title: Improving the sensitivity of the Xpert MTB/RIF assay on sputum pellets by decreasing the amount of added sample reagent: a laboratory and clinical evaluation.
abstract: The Xpert MTB/RIF (Xpert) assay permits rapid near-patient detection of Mycobacterium tuberculosis in sputum; however, the test sensitivity remains suboptimal in paucibacillary specimens that are negative for acid-fast bacilli using smear microscopy. Xpert testing includes dilution with sample reagent, and when processed sputum pellets are tested, the recommended sample reagent/pellet ratio is 3:1. We evaluated whether a decreased sample reagent/pellet ratio of 2:1 increased Xpert sensitivity compared to the recommended 3:1. The limit of detection was determined by inoculating serial dilutions of M. tuberculosis into sputum samples, preparing sputum pellets, and testing each pellet by Xpert at both sample reagent ratios. Processed sputum pellets obtained from M. tuberculosis culture-positive clinical specimens were also tested by Xpert at both ratios. Among spiked sputum pellets, the limit of detection was 1,478 CFU/ml (95% confidence interval [CI], 1,211 to 1,943) at a 3:1 ratio and decreased to 832 CFU/ml (95% CI, 671 to 1,134) at 2:1. The proportion of specimens in which M. tuberculosis was detected was greater at 2:1 than at 3:1 for almost all numbers of CFU/ml; this difference was most prominent at lower numbers of CFU/ml. Among 134 concentrated sputum pellets from the clinical study, the sensitivity of Xpert at 2:1 was greater than at 3:1 overall (80% versus 72%; P=0.03) and for smear-negative specimens (67% versus 58%; P=0.12). For Xpert testing of sputum pellets, using a lower sample reagent/pellet ratio increased M. tuberculosis detection, especially for paucibacillary specimens. Our study supports use of a 2:1 sample reagent/pellet dilution for Xpert testing of sputum pellets.
authors: New Jersey Medical School-Rutgers, the State University of New Jersey, Newark, New Jersey, USA dharannj@njms.rutgers.edu.
authors: Nila J
authors: NJ
authors: Dharan
authors: New Jersey Medical School-Rutgers, the State University of New Jersey, Newark, New Jersey, USA.
authors: Danielle
authors: D
authors: Amisano
authors: Makerere University College of Health Sciences Mycobacteriology Laboratory, Kampala, Uganda.
authors: Gerald
authors: G
authors: Mboowa
authors: Makerere University College of Health Sciences Mycobacteriology Laboratory, Kampala, Uganda.
authors: Willy
authors: W
authors: Ssengooba
authors: New Jersey Medical School-Rutgers, the State University of New Jersey, Newark, New Jersey, USA.
authors: Robert
authors: R
authors: Blakemore
authors: Boston Medical Center, Boston, Massachusetts, USA.
authors: Rachel W
authors: RW
authors: Kubiak
authors: Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors: Derek T
authors: DT
authors: Armstrong
authors: Cepheid, Sunnyvale, California, USA.
authors: Martin
authors: M
authors: Jones
authors: Johns Hopkins University School of Medicine, Baltimore, Maryland, USA Johns Hopkins University Bloomberg School of Public Health, Baltimore, Maryland, USA.
authors: Yukari C
authors: YC
authors: Manabe
authors: Makerere University College of Health Sciences Mycobacteriology Laboratory, Kampala, Uganda.
authors: Moses L
authors: ML
authors: Joloba
authors: Boston Medical Center, Boston, Massachusetts, USA Boston University School of Medicine, Boston, Massachusetts, USA.
authors: Jerrold J
authors: JJ
authors: Ellner
authors: Johns Hopkins University School of Medicine, Baltimore, Maryland, USA Johns Hopkins University Bloomberg School of Public Health, Baltimore, Maryland, USA.
authors: Susan E
authors: SE
authors: Dorman
authors: New Jersey Medical School-Rutgers, the State University of New Jersey, Newark, New Jersey, USA.
authors: David
authors: D
authors: Alland
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D024881:Drug Resistance, Bacterial; D005260:Female; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013183:Sputum / Q000382:microbiology*; D014397:Tuberculosis, Pulmonary / Q000175:diagnosis* / Q000382:microbiology; D055815:Young Adult
chemical_list: D007202:Indicators and Reagents
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