Evaluation of several biochemical and molecular techniques for identification of Streptococcus pneumoniae and Streptococcus pseudopneumoniae and their detection in respiratory samples

Abstract

Identification and detection of Mitis group streptococci, which contains Streptococcus pneumoniae, has been hampered by the lack of sensitive and specific assays. In this study we evaluated several biochemical and molecular assays for identification of S. pneumoniae and S. pseudopneumoniae and their distinction from other Mitis group streptococci using a collection of 54 isolates obtained by routine culture of 53 respiratory specimens from patients with community-acquired pneumonia. The combined results of the biochemical and molecular assays indicated the presence of 23 S. pneumoniae, two S. pseudopneumoniae and 29 other Mitis group streptococci isolates. The tube bile solubility test that is considered as gold standard for identification of S. pneumoniae showed concordant results with optochin susceptibility testing (CO(2) atmosphere) and a real-time multiplex PCR assay targeting the Spn9802 fragment and autolysin gene. Optochin susceptibility testing upon incubation in O(2) atmosphere, bile solubility testing by oxgall disk, matrix-assisted laser desorption ionization-time of flight mass spectrometry and sequence analysis of the tuf and rpoB genes resulted in several false positive, false negative or inconclusive results. The S. pseudopneumoniae isolates could only be identified by molecular assays and the multiplex real-time PCR assay was concluded to be most convenient for identification of S. pneumoniae and S. pseudopneumoniae. Using this method S. pneumoniae and S. pseudopneumoniae DNA could be detected in the respiratory samples from which they were isolated and in an additional 11 samples from which only other streptococci were isolated.