In order to study the inhibitory effect of the adhesion protein of Streptococcus pneumoniae,pneumococcal surface adhesin A(PsaA),and anti-rPsaA serum on the adhesion to bovine embryonic tracheal epithelial cells,this study amplified psaA gene of S.pneumoniae by PCR.The recombinant plasmid pET32a-PsaA was constructed for prokaryotic expression.The recombinant plasmid identified as positive by double digestion was transformed into BL21(DE3)competent cells and expressed by IPTG induction.SDS-PAGE and western blot determined the solubility and antigenicity of the recombinant protein.The results showed that the recombinant PsaA protein(rPsaA)was expressed in soluble form and showed good antigenicity.The adhesion of S.pneumoniae to bovine duct epithelial cells was detected by indirect immunofluorescence assay after bovine respiratory tract epithelial cells were incubated with 1.3×108cfu/mL S.pneumoniae dilution.The results showed that S.pneumoniae could adhere to bovine embryonic trachea epithelial cells.Bovine embryonic tracheal epithelial cells treated with different concentrations of rPsaA proteins were incubated with S.pneumoniae diluted solution,and the number of adhesion bacteria was statistically analyzed.The results showed that the rPsaA protein could significantly inhibit the adhesion of S.pneumoniae to bovine embryonic tracheal epithelial cells.The bovine embryonic tracheal epithelial cells were infected with S.pneumoniae that pre-incubated with the anti-rPsaA polyclonal antibodies or anti-S.pneumoniae serum,respectively.Results showed that anti-rPsaA serum and anti-S.pneumoniae serum could effectively inhibit the adhesion of S.pneumoniae to bovine embryonic tracheal epithelial cells.In this study,the rPsaA protein of S.pneumoniae bovine was successfully expressed.For the first time,the rPsaA protein of S.pneu-moniae and anti-rPsaA protein serum were confirmed to inhibit the adhesion of S.pneumoniae to the trachea epithelial cells from the bovine embryo.This study laid foundation for exploring the mechanism of inhibiting the adhesion of S.pneumoniae to respiratory epithelial cells and the clinical application of S.pneumoniae subunit vaccine.
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