molecular detection of tuberculosis and rifampin resistance.
1. Rapid Molecular Detection of
Tuberculosis and Rifampin
Resistance.
Presented By : Khaled Alkhodari
Supervised By :Dr. saeb aliwaini
Due to : 26/11/2014
3. Introduction
Tuberculosis (TB): is a bacterial infection that can spread through the lymph
nodes and bloodstream to any organ in the body. It is most often found in the
lungs.
Rifampin : is a drug used to treat TB
inhibits bacterial RNA polymerase.
Bacterial resistance to it is caused by mutations in rpoB gene leading to a change
in the structure of the β subunit of RNA polymerase.
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4. Methods
Xpert MTB/RIF : an automated molecular test for Mycobacterium
tuberculosis(MTB) and resistance to rifampin (RIF).
Three sputum specimens:
Two specimens were processed with:
N-acetyl-l-cysteine ( NALC ) and sodium hydroxide (NaOH ) before microscopy, solid and
liquid culture.
The MTB/RIF test.
One specimen was used for direct testing with microscopy and the MTB/RIF test.
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5. Xpert MTB/RIF Test
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an automated molecular test for
Mycobacterium tuberculosis(MTB) and
resistance to rifampin (RIF),
polymerasechain-reaction (PCR) assay to
amplify an MTB specific sequence of the
rpoB gene , gene encodes the β subunit of
bacterial RNA-polymerase.
11. Conclusion
The MTB/RIF test provided sensitive detection of tuberculosis and rifampin
resistance directly from untreated sputum in less than 2 hours with minimal
hands-on time.
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13. The authors
Catharina C. Boehme, M.D., Pamela Nabeta, M.D., Doris Hillemann, Ph.D.,
Mark P. Nicol, Ph.D.,
Shubhada Shenai, Ph.D., Fiorella Krapp, M.D., Jenny Allen, B.Tech., Rasim
Tahirli, M.D., Robert Blakemore, B.S.,
Roxana Rustomjee, M.D., Ph.D., Ana Milovic, M.S., Martin Jones, Ph.D., Sean
M. O’Brien, Ph.D.,
David H. Persing, M.D., Ph.D., Sabine Ruesch-Gerdes, M.D., Eduardo Gotuzzo,
M.D., Camilla Rodrigues, M.D.,
David Alland, M.D., and Mark D. Perkins, M.D.
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Editor's Notes
1- TB never develop symptoms because the bacteria can live in an inactive form in the body.
But if the immune system weakens, such as in people with HIV or elderly adults, TB bacteria can become active.
In their active state, TB bacteria cause death of tissue in the organs they infect. Active TB disease can be fatal if left untreated.
2-RNA polymerase, the enzyme responsible for DNA transcription to form mRNA
***Early detection is essential to reduce the death rate but the complexity and infrastructure needs of sensitive methods limit their accessibility and effect.
NALC: mucolytic agent
PCR is used to amplify a single copy or a few copies of a piece of DNA 7- for rpoB gene encodes the β subunit of bacterial RNA polymerase. It is the site of mutations that confer rifampin
provide three sputum specimens over a 2 days+1 of them in morning
two of the three samples were processed with ( N A L C – N a OH ):for decontamination of a sputum
followed by centrifugation, and then were resuspended in 1.5 ml of phosphae buffer and subjected to microscopy with Ziehl–Neelsen staining-identify acid-fast organisms-, and cultivation on solid medium & and liquid medium & and the MTB/RIF test
MGIT :liquid medium (BACTEC MGIT [mycobacteria growth indicator tube]
LJ : solid medium
(egg-based Löwenstein–Jensen
clinical tuberculosis smear- and culture-negative for pulmonary tuberculosis who nonetheless were treated for tuberculosis on the basis of clinical and radiologic findings
Smear + case was defined as >1 bacilli per 100 fields)
Phenotypic Drug-Susceptibility Testing: which drugs the TB bacteria in a person are sensitive to, and therefore whether the person has got drug resistant TB.