Quantitative gene expression and in situ localization of scygonadin potentially associated with reproductive immunity in tissues of male and female mud crabs, Scylla paramamosain.

Scygonadin (Scy) is an important antimicrobial peptide which was first isolated from the seminal plasma of Scylla serrata (now renamed as Scylla paramamosain). Elucidation of the Scy expression pattern in tissues will help in understanding its potential function associated with the reproductive immunity. In our study, Scy mRNA transcripts and its protein were found widely distributed in mature male and female crabs. Scy mRNA transcripts were significantly demonstrated in the ejaculatory duct and hemocytes of males but were much less expressed in the other tissues tested. In addition, Scy mRNA transcripts were discerned in a number of cells in the glandular epithelium of the inner wall and in the secretion inside the ejaculatory duct using the in situ hybridization method. In females, Scy mRNA transcripts were obviously demonstrated in the hemocytes and gills but weakly detected in other tissues tested. The copy number of scygonadin mRNA transcripts in the ejaculatory duct of males was greatly higher than those in other tissues, in particular, was over 60,000 fold that in the hemocytes of females. Using immunohistochemistry, the Scy protein was found at higher levels in male tissues than in female ones, particularly in the reproductive duct of males. It was also interesting to note that Scy gene expression was not significantly induced with lipopolysaccharide challenge. However, it was highly expressed in the ejaculatory duct and the seminal vesicle of pre-copulatory males and in the spermathecae of post-copulatory females under mating conditions. The results suggested that Scy, as an important antimicrobial component, probably performed more functions in males, and was likely to be involved in a function associated with crab fertilization and reproduction in both males and females during mating.