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Objective: This study was to clone, identify and analyze the characteristics of egG1Y162 gene from Echinococcus granulosus. Methods: Genomic DNA and total RNAs were extracted from four different developmental stages of protoscolex, germinal layer, adult and egg of Echinococcus granulosus, respectively. Fluorescent quantitative PCR was used for analyzing the expression of egG1Y162 gene. Prokaryotic expression plasmid of pET41a-EgG1Y162 was constructed to express recombinant His-EgG1Y162 antigen. Western blot analysis was performed to detect antigenicity of EgG1Y162 antigen. Gene sequence, amino acid alignment and phylogenetic tree of EgG1Y162 were analyzed by BLAST, online Spidey and MEGA4 software, respectively. Results: EgG1Y162 gene was expressed in four developmental stages of Echinococcus granulosus. And, egG1Y162 gene expression was the highest in the adult stage, with the relative value of 19.526, significantly higher than other three stages. Additionally, Western blot analysis revealed that EgG1Y162 recombinant protein had good reaction with serum samples from Echinococcus granulosus infected human and dog. Moreover, EgG1Y162 antigen was phylogenetically closest to EmY162 antigen, with the similarity over 90%. Conclusion: Our study identified EgG1Y162 antigen in Echinococcus granulosus for the first time. EgG1Y162 antigen had a high similarity with EmY162 antigen, with the genetic differences mainly existing in the intron region. And, EgG1Y162 recombinant protein showed good antigenicity.

Objective: This study is to clone and identify B- and T-cell combined epitopes from Em95 antigen. Methods: The B- and T-cell combined epitopes were predicted using bioinformatic software. Two DNA sequences of Em95-1 (which contained the coding region of one B- and T-cell combined epitope) and Em95-2 (which contained the coding regions of two B- and T-cell combined epitopes) were amplified by PCR. Em95-1 and Em95-2 were cloned into pET32a vector for protein expression. Rabbit was immunized with the expressed proteins of rEm95-1 and rEm95-2 to produce polyclonal antibodies. The immunogenicity and antigenicity of rEm95-1 and rEm95-2 were examined by Western blot analysis. Results: The three B- and T-cell combined epitopes were successfully cloned and expressed in PET32a vector. The recombinant antigens of rEm95-1 and rEm95-2 could specifically bind the human serum from patients with alveolar echinococcosis and specifically bind the prepared polyclonal antibodies. Conclusion: Three B- and T-cell combined epitopes were successfully cloned with good immunogenicity and antigenicity. Our data suggest that B- and T-cell combined epitopes predicted from the Em95 antigen may be used for the construction of high-valence vaccines and as targets for prevention of echinococcosis.

Medical microbiology is an important part in microbiology teaching. However, there are some problems in our experiment teaching now, we endeavor to work out countermeasures and believe that the experiment teaching level and teaching quality will be improved after improve.