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Diagn Microbiol Infect Dis. 2019 Apr;93(4):287-292. doi: 10.1016/j.diagmicrobio.2018.11.006. Epub 2018 Nov 20.

High-throughput screening of bacterial pathogens in clinical specimens using 16S rDNA qPCR and fragment analysis.

Author information

1
Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland. Electronic address: karoline.wagner@usb.ch.
2
Institute of Medical Microbiology and Hygiene, Austrian Agency for Health and Food Safety, Graz, Austria.
3
Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland.

Abstract

Molecular-based detection of bacterial pathogens directly from clinical specimens permits rapid initiation of effective antimicrobial treatment and adequate patient management. Broad-range polymerase chain reaction (PCR) amplification of the 16S rRNA gene (16S rDNA qPCR) is used in many diagnostic laboratories as a complement to cultural identification of bacterial pathogens. However, efforts for automation of 16S rDNA PCR workflows are needed in order to reduce turnaround times and to enhance reproducibility and standardization of the technique. In this retrospective method evaluation study, clinical specimens (N = 499) from patients with suspected bacterial infections were used to evaluate 2 diagnostic semiautomated workflows for rapid bacterial pathogen detection. The workflows included automated DNA extraction (QIASymphony), 16S rDNA qPCR, fragment or melting curve analysis, and amplicon sequencing. Our results support the use of the 16S rDNA qPCR and fragment analysis workflow as it enabled rapid and accurate identification of bacterial pathogens in clinical specimens.

KEYWORDS:

16S; Bacterial broad-range qPCR; Bacterial infection; Fragment analysis; High throughput screening

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