Highly sensitive detection of Staphylococcus aureus directly from patient blood.

PloS One
Padmapriya P BanadaDavid Alland

Abstract

Rapid detection of bloodstream infections (BSIs) can be lifesaving. We investigated the sample processing and assay parameters necessary for highly-sensitive detection of bloodstream bacteria, using Staphylococcus aureus as a model pathogen and an automated fluidic sample processing-polymerase chain reaction (PCR) platform as a model diagnostic system. We compared a short 128 bp amplicon hemi-nested PCR and a relatively shorter 79 bp amplicon nested PCR targeting the S. aureus nuc and sodA genes, respectively. The sodA nested assay showed an enhanced limit of detection (LOD) of 5 genomic copies per reaction or 10 colony forming units (CFU) per ml blood over 50 copies per reaction or 50 CFU/ml for the nuc assay. To establish optimal extraction protocols, we investigated the relative abundance of the bacteria in different components of the blood (white blood cells (WBCs), plasma or whole blood), using the above assays. The blood samples were obtained from the patients who were culture positive for S. aureus. Whole blood resulted in maximum PCR positives with sodA assay (90% positive) as opposed to cell-associated bacteria (in WBCs) (71% samples positive) or free bacterial DNA in plasma (62.5% samples positive). Both the assays we...Continue Reading

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Citations

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Nov 17, 2016·Journal of Clinical Microbiology·Padmapriya P BanadaDavid Alland
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Methods Mentioned

BETA
nucleic acid extraction
PCR

Software Mentioned

Excel
GeneXpert
GraphPad
GraphPad Prism
GeneXpert®
Primer3
PrimerSelect DNASTAR Lasergene
Mfold

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