Xpert MTB/RIF: Introduction, Principle, Procedure, Result-Interpretation, Uses, and Keynotes
GeneXpert® Xpert® MTBRIF Test Result- MTB Detected (Very Low) and Rifampicin Resistance Not Detected
Introduction
Table of Contents
The Xpert MTB/RIF assay is an automated, cartridge-based molecular test used to detect Mycobacterium tuberculosis (MTB) and rifampicin (RIF) resistance, the most potent first-line anti-TB drug.
Fig. GeneXpert IV 4-module system
Endorsed by the WHO in 2010, the Xpert MTB/RIF assay revolutionized TB diagnosis by providing results in less than two hours. It is performed on the Cepheid GeneXpert System, which integrates sample processing and real-time PCR into a single-use disposable cartridge.
Principle
The assay uses nested real-time PCR and molecular beacons to amplify and detect a specific DNA sequence of the rpoB gene.
Target: An 81-base pair “core region” (RRDR) of the rpoB gene.
Mechanism: Five molecular beacons (Probes A–E) are used; each is labeled with a different fluorophore. They are designed to differentiate between the wild-type (normal) sequence and mutations associated with rifampicin resistance.
Procedure
Sample Collection: Collect at least 1 mL of raw sputum or 0.5 mL of sputum sediment.
Liquefaction: Add twice the volume of Xpert Sample Reagent to the sample.
Incubation: Shake vigorously and incubate at room temperature for 15 minutes (with a second shake after 10 minutes) until the sample is completely liquefied.
Loading: Use a sterile pipette to transfer 2 mL of the treated sample into the Xpert MTB/RIF cartridge.
Running the Test: Scan the cartridge barcode, insert the cartridge into the GeneXpert module, and start the run via the software. Results are ready in approximately 100 minutes.
Positive for TB; likely susceptible to rifampicin.
MTB Detected; RIF Resistance Detected
Positive for TB; indicates multidrug-resistant TB (MDR-TB).
MTB Detected; RIF Resistance Indeterminate
TB is present, but the load is too low to determine RIF status.
MTB Not Detected
No M. tuberculosis DNA found.
Invalid / Error / No Result
Test failed due to sample processing error or instrument issues; must be repeated.
Fig. GeneXpert® Xpert® MTBRIF Test Result- MTB Detected (Very Low) and Rifampicin Resistance Not Detected
Uses
Rapid Diagnosis: Primary diagnostic tool for individuals with clinical suspicion of pulmonary TB.
MDR-TB Screening: Quickly identifies patients likely to have multidrug-resistant strains.
HIV/TB Co-infection: Preferred test for people living with HIV, as they often have low bacterial loads (smear-negative).
Infection Control: Aids in deciding if a patient should be placed in or removed from airborne isolation.
Keynotes
Not for Monitoring: It cannot be used to monitor treatment response because it detects both live and dead DNA.
Controls: Each cartridge includes a Sample Processing Control (SPC) to monitor PCR inhibitors and a Probe Check Control (PCC) to verify reagent integrity.
Sensitivity: Significantly more sensitive than AFB smear microscopy, especially in smear-negative, culture-positive cases.
Culture Still Required: Culture remains necessary for definitive drug susceptibility testing for other drugs (e.g., isoniazid).