title: Sensitive Detection of Francisella tularensis Directly from Whole Blood by Use of the GeneXpert System.
abstract: Francisella tularensis is a potential bioterrorism agent that is highly infectious at very low doses. Diagnosis of tularemia by blood culture and nucleic acid-based diagnostic tests is insufficiently sensitive. Here, we demonstrate a highly sensitive F. tularensis assay that incorporates sample processing and detection into a single cartridge suitable for point-of-care detection. The assay limit of detection (LOD) and dynamic range were determined in a filter-based cartridge run on the GeneXpert system. F. tularensis DNA in buffer or CFU of F. tularensis was spiked into human or macaque blood. To simulate detection in human disease, the assay was tested on blood drawn from macaques infected with F. tularensis Schu S4 at daily intervals. Assay detection was compared to that with a conventional quantitative PCR (qPCR) assay and blood culture. The assay LOD was 0.1 genome equivalents (GE) per reaction and 10 CFU/ml F. tularensis in both human and macaque blood. In infected macaques, the assay detected F. tularensis on days 1 to 4 postinfection in 21%, 17%, 60%, and 83% of macaques, respectively, compared to conventional qPCR positivity rates of 0%, 0%, 30%, and 100% and CFU detection of blood culture at 0%, 0%, 0%, and 10% positive, respectively. Assay specificity was 100%. The new cartridge-based assay can rapidly detect F. tularensis in bloodstream infections directly in whole blood at the early stages of infection with a sensitivity that is superior to that of other methods. The simplicity of the automated testing procedures may make this test suitable for rapid point-of-care detection.
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: Padmapriya P
authors.initials: PP
authors.lastname: Banada
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: Srinidhi
authors.initials: S
authors.lastname: Deshpande
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: Soumitesh
authors.initials: S
authors.lastname: Chakravorty
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: Riccardo
authors.initials: R
authors.lastname: Russo
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: James
authors.initials: J
authors.lastname: Occi
authors.affiliation: Battelle Memorial Institute, Columbus, Ohio, USA.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Meister
authors.affiliation: Battelle Memorial Institute, Columbus, Ohio, USA.
authors.forename: Kelly J
authors.initials: KJ
authors.lastname: Jones
authors.affiliation: Battelle Memorial Institute, Columbus, Ohio, USA.
authors.forename: Carl H
authors.initials: CH
authors.lastname: Gelhaus
authors.affiliation: Lovelace Respiratory Research Institute (LBERI), Albuquerque, New Mexico, USA.
authors.forename: Michelle W
authors.initials: MW
authors.lastname: Valderas
authors.affiliation: Cepheid, Sunnyvale, California, USA.
authors.forename: Martin
authors.initials: M
authors.lastname: Jones
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA.
authors.forename: Nancy
authors.initials: N
authors.lastname: Connell
authors.affiliation: Center for Emerging Pathogens, Department of Medicine, Rutgers Biomedical and Health Sciences (RHBS), Rutgers University, Newark, New Jersey, USA allandda@njms.rutgers.edu.
authors.forename: David
authors.initials: D
authors.lastname: Alland
mesh_terms: D000818:Animals; D057205:Automation, Laboratory / Q000379:methods*; D001431:Bacteriological Techniques / Q000379:methods*; D001769:Blood / Q000382:microbiology*; D005604:Francisella tularensis / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D008251:Macaca; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D014406:Tularemia / Q000175:diagnosis*
keywords: Francisella tularensis; GeneXpert; PCR; diagnostic; macaque blood; whole blood