Detection of bacteremia in emergency department patients at risk for infective endocarditis using universal 16S rRNA primers in a decontaminated polymerase chain reaction assay.
Prompt definitive diagnosis of acute bacterial endocarditis in febrile injection drug users (IDUs) remains problematic because of delays associated with blood culture. Rapid detection of bacteremia by polymerase chain reaction (PCR) by use of "universal" primers has been hampered by background bacterial contamination. Broad-range eubacterial primers selected from the 16S rRNA gene were used in a PCR assay coupled with a simple pre-PCR decontamination step. All PCR reagents were pretreated with the restriction enzyme AluI, which has multiple digestion sites in the amplicon but none in the primer sets. When 4 different bacterial species were spiked into healthy human blood specimens, the assay identified each pathogen with an analytic sensitivity of 5 bacteria/PCR reaction. A clinical trial with 51 febrile IDUs revealed that PCR had a sensitivity and specificity of 86.7% and 86.9%, respectively, versus blood culture. Importantly, all (8/8) patients with blood culture-positive infective endocarditis were determined to be positive by PCR. This assay provides a promising diagnostic for rapid identification of bacteremia, particularly valuable in acute care settings.
Rothman
Richard E
RE
Department of Emergency Medicine, Johns Hopkins University, Bethesda, Maryland 21205, USA. rrothman@jhmi.edu
Majmudar
Maulik D
MD
Kelen
Gabor D
GD
Madico
Guillermo
G
Gaydos
Charlotte A
CA
Walker
Tarik
T
Quinn
Thomas C
TC
D016470:Bacteremia / Q000175:diagnosis* / Q000382:microbiology; D017931:DNA Primers; D004269:DNA, Bacterial / Q000097:blood / Q000302:isolation & purification; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004636:Emergency Service, Hospital; D004697:Endocarditis, Bacterial / Q000175:diagnosis* / Q000382:microbiology; D004866:Equipment Contamination; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012307:Risk Factors; D012680:Sensitivity and Specificity; D015819:Substance Abuse, Intravenous / Q000150:complications*
D017931:DNA Primers; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; C024669:endodeoxyribonuclease AluI; D015252:Deoxyribonucleases, Type II Site-Specific