CRISPR technology - innovative diagnostics for multidrug-resistant tuberculosis
DOI:
https://doi.org/10.52692/1857-0011.2025.2-82.35Keywords:
MDR-TB, diagnosis, CRISPR-Cas, mutations, Mycobacterium tuberculosisAbstract
Multidrug-resistant tuberculosis (MDR-TB) remains a major public health challenge, with approximately 400,000 cases reported globally in 2023. The Republic of Moldova is among the countries with the highest incidence of MDR- TB in the European region. In this context, the application of a faster diagnostic method, with higher sensitivity and specificity compared to existing tuberculosis diagnostic techniques-such as bacteriological or molecular methods like GeneXpert-which could enable the detection of Mycobacterium tuberculosis (MTB) even in modest samples or in the absence of sputum, becomes increasingly urgent.This paper synthesizes recent data from the scientific literature regarding the use of CRISPR-Cas in tuberculosis (TB) diagnostics, with a focus on diagnostic accuracy, amplification methods, and its potential applicability in resource-limited settings.The CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology offers a new, precise, and efficient approach to the rapid diagnosis of MDR-TB, which is crucial for early diagnosis and timely initiation of appropriate treatment-key steps in preventing the global spread of multidrug-resistant tuberculosis.This article explores the CRISPR diagnostic mechanism, its advantages, and a comparative analysis with currently used tuberculosis (TB) diagnostic methods.
References
Houben R.M.G.J., Dodd P.J. The Global Burden of Latent Tuberculosis Infection: A Re-estimation Using Mathematical Modelling. În: PLoS Med 2016, nr. 13, pp. e1002152. ISSN15491676.
Global tuberculosis report. 2024.
Pandya K., Jagani D., Singh N. CRISPR-Cas Systems: Programmable Nuclease Revolutionizing the Molecular Diagnosis. În: Mol Biotechnol 2024, nr. 66, pp. 1739–1753. ISSN15590305.
Abavisani M. et al. Mycobacterium tuberculosis Detection Using CRISPR Technology: An Updated Systematic Review and Meta-analysis. În Mol Diagn Ther 2024, nr. 28, pp. 777–790. ISSN11792000.
Qi Y. et al. CRISPR-Based Diagnostics: A Potential Tool to Address the Diagnostic Challenges of Tuberculosis. În Pathogens, nr. 11. Epub ahead of print 20 October 2022. DOI: 10.3390/pathogens11101211. ISSN2076-0817.
Mukherjee S. et al. Evolution of tuberculosis diagnostics: From molecular strategies to nanodiagnostics. În Tuberculosis, nr. 140. Epub ahead of print 1 May 2023. DOI: 10.1016/j.tube.2023.102340. ISSN1873281X.
Zhang X. et al. A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/ Cas system. În: BMC Infect Dis 2023, nr. 23, pp. 1–10. ISSN14712334.
Shmakov S. et al. Diversity and evolution of class 2 CRISPR-Cas systems. În Nat Rev Microbiol 2017, nr. 15, pp. 169–182. ISSN17401534.
Augustin L., Agarwal N. Designing a Cas9/ gRNA-assisted quantitative Real-Time PCR (CARP) assay for identification of point mutations leading to rifampicin resistance in the human pathogen Mycobacterium tuberculosis. În Gene 2023, nr. 857, pp. 147173. ISSN0378- 1119.
Li Q. et al. Rapid and Highly Sensitive Detection of Mycobacterium tuberculosis Utilizing the Recombinase Aided Amplification-Based CRISPR-Cas13a System. În Microorganisms 2024, nr. 12, pp. 1507. ISSN20762607.
Taufiq S. et al. An electrochemical biosensor for the detection of tuberculosis specific DNA with CRISPR- Cas12a and redox-probe modified oligonucleotide. În Heliyon 2024, nr. 10, pp. e40754. ISSN24058440.
Liu P. et al. A Recombinase Polymerase Amplification-Coupled Cas12a Mutant-Based Module for Efficient Detection of Streptomycin-Resistant Mutations in Mycobacterium tuberculosis. În Front Microbiol 2022, nr. 12, pp. 796916. ISSN1664302X.
Xiao G. et al. Direct detection from sputum for drug-resistant Mycobacterium tuberculosis using a CRISPR-Cas14a-based approach. În BMC Microbiol 2025, nr. 25, pp. 1–11. ISSN14712180.
Peng L. et al. Rapid detection of Mycobacterium tuberculosis in sputum using CRISPR-Cas12b combined with cross-priming amplification in a single reaction. În J Clin Microbiol, nr. 62. Epub ahead of print 1 January 2024. DOI: 10.1128/JCM.00923-23. ISSN1098660X.
Lin Z. et al. Ultra-sensitive in situ detection of intracellular Mycobacterium tuberculosis with CRISPR/ Cas12a. În Front Immunol, nr. 16. Epub ahead of print 2025. DOI: 10.3389/FIMMU.2025.1597654. ISSN16643224.
