TrueNat for Tuberculosis: Introduction, Principle, Procedure, Result

Introduction

Table of Contents

TrueNat is a rapid, chip-based, and portable molecular diagnostic test. It is developed by Molbio Diagnostics for the detection of tuberculosis (TB) and rifampicin (RIF) resistance. It is the first molecular point-of-care test for TB recommended by the World Health Organization (WHO).

Principle

The TrueNat system utilizes Real-time Polymerase Chain Reaction (PCR) technology. It amplifies specific DNA sequences of the Mycobacterium tuberculosis complex (MTBC) on a disposable micro-PCR chip.

Detection Targets: The test targets the nrdB gene (standard MTB assay) or a combination of nrdZ and IS6110 (MTB Plus assay) for higher sensitivity.
Resistance Testing: Rifampicin resistance is identified by targeting the rpoB gene (RRDR region) using a probe melt assay on a separate reflex chip.

Procedure

The entire process takes approximately one hour for TB detection, with an additional hour for resistance testing.

Sample pre-treatment (20 mins): Sputum or extrapulmonary specimens are mixed with a liquefaction and lysis buffer to release DNA.
DNA Extraction (20 minutes): The pre-treated sample is loaded into a single-use cartridge and processed in the Trueprep AUTO v2 device to extract and purify the DNA.
Amplification (35 minutes): 6µl of purified DNA is mixed with freeze-dried PCR reagents. It is loaded onto a TrueNAT MTB chip. It is then placed in the Truelab Real-time micro–PCR Analyzer.
Reflex Testing (55 minutes): If TB is detected, a reflex test for rifampicin resistance is performed using the same extracted DNA on a Truenat MTB-RIF Dx chip.

Result Interpretation

Results are displayed on the device screen as follows:

MTB Detection:
- DETECTED: Positive for TB. The screen also displays the bacterial load (e.g., High, Medium, Low, Very Low).
- NOT DETECTED: Negative for TB.
Rifampicin Resistance:
- Resistance Detected: Mutation in the rpoB gene identified.
- Resistance Not Detected: No resistance mutations found.
- Indeterminate: Low bacterial load or error; requires a repeat test.
Validity:
- VALID/INVALID: Based on the Internal Positive Control (IPC) to ensure the test run was successful.
- ERROR: Indicates a hardware or process failure.

Application

Point-of-Care (POC): It is ideal for peripheral laboratories and primary health centers. It has its portability and battery operation.
Broad Diagnosis: Applicable for for both pulmonary and extrapulmonary tuberculosis (EPTB).
Active Case Finding: Deployed in mobile units and health camps in remote or hard-to-reach areas.
Initial Diagnostic: Recommended as the first-line test for anyone with TB symptoms and for rapid rifampicin resistance screening.

Keynotes

Infrastructure: Requires minimal setup; it does not need air conditioning and can operate up to 40°C.
Battery Power: It can run for up to 8–10 hours on a single charge.
Storage: Reagents and chips are stable at room temperature.
Sensitivity: The MTB Plus assay has a Limit of Detection (LOD) of 30 CFU/ml, making it more sensitive than standard microscopy (which needs ~5000 CFU/ml).

TrueNat for Tuberculosis: Introduction, Principle, Procedure, Result-Interpretation, application, and Keynotes

Introduction

Principle

Procedure

Result Interpretation

Application

Keynotes

Further Readings

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