A novel and highly innovative method for visualization and characterization of the insect surface inward barriers

Abstract

In insects, cuticular hydrocarbons (CHCs) play a critical role in the establishment of the waterproof barrier that prevents both dehydration and penetration of environmental hazards such as pathogens and toxic molecules. While rich data is available on CHC composition in different species and their relationship with insect dehydration resistance, we know little about their distribution, organization and their functional roles in the formation of the insect surface inward barrier. Here, I present my work on the development of an Eosin Y penetration method to visualize and characterize the insect surface inward barrier. In the Eosin Y penetration method, the investigated insect is incubated in Eosin Y solution at certain temperatures. The dye penetrates different regions of the insect cuticle at distinct temperatures. The temperatures, on which the Eosin Y penetrated the certain cuticle region and stain the tissue under such regions, can be recorded as Tpe of such region. Differences between Tpe values indicate the differences in lipids composition in the respective regions. This Eosin Y penetration method is easy, quick, cost limited and can be commonly used in different insect species. We have used this method already to analyse the barrier distribution in fruit fly (Drosophila melanogaster) adults and larvae, locust (Locusta migratoria) nymphs, mealworm (Tenebrio molitor) larvae, whitefly (Trialeurodes vaporariorum) adults and bedbug (Cimex lectularius) nymphs. The results indicate that the deposition of surface lipids of most insects is regionalized. Furthermore, on some species, such as whitefly and bedbug, exist two barriers parallel in same regions of cuticle with distinct temperature-sensitive and lipid-based physico-chemical material properties. We also applied this method to determine the effect of certain factors such as ABCH-9C transporter Snu and cuticle protein Snsl in the establishment of cuticular inward barrier. We believe that this new method gives us new insights into theme cuticle permeability, and can be widely used in designing new insecticides which are more efficiently taken up by specific target species. Additionally, the Tpe alteration is also a good readout for genome screen for study the molecular principle of cuticle hydrocarbon layer establishment.