Detection of group B streptococci (GBS) in vaginal swabs using real-time PCR with TaqMan probe hybridization.
OBJECTIVE
Implementation of a screening based strategy for the prevention of perinatal group B streptococcus (GBS) disease is anticipated to increase the demand of fast laboratory techniques. The aim of the present study was to develop a real-time PCR method for the specific detection of GBS in vaginal swabs.


METHODS
Based on partial sequencing of the sip gene, primers and a TaqMan hybridization probe were constructed for a real-time PCR assay. The performance of the assay was tested on 100 consecutive vaginal specimens submitted to the laboratory for culture. Lysis of bacteria and DNA extraction was performed by lysozyme, mutanolysin, proteinase K and Quiagen spin columns. PCR was performed using LightCycler. Highly purified DNA from GBS was used as positive control.


RESULTS
Of the 100 samples investigated, 25 were positive by culture (16 abundant/moderate growth, 6 sparse growth and 3 after enrichment only). At a cut-off of 12 fg per reaction (corresponding to 4 bacterial cells), PCR was positive in 32 samples. A complete concordance was noted between PCR positivity and abundant/moderate and sparse growth by culture. One of 3 samples that were positive by culture only after enrichment was negative in PCR. On the other hand, 8 samples were positive by PCR and negative by culture.


CONCLUSIONS
The real-time PCR assay was sensitive and rapid and enabled detection of GBS in less than 2 h after collection. Due to the rapidity of the assay by which results could be obtained, the assay harbors the potential for intrapartum detection of GBS.
Bergh
K
K
Norwegian University of Science & Technology, Department of Laboratory Medicine, N-7006 University Hospital of Trondheim, Norway. kare.bergh@medisin.ntnu.no
Stoelhaug
A
A
Loeseth
K
K
Bevanger
L
L
D001483:Base Sequence; D017931:DNA Primers; D005260:Female; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods*; D013292:Streptococcus agalactiae / Q000235:genetics / Q000302:isolation & purification*; D014621:Vagina / Q000382:microbiology*; D014626:Vaginal Smears
D017931:DNA Primers