The Cepheid Xpert MTB/RIF research-use-only (RUO) assay and a laboratory-developed test (LDT) targeting IS6110 were evaluated and compared to mycobacterial culture as the gold standard. The performance characteristics of both molecular assays were determined by using 112 specimens from 90 patients, including 89 pulmonary specimens and 23 extrapulmonary specimens. Of the specimens tested, 37 (33%) were culture positive for the Mycobacterium tuberculosis complex; 29 were pulmonary, and 8 were extrapulmonary. Of these culture-positive specimens, 83% of the pulmonary specimens and 50% of the extrapulmonary specimens were smear positive. There was complete concordance between the smear-positive culture-positive specimens, independent of the anatomical site (100% sensitivity). The sensitivity of the MTB/RIF RUO assay for smearnegative specimens was 60% for pulmonary and 75% for extrapulmonary specimens, while the IS6110 LDT sensitivities were 40% and 0%, respectively. There was also complete concordance among the culture-negative specimens tested. Both assays showed 95% specificity, with four culture-negative specimens testing as positive. A review of patient records indicated that there was a high likelihood of the presence of M. tuberculosis complex DNA in the false-positive specimens. Biosafety analysis was performed and showed an acceptable reduction in organism viability using the processing methods described above. Both molecular assays are suitable for the detection of M. tuberculosis isolates in smear-positive pulmonary and extrapulmonary specimens, while the sensitivity of the detection of M. tuberculosis isolates in smear-negative specimens was variable.Tuberculosis remains an important worldwide health concern, with over 9 million new cases and approximately 2 million deaths annually. Currently in the United States, there is only one FDA-approved nucleic acid amplification test (NAAT) for the Mycobacterium tuberculosis complex, the Gen-Probe Amplified Mycobacterium Tuberculosis Direct (MTD) test (San Diego, CA). The Roche Amplicor Mycobacterium tuberculosis complex (MTB) test was discontinued in 2010. A recently introduced research-use-only (RUO) NAAT for the M. tuberculosis complex is the GeneXpert MTB/RIF assay (Cepheid, Sunnydale, CA), which simultaneously detects the presence of the M. tuberculosis complex and rifampin resistance directly from respiratory specimens. The Xpert MTB/RIF assay is a semiquantitative, nested, real-time PCR designed to amplify a 192-bp segment of the M. tuberculosis complex rpoB gene using five overlapping molecular beacon probes that span the entire 81-bp rifampin-resistance-determining region. The MTB/RIF assay also contains an internal control in which the detection of lyophilized Bacillus atrophaeus subsp. globigii spores serves as an internal processing and amplification control. Previous work has demonstrated that the Xpert MTB/RIF assay displays high percentages of sensitivity and specificity for the detection of M. tuberculosis complex isolates, particularly in smear-p...

These results suggest that PDE 5 inhibitors may play a more important role in early postoperative skin flap viability rather than at later time points and may be beneficial for skin flap viability as shown in the rat model. PDE 5 inhibitors may reduce the extent of necrosis after reconstructive surgeries.

We present the evolution of testing algorithms at our institution in which the C. Diff Quik Chek Complete immunochromatographic cartridge assay determines the presence of both glutamate dehydrogenase and Clostridium difficile toxins A and B as a primary screen for C. difficile infection and indeterminate results (glutamate dehydrogenase positive, toxin A and B negative) are confirmed by the GeneXpert C. difficile PCR assay. This two-step algorithm is a cost-effective method for highly sensitive detection of toxigenic C. difficile.

dClostridium difficile can carry a genetically variable pathogenicity locus (PaLoc), which encodes clostridial toxins A and B. In hospitals and in the community at large, this organism is increasingly identified as a pathogen. To develop a diagnostic test that combines the strengths of immunoassays (cost) and DNA amplification assays (sensitivity/specificity), we targeted a genetically stable PaLoc region, amplifying tcdB sequences and detecting them by hybridization capture. The assay employs a hot-start isothermal method coupled to a multiplexed chip-based readout, creating a manual assay that detects toxigenic C. difficile with high sensitivity and specificity within 1 h. Assay automation on an electromechanical instrument produced an analytical sensitivity of 10 CFU (95% probability of detection) of C. difficile in fecal samples, along with discrimination against other enteric bacteria. To verify automated assay function, 130 patient samples were tested: 31/32 positive samples (97% sensitive; 95% confidence interval [CI], 82 to 99%) and 98/98 negative samples (100% specific; 95% CI, 95 to 100%) were scored correctly. Largescale clinical studies are now planned to determine clinical sensitivity and specificity.

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