Antimicrobial activities of chitosan nanoparticles against pathogenic microorganisms in Nile tilapia, Oreochromis niloticus

Nanotechnology is a recent unique technique generally used for nutrition and therapy purposes among others. In this respect, the present study was carried out to evaluate the antimicrobial activity of chitosan nanoparticles (CNP) against various microorganisms (fungi and bacteria) isolated from diseased or health Nile tilapia, Oreochromis niloticus. The CNP was prepared based on the ionic gelation of chitosan with tripolyphosphate anion and its mean size was 35 nm with a narrow size distribution and zeta potential of 61.2 mV. The lethal dose of pathogenic bacterial isolates for Nile tilapia was successfully standardized. Clinical signs including weakness, slower movement, swimming closer to the surface, fin hemorrhages, and red patches at the gut regions were observed. Enlargement of spleen followed by tissue necrosis along with signs of hemorrhagic septicemia was also seen in infected fish. Fungal and bacterial isolates were exposed to different CNP doses and it is noticed that CNP inhibited all examined fungal and bacterial isolates in a dose-dependent manner. However, high CNP doses (80 mu g/ml) gave highest inhibition zones where Aspergillus flavus, Mucor sp., and Candida sp. were more susceptible, whereas Aspergillus niger, A. fumigatus, and Fusarium sp. were more resistant. Similarly, largest inhibition zones of tested bacteria were obtained at high CNP dose (20 mu g/ml). And Aeromonas sobria, A. hydrophila, and Pseudomonas aeruginosa were the most susceptible bacterial strain; meanwhile, Staphylococcus aureus and Pseudomonas fluorescens were the most resistant ones. The minimal inhibitory concentration of CNP against the examined bacteria ranged from 0.156 to 2.5 mu g/ml causing their minimal counts. The transmission electron microscope images revealed that CNP showed antagonistic action against A. hydrophila causing disruption of cell membranes and the leakage of cytoplasm. In a practical experiment, Nile tilapia fed dietary CNP at levels of 0.0 and 1.0 g/kg diet for 3 weeks and post-challenged with different pathogenic bacteria via intraperitoneal injection. It is noticed that fish fed a CNP-enriched diet showed less mortality with all bacterial strains (6.7-20%), while, fish fed a CNP-free diet showed highest mortality (66.7-100%). The dietary CNP protected Nile tilapia efficiently against A. hydrophila, A. sobria, and Streptococcus agalactiae infections with relative level of protection (RLP) value of 93.3%, while the RLP against Staphylococcus aureus was 70.0%.