Context:
Researchers at the Sree Chitra Tirunal Institute for Medical Sciences and Technology (SCTIMST), Thiruvananthapuram, have developed an innovative and cost-effective real-time LAMP (rt-LAMP) assay for the early diagnosis of Tuberculosis (TB). This molecular test offers high sensitivity and specificity, making it comparable to existing tests like GeneXpert and Truenat.
Key Features of the rt-LAMP Assay
- High Sensitivity:
- The rt-LAMP assay can detect TB DNA even in samples with as few as 10 copy numbers per microlitre, making it highly sensitive.
- Single Temperature Testing:
- Unlike RT-PCR, which requires three different temperature settings, the rt-LAMP test operates at a single temperature, making the process simpler and quicker.
- Real-time Monitoring:
- Using the fluorescent dye Syto 16, the amplification process can be monitored every minute during the test, as opposed to waiting until the end, offering real-time results.
- Quick Results:
- With six primers used for amplification (compared to only two in RT-PCR), the assay provides positive results in just 10-20 minutes.
- High Specificity:
- The requirement for all six primers to bind to the DNA before amplification begins ensures high specificity. The test avoids using probes, which reduces complexity and cost.
- Affordability:
- Since the fluorescent dye and primers are inexpensive, the rt-LAMP test is cost-effective, offering an affordable alternative to existing molecular tests.
Testing and Evaluation
- The assay was evaluated using 350 presumptive pulmonary TB sputum samples in two phases (October 2019 – March 2020 and January 2023 – March 2024).
- Comparison with Other Tests:
The rt-LAMP assay was compared against a microbiological reference standard (MRS), GeneXpert, and smear test. It showed 89.36% sensitivity and 94.06% specificity compared to MRS.
Advantages Over Existing Tests
- Repurposing RT-PCR Machines:
- The rt-LAMP kit can be used with existing RT-PCR machines, which can be reprogrammed to operate at a single temperature, enabling high-throughput testing (up to 96 or 384 samples in a single run).
- Open Platform System:
- The assay is designed as an open platform, allowing for easy integration with existing equipment and making it accessible for larger-scale use.
Regulatory Approval and Ongoing Validation
- The technology has been licensed to industry, received approval from the Central Drugs Standard Control Organization (CDSCO), and is currently being validated by the Indian Council of Medical Research (ICMR).
- The WHO Health Technology Access Pool program is also evaluating the technology, awaiting ICMR validation.
Impact on TB Diagnosis in India
- Current Diagnostics:
- As of 2023, approximately 79% of presumptive TB cases in India are diagnosed using sputum smear microscopy, while only 21% are tested with molecular assays.
- Increasing Molecular Testing:
- Despite the increase in molecular testing facilities (from 5,090 in 2022 to 6,496 in 2023), India still faces challenges in meeting the National Strategic Plan 2017-2025 to reduce reliance on smear microscopy.
The rt-LAMP assay offers a cost-effective and rapid diagnostic solution for TB, with high sensitivity and specificity. Its ability to integrate with existing RT-PCR machines and its potential for high-throughput testing makes it an important advancement in TB diagnostics. If validated and widely adopted, this assay could play a significant role in improving TB diagnosis and treatment, particularly in resource-limited settings.
