Proteome-Wide Prediction of Putative Bemisia tabaci Effector Candidates and Transient Validation of BtApe-Mediated Suppression of Tomato Immune Responses

Abstract

Phloem-feeding insects such as aphids, planthoppers, and whiteflies use salivary proteins to aid feeding, adapt to hosts, and manipulate plant defenses, shaping a dynamic plant-herbivore conflict. Among them, the sweet potato whitefly, Bemisia tabaci, is a major global pest that threatens food security by causing feeding damage and transmitting plant viruses. The salivary secretory proteins of B. tabaci remain poorly characterized. We conducted a proteome-wide hypothesis-based screening of the B. tabaci MEAM1/B-biotype reference proteome using secretion, membrane-topology, localization, and effector-likelihood filters to prioritize candidate secreted proteins. Among 1404 proteins screened in EffectorP 3.0, 884 were identified as putative effector candidates and reported as computationally predicted rather than a validated effectorome. Most of these candidates were cytoplasmic (741), with fewer following the classical extracellular secretory pathway (143). Functional annotation showed strong enrichment of proteases (cathepsin B, cathepsin L, and serine proteases) and thioredoxin-related proteins, some of which resemble proteins implicated in feeding systems among hemipteran insects, supporting their prioritization for validation. Protein-association analyses highlighted protease-related candidates as central predicted network hubs. Transient validation of one prioritized candidate, BtApe, showed attenuation of chitin-associated oxidative staining and normalized defense-marker responses in tomato, supporting BtApe as an immune-modulatory candidate and defining a focused framework to validate salivary gland expression and secretion, as well as loss of function effects.