Culex pipiens pallens cuticular protein CPLCG5 participates in pyrethroid resistance by forming a rigid matrix

Abstract

BackgroundChemical insecticides have hugely reduced the prevalence of vector-borne diseases around the world, but resistance threatens their continued effectiveness. Despite its importance, cuticle resistance is an under-studied area, and exploring the detailed molecular basis of resistance is critical for implementing suitable resistance management strategies.MethodsWe performed western blotting of cuticular protein CPLCG5 in deltamethrin-susceptible (DS) and laboratory-produced deltamethrin-resistant (DR) strains of Culex pipiens pallens. Immunofluorescence assays using a polyclonal antibody to locate cuticular CPLCG5 in mosquitoes. EM immunohistochemical analysis of the femur segment was used to compare the cuticle in control and CPLCG5-deficient siRNA experimental groups.ResultsThe gene CPLCG5 encodes a cuticle protein that plays an important role in pyrethroid resistance. Based on a prior study, we found that expression of CPLCG5 was higher in the resistant (DR) strain than the susceptible (DS) strain. CPLCG5 transcripts were abundant in white pupae and 1-day-old adults, but expression was dramatically decreased in 3-day-old adults, then remained stable thereafter. Western blotting revealed that the CPLCG5 protein was ~2.2-fold higher in the legs of the DR strain than the DS strain. Immunofluorescence assays revealed CPLCG5 expression in the head, thorax, abdomen, wing, and leg, and expression most abundant in the leg and wing. EM immunohistochemical analysis suggested that the exocuticle thickness of the femur was significantly thinner in the CPLCG5-deficient siCPLCG5 strain (0.717 ± 0.110 μm) than the siNC strain (0.946 ± 0.126 μm). Depletion of CPLCG5 by RNA interference resulted in unorganised laminae and a thinner cuticle.ConclusionsThe results suggest CPLCG5 participates in pyrethroid resistance by forming a rigid matrix and increasing the thickness of the cuticle.