Real-time PCR of the 16S-rRNA gene in the diagnosis of neonatal bacteraemia.
OBJECTIVE
To evaluate a real-time PCR assay for the diagnosis of neonatal bacteraemia.


METHODS
Two hundred ninety-five plasma samples from 288 newborns with suspected neonatal sepsis were collected prospectively for the purpose of polymerase chain reaction (PCR)-based bacterial detection. A real-time PCR targeting the bacterial gene for 16S-rRNA gene combined with four specific probes designed to detect Gram-negative bacteria, Staphylococcus aureus and coagulase-negative staphylococci (CoNS) was developed. All samples positive in the universal PCR were further sequenced for bacterial identification.


RESULTS
When applied to a material from 50 patients with positive blood culture and 245 patients with negative blood culture, the universal PCR showed a sensitivity of 42% (28-57), a specificity of 95% (92-97), a positive predictive value of 64% (45-80), and a negative predictive value of 89% (84-92) (95% confidence intervals in brackets).


CONCLUSIONS
A new real-time PCR technique was for the first time applied to a well-defined prospectively and consecutively enrolled material of newborns with suspected sepsis, combining the benefits of real-time PCR with specific probes and sequencing. The method managed to detect bacteraemia with high specificity even though the sensitivity was low. Factors causing the low sensitivity are identified and further strategies to develop the method are described.
Ohlin
Andreas
A
Department of Paediatrics, Orebro University Hospital, Orebro, Sweden. andreas.ohlin@orebroll.se
Bäckman
Anders
A
Björkqvist
Maria
M
Mölling
Paula
P
Jurstrand
Margaretha
M
Schollin
Jens
J
D016470:Bacteremia / Q000175:diagnosis* / Q000235:genetics*; D001483:Base Sequence; D003955:Diagnostic Tests, Routine; D006801:Humans; D007231:Infant, Newborn; D010865:Pilot Projects; D015347:RNA Probes; D012336:RNA, Ribosomal, 16S / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018570:Risk Assessment; D012680:Sensitivity and Specificity; D013997:Time Factors
D015347:RNA Probes; D012336:RNA, Ribosomal, 16S