Sensitive and specific method for rapid identification of Streptococcus pneumoniae using real-time fluorescence PCR.
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.
McAvin
J C
JC
Molecular Epidemiology Branch, Air Force Institute for Environment and Occupational Health Risk Analysis/Epidemiology Surveillance Division, Brooks Air Force Base, San Antonio, Texas 78235, USA.
Reilly
P A
PA
Roudabush
R M
RM
Barnes
W J
WJ
Salmen
A
A
Jackson
G W
GW
Beninga
K K
KK
Astorga
A
A
McCleskey
F K
FK
Huff
W B
WB
Niemeyer
D
D
Lohman
K L
KL
D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D005453:Fluorescence; D006801:Humans; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics; D011018:Pneumonia, Pneumococcal / Q000175:diagnosis* / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013296:Streptococcus pneumoniae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D019914:Taq Polymerase / Q000378:metabolism
D017931:DNA Primers; D004269:DNA, Bacterial; D019914:Taq Polymerase; C076332:EJL amidase; D009238:N-Acetylmuramoyl-L-alanine Amidase