Elucidation of innate immune components in the epidermal mucus of different freshwater fish species

Abstract

Background. Fish epidermal mucus is a key component of innate immune system and plays a major role in protecting fish against invading pathogenic microbes. The information on the role of epidermal mucus components is insufficient for many commercially important freshwater fish species. Hence, this study was directed to understand the innate immune components of epidermal mucus in these fish species. Materials and methods. Mucus samples were obtained from African catfish, Clarias gariepinus (Burchell, 1822); Indonesian snakehead, Channa micropeltes (Cuvier, 1831); striped snakehead, Channa striatus (Bloch, 1793); marble goby, Oxyeleotris marmorata (Bleeker, 1852); Nile tilapia, Oreochromis niloticus (Linnaeus, 1758); and Asian redtail catfish, Hemibagrus nemurus (Valenciennes, 1840). The variation in the protein profile of the mucus were observed using SDS-PAGE analysis. The specific activities of various hydrolytic enzymes including lysozyme, alkaline phosphatase, esterase, and protease were analysed in the mucus and the levels of enzyme activities were compared among the experimental fishes. We also elucidated the effect of various protease inhibitors on the protease activity and identified the type of proteases in the fish mucus by azocasein hydrolysis assay and zymography. Results. Significantly (P < 0.05) highest levels of lysozyme-, alkaline phosphatase-, and esterase activities were observed to be 15.77, 18.96, and 10.65 U · mg–1 protein respectively in C. striatus followed by 13.67, 15.06, and 16.67 U · mg–1 protein, respectively in C. gariepinus. The highest level of protease activity was recorded in H. nemurus (1630.97 U · mg–1 protein) and C. gariepinus (1596.4 U · mg–1 protein). A wide range of variation was noticed in the enzyme activities of fishes, even from the same genus, for example C. micropeltes and C. striatus. Specific inhibitors were added to azocasein hydrolysis assay and zymography experiments to characterize the individual proteases present in the mucus of each fish species. This result revealed the highest level of serine proteases in mucus of redtail catfish and African catfish compared to cysteine and metalloproteases. In contrast, Indonesian snakehead and marble goby had similar levels of serine, cysteine and metalloproteases. Conclusion. This study provides an insight into the presence of various epidermal mucus enzymes in fishes and therefore believed to be the components of innate immune system that may possibly provide protection for the fish against invading pathogenic microbes.

Keywords

• fish mucus
• innate immunity
• enzyme activity
• azoc