二代测序技术在骨关节结核临床诊断中的应用价值

doi:10.19983/j.issn.2096-8493.20230094

摘要/Abstract

摘要：

目的: 评估二代测序技术(next generation sequencing,NGS)在骨关节结核诊断中的应用价值。方法: 采用回顾性研究方法,参照入组标准收集2019年12月至2022年12月河北省胸科医院骨科收治的185例疑似骨关节结核患者临床资料,根据最终临床诊断将患者分为结核组(骨关节结核155例)和非结核组(非骨关节结核30例)。所有患者病灶标本均经手术或穿刺途径获取(包括51份脓液、89份肉芽组织和45份骨组织标本),且均同时送检NGS、BACTEC MGIT 960分枝杆菌液体培养(简称“培养”)及利福平耐药实时荧光定量核酸扩增(GeneXpert MTB/R1F,简称“Xpert”)检测。以最终临床诊断为参照标准,比较3种检测方法诊断骨关节结核的检测效能,并分析3种方法检测脓液、肉芽组织和骨组织标本的阳性检出情况。结果: 185例疑似骨关节结核患者中,NGS检测骨关节结核的阳性率[63.24%(117/185)]明显高于Xpert[54.05%(100/185)]和分枝杆菌培养[35.68%(66/185)],差异均有统计学意义(χ²=24.982,P<0.001;χ²=37.934,P<0.001)。以最终临床诊断为参照标准,NGS、Xpert和培养法诊断骨关节结核的敏感度分别为74.84%(116/155)、64.52%(100/155)和42.58%(66/155),特异度分别为 96.67%(29/30)、100.00%(30/30)和100.00%(30/30),诊断符合率分别为78.38%(145/185)、70.27%(130/185)和51.89%(96/185),Kappa值分别为0.799、0.590和0.504,AUC(95%CI)值分别为0.867(0.693~0.941)、0.703(0.612~0.784)和0.623(0.529~0.717)。NGS检测脓液的阳性率[80.39%(41/51)]明显高于检测肉芽组织[62.92%(56/89)]和骨组织[44.44%(20/45)],差异均有统计学意义(χ²=4.560,P=0.031;χ²=13.335,P<0.001)。结论: NGS技术可明显提高骨关节结核患者病原学阳性检出率,具有较高的检测效能,并以脓液标本的检测诊断价值最高。

关键词: 结核,骨关节, 宏基因组二代测序, 诊断技术和方法, 对比研究

Abstract:

Objective: To evaluate the application value of next generation sequencing (NGS) technology in the diagnosis of osteoarticular tuberculosis. Methods: A retrospective study method was used to collect the clinical data of 185 patients with suspected osteoarticular tuberculosis admitted to the Department of Orthopedics of Hebei Provincial Chest Hospital from Dec 2019 to Dec 2022 with reference to the enrollment criteria. The patients were categorized into the tuberculosis group (osteoarticular tuberculosis, 155 cases) and the non-tuberculosis group (non-osteoarticular tuberculosis, 30 cases) according to the final clinical diagnosis. All patients’ lesion specimens were obtained during surgery or puncture (including 51 pus, 89 granulation tissue and 45 bone tissue specimens), and all of them were sent for NGS, BACTEC MGIT 960 mycobacterial liquid culture (culture), and rifampicin resistance real-time fluorescence quantitative nucleic acid amplification assay (GeneXpert MTB/RIF, Xpert). