Tuberculosis (TB), an infectious disease caused by bacteria Mycobacterium tuberculosis, remains a significant global health issue, with millions of new cases and deaths reported each year. The World Health Organization (WHO) classifies TB as a priority infectious disease as it is so difficult to diagnose and treat. According to WHO Global Tuberculosis Report 2023, an estimated global total of 10.6 million people fell ill with TB in 2022, equivalent to 133 incident cases per 100,000 population. According to the same report, 1.3 million people succumbed to the disease in 2022.
TB BURDEN
With an estimated 2.64 million cases in 2021, India accounts for about 25% of the world’s TB burden. In 2023, India saw an improvement in its TB notification numbers compared to 2022, with 2.5 m TB cases reported. Of these, 1.6 m cases were notified by the public sector, while the private sector reported 838,116 cases. The public sector achieved 93% of its TB notification target, whereas the private sector reached 89%. According to WHO, India has the highest burden of TB , with two deaths occurring every three minutes from TB. These numbers underscore the ongoing challenge of TB control and the need for sustained efforts to prevent, diagnose, and treat the disease effectively.
Tuberculosis predominantly impacts the pulmonary system, although it can also affect other anatomical regions, including the kidneys, spine, and brain. TB involves airborne transmission, wherein an individual afflicted with the disease expels minuscule droplets containing the bacterium through coughing or sneezing. When an individual breathes in these tiny liquid particles, the bacteria can establish themselves in the lungs and commence reproduction. The majority of individuals who contract TB do not immediately develop active disease. Instead, they acquire latent TB infection, wherein the bacteria reside in their bodies but remain dormant and do not produce any symptoms. However, in certain circumstances, particularly when the immune system is compromised, the bacteria might become active and result in the development of tuberculosis disease. TB is a significant public health issue. However, it can be prevented and treated with suitable measures such as immunisation (using the Bacille Calmette-Guérin or BCG vaccine), early detection, and successful treatment.
DETECTION AND PREVENTION
Conventional detection procedures, including smear microscopies, are frequently inconclusive due to their extremely low sensitivity. The nucleic acid amplification test (NAAT) is a fast TB screening test that can identify TB bacteria from sputum samples in under three hours. PCR-based tests, such as CBNAAT and Truenat, can be used to confirm instances of tuberculosis that have tested positive. While these technologies exhibit excellent accuracy, their capacity is limited to simultaneously processing only a few samples. Additionally, they are designed as closed systems, meaning they require a specific PCR machine to complete the process. Furthermore, these technologies are expensive.
Dr Anoop Kumar Thekkuveettil and his team at the Division of Molecular Medicine, BMT wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology (SCTIMST) in Thiruvananthapuram have made a significant breakthrough in making early detection cost-effective. They have created a new nucleic acid amplification test to use the current PCR testing infrastructure. This has the potential to revolutionise the screening process for pulmonary TB on a big scale. The AG Chitra TB diagnosis kit employs a loop-mediated amplification methodology (LAMP) to amplify DNA isothermally. Contrary to polymerase chain reaction (PCR), which necessitates cycling through several temperature stages, LAMP amplifies DNA or RNA at a consistent temperature, usually about 60-65°C.
The LAMP reaction depends on a DNA polymerase that can displace strands and a group of four to six primers designed to identify six to eight places on the target sequence. These primers trigger the process of DNA synthesis and the subsequent synthesis of a displacement strand, resulting in the formation of stem-loop DNA structures known as amplicons. These amplicons act as templates for subsequent amplification, leading to exponential target sequence amplification.
An important benefit of LAMP is its exceptional specificity and sensitivity, enabling the identification of very minute quantities of target nucleic acid in a given sample. In addition, LAMP reactions can be visually examined for amplification using methods such as observing turbidity generated by the precipitation of magnesium pyrophosphate or utilising fluorescence-based detection techniques.
The researchers at SCTIMST altered the LAMP assay methodology to develop a real-time assay. Consequently, each sample will undergo analysis at one-minute intervals to examine the amplification profile, yielding exact outcomes. Throughout the experiment, 40 data points will be gathered from each sample to identify TB bacteria precisely. This test is a simplified nucleic acid amplification procedure compared to PCR. The test can be effortlessly conducted with minimum instruction and executed on any PCR machine with fluorescence detection.
The AG Chitra Tuberculosis diagnostic kit possesses the following characteristics:
1. This test is both fast and highly responsive, capable of being conducted in a laboratory setting in under 1 hour.
2. The test demonstrates a precision of 97.71%.
3. The device also incorporates an automated protocol for isolating DNA from TB germs in sputum samples.
4. It is more economical in comparison to PCR-based diagnostics.
5. The implementation of open-platform technologies for TB diagnostics will greatly contribute to the early detection of undiagnosed cases and help achieve the goal of eliminating TB by 2030.
6. The technology has been created indigenously and is safeguarded by intellectual property rights.
The kit includes all the essential reagents required to conduct the assay. Automated DNA isolation technology is available to extract DNA from the sputum sample. The cutting-edge technology has been officially authorised to M/S Agappe Diagnostics in Kochi. The AG Chitra TB testing kit has obtained manufacture and marketing approval from the Drugs Standard Control Organisation (CDSCO).
Images Courtesy: Dr Biju Dharmapalan
According to the lead researcher, Dr Anoop Kumar Thekkuveettil, the AG Chitra TB kit may be used with any PCR equipment, significantly decreasing the expenses associated with setting up TB screening centres. Furthermore, it will substantially affect the worldwide endeavours to attain a tuberculosis-free state by 2030. The countries can utilise their current infrastructure facilities for TB detection to identify undetected cases and establish an efficient treatment plan. It would be a commendable achievement if countries could successfully adopt TB screening and reduce the number of cases to less than 50 per 100,000 people during the next two-to-three years. In that case, we will achieve a world free from tuberculosis.
*The writer is a science communicator and an adjunct faculty at the National Institute of Advanced Studies, Bangalore. He can be reached at bijudharmapalan@gmail.com.
