Potato aphid salivary proteome: enhanced salivation using resorcinol and identification of aphid phosphoproteins

Aphids deliver saliva into plants and acquire plant sap for their nourishment using a specialized mouthpart or stylets. Aphid saliva is of great importance because it contains effectors that are involved in modulating host defense and metabolism. Although profiling aphid salivary glands and identifying secreted proteins have been successfully used, success in direct profiling of aphid saliva have been limited due to scarcity of saliva collected in artificial diets. Here we present the use of a neurostimulant, resorcinol, for inducing aphid salivation. Saliva of potato aphids (Macrosiphum euphorbiae), maintained on tomato, was collected in resorcinol diet. Salivary proteins were identified using mass spectrometry and compared with the existing M. euphorbiae salivary proteome collected in water. Comparative analysis was also performed with existing salivary proteomes from additional aphid species. Most of the proteins identified in the resorcinol diet were also present in the water diet and represented proteins with a plethora of functions in addition to a large number of unknowns. About half of the salivary proteins were not predicted for secretion or had canonical secretion signal peptides. We also analyzed the phosphorylation states of M. euphorbiae salivary proteins and identified three known aphid effectors, Me_WB01635/Mp1, Me10/Mp58, and Me23 that carry phosphorylation marks. In addition to insect proteins, tomato host proteins were also identified in aphid saliva. Our results indicate that aphid saliva is complex and provides a rich resource for functional characterization of effectors.