New and Simple Methods for Studying Hemipteran Stylets, Bacteriomes, and Salivary Sheaths in Host Plants

Many hemipteran insects are important agricultural pests because they cause direct feeding damage to their host plants and / or transmit plant disease agents including viruses and bacteria. Microscopic and behavioral studies on five hemipteran species from four families (Psyllidae, Aphididae, Cicadellidae, and Aleyrodidae) showed that their exuviae (molted skins) normally had either fully or partially extended stylets in a feeding-like position. In most cases these stylets were still partially embedded in their host plants after ecdysis, which indicated that plant-feeding hemipteran nymphs use their stylets to anchor themselves to host plants during molting. This phenomenon was used here to study the stylet length and ultrastructure in exuviae of various instars, which is normally more difficult in nymphs than in adults because of the fragility and smaller size of nymphs. Additionally, autofluorescence was used for studying the hemipteran salivary sheaths of nymphs and adults in their host plants. This method is based on fixation of free hand sections of plant parts on which hemipteran insects have been feeding, then mounting and examination of these sections with epifluorescence or confocal microscopy. No embedding, microtomy, or staining is necessary for this method that makes it much faster and simpler than other methods. Autofluorescence was also used to study the location and size of bacteriomes/mycetomes (organs containing symbionts) in hemipteran eggs and nymphs. The above methods were applied successfully with the Asian citrus psyllid (Diaphorina citri Kuwayama), melaleuca psyllid (Boreioglycaspis melaleucae Moore), oleander aphid (Aphis nerii Boyer de Fonscolomb), the whitefly Bemisia tabaci (Gennadius), and/or the glassy-winged sharpshooter leafhopper (Homalodisca vitripennis Germar).