Souradip Mallick, National Institute of Technology, Rourkela
Tuberculosis Disease (TB) is a major infectious disease with a high rate of morbidity and mortality. TB is one of the tenth leading causes of death worldwide ranking above HIV/AIDS. The World Health Organization (WHO) estimated that about 1.66 million people died from TB in 2020. Thus, various health care programs are being implemented to control TB spread and infection under the flagship of “End TB Strategy” by WHO. The accurate and timely diagnosis of TB is essential for the initiation of the treatment of the disease. However, not all TB cases are successfully identified, and about 40% of TB cases are estimated to be undiagnosed.
The timely diagnosis of active tuberculosis disease (TB) is most important to combat the transmission as well as the spread of Mycobacterium tuberculosis (Mtb), the causative agent for TB. A new specimen-direct rapid diagnostic method for TB was discovered, which consists of an isothermal amplification device such as Tiny Isothermal Nucleic acid quantification system (TINY) along with helicase-dependent amplification (HDA). HAD is an isothermal amplification technique along with TINY platform which used pUCIDT-AMP vector having the target DNA sequence for Mtb i.e. IS6110.
The limit of detection of this technique for detecting the IS6110 within a threshold time of 50 min is 2.5 × 105 copies of IS6110. HDA in TINY can detect TB using three IS6110-positive Mtb strains such as H37Rv, CDC 1551, and Erdman wild-type and one IS6110-negative Mycobacterium avium. TINY does not require a continuous power supply and is portable. Moreover, it has easier operation in clinical settings with poor infrastructure. The detection limit of TINY for the detection of IS6110 in Mtb is about 2.5 × 105 copies/µl. The GeneXpert Assay has a detection limit is about 131 CFU/ml of clinical specimen.
HDA in TINY platform can detect the presence of IS6110 in Mtb strains without any non-specific identification of the negative control strain. Moreover, it could detect the IS6110 directly from the spiked oral swabs without any non-specific amplification demonstrating its utility for oral specimens. Since there is no specialized sample preparation process for DNA isolation from oral swabs, it can be a more reliable diagnostic for TB. The specimen and HDA reaction mixture preparation take at most 30 min. HDA reaction over the TINY platform and result interpretation takes less than 60 minutes. Therefore, for overall detection, from specimen preparation to result in interpretation, HDA in TINY requires less than 90 minutes to detect the presence of Mtb in oral swabs. Moreover, TINY can be used for routine clinical examination and also in the community setting. Hence, HDA in TINY has the potential for the qualitative detection of TB rapidly and efficiently.
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References:
- Shanmugakani, R. K., Bonam, W., Erickson, D., & Mehta, S. (2021). An isothermal amplification-based point-of-care diagnostic platform for the detection of Mycobacterium tuberculosis: A proof-of-concept study. Current Research in Biotechnology, 3, 154–159. https://doi.org/10.1016/j.crbiot.2021.05.004
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