A new method for the detection of Mycobacterium tuberculosis based on the CRISPR/Cas system

BMC Infectious Diseases

Research

Yueying Huang¹  Xiaoying He²  Lei Chen²  Zhaobao Pan²  Heng Li³  Xiaoyu Zhang³  Yubo Zhang³ 

[1] Department of Laboratory Medicine, Peking University Shenzhen Hospital, Shenzhen, China;Department of Laboratory Medicine, the Second People’s Hospital of Weifang 261041, 261041, Weifang, Shandong, China;Department of Medical Laboratory, Weifang Medical University, 261041, Weifang, Shandong, China;

关键词: CRISPR;    RAA;    Mycobacterium tuberculosis;    Molecular testing;    Diagnosis;   

DOI  :  10.1186/s12879-023-08656-4

 received in 2023-02-15, accepted in 2023-09-28,  发布年份 2023

来源: Springer

【 摘 要 】

ObjectMycobacterium 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.MethodsIn 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.ResultsMTB 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).ConclusionTB-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.

【 授权许可】

CC BY   
© BioMed Central Ltd., part of Springer Nature 2023

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