Simplified and rapid staining of leafhopper salivary sheaths in plant tissues for electrical penetration graph waveform correlations

Herbivorous insects in the order Hemiptera use piercing-sucking mouthparts to utilize plant sap. Among them salivary sheath feeders penetrate into by their flexible stylets reach vascular elements. Manoeuvering tissues is aided creation of sheaths, which solidify from proteinous gelling saliva and remain as lasting artefacts tissues. Studying structure reveals hidden details feeding behaviour transmission pathogens case vector insects. One important aspect studying sheaths that it can be used confirm biological function electropenetrography (EPG) waveform patterns. Previously, complex vaguely documented histological methods have been observe structure. Building on existing methodologies, we report a simplified procedure where each step was optimized offer rapid process does not require special equipment, applied many samples, has good success rate low cost errors terms time materials. We describe procedure, using Psammotettix alienus – barley model system, three steps. (i) Clarification entire parts pre-staining with aqueous fuchsin. This allows identify starting points surface. (ii) Knowing location, hand sectioning, produce single c. 60 ?m section contains sheath. (iii) Counterstain methylene green and, after further clarification, study under light microscope glycerol - ethanol embedding solution, without fixed mounting.