Molecular characterization of a novel aspartyl protease-1 from Trichinella spiralis.

ABSTRACT

The aim of this study was to characterize the biological properties of a novel aspartic protease-1 from Trichinella spiralis (TsASP1) and evaluate its potential in inducing immune response. TsASP1 gene was cloned and expressed in Escherichia coli BL21 (DE3). On Western blotting analysis with anti-rTsASP1 serum, native TsASP1 was detected in various T. spiralis phases other than newborn larvae (NBL). qPCR results showed that TsASP1 transcription was the highest in intestinal infective larvae (IIL) and the lowest in the NBL stage. Immunofluorescence test result shows that native TsASP1 was principally localized in stichosome, muscle cells of muscle larvae (ML) and IIL, and surrounded intrauterine embryos in female adult worms (AW). After silencing TsASP1 gene of the ML by siRNA, the worm development was significantly inhibited, showed by shorter AW and more wrinkles and longitudinal crack on epicuticle of AW on scanning electron microscopy; the AW and ML burdens were reduced by 41.82 and 56.36% respectively, compared with the control siRNA or PBS group (P < 0.001). Immunization of mice with rTsASP1 elicited an evident antibody response (serum IgG, IgG1/IgG2a and enteral sIgA), and systemic (spleen) and intestinal local mucosal (mesenteric lymph node) cellular immune response, demonstrated by a prominent elevation of IFN-γ and IL-4. The results suggested TsASP1 participated in T. spiralis development and survival in host, and immunization of mice with rTsASP1 induced systemic/intestinal local mucosal humoral and cellular immune response against Trichinella.