Universal 16S rDNA PCR and sequencing in the diagnosis of infective endocarditis directly from heart valve tissue
Abstract number: 902_p701
The microbiological diagnosis of infective endocarditis (IE) is based on positive culture of heart valve tissue or blood culture but cultures remain negative when IE is caused by fastidious micro-organisms or antimicrobial treatment is started before cultures are obtained.
To evaluate the usefulness of a universal 16S rDNA PCR method followed by direct sequencing in heart valve tissue for IE diagnosis in the routine of a clinical microbiology laboratory, compared with traditional heart valve culture (HVC) and blood culture (BC).
Heart valves received for culture over a ten-month period were studied by 16S rDNA PCR with primers PSL and P13P. Positive samples were subsequently sequenced for identification. HV were cultured by conventional methods. BCs were performed by the BACTEC 9240 system. Sensitivity of the assay was assessed by obtaining DNA from 10-fold dilutions of Streptococcus oralis. After molecular analysis, clinical records of patients and results of conventional cultures were consulted.
Twenty-four samples of HV (24 patients) were studied. In 10 patients IE was clinically rejected and their valves were included in the study as negative controls. Their HVC, BCs and PCR were negative. The remaining 14 cases had either proven (13) or possible (1) IE. Overall, BCs were positive in 11 patients but HVC remained positive at the moment of resection in only four patients. Of the 14 cases, 12 were microbiologically documented by conventional cultures. PCR was positive in the 12 confirmed cases. Micro-organisms identified by PCR matched those cultured by conventional cultures except in one case in which the valve was inadequately remitted to the microbiology laboratory. In the 2 cases with IE and with no micro-organisms demonstrated by conventional cultures, PCR was also negative. The median time of analysis to a PCR result was 1 day and to a sequence and bacterial identification in PCR-positive samples, 3 days. The analytical sensitivity of this assay was 100 CFU/mL.
Universal 16S rDNA PCR followed by sequencing applied to resected heart valves seems to be a reliable test for diagnosis of IE. Our study suggests its applicability to patients with conventional negative microbiology, mainly those studied during the course of antimicrobial therapy."
|Session name:||XXIst ISTH Congress|
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