Evaluation of real-time PCR and pyrosequencing for screening incubating blood culture bottles from adults with suspected bloodstream infection

• Culture fluids screened by PCR after 8 h had >90% sensitivity compared to culture.
• Molecular diagnostic (MDx) testing provided results more rapidly than automated blood culturing.
• Lab logistics must be considered before implementing MDx testing for bloodstream infection.

Several molecular platforms can identify bacteria associated with bloodstream infections but require positive culture bottles as starting material. Here, we describe results of screening 1140 blood cultures at 8 h postinoculation, from 918 eligible adults being evaluated for bloodstream infection. DNA was extracted and analyzed by 16S and/or 23S rRNA real-time PCR/pyrosequencing. Compared to culture, PCR/pyrosequencing displayed 90.9% sensitivity, 99.6% specificity, 95.7% positive predictive value, and 99.1% negative predictive value. Overall concordance rate was 98.9% (1127/1140). In 4 cases with molecular-positive/culture-negative results, medical chart reviews provided evidence of identical bacteria from subsequent blood or concomitant urine/sputum cultures. Nine culture-positive/molecular-negative cases were associated with either polymicrobial growth, grew only in the anaerobic bottle of the clinical pair, and/or were detected by PCR/pyrosequencing after 8 h. In summary, this approach accurately detected and identified bacteria in ~91% of culture-confirmed cases significantly sooner than the phenotypic identification was available, having the potential to improve antibiotic stewardship.