RESUMO
Molecular surveillance of pathogens has shown the need for rapid and dependable methods for the identification of organisms of clinical and epidemiological importance. As the leading cause of community-acquired pneumonia, Streptococcus pneumoniae was used as a model organism to develop and refine a real-time fluorescence PCR assay and enhanced DNA purification method. Seventy clinical isolates of S. pneumoniae, verified by latex agglutination, were screened against 26 negative control clinical isolates employing a TaqMan assay on a thermocycler (LightCycler). The probe, constructed from the lytA gene, correctly detected all S. pneumoniae genomes without cross-reaction to negative controls. The speed and ease of this approach will make it adaptable to identification of many bacterial pathogens and provide potential for adaptation to direct detection from patient specimens.
Assuntos
Pneumonia Pneumocócica/diagnóstico , Pneumonia Pneumocócica/microbiologia , Reação em Cadeia da Polimerase/métodos , Streptococcus pneumoniae/classificação , Primers do DNA/genética , DNA Bacteriano/análise , DNA Bacteriano/isolamento & purificação , Fluorescência , Humanos , N-Acetil-Muramil-L-Alanina Amidase/genética , Sensibilidade e Especificidade , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/isolamento & purificação , Taq Polimerase/metabolismoRESUMO
Enterococcus faecium has recently emerged as a serious nosocomial pathogen. The prevalence and severity of enterococcal infections, the mortality rate from such infections, and the antibiotic resistance of enterococci are often species dependent. Since conventional biochemical methods fail to differentiate E. faecium from certain newly described enterococcal species, a PCR-based assay was developed for the rapid identification of E. faecium.