The Preliminary Study On Schistosoma Japonicum Tegumental Protein SjOST48

Schistosomiasis is a serious parasitic zoonosis which does great harm to human health and livestock production. Praziquantel(PZQ) is efficient against adult worms, but does not kill larvae or prevent reinfection, then screening and identifying efficient and safe vaccine candidate antigens against schistosomiasis have been the main targets for researchers. The tegument proteins of S. japonicum are considered to be good vaccine candidates and drug targets, SJCHGC01743 was picked out after proteomics analysis of tegument-exposed proteins from schistosomes. In this study, the gene coding this protein was cloned and expressed, and a preliminary study of basic characteristics and biological function of SjOST48 was carried out.To identify SJCHGC01743 gene and evaluate the potentiality of the recombinant protein as a new vaccine candidate for schistosomiasis, a cDNA with 1248 nucleotides was isolated from 28-day-old schistosomes cDNAs by PCR. Sequence analysis revealed that SJCHGC01743 was a 48-kDa subunit of the oligosaccharyltransferase complex (OST48) and named as SjOST48. Real-time PCR analysis indicated that this gene was expressed in all developmental stages of schistosomes and had the highest expression level in 28d worms, the level of gene transcription in female worms was significantly higher than that of male worms. Then recombinant plasmid pET28a(+)-SjOST48 was successfully constructed and expressed in Escherichia coli BL21 (DE3). The purified recombinant protein (rSjOST48) emulsified with ISA206 adjuvant was used to immunize BALB/c mice for three times to produce immune sera. Western blotting analysis showed that rSjOST48 had good immunogenicity. Indirect immunofluorescent analysis revealed that SjOST48 was mainly distributed on the tegument of the worms. The result of ELISA indicated that the rSjOST48 vaccinated group could induce a significant increase in the level of specific IgG, IgGl and IgG2a. Two immunoprotection experiments showed that the vaccination of rSjOST48 in mice induced 21.58%-32.62%(P< 0.05) and 50.94%-57.61%(P< 0.01) reduction in the numbers of worm and eggs in liver respectively, compared with that of the blank control group.RNAi was used to study the biological function of SJOST48 in this study. Fore specific siRNAs targeting at SjOST48 gene were designed to silence the gene by soaking the parasite in medium for 3 days in vitro. The result of real time PCR result indicated that siRNA-713 induced the best inhibitory effect and it led to 40% reduction in the transcriptional level of SJOST48 compared to the control group. Then siRNA-713 was injected into the mice infected with S.japonicum to silence SjOST48 gene in vivo, the result showed that the group treated with siRNA-713 induced 29.50% and 30.08% reduction in worms burden and liver eggs count respectively, compared with that of the irrelevant control group after injection 7 times.This study provides the foundation for proceeding further research on the biological function of SjOST48 and screening new vaccine candidates for schistosomiasis.