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Multiplex Detection of Three Select Agents Directly from Blood by Use of the GeneXpert System

Padmapriya P Banada1, Srinidhi Deshpande2, Sukalyani Banik2

  • 1Center for Emerging Pathogens, Division of Infectious Diseases, New Jersey Medical School, Rutgers Biomedical and Health Sciences University, Newark, New Jersey, USA Priya.banada@rutgers.edu allandda@njms.rutgers.edu.

Journal of Clinical Microbiology
|March 8, 2019
PubMed

Insights

A new GeneXpert assay rapidly detects three dangerous bacteria: Francisella tularensis, Bacillus anthracis, and Yersinia pestis. This point-of-care test offers high sensitivity and specificity in blood samples within 100 minutes.

Area of Science:

  • Microbiology and Infectious Diseases
  • Biotechnology and Molecular Diagnostics
  • Public Health and Biodefense

Background:

  • Tier 1 select agents like *Francisella tularensis*, *Bacillus anthracis*, and *Yersinia pestis* pose significant bioterrorism threats.
  • Timely detection of these pathogens is crucial for effective patient management and public health response.
  • Current diagnostic methods may lack the speed and point-of-care suitability required for bioterrorism scenarios.

Purpose of the Study:

  • To develop and validate a rapid, multiplex, cartridge-based assay for the simultaneous detection of *F. tularensis*, *B. anthracis*, and *Y. pestis*.
  • To assess the assay's analytical performance, including limit of detection (LOD), sensitivity, and specificity, directly from patient blood samples.
  • To evaluate the suitability of the assay for point-of-care use in bioterrorism preparedness.

Main Methods:

  • Development of a multiplex nested PCR assay on the GeneXpert system.
  • Determination of analytical limit of detection (LOD) and dynamic range using spiked blood samples.
  • Evaluation of assay sensitivity and specificity using spiked blood, negative controls, and clinical samples.

Main Results:

  • The assay achieved limits of detection of 8.5 CFU/ml for *F. tularensis*, 10 CFU/ml for *B. anthracis*, and 4.5 CFU/ml for *Y. pestis*.
  • 100% sensitivity was observed at the LOD in spiked patient blood samples for all three target organisms.
  • 100% specificity was confirmed against nontarget pathogens and clinical blood samples, with a total assay time of approximately 100 minutes.

Conclusions:

  • The automated, cartridge-based GeneXpert assay enables rapid, sensitive, and specific detection of three critical select agent bacteria directly from blood.
  • This point-of-care assay provides results within 100 minutes without the need for sample enrichment.
  • The developed assay holds significant potential for timely diagnosis and treatment initiation in bioterrorism events.
Keywords:
Bacillus anthracisFrancisella tularensisGeneXpertYersinia pestisanthraxdiagnosticmultiplex PCRplaguetularemiawhole blood

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