A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system
Abstract Object Mycobacterium tuberculosis (MTB) is a bacterium that can cause zoonoses by aerosol transmission. Tuberculosis (TB) caused by MTB heavily burdens world public health security. Developing efficient, specific, convenient, and inexpensive MTB assays are essential for preventing and contr...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2023-10-01
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| Series: | BMC Infectious Diseases |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12879-023-08656-4 |
| _version_ | 1797453768948711424 |
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| author | Xiaoyu Zhang Xiaoying He Yubo Zhang Lei Chen Zhaobao Pan Yueying Huang Heng Li |
| author_facet | Xiaoyu Zhang Xiaoying He Yubo Zhang Lei Chen Zhaobao Pan Yueying Huang Heng Li |
| author_sort | Xiaoyu Zhang |
| collection | DOAJ |
| description | Abstract Object Mycobacterium tuberculosis (MTB) is a bacterium that can cause zoonoses by aerosol transmission. Tuberculosis (TB) caused by MTB heavily burdens world public health security. Developing efficient, specific, convenient, and inexpensive MTB assays are essential for preventing and controlling TB. Methods In this study, we established a specific detection method for MTB using the Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) system, combined with recombinase mediated isothermal nucleic acid amplification (RAA) to improve the sensitivity of the detection system and achieve “two-level” amplification of the detection signal. The sensitivity and specificity of RAA combined with the CRISPR/Cas system were analyzed. Using BACTEC 960 culture as the gold standard for detecting MTB, we established the TB-CRISPR technique by testing 504 samples from patients with suspected TB. Results MTB H37Ra could be seen as low as 3.13 CFU/mL by the CRISPR-Cas12a system targeting IS6110. With BACTEC960 culture (120 positives and 384 negatives) as the gold standard, the sensitivity of the TB-CRISPR technique was 0.883 (0.809–0.932), and the specificity was 0.940 (0.910–0.961). According to the receiver operating characteristic (ROC) curve analysis, the area under the curve (AUC) reached 0.944 (0.914–0.975) within 95% CI. The positive likelihood ratio (PLR) was 14.747 (9.870-22.035), and the negative likelihood ratio (NLR) was 0.124 (0.076–0.203). The positive predictive value (PPV) was 0.822 (0.742–0.881), and the negative predictive value (NPV) was 0.963 (0.937–0.979). Conclusion TB-CRISPR plays an essential role in the molecular diagnosis of TB. The whole detection time is less than 1.5 h. It is easy to operate and does not need complex instruments. It is of great significance for the rapid detection of MTB and the clinical diagnosis of TB. |
| first_indexed | 2024-03-09T15:27:39Z |
| format | Article |
| id | doaj.art-cbe05bbc1f5a412285a4feecd5162444 |
| institution | Directory Open Access Journal |
| issn | 1471-2334 |
| language | English |
| last_indexed | 2024-03-09T15:27:39Z |
| publishDate | 2023-10-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Infectious Diseases |
| spelling | doaj.art-cbe05bbc1f5a412285a4feecd51624442023-11-26T12:27:09ZengBMCBMC Infectious Diseases1471-23342023-10-0123111010.1186/s12879-023-08656-4A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas systemXiaoyu Zhang0Xiaoying He1Yubo Zhang2Lei Chen3Zhaobao Pan4Yueying Huang5Heng Li6Department of Medical Laboratory, Weifang Medical UniversityDepartment of Laboratory Medicine, the Second People’s Hospital of Weifang 261041Department of Medical Laboratory, Weifang Medical UniversityDepartment of Laboratory Medicine, the Second People’s Hospital of Weifang 261041Department of Laboratory Medicine, the Second People’s Hospital of Weifang 261041Department of Laboratory Medicine, Peking University Shenzhen HospitalDepartment of Medical Laboratory, Weifang Medical UniversityAbstract Object Mycobacterium tuberculosis (MTB) is a bacterium that can cause zoonoses by aerosol transmission. Tuberculosis (TB) caused by MTB heavily burdens world public health security. Developing efficient, specific, convenient, and inexpensive MTB assays are essential for preventing and controlling TB. Methods In this study, we established a specific detection method for MTB using the Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) system, combined with recombinase mediated isothermal nucleic acid amplification (RAA) to improve the sensitivity of the detection system and achieve “two-level” amplification of the detection signal. The sensitivity and specificity of RAA combined with the CRISPR/Cas system were analyzed. Using BACTEC 960 culture as the gold standard for detecting MTB, we established the TB-CRISPR technique by testing 504 samples from patients with suspected TB. Results MTB H37Ra could be seen as low as 3.13 CFU/mL by the CRISPR-Cas12a system targeting IS6110. With BACTEC960 culture (120 positives and 384 negatives) as the gold standard, the sensitivity of the TB-CRISPR technique was 0.883 (0.809–0.932), and the specificity was 0.940 (0.910–0.961). According to the receiver operating characteristic (ROC) curve analysis, the area under the curve (AUC) reached 0.944 (0.914–0.975) within 95% CI. The positive likelihood ratio (PLR) was 14.747 (9.870-22.035), and the negative likelihood ratio (NLR) was 0.124 (0.076–0.203). The positive predictive value (PPV) was 0.822 (0.742–0.881), and the negative predictive value (NPV) was 0.963 (0.937–0.979). Conclusion TB-CRISPR plays an essential role in the molecular diagnosis of TB. The whole detection time is less than 1.5 h. It is easy to operate and does not need complex instruments. It is of great significance for the rapid detection of MTB and the clinical diagnosis of TB.https://doi.org/10.1186/s12879-023-08656-4CRISPRRAAMycobacterium tuberculosisMolecular testingDiagnosis |
| spellingShingle | Xiaoyu Zhang Xiaoying He Yubo Zhang Lei Chen Zhaobao Pan Yueying Huang Heng Li A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system BMC Infectious Diseases CRISPR RAA Mycobacterium tuberculosis Molecular testing Diagnosis |
| title | A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system |
| title_full | A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system |
| title_fullStr | A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system |
| title_full_unstemmed | A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system |
| title_short | A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system |
| title_sort | new method for the detection of mycobacterium tuberculosis based on the crispr cas system |
| topic | CRISPR RAA Mycobacterium tuberculosis Molecular testing Diagnosis |
| url | https://doi.org/10.1186/s12879-023-08656-4 |
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