title: Detection of Mycobacterium tuberculosis in blood by use of the Xpert MTB/RIF assay.
abstract: We have developed a novel blood lysis-centrifugation approach for highly sensitive Mycobacterium tuberculosis detection in large volumes of blood with the Xpert MTB/RIF assay. One through 20 ml of blood was spiked with 0.25 to 10 CFU/ml of the M. tuberculosis surrogate M. bovis BCG. Multiple replicates of each sample were processed by a new lysis-centrifugation method and tested with the Xpert MTB/RIF assay. The assay was very sensitive with increased blood volumes. In the 20-ml samples, BCG was detected in blood spiked with 10, 5, 1, and 0.25 CFU/ml 100, 100, 83, and 57% of the time, respectively, compared to 100, 66, 18, and 18%, of the time, respectively, in 1-ml blood samples. Assay sensitivity was influenced by the type of anticoagulant used, with acid-citrate-dextrose solution B (ACD-B) providing the best results. A limit of detection of 10 CFU/ml was established with BCG spiked into ACD-B-treated blood, and 92, 36, and 33% of the samples with 5, 1, and 0.5 CFU/ml, respectively, were assay positive. The lysis buffer was stable both at room temperature and at 4°C for 2 months. The assay was tested with blood stored for 8 days without a change in sensitivity as measured by cycle threshold. This new assay format extends the capability of the Xpert MTB/RIF test, enabling up to 20 ml of blood to be tested rapidly for the presence of M. tuberculosis. This approach may be a useful method to detect extrapulmonary tuberculosis and the risk of death in immunocompromised patients.
authors.affiliation: Center for Infectious Diseases, New Jersey Medical School-UMDNJ, Newark, New Jersey, USA.
authors.forename: Padmapriya P
authors.initials: PP
authors.lastname: Banada
authors.forename: Ranie
authors.initials: R
authors.lastname: Koshy
authors.forename: David
authors.initials: D
authors.lastname: Alland
mesh_terms: D016470:Bacteremia / Q000175:diagnosis*; D001431:Bacteriological Techniques / Q000379:methods*; D002498:Centrifugation / Q000379:methods; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods; D014376:Tuberculosis / Q000175:diagnosis*