To explore the sex difference in proteins between male and female adult worm of Schistosoma japonicum.Two-electrophoresis was used to analyse the difference of protein between the male and female adult worm of S. japonicum (Chinese strain).Two-dimensional electrophoresis analysis revealed that at the site of 43 kDa and an isoelectric point (pI) of 5.60-5.90 the male worm exhibited a band carrying a number of spots and dots, being longer and wider than that exhibited by the female worm. The female worm exhibited 7 specific dots.The sex differences in proteins between male and female adult worms of S. japonicum are significant.
A 558 bp cDNA fragment was amplified by RT-PCR from adult Schistosoma japonicum(Chinese strain) mRNA with a pair of primers that were designed according to published Sj21.7p gene encoding 21.7 kD protein of Schistosoma japonicum(Philippines strain). Sequence analysis indicated that this frame, named Sj21.7 (Ch), with 99% homology to Sj21.7 p, contained a complete open reading fragment (ORF) of 21.7 kD protein gene of Schistosoma japonicum(Chinese strain). The amino acid sequence shared 98% homology with 21.7 kD protein of Schistosoma japonicum. This fragment was cloned into the expression vector pET28a (+) and subsequently expressed in Escherichia coli with IPTG induction. SDS-PAGE analysis revealed that the molecular weight of this expressed product was 25.4 kD. Western blotting showed that the recombinant protein reacted well with the rabbit serum immunized with Sj worm antigen, indicating that this expressed product had good antigenicity.
To understand the characteristics of pro-apoptotic gene SjBAD of Schistosoma japonicum, such as its biology, immunology, and transcriptional expression, and evaluate its potential of the recombinant protein as a vaccine candidate for schistosomiasis.SjBAD was amplified by PCR and subeloned into a pET-28a(+) vector, and the recombinant plasmid was transformed into competent E. coli BL21 for producing recombinant protein. The expressions of SjBAD in different development stages of schistosomula and 42-day male and female worms were determined by real-time PCR. The immunogenicity of the recombinant protein was analyzed by Western blotting and ELISA. The potential of this protein as a vaccine candidate molecule was assessed by testing the worm reduction rate and liver egg reduction rate in the BALB/c mice immunized by the recombinant antigen SjBAD.SjBAD was successfully cloned, the recombinant plasmid pET-28a(+)-SjBAD was successfully expressed in E. coli, and the molecular weight of the recombinant protein was around 22 kDa. Western-blotting showed that the recombinant protein had good immunogenicity. The recombinant protein could induce high level of specific IgG antibodies in the BALB/c mice. SjBAD was expressed in all tested 7-, 14-, 21-, 28-, 35- and 42-day worms, and was highly expressed in 14-day schistosomula, while the expression level in 42-day male worms was higher than that in 42-day female worms. Two in- dependent animal trials showed that 30.82% and 27.87% worm reduction rates, as well as 42.52% and 45.84% liver eggs reduction rates were obtained in the rSjBAD vaccinated group compared with those of the blank control group (both P < 0.05).The proapoptotic gene SjBAD is successfully cloned and expressed. The gene is expressed in different development stages of S. japonicum. The rSjBAD vaccinated BALB/c mice can obtain a partial protective immunity against S. japonicum infection.
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