Qiao Y. et al. A chemiluminescent sensor based on CRISPR-HCR technology for the hypersensitive detection of Mycobacterium tuberculosis. În Analytical Methods 2024, nr. 16, pp. 7927–7939. ISSN17599679.
Wang X.Y. et al. Whole genome CRISPRi screening identifies druggable vulnerabilities in an isoniazid resistant strain of Mycobacterium tuberculosis. În Nature Communications , nr. 15. Epub ahead of print 1 December 2024. DOI: 10.1038/S41467-024-54072-W ISSN20411723.
Padda I.S., Reddy K.M. Antitubercular Medications. În StatPearls.
Xiong X.S. et al. Identification of Mycobacterium tuberculosis Resistance to Common Antibiotics: An Overview of Current Methods and Techniques. În Infect Drug Resist 2024, nr. 17, pp. 1491–1506. ISSN11786973.
Liu Q. et al. Drug resistance gene mutations and treatment outcomes in MDR-TB: A prospective study in Eastern China. În PLoS Negl Trop Dis 2021, nr. 15, pp. e0009068. ISSN19352735.
Khan A.S. et al. Genetic mutations underlying isoniazid-resistant Mycobacterium tuberculosis in Khyber Pakhtunkhwa, Pakistan. În Tuberculosis, nr. 138. Epub ahead of print 1 January 2023. DOI: 10.1016/j. tube.2022.102286. ISSN1873281X.
Islam M.M. et al. Phenotypic and Genotypic Characterization of Streptomycin-Resistant Multidrug- Resistant Mycobacterium tuberculosis Clinical Isolates in Southern China. În Microbial Drug Resistance 2020, nr. 26, pp. 766–775. ISSN19318448.
Cao G. et al. Completely Free from PAM Limitations: Asymmetric RPA with CRISPR/Cas12a for Nucleic Acid Assays. În ACS Sens 2023, nr. 8, pp. 4655– 4663. ISSN23793694.
Dai J. et al. Exploiting the Potential of Spherical PAM Antenna for Enhanced CRISPR-Cas12a: A Paradigm Shift toward a Universal Amplification-Free Nucleic Acid Test Platform. În Anal Chem 2025, nr. 97, pp. 1236–1245. ISSN15206882.
Riordan S.M. et al. Application of CRISPR/Cas9 for biomedical discoveries. În Cell Biosci 2015, nr. 5, pp. 1–11. ISSN20453701.
Zhou M. et al. The construction of CRISPR/ Cas9-mediated FRET 16S rDNA sensor for detection of Mycobacterium tuberculosis. În Analyst 2023, nr. 148, pp. 2308–2315. ISSN1364-5528.
View of CRISPR-Cas9-based electrochemical biosensor for the detection of katG gene mutations in isoniazid-resistant tuberculosis, https://pub.iapchem.org/ojs/index.php/admet/article/view/2766/2519 (accessed 29 June 2025).
Le T.H.U.T.H.U.Y., Huang Y., Xie J.P. Application and optimization of CRISPRi to the biology of Mycobacterium tuberculosis. În Zhonghua Jie He He Hu Xi Za Zhi 2024, nr. 47, pp. 376–382. ISSN1001-0939.
Gan T. et al. ERA-CRISPR/Cas12a system: a rapid, highly sensitive and specific assay for Mycobacterium tuberculosis. În Front Cell Infect Microbiol, nr. 14. Epub ahead of print 2024. DOI: 10.3389/FCIMB.2024.1454076 ISSN22352988.
Yang L. et al. CRISPR-Cas12a-Based Rapid and Sensitive Detection of rpoB L378R in Mycobacterium tuberculosis. Epub ahead of print 7 June 2023. DOI: 10.1101/2023.06.06.543922.
Deng Z. et al. Rapid and sensitive detection of Mycobacterium tuberculosis using the RPA/Cas12f1_ge4.1 system with fluorescence and lateral flow readouts. În Microbiol Spectr, nr. 13. Epub ahead of print 9 July 2025. DOI: 10.1128/SPECTRUM.02652-24. ISSN2165-0497.
Qian X. et al. CRISPR for companion diagnostics in low-resource settings. În Lab Chip 2024, nr. 24, pp. 4717–4740. ISSN1473-0189.
Lyu C. et al. CRISPR-based biosensing is prospective for rapid and sensitive diagnosis of pediatric tuberculosis. În International Journal of Infectious Diseases 2020, nr. 101, pp. 183–187. ISSN1201-9712.
Bai X. et al. A Highly Sensitive and Specific Detection Method for Mycobacterium tuberculosis Fluoroquinolone Resistance Mutations Utilizing the CRISPR-Cas13a System. În Front Microbiol 2022, nr. 13, pp. 847373. ISSN1664302X.
Ren W. et al. Development and clinical evaluation of a CRISPR/Cas13a-based diagnostic test to detect Mycobacterium tuberculosis in clinical specimens. În Front Microbiol 2023, nr. 14, pp. 1117085. ISSN1664302X.
Harrington L.B. et al. Programmed DNA destruction by miniature CRISPR-Cas14 enzymes. În Science (1979) 2018, nr. 362, pp. 839–842. ISSN10959203.
Kostyusheva A. et al. CRISPR-Cas systems for diagnosing infectious diseases. În Methods 2022, nr. 203, pp. 431–446. ISSN10959130.
Padmanaban V., Ranganathan U.D.K. CRISPR– Cas system and its use in the diagnosis of infectious diseases. În Microbiol Res, nr. 263. Epub ahead of print 1 October 2022. DOI: 10.1016/j.micres.2022.127100. ISSN09445013.
Chakraborty J. et al. CRISPR/Cas-Based Biosensor As a New Age Detection Method for Pathogenic Bacteria. În ACS Omega 2022, nr. 7, pp. 39562–39573. ISSN24701343.
Shi M. et al. EspB and HtpG interact with the type III-A CRISPR/Cas system of Mycobacterium tuberculosis. În Front Mol Biosci 2023, nr. 10, pp. 1261613. ISSN2296889X.
Wang Y. et al. LAMP-CRISPR-Cas12-based diagnostic platform for detection of Mycobacterium tuberculosis complex using real-time fluorescence or lateral flow test. În Microchimica Acta, nr. 188. Epub ahead of print 1 October 2021. DOI: 10.1007/S00604-021-04985-W ISSN14365073.
Wu K., Wu Z., Li X. Clinical diagnostic value of CRISPR-Cas13a-based molecular technology for tuberculosis. În Ir J Med Sci 2024, nr. 193, pp. 2889–2895. ISSN1863-4362.
Wang X.Y. et al. Whole genome CRISPRi screening identifies druggable vulnerabilities in an isoniazid resistant strain of Mycobacterium tuberculosis. În Nat Commun 2024, nr. 15, pp. 9791. ISSN20411723.
Thakku S.G. et al. Genome-wide tiled detection of circulating Mycobacterium tuberculosis cell-free DNA using Cas13. În Nat Commun, nr. 14. Epub ahead of print 1 December 2023. DOI: 10.1038/S41467-023-37183-8 ISSN20411723.
Wang J. et al. High-Performance Detection of Mycobacterium bovis in Milk Using Recombinase-Aided Amplification–Clustered Regularly Interspaced Short Palindromic Repeat–Cas13a–Lateral Flow Detection. În Foods, nr. 13. Epub ahead of print 1 June 2024. DOI: 10.3390/FOODS13111601 ISSN23048158.
Lyu C. et al. CRISPR-based biosensing is prospective for rapid and sensitive diagnosis of pediatric tuberculosis. În International Journal of Infectious Diseases 2020, nr. 101, pp. 183–187. ISSN18783511.
Cabibbe A.M. et al. Application of targeted next- generation sequencing assay on a portable sequencing platform for culture-free detection of drug-resistant tuberculosis from clinical samples. În J Clin Microbiol, nr. 58. Epub ahead of print October 2020. DOI: 10.1128/ JCM.00632-20/SUPPL_FILE/JCM.00632-20-SD002. XLSX. ISSN1098660X.
Cates L. et al. Laboratory costs of diagnosing TB in a high multidrug-resistant TB setting. În The International Journal of Tuberculosis and Lung Disease 2021, nr. 25, pp. 228. ISSN18157920.
Shetty A. et al. Revolutionizing Tuberculosis Management With Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas Technology: A Comprehensive Literature Review. În Cureus, nr. 16. Epub ahead of print 17 October 2024. DOI: 10.7759/CUREUS.71697 ISSN2168-8184.
Zein-Eddine R. et al. The future of CRISPR in Mycobacterium tuberculosis infection. În J Biomed Sci, nr. 30. Epub ahead of print 1 December 2023. DOI: 10.1186/ S12929-023-00932-4 ISSN14230127.
Akaçin İ. et al. Comparing the significance of the utilization of next generation and third generation sequencing technologies in microbial metagenomics. În Microbiol Res 2022, nr. 264, pp. 127154. ISSN0944-5013.
Hassan Y.M. et al. Recent developments and future directions in point-of-care next-generation CRISPR-based rapid diagnosis. În Clin Exp Med, nr. 25. Epub ahead of print 1 December 2025. DOI: 10.1007/S10238-024-01540- 8 ISSN15919528.
Kaushik A. et al. Establishment and evaluation of a naked-eye diagnostic assay for tuberculosis utilizing reverse isothermal amplification-assisted CRISPR-Cas in resource-limited settings. În Drug Target Insights 2025, nr. 19, pp. 31–40. ISSN11773928.
Li H. et al. Amplification-free CRISPR/ Cas detection technology: challenges, strategies, and perspectives. În Chem Soc Rev 2023, nr. 52, pp. 361–382. ISSN1460-4744.
Downloads
Published
License
Copyright (c) 2026 Bulletin of the Academy of Sciences of Moldova. Medical Sciences
This work is licensed under a Creative Commons Attribution 4.0 International License.
