脑脊液宏基因组二代测序对结核性脑膜炎诊断价值

Value of metagenomic next-generation sequencing to the diagnosis of tuberculous meningitis

目的分析比较结核性脑膜炎患者应用宏基因组二代测序(metagenomic next-generation sequencing,mNGS)、脑脊液抗酸染色、结核分枝杆菌培养、结核分枝杆菌/利福平耐药实时荧光定量核酸扩增(Xpert MTB/RIF)检测结核分枝杆菌的结果,探讨mNGS对结核性脑膜炎的诊断价值。方法临床确诊的结核性脑膜炎患者34例,收集患者临床特征及脑脊液mNGS、常规检测、生化检测、抗酸染色、结核分枝杆菌培养、Xpert MTB/RIF检测等结果,比较脑脊液mNGS与病原学检测结核分枝杆菌检出率。结果34例结核性脑膜炎患者发热29例,头痛27例,颈项强直14例,精神行为异常9例,意识障碍11例,认知功能障碍4例,继发性癫痫5例。脑脊液压力升高26例[240(150,330)mm H_(2)O],白细胞计数升高32例[194.0(92.5,453.5)×10^(6)/L],均以单核细胞为主,蛋白质水平升高32例[1.58(0.95,2.57)g/L],葡萄糖水平降低20例[1.93(1.19,2.95)mmol/L],氯离子水平降低21例[115.20(106.85,121.30)mmol/L]。34例结核性脑膜炎患者脑脊液Xpert MTB/RIF检出结核分枝杆菌阳性12例,抗酸染色检出结核分枝杆菌阳性10例,结核分枝杆菌培养阳性11例,其中4例为3种检测方法阳性,6例为抗酸染色和结核分枝杆菌培养阳性,脑脊液病原学检测共检出结核分枝杆菌阳性12例,阳性检出率35.3%;脑脊液mNGS检出结核分枝杆菌阳性24例,序列数2~49条,未检测到其他可疑病毒、细菌、弓形虫等微生物序列,阳性检出率70.6%。脑脊液mNGS检测结核分枝杆菌阳性检出率高于脑脊液病原学检测(P=0.027)。结论结核性脑膜炎患者临床表现及脑脊液常规、生化表现不典型,脑脊液抗酸染色、结核分枝杆菌培养、Xpert MTB/RIF阳性检出率低,mNGS可提高阳性检出率,为临床治疗方案制订提供依据。

Objective To analyze and compare the results of Mycobacterium tuberculosis by metagenomic next-generation sequencing(mNGS), acid-fast staining, Mycobacterium tuberculosis culture, and real-time fluorescence quantitative nucleic acid amplification assay detection of Mycobacterium tuberculosis/rifampicin resistance(Xpert MTB/RIF) in patients with tuberculous meningitis, and to investigate the value of mNGS to the diagnosis of tuberculous meningitis. Methods The clinical features and results of cerebrospinal fluid mNGS, routine test, biochemical test, acid-fast staining, Mycobacterium tuberculosis culture and Xpert MTB/RIF test were collected in 34 patients with clinically diagnosed tuberculous meningitis to compare the detection rates of Mycobacterium tuberculosis by cerebrospinal fluid mNGS and pathogenic test. Results In 34 patients with tuberculous meningitis, there were 29 cases of fever, 27 of headache, 14 of cervical tonicity, 9 of mental behavioral abnormalities, 11 of consciousness disturbance, 4 of cognitive dysfunction, and 5 of secondary epilepsy. Cerebrospinal fluid pressure was elevated to 240(150, 330) mm H_2O in 26 cases and the white blood cell count was elevated to 194.0(92.5, 453.5)×10^(6)/L in 32 cases, with a predominance of monocytes;the protein level was elevated to 1.58(0.95, 2.57) g/L in 32 cases, glucose level decreased to 1.93(1.19, 2.95) mmol/L in 20 cases and chloride level decreased to 115.20(106.85, 121.30) mmol/L in 21 cases. In 34 patients with tuberculous meningitis, Mycobacterium tuberculosis was detected by cerebrospinal fluid Xpert MTB/RIF in 12 cases, by acid-fast staining in 10 cases, and by Mycobacterium tuberculosis culture in 11 cases, in which 4 cases were positive by the combination of the above three methods, and 6 were positive by acid-fast staining and Mycobacterium tuberculosis culture. Mycobacterium tuberculosis was detected by pathogenic test in 12 cases(35.3%). Mycobacterium tuberculosis was detected by mNGS in 24cases(70.6%),with 2to 49sequences.No other suspicious virus,bacteria,Toxoplasma gondii or other microbial sequences were detected.The detection rate of Mycobacterium tuberculosis by mNGS was higher than that by pathogenic test in cerebrospinal fluid(P=0.027).Conclusion The clinical manifestations and cerebrospinal fluid routine and biochemical tests are atypical in patients with tuberculous meningitis,and acid-fast staining,Mycobacterium tuberculosis culture and Xpert MTB/RIF detection in cerebrospinal fluid have low detection rates for Mycobacterium tuberculosis,while mNGS can improve the detection rate and provides a basis for clinical treatment planning.