Abstract

Nosema bombycis , the causal agent of silkworm pébrine disease, causes substantial economic losses to sericulturists annually. Previously, 19 serpin genes (NbSPNs) were identified in this parasite, but most of their functions remain unidentified yet. Here, we provide a functional and cellular characterization of NbSPN4 and NbSPN5. Bioinformatics tools predicted four cis -regulatory motifs in the promoter region of NbSPN genes. A yeast signal sequence trap (YSST) assay confirmed the computationally predicted N-terminal signal peptide for NbSPN4 but not for NbSPN5. Immunofluorescence assay revealed that NbSPN4 was localized to the nucleus and NbSPN5 to the cytoplasm of infected host BmE cells. Recombinant NbSPN4/5 proteins significantly inhibited host hemolymph melanization and phenoloxidase activity in vitro, demonstrating their immune-regulatory roles. These findings provide essential insights into the roles of NbSPNs in host–pathogen interactions during N. bombycis infection.