Psp140: an immunodominant antigen in the supernatant of Streptococcus pneumoniae culture

Abstract

Background and objectives: Streptococcus pneumoniae causes many lethal infections. Due to its reduced sensitivity to commonly used antibiotics, development of new strategies against pneumococcal infections seems to be necessary. We aimed to investigate immunodominant antigens in S. pneumoniae culture supernatant in order to develop novel targets for pneumococcal vaccines.

Materials and methods: In this study S. pneumoniae ATCC49619 was sub-cultured into BHI broth from overnight culture at 37°C for 4 h. The supernatant proteins were precipitated using acetone precipitation method. A rabbit was intramuscularly immunized with alum adjuvant and 100 μg pneumococcal supernatant proteins, 6 times at 14 days' intervals to produce hyperimmune serum. ELISA assay was performed to determine the antibody level response to pneumococcal secretory proteins. Then dot blot applied for rapid evaluation of hyperimmune serum reactivity to pneumococcus supernatant proteins. The western blot was also used to determine the interaction of supernatant proteins with immunogenic rabbit's hyperimmune-serum.

Results: According to the western blot analysis, the immunodominant protein had 140KDa molecular weight and designated as pneumococcal secretory protein140 (Psp140).

Conclusion: The Psp140 protein in the supernatant of S. pneumoniae culture is an immunodominant protein and it is likely related to pneumococcal secretory protein or surface exposed protein which released into culture supernatant during bacterial growth.

Keywords: Enzyme-linked immunosorbent assay; Immunodominant antigen; Streptococcus pneumoniae; Western blot.

Fig. 1.

Titration of rabbit non immune serum and hyperimmune serumafter last secretory proteins injection. The numbers 1–8 are 1:100, 1:200, 1:400, 1:800, 1:1600, 1:3200, 1:6400 and 1:12800 sera dilutions, respectively.The vertical line show optical density (OD).

Fig. 2.

Immunoblotting pneumococcal secretory proteins. NC1; Negative control with non-hyperimmune reaction, NC2; Negative control without secretory proteins and test lane a positive reaction with rabbit hyperimmune serum.

Fig. 3.

Western blot analysis of secretory proteins of S. pneumoniae by hyperimmune serum. lane 1; protein marker, lane 2; the interaction of secretory proteins and rabbit’s hyperimmune serum, lane 3 negative control without secretory proteins, lane 4 negative control with rabbit non-immune serum, lane 5; polyacrylamide gel electrophoresis of secretory proteins.