Poly(DADMAC) incorporated lipid nanoparticles enhance the delivery of antimicrobial peptides into plant cells

Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium that causes the citrus greening disease. One of the main challenges in treating citrus greening disease is delivering the therapeutic agents to the vascular system where CLas bacteria reside. This study proposes Polydiallyldimethylammonium chloride (poly(DADMAC)) incorporated lipid nanoparticles as a potential method to deliver the drugs into the plant vascular system via foliar application. The positive charge of poly(DADMAC) is expected to permeabilize the negatively charged cell surface, allowing therapeutic particles to permeate through the cell surface. Glyceryl monostearate-based lipid nanoparticles were synthesized using a high-shear homogenization method using Tween 80 and poly(DADMAC) as surfactants, and were characterized using a particle size analyzer. Nanoparticles containing 0.5% Tween 80 and 0.5% poly(DADMAC) displayed the most desirable characteristics, with an average particle size of 513.6 nm, an average zeta potential of + 13.78 mV, and an encapsulation efficiency of 94.8%. Successful encapsulation of anti-microbial peptides in the nanoparticles was verified using fluorescence imaging and fluorescence lifetime analysis. Fluorescence imaging of leaf vascular tissues showed significantly higher fluorescence in samples treated with nanoparticles encapsulating fluorescent-tagged peptides, compared to untreated samples and those treated with aqueous peptides. The presence of the peptide in nanoparticle-treated leaves was also validated using Matrix-Assisted Laser Desorption/Ionization - Time of Flight (MALDI-TOF) mass spectrometry. Thus, these results indicate that poly(DADMAC) incorporated nanoparticles can enhance the migration of peptides into vascular tissues. This can be an effective and convenient technique to deliver therapeutics against citrus greening disease.