title: Molecular detection of culture-confirmed bacterial bloodstream infections with limited enrichment time.
abstract: Conventional blood culturing using automated instrumentation with phenotypic identification requires a significant amount of time to generate results. This study investigated the speed and accuracy of results generated using PCR and pyrosequencing compared to the time required to obtain Gram stain results and final culture identification for cases of culture-confirmed bloodstream infections. Research and physician-ordered blood cultures were drawn concurrently. Aliquots of the incubating research blood culture fluid were removed hourly between 5 and 8 h, at 24 h, and again at 5 days. DNA was extracted from these 6 time point aliquots and analyzed by PCR and pyrosequencing for bacterial rRNA gene targets. These results were then compared to those of the physician-ordered blood culture. PCR and pyrosequencing accurately identified 92% of all culture-confirmed cases after a mean enrichment time of 5.8 ± 2.9 h. When the time needed to complete sample processing was included for PCR and pyrosequencing protocols, the molecular approach yielded results in 11.8 ± 2.9 h compared to means of 27.9 ± 13.6 h to obtain the Gram stain results and 81.6 ± 24.0 h to generate the final culture-based identification. The molecular approach enabled accurate detection of most bacteria present in incubating blood culture bottles on average about 16 h sooner than Gram stain results became available and approximately 3 days sooner than the phenotypic identification was entered in the Laboratory Information System. If implemented, this more rapid molecular approach could minimize the number of doses of unnecessary or ineffective antibiotics administered to patients.
authors.affiliation: Department of Biology, Georgetown University, Washington, DC, USA.
authors.forename: Miranda S
authors.initials: MS
authors.lastname: Moore
authors.forename: Chase D
authors.initials: CD
authors.lastname: McCann
authors.forename: Jeanne A
authors.initials: JA
authors.lastname: Jordan
mesh_terms: D016470:Bacteremia / Q000175:diagnosis* / Q000382:microbiology*; D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D001431:Bacteriological Techniques / Q000379:methods*; D004275:DNA, Ribosomal / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D011446:Prospective Studies; D017422:Sequence Analysis, DNA / Q000379:methods; D013997:Time Factors
chemical_list: D004275:DNA, Ribosomal