In vitro immunostimulatory potential of fungal β-glucans in pacific red snapper (Lutjanus peru) cells

Abstract

This study attempts to describe the immunostimulatory effects of three fungal glucans on innate immunity responses in an in vitro assays using Pacific red snapper leukocytes. First, the yield glucans obtained was higher in Aspergillus niger, follow by Aspergillus ochraceus and Alternaria botrytis (40, 20 and 10%, respectively). Structural characterization of these fungal glucans by proton nuclear magnetic resonance (NMR) indicated structures containing (1–6)-branched (1–3)-β-D-glucan. The immunostimulatory activity of fungal glucans were assessed in head-kidney leukocytes at 24 h using colorimetric assays and molecular gene expression. In addition, the response against bacterial infection using Aeromonas hydrophila was evaluated by flow cytometry with annexin V/propidium iodide. Leukocytes responded positively to fungal glucans where the viability was higher than 80%. Interestingly, A. niger β-glucans enhanced the phagocytic ability and capacity in head-kidney leukocytes. Immunological assays reveled an increased in nitric oxide production, myeloperoxidase, superoxide dismutase and catalase activities, in fish stimulated with A. niger β-glucans. Induction of cytokines (IL-1β, TNF-α, IL-6, IL-8 and IL-12) were more pronounced in A. niger β-glucans leukocytes stimulated compared to other group. Finally, flow cytometry assay showed that A. botrytis and A. niger β-glucans were able to inhibit apoptosis caused by Aeromonas hydrophila in the Pacific red snapper leukocytes indicating an immunostimulant potent response by fungi derived-glucans. These results strongly support the idea that fungal β-glucans can stimulate the immune mechanism in head-kidney leukocytes and that Aspergillus niger β-glucan possess immunostimulatory properties cell increasing viability, and reducing necrotic cell death caused by Aeromonas hydrophila.