Application of the rapid detection system for M.tuberculosis complex and rifampicin resistance Xpert MTB/RIF in decontaminated respiratory specimens, non-respiratory specimens and cultures
Abstract number: O235
Deforges L., Le Glaunec J.M., Launay N., Vergne R., Minaret A., Marzouk M., Marmiesse M., Finnström N., Legrand P.
Objectives: GeneXpert (Cepheid) is an automated real-time PCR system which is very easy to use and suitable for emergency use. The principle of the Xpert MTB/RIF test, running on GeneXpert, is to detect M. tuberculosis complex and mutations in the gene rpoB that cause resistance to rifampicin. This is done directly from at minimum of 500 ml of a respiratory sample. After 15 min treatment of the sample all stages of the sample preparation and PCR takes place within the instrument in 1 h30.
Some restrictions of use led us to validate this system on various types of samples.
Methods: Respiratory samples were decontaminated with NACl/NaOH and 500 ml of decontaminated product added to 1.5 ml of lysis buffer (provided with the Xpert product). For non-respiratory non-decontaminated samples (CSF, pleural liquid and biopsies) the sample volume was sometimes too low. In these cases distilled water was added up to 500 ml before addition of lysis buffer. For solid culture one colonie was resuspended in 500 ml of distilled water, for liquid culture 500 ml of medium were centrifuged prior to addition of lysis buffer. Fifteen decontaminated respiratory samples (PPD) and 24 non-pulmonary samples (PNP) were tested in the Xpert MTB/RIF assay, solid culture, Cobas TaqMan MTB test and cultures were tested with probes from GenProbe for identification.
Results: Twelve PPD of 15 were negative in Xpert MTB/RIF and the COBAS TaqMan MTB Test. Culture was also negative for 10 of these. In two samples atypical mycobacterium was isolated. 3 PPD were positive in Xpert and COBAS and showed no mutations in rpoB. The culture confirmed M. tuberculosis sensitive to rifampicin in these 3 cases. 20 PNP of 24 were negative by both PCR techniques and culture. Four of 24 (1 CSF, 1 pleural, 2 lymph node biopsies) were positive by both techniques and showed no mutations in the rpoB gene. The culture confirmed M. tuberculosis sensitive to rifampicin in these 4 cases. Nine culture isolates tested were identified as M. tuberculosis by Xpert and GenProbe. One isolate was determined resistant to rifampicin in Xpert. This was confirmed by susceptibility testing.
Conclusion: These preliminary results obtained on a limited number of samples show a perfect match for Xpert MTB/RIF with conventional laboratory tests irrespective of the nature of the sample tested.
|Session name:||Abstracts 20th European Congress of Clinical Microbiology and Infectious Diseases|
|Location:||Vienna, Austria, 10 - 13 April 2010|
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