Identification and characterization of paramyosin from cyst wall of metacercariae implicated protective efficacy against Clonorchis sinensis infection

• Published in: PloS one 2012-03, Vol.7 (3), p.e33703-e33703
• Main Authors: Wang, Xiaoyun, Chen, Wenjun, Lv, Xiaoli, Tian, Yanli, Men, Jingtao, Zhang, Xifeng, Lei, Huali, Zhou, Chenhui, Lu, Fangli, Liang, Chi, Hu, Xuchu, Xu, Jin, Wu, Zhongdao, Li, Xuerong, Yu, Xinbing
• Format: Article
• Language: English
• Subjects: Acids; Amino Acid Sequence; Amino acids; Analysis; Animals; Antibodies; Antibodies - analysis; Antibodies - immunology; Bacillus subtilis; Bioinformatics; Biology; Bladder; Cell-mediated immunity; Cercaria; Clonorchiasis - parasitology; Clonorchiasis - prevention & control; Clonorchis sinensis; Clonorchis sinensis - immunology; Cytokines; Deoxyribonucleic acid; Dictyocaulus; DNA; DNA vaccines; Epidemics; Epitopes; Health aspects; Health risks; Humoral immunity; Immune response; Immunity; Immunity, Cellular; Immunization; Immunogenicity; Immunoglobulin G; Immunoglobulin G - blood; Interferon; Interferon-gamma - metabolism; Interleukin 4; Interleukin-4 - metabolism; Leishmania donovani; Localization; Lymphocytes B; Lymphocytes T; Medical research; Medicine; Metacercariae - metabolism; Molecular Sequence Data; Nucleic acids; Parasites; Parasitic diseases; Pets; Proteins; Proteome - metabolism; Public health; Rats; Rats, Sprague-Dawley; Recombinant proteins; Recombinant Proteins - genetics; Recombinant Proteins - immunology; Recombinant Proteins - metabolism; Schistosoma japonicum; Schistosoma mansoni; Sequence Alignment; Splenocytes; Tegument; Trichinella spiralis; Tropical diseases; Tropomyosin - genetics; Tropomyosin - immunology; Tropomyosin - metabolism; Urinary bladder; Vaccines; Vaccines, DNA - immunology; γ-Interferon
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0033703

Human clonorchiasis has been increasingly prevalent in recent years and results in a threat to the public health in epidemic regions, motivating current strategies of vaccines to combat Clonorchis sinensis (C. sinensis). In this study, we identified C. sinensis paramyosin (CsPmy) from the cyst wall proteins of metacercariae by proteomic approaches and characterized the expressed recombinant pET-26b-CsPmy protein (101 kDa). Bioinformatics analysis indicated that full-length sequences of paramyosin are conserved in helminthes and numerous B-cell/T-cell epitopes were predicted in amino acid sequence of CsPmy. Western blot analysis showed that CsPmy was expressed at four life stages of C. sinensis, both cyst wall proteins and soluble tegumental components could be probed by anti-CsPmy serum. Moreover, immunolocalization results revealed that CsPmy was specifically localized at cyst wall and excretory bladder of metacercaria, as well as the tegument, oral sucker and vitellarium of adult worm. Both immunoblot and immunolocalization results demonstrated that CsPmy was highly expressed at the stage of adult worm, metacercariae and cercaria, which could be supported by real-time PCR analysis. Both recombinant protein and nucleic acid of CsPmy showed strong immunogenicity in rats and induced combined Th1/Th2 immune responses, which were reflected by continuous high level of antibody titers and increased level of IgG1/IgG2a subtypes in serum. In vaccine trials, comparing with control groups, both CsPmy protein and DNA vaccine exhibited protective effect with significant worm reduction rate of 54.3% (p