Improved sensitivity for molecular detection of bacterial and candida infections in blood

Andrea Bacconi, Gregory S. Richmond, Michelle A. Baroldi, Thomas G. Laffler, Lawrence B. Blyn, Heather E. Carolan, Mark R. Frinder, Donna M. Toleno, David Metzgar, Jose R. Gutierrez, Christian Massire, Megan Rounds, Natalie J. Kennel, Richard E. Rothman, Stephen Peterson, Karen C. Carroll, Teresa Wakefield, David J. Ecker, Rangarajan Sampath

Research output: Contribution to journalArticlepeer-review

Abstract

The rapid identification of bacteria and fungi directly from the blood of patients with suspected bloodstream infections aids in diagnosis and guides treatment decisions. The development of an automated, rapid, and sensitive molecular technology capable of detecting the diverse agents of such infections at low titers has been challenging, due in part to the high background of genomic DNA in blood. PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) allows for the rapid and accurate identification of microorganisms but with a sensitivity of about 50% compared to that of culture when using 1-ml wholeblood specimens. Here, we describe a new integrated specimen preparation technology that substantially improves the sensitivity of PCR/ESI-MS analysis. An efficient lysis method and automated DNA purification system were designed for processing 5 ml of whole blood. In addition, PCR amplification formulations were optimized to tolerate high levels of human DNA. An analysis of 331 specimens collected from patients with suspected bloodstream infections resulted in 35 PCR/ESI-MS-positive specimens (10.6%) compared to 18 positive by culture (5.4%). PCR/ESI-MS was 83% sensitive and 94% specific compared to culture. Replicate PCR/ESI-MS testing from a second aliquot of the PCR/ESI-MS-positive/ culture-negative specimens corroborated the initial findings in most cases, resulting in increased sensitivity (91%) and specificity (99%) when confirmed detections were considered true positives. The integrated solution described here has the potential to provide rapid detection and identification of organisms responsible for bloodstream infections.

Original languageEnglish (US)
Pages (from-to)3164-3174
Number of pages11
JournalJournal of clinical microbiology
Volume52
Issue number9
DOIs
StatePublished - Sep 2014

ASJC Scopus subject areas

  • Microbiology (medical)

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