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Indian Scientists Develop Highly Sensitive and Rapid Tests for Detection of Tuberculosis

Tuberculosis (TB) continues to be one of the leading causes of death worldwide. According to the World Health Organisation (WHO), some 10 million people across the world fell ill with TB in 2017, and 1.6 million died from the disease. India, which saw an estimated 28 lakh cases occurred and 4.8 lakh people died due to the disease in 2015, accounts for one-fourth of the global TB burden. Offering hope of finding tuberculosis early, scientists at the Translational Health Science and Technology Institute (THSTI) and All India Institute of Medical Sciences, New Delhi, have jointly developed highly sensitive and rapid tests for detection of TB infection in lungs and surrounding membranes, says a report in India Science Wire. Early detection and treatment are crucial to prevent the spread, outbreaks, and development of resistance.

Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis. It commonly affects the lungs, but can also affect other parts of the body. It spreads from person to person through inhalation of infected air - when people who are infected with TB cough, sneeze they propel the germs into the air. According to the report, when the bacteria attack the lungs, the disease manifests itself as pulmonary form or pulmonary TB - the most common form of TB. But in 2016,  about 15% of new patients were found infected with extrapulmonary TB where organs other than the lungs may be affected.

Till date, the detection of all forms of TB is mostly based on sputum smear microscopy and culture tests. Although smear microscopy is inexpensive and simple, it has low sensitivity. A culture test is highly sensitive but takes 2 to 8 weeks to get results. For detection of bacterial proteins in sputum samples, conventional diagnostic tests use antibodies. And such tests suffer from limitations including batch-to-batch variability, limited shelf-life, and cost said the report.

However, to help address the shortcomings in the present testing protocol, the researchers developed two DNA aptamer-based tests - Aptamer Linked Immobilized Sorbent Assay (ALISA) and Electrochemical Sensor (ECS) for detecting a bacterial protein in the sputum. Aptamers are DNA, RNA or peptide molecules that bind to a specific target molecule. They can be used for both basic research and clinical purposes as macromolecular drugs. Aptamers bind the right target (which defines sensitivity) and at the same time rule out any non-specific binding to other targets.

In the present work, the researchers compared the performance of the newly developed tests with antibody-based tests in 314 sputum samples. The found that ALISA produced 92% sensitivity compared to the antibody-based method that showed 68% sensitive. The team used ALISA to detect a bacterial protein, HspX. But it took five hours for the researchers to yield results as the method requires sputum immobilisation which is a time-consuming step. So the team developed a simplified ECS test. In this technique, the team immobilized the aptamer with an electrode, and upon binding to HspX in the sputum sample, a drop in the electrical signal was recorded.

According to the researchers, the ECS test can be used for screening of samples in the field. It takes as less as 30 minutes to deliver results and is highly sensitive. The ECS test could also detect HspX protein in 91% of the samples tested in the study. In addition, there is no need for sputum sample preparation which is a complex and time-consuming process, said the report.

“We aim to develop aptamers for detection of multiple bacterial proteins simultaneously which is expected to lead a more robust test,” said Tarun Sharma, a member of the research team.

“The aptamer-based screening tests for pulmonary TB, pleural TB, and TB meningitis hold immense promise for a country like India, where the disease burden in high and primary health care is only a dream for many. The ECS platform could be used in a mobile screening van at the point-of-care,” added Jaya Tyagi, who led the research team AIIMS. “We hope that these tests are adopted by the TB programs in the country,” she added.

In the study, the team used the aptamer-based test also for detection of pleural TB, the second most prevalent form of extrapulmonary TB. The researchers note the early diagnosis of pleural TB is limited by the availability of a sensitive and rapid test. The performance of existing DNA-based tests varies widely due to low bacterial load in the pleural fluid sample.

“There is no test for making a confirmed diagnosis of pleural TB. Even WHO-endorsed Gene Xpert has a poor sensitivity of 22%. On the contrary, our aptamer-based test for pleural TB showed 93% sensitivity and is cost-effective,” explained Sagarika Haldar, a member of the research team, was quoted as saying by India Science Wire.

The results were published in the journals ACS Infectious Diseases and Analytical Biochemistry.

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