›› 2011, Vol. 29 ›› Issue (3): 1-161-166.

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Cloning, Expression and Stage-specific Analysis of Schistosoma japonicum P7 Antigen and Evaluation of its Value in Early Diagnosis

 XU  Bin, DUAN  Xin-Wei, LU  Yan, CHEN  Shen-bo, FENG  Zheng, HU  Wei   

  1. 1 National Institute of Parasitic Diseases，Chinese Center for Disease Control and Prevention；Key Laboratory of Parasite and Vector Biology，MOH；WHO Collaborating Center of Malaria，Schistosomiasis and Filariasis，Shanghai 200025，China；2 Biotechnology School，East China University of Science and Technology，Shanghai 200237，China
  • Online:2011-06-30 Published:2012-09-27

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Abstract:   Objective   To clone and express Schistosoma japonicum P7 antigen（GenBank accession No. EU121231）, analyze stage-specific transcription and expression of the antigen, and evaluate its value in early diagnosis.  Methods  The positive clone （P7） screened from schistosomula cDNA library was amplified by PCR. The PCR product was subcloned into prokaryotic expression vector pET28a. The recombinant plasmids were identified by restrictive enzymes digestion. The positive recombinant plasmids were transformed into E. coli BL21（DE3）, induced by IPTG for expression and purified. The diagnostic value of P7 recombinant protein was evaluated by Western blotting analysis. RT-PCR and Western blotting were used to investigate the differential transcription and expression of P7 during the developmental stages. The specific antibodies against P7 recombinant protein in the sera of S. japonicum-infected rabbits at 14 d post-infection, sera of schistosomiasis （28 cases）, clonorchiasis （30 cases） and paragonimiasis （20 cases） patients, and sera of healthy people (30 cases) were detected by ELISA, respectively.  Results   The expression vector of p7/pET28a was established and the P7 recombinant protein （about Mr 20 100） was expressed in E. coli. Western blotting analysis showed that the recombinant protein was specifically recognized by immunized rabbit sera, and sera from mice on the 14th day post infection, but was not recognized by the sera of mice at 42 d post-infection. P7 mRNA was detected in cercariae, schistosomula and adult worms, while the protein was only found in schistosomula. The positive rate of rabbit sera collected at 14 d post-infection was 83.3%（15/18）. The sensitivity and specificity of ELISA for diagnosis of schisto-somiasis japonica were 75.0% （21/28） and 93.8% （75/80）, respectively. And the P7 protein showed cross reaction with sera of clonorchiasis and paragonimiasis patients with positive rates of 6.7%（2/30） and 5.0%（1/20）, respectively.  Conclusion   P7 antigen might be a potential candidate for early diagnosis of schistosomiasis.

Key words: Schistosoma japonicum, P7 antigen, Stage-specific analysis, Diagnosis