Rapid identification of Streptococcus pneumoniae strains using 16S rRNA gene based PCR method.

Abstract

Streptococcus pneumoniae strains are responsible for severe diseases, including pneumonia, bronchitis, otitis media, septicemia, meningitis. Early identification of etiologic agent of pneumococcal diseases is essential and usually performed by “Gold Standard” culture based method which is time-consuming and has poor sensitivity. The aim of this study was to develop a PCR based method for rapid diagnosis of pneumococcal diseases by targeting 16S rRNA region of Streptococcus pneumoniae. In this study, 16S rRNA based primers were designed using CLUSTALW and Primer-BLAST. Appropriate laboratory tests such as optochin disk test, bile solubility test, gram-staining and catalase test as well as PCR, gel run electrophoresis and nucleotide sequencing was performed to examine the specificity and utility of the primers in Streptococcus pneumoniae detection using eighteen stored Streptococcus pneumoniae strains. For further validation, the same primers were used against the DNA extracted from six throat swab samples pre-incubated in enrichment broth and DNA extracted directly from same samples without using enrichment broth. It took total 8 hours and 6 hours respectively to achieve the results. After nucleotide sequencing and subsequent BLASTn analysis, it was substantiated; this approach is specific and faster than present diagnostic approach for Streptococcus pneumoniae detection.