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J Mol Diagn. 2014 Mar;16(2):261-6. doi: 10.1016/j.jmoldx.2013.10.011. Epub 2013 Dec 21.

Sensitive detection and serovar differentiation of typhoidal and nontyphoidal Salmonella enterica species using 16S rRNA Gene PCR coupled with high-resolution melt analysis.

Author information

1
Department of Emergency Medicine, School of Medicine, The Johns Hopkins University, Baltimore, Maryland. Electronic address: bmasek@jhmi.edu.
2
Division of Infectious Diseases, School of Medicine, The Johns Hopkins University, Baltimore, Maryland.
3
Department of Emergency Medicine, School of Medicine, The Johns Hopkins University, Baltimore, Maryland; Memorial and the Sloan-Kettering Cancer Center, New York, New York.
4
Department of Emergency Medicine, School of Medicine, The Johns Hopkins University, Baltimore, Maryland.
5
Department of Emergency Medicine, School of Medicine, The Johns Hopkins University, Baltimore, Maryland; Division of Infectious Diseases, School of Medicine, The Johns Hopkins University, Baltimore, Maryland.

Abstract

Salmonella enterica species infections are a significant public health problem causing high morbidity rates worldwide and high mortality rates in the developing world. These infections are not always rapidly diagnosed as a cause of bloodstream infections because of the limitations of blood culture, which greatly affects clinical care as a result of treatment delays. A molecular diagnostic assay that could rapidly detect and identify S. enterica species infections as a cause of sepsis is needed. Nine typhoidal and nontyphoidal S. enterica serovars were used to establish the limit of detection (LOD) of a previously published 16S rRNA gene PCR (16S PCR) in mock whole blood specimens. In addition, 16 typhoidal and nontyphoidal S. enterica serovars were used to evaluate the serovar differentiation capability of 16S PCR coupled with high-resolution melt analysis. The overall LOD of 16S PCR for the nine typhoidal and nontyphoidal S. enterica serovars analyzed was <10 colony-forming units per milliliter (CFU/mL) in mock whole blood specimens, with the lowest and highest LOD at <1 CFU/mL and 9 CFU/mL, respectively. By high-resolution melt analysis, the typhoidal and nontyphoidal S. enterica serovar groups analyzed each generated a unique grouping code, allowing for serovar-level identification. 16S PCR coupled with high-resolution melt analysis could be a useful molecular diagnostic that could enhance the current diagnostic, treatment, and surveillance methods of S. enterica bloodstream infections.

PMID:
24365382
PMCID:
PMC3937533
DOI:
10.1016/j.jmoldx.2013.10.011
[Indexed for MEDLINE]
Free PMC Article

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