Chitosan nanoparticles: A positive immune response modulator as display in zebrafish larvae against Aeromonas hydrophila infection.

Chitosan nanoparticles (CNPs) were synthesized by ionic gelation method and its immunomodulatory properties were investigated in zebrafish larvae. Average particle size and zeta potential were 181.2 nm and +37.2 mv, respectively. Initially, toxicity profile was tested in zebrafish embryo at 96 h post fertilization (hpf) stage using medium molecular weight chitosan (MMW-C) and CNPs. At 5 μg/mL, the hatching rate was almost similar in both treatments, however, the survival rate was lower in MMW-C compared to CNPs exposure, suggesting that toxicity effect of CNPs in hatched larvae was minimal at 5 μg/mL compared to MMW-C. Quantitative real time PCR results showed that in CNPs exposed larvae at 5 days post fertilization (5 dpf) stage, immune related (il-1β, tnf-α, il-6, il-10, cxcl-18b, ccl34a.4, cxcl-8a, lyz-c, defβl-1, irf-1a, irf-3, MxA) and stress response (hsp-70) genes were induced. In contrast, basal or down regulated expression of antioxidant genes (gstp-1, cat, sod-1, prdx-4, txndr-1) were observed. Moreover, zebrafish larvae (at 5 dpf stage) exposed to CNPs (5 μg/mL) showed higher survival rate at 72 h post infection stage against pathogenic Aeromonas hydrophila challenge compared to controls. These results suggest that although CNPs can have toxic effects to the larvae at higher doses, CNPs exposure at 5 μg/mL could enhance the immune responses and develop the disease resistance against A. hydrophila, which could be attributed to its strong immune modulatory properties.