Real-time pneumolysin polymerase chain reaction with melting curve analysis differentiates pneumococcus from other α-hemolytic streptococci

• Published in: Diagnostic microbiology and infectious disease 2005-12, Vol.53 (4), p.293-299
• Main Authors: Kaijalainen, Tarja, Saukkoriipi, Annika, Bloigu, Aini, Herva, Elja, Leinonen, Maija
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - genetics; Bacteriology; Base Sequence; Biological and medical sciences; DNA, Bacterial - genetics; Fundamental and applied biological sciences. Psychology; Humans; Infant; Infectious diseases; Medical sciences; Microbiology; Miscellaneous; Molecular Sequence Data; Nasopharyngeal specimen; Nasopharynx - microbiology; Pneumococcal Infections - diagnosis; Polymerase Chain Reaction - methods; Real-time PCR; Sequence Analysis, DNA; Streptococcal Infections - microbiology; Streptococcus; Streptococcus - classification; Streptococcus - isolation & purification; Streptococcus pneumoniae; Streptococcus pneumoniae - classification; Streptococcus pneumoniae - genetics; Streptolysins - genetics; Transition Temperature; α-hemolytic streptococcus
• ISSN: 0732-8893; 1879-0070
• DOI: 10.1016/j.diagmicrobio.2005.07.005

The majority of pneumococcal isolates can be identified by the conventional methods based on phenotypic characteristics. Occasionally, however, the differentiation of α-hemolytic streptococci from pneumococci, especially those isolated from nasopharynx, is problematic due to the discrepant results obtained by the conventional identification methods. Several gene technological methods based on the amplification of genes encoding pneumococcal virulence factors, such as pneumolysin ( ply) and autolysin, have been used as additional identification methods. Recent studies have shown that especially the ply gene is frequently also present in nonpneumococcal α-hemolytic streptococci. In this study, we compared the commonly used phenotypic identification methods with nucleic acid–based methods, commercial AccuProbe™, conventional pneumolysin polymerase chain reaction (Ply-PCR), and real-time Ply-PCR in the identification of α-hemolytic streptococcal strains isolated from 100 consecutive nasopharyngeal specimens. We also studied if melting curve analysis and sequencing of the amplification products of a ply gene fragment could be helpful in the identification. Our results suggest that the ply gene present in α-hemolytic streptococci differs from that present in pneumococcus, and that melting curve analysis would prove useful in the differentiation of these bacteria.