Stress Response of Citrus Leaves under Mechanical Damage and Huanglongbing Disease Infection Using Plasmonic TiO2 Nanotube Substrate-Based Imprinting Mass Spectrometry Imaging

Mapping the molecular signatures and metabolic regulation of plant tissues under biotic/abiotic stresses and defensive responses has become a subject of increasing interest in plant biology and systems biology, but determining when and where specialized metabolites are produced and accumulated currently remains a somewhat elusive goal. Herein, we demonstrated the use of a TiO2 nanotube-based composite substrate modified with plasmonic gold nanoparticles and hydrophobic polydopamine (AuNP-hPDA-TDNT) for surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) analysis of a wide range of pesticides and for visualizing the stress-responsive metabolites of citrus leaves during various plant defense processes. This method enabled the visualization of non-uniform and tissue-specific distribution patterns of functional metabolites of citrus leaves that were mechanically damaged, fed to larvae, and infected by Huanglongbing disease. Interestingly, some specialized metabolites exhibited different accumulation and regulation patterns for mechanical damage and larval feeding, suggesting that plant-derived secondary metabolites exercise specific defensive functions with respect to various damage processes. Moreover, the early diagnosis and detection of HLB disease-associated biomarkers can facilitate the prevention of citrus HLB diseases. Overall, this imprinting MS imaging strategy will expand the scope of MS techniques in plant biology, providing more biologically relevant insights into the biosynthesis, accumulation, and defensive role of bioactive metabolites in economically important plants.