Molecular Cloning And Function Identification Of Adhesion Protein Gene Family From Taenia Multiceps

Coenurosis, caused by the larval stage(Coenurus cerebralis) of Taenia multiceps, is a common parasitic infection in sheep and goats, and occasionally in humans. The parasite can usually lead to encephalitis and neurological symptoms in infected hosts, or even death sometimes. Coenurosis cerebralis infections in humans have been widely reported in Asia, Europe, USA and Africa, which raise the public health concerns in the world. With the development of molecular biology and immunology technique, recombinant antigen vaccines have been developed against infection with taeniid species. But no such studies could be available on vaccines against Taenia multiceps infection.In this study, the genomic DNA sequence and c DNA sequence of Tm Adh genes were amplified by PCR and analyzed by bioinformatics softwares. The structures of Tm Adh1-6 genes were comprised of N terminal signal peptide and one Fn III domain and shared 44.9-99.2% amino acid sequence similarity. Tm Adh1, Tm Adh2 and Tm Adh4 c DNA sequences have been cloned and sequenced and Tm Adh3 c DNA sequence was artificially synthesized. The Tm Adh1-4 gene fragments were then ligated into p GEX4T-1 vector and successfully expressed in Escherichia coli. The weights of recombinant proteins Tm Adh1-4 were 39.0, 38.1, 38.9 and 37.7 ku, respectively. Western-blot analysis showed the recombinant Tm Adh3 protein could react with sheep serum infected with Coenurus cerebralis, but the recombinant Tm Adh1, Tm Adh2 and Tm Adh4 proteins can not react with sheep serum infected with Coenurus cerebralis. The adhesion assay showed that Tm Adh1-4 could adhere to NRK and CHO cells. Immunohistochemistry assay showed that the Tm Adh1 protein was mainly distributed on epithelium, rostellum(most in small hook) and a little in organization. The Tm Adh2 protein was mainly distributed in organization, rostellum, a little on epithelium. The Tm Adh3 protein was mainly distributed on rostellum, small hook and epithelium. The Tm Adh4 protein was mainly distributed in organization, rostellum and small hook. Sheep immunized with recombinant Tm Adh1, Tm Adh3 and Tm Adh4 protein produced higher antibody levels. The Tm Adh1 group necropsy result with 61.17% reduction rate of cyst, Tm Adh3 group necropsy result with 53.17% reduction rate of cyst, Tm Adh4 group necropsy result with 50.76% reduction rate of cyst after challenged with T. multiceps eggs compired those in controls. These results indicated that Tm Adh1, Tm Adh3 and Tm Adh4 recombinant proteins would be promising vaccine candidates against T. multiceps infection. The present study will lay a foundation for the further development of recombinant antigen vaccine of cerebral conurosis.