Cloning and Characterization of the Differentially Expressed Genes in Streptococcus pneumoniae

ABSTRACT

In spite of the extensive use of antibiotics over several decades, Streptococcus pneumoniae still remains as one of the most serious human bacterial pathogens. In order to clone the pneumococcal genes whose expression is induced when pneumococcus causes infection in mice, S. pneumoniae strain ATCC 6303 was subcultured on blood agar plates (BAP) ten times to reduce the virulence first, and then passaged through BALB/c mice three times to restore the virulence. Subtractive hybridization was performed using the total RNA preparations isolated from BAP-cultured and mouse-passaged strains. Complementary DNAs corresponding to any mRNA species that were differentially expressed in the mouse- passaged strain were used as the probes to screen the genomic library of pneumococcus. Positive recombinants were selected and sequenced partially to identify the genes located within the cloned DNA. GenBank search of the sequence data has identified several genes including two heat shock genes (dnaK and dnap, a transposase-encoding gene, and a sequence which is very homologous to that of the ftsH gene.