Biological effects of Thymol loaded chitosan nanoparticles (TCNPs) on bacterial plant pathogen Xanthomonas campestris pv. campestris

Engineered nanomaterials can provide eco-friendly alternatives for crop disease management. Chitosan based nanoparticles has shown beneficial applications in sustainable agricultural practices and effective healthcare. Previously we demonstrated that Thymol loaded chitosan nanoparticles (TCNPs) showed bactericidal activity against Xanthomonas campestris pv campestris (Xcc), a bacterium that causes black rot disease in brassica crops. Despite the progress in assessing the antibacterial action of TCNPs, the knowledge about the molecular response of Xcc when exposed to TCNPs is yet to be explored. In the present study, we combined physiological, spectroscopic and untargeted metabolomics studies to investigate the response mechanisms in Xcc induced by TCNPs. Cell proliferation and membrane potential assays of Xcc cells exposed to sub-lethal concentration of TCNPs showed that TCNPs affects the cell proliferation rate and damages the cell membrane altering the membrane potential. FTIR spectroscopy in conjunction with untargeted metabolite profiling using mass spectrometry of TCNPs treated Xcc cells revealed alterations in amino acids, lipids, nucleotides, fatty acids and antioxidant metabolites. Mass spectroscopy analysis revealed a 10-25% increase in nucleic acid, fatty acids and antioxidant metabolites and a 20% increase in lipid metabolites while a decrease of 10-20% in amino acids and carbohydrates was seen in in TCNP treated Xcc cells. Overall, our results demonstrate that the major metabolic perturbations induced by TCNPs in Xcc are associated with membrane damage and oxidative stress, thus providing information on the mechanism of TCNPs mediated cytotoxicity. This will aid towards the development of nano- based agrochemicals as an alternative to chemical pesticides in future.