats. These results suggest that changes in cuticle composition which increase its waterproofing function were one of the key mechanisms that enabled improved tolerance to salinity and desiccation and the diversification of saline species in the studied genera. Since maintaining standing defences incurs significant energetic costs, adaptation to saline environments in these taxa may have entailed a trade-off between physiological mechanisms to cope with osmotic stress and investment in immune defences. The lower immune capacity of saline species could be one of the factors accounting for their absence in freshwater.