Value of CRISPR-Cas13a detection in the diagnosis of pulmonary tuberculosis

doi:10.19982/j.issn.1000-6621.20230014

Abstract

Abstract:

Objective: To assess the diagnostic value of CRISPR-Cas13a detection in pulmonary tuberculosis (TB) using clinical samples. Methods: A retrospective study was conducted at Beijing Chest Hospital Affiliated to Capital Medical University. The bronchoalveolar lavage fluid (BALF) specimens of 102 patients with pulmonary TB confirmed by GeneXpert MTB/RIF test (Xpert test) from September 2022 to December 2022 were enrolled as TB group, and the BALF specimens of 100 patients (including lung cancer, bacterial pneumonia, bronchiectasis, and chronic obstructive pulmonary disease) during the same period were recruited as non-TB group. The BALF samples of all patients were subjected to acid-fast bacilli smear microscopy, culture, Xpert test and CRISPR-Cas13a test. The diagnostic value of CRISPR-Cas13a in the detection of BALF for pulmonary TB was evaluated based on the results of Xpert test. Results: Among 102 specimens in the TB group, 99 (97.1%) were positive for CRISPR-Cas13a, 40 (39.2%) were positive for smear, and 68 (66.7%) were positive for culture. The positive detection rate of CRISPR-Cas13a was significantly higher than those of smear and culture (smear: χ²=58.141, P<0.001; culture: χ²=30.250, P<0.001). Compared with traditional methods, CRISPR-Cas13a was able to identify 28 clinically confirmed TB patients which would be otherwise missed by either smear or culture. The positive detection rate of Xpert was 100.0% in TB group, and the negative detection rate of Xpert was 100.0% in non-TB group. Taking the results of Xpert as a reference, the sensitivity, specificity, accuracy, positive predictive value and negative predictive value of CRISPR-Cas13a in detecting pulmonary TB were 97.1% (99/102), 96.0% (96/100), 96.5% (195/202), 96.1% (99/103) and 97.0% (96/99), respectively. Conclusion: Compared with traditional tests, CRISPR-Cas13a detection is simple and fast, with higher sensitivity and higher specificity in the detection of BALF for the diagnosis of pulmonary TB, and has a good application prospect.

Key words: Repetitive sequences, nucleic acid, Tuberculosis, pulmonary, Bronchoalveolar lavage fluid, CRISPR-Cas system

CLC Number:

R521
R8

Zhang Zhiguo, Shang Yuanyuan, Zhang Xuxia, Liu Rongmei, Ma Liping, Qin Lin, Kong Zhongshun, Ren Weicong. Value of CRISPR-Cas13a detection in the diagnosis of pulmonary tuberculosis[J]. Chinese Journal of Antituberculosis, 2023, 45(6): 589-593. doi: 10.19982/j.issn.1000-6621.20230014

Figures/Tables 2

References 19

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名称	用量
PCR 产物	2μl
LwCas13a 蛋白	45nmol/L
NTP混合物	1μL (每种NTP终浓度为1mmol/L)
T7 RNA聚合酶	0.4μl
鼠RNA酶抑制剂	0.5μl
crRNA	100ng
RNase Alert v2	125nmol/L
10×缓冲液	2.5μl
无霉无菌水	补足体系至25μl