The role of V-ATPase in regulating pH in the digestive tract of Tetranychus urticae (Koch)

Abstract: Spider mites are phytophagous organisms and resilient pests in plant-growing
industries. One of their main qualities that allows for successful herbivory on plant hosts is their ability to detoxify xenobiotic compounds, defensive allelochemicals and pesticides. Previous research has identified genes/enzymes involved in the digestion of ingested plant cell content and detoxification of xenobiotic compounds. However, the cell types within the digestive track where digestion and detoxification occur are not well understood. In particular, the conditions of pH in digestive compartments are not known, which is important for optimal performance of enzymes carrying out these processes. The purpose of this study was to a) establish pH in intercellular and intracellular compartments of the spider mite midgut, and b) to examine the effects of silencing the expression of a gene coding for the pH-regulating protein complex, VATPase, on the pH status of digestive compartments. Indicator dyes were applied in adult female two-spotted spider mites (Tetranychus urticae Koch) to establish the pH environment present in the midgut ventriculus, gastric caeca and posterior midgut in both fed and unfed states. A slightly alkaline pH range of 7.3 to 7.5 was observed in intercellular midgut compartments of unfed mites whereas the vesicles in free-floating digestive cells where digestive activity is predicted to occur were more acidic at 5.2. There were no significant changes in pH in any compartment when mites were fed. The expression of the gene tetur09g004140 (coding for subunit A of V-ATPase) was subsequently downregulated by RNA interference in fed mites. The pH in the intercellular midgut compartments of knockdown mites showed significant increase in pH whereas no significant increase was observed in the digestive cell vesicles. This study reports for the first time the pH environment in the midgut of spider mites. It provides a basis for determining where enzymes associated with digestive and/or detoxification activities may operate in the spider mite midgut based on their pH optima.