Sensitivity to chitosan and chitosan nanoparticles by three Colletotrichum species belonging to C. acutatum species complex

Chitosan (CH) is one of the most studied biomaterials in nanotechnology due to its unconstrained biological properties like antimicrobial and plant growth regulatory activity, biodegradability, biocompatibility and non-toxicity to humans. Although many reports of the application of chitosan in agriculture are available, much work has to be done using chitosan nanoparticles (CHNPs) to fully understand their true potential. The advantages of CHNPs over bulk materials relies in their small size and large surface area. CHNPs exhibited higher antimicrobial activity than bulk materials and their cellular uptake into cells was higher than that of chitosan molecules. In this study, antifungal effects of chitosan nanoparticles (CHNPs) and commercial chitosan (ChitoPlant®) against Colletotrichum lupini, C. nymphaeae and C. acutatum sensu stricto were compared using a microdilution assay in 96-well microtiter plates. Fungi were isolated from lupin, laurel and fig respectively and were molecularly identified using ITS and GAPDH sequences. CHNPs were synthesized via the ionotropic gelation method using sodium tripolyphosphate (TPP) as cross-linking agent and were characterized by size (DLS), and zeta potential analysis. The size of CHNPs was approximately 98 nm with a zeta potential of 12mV. CHNPs (5000 ppm) were found most effective showing 90-100% growth inhibition of the three Colletotrichum spp. which showed a tolerance to CH (5000 ppm) with percentage of growth inhibition ranging from 5 to 29%. The 260 nm absorbing cellular material increased significantly when compared to untreated control samples indicating a membrane damage mediated by CHNPs. The results suggest that CHNPs have remarkable potential for further in vivo applications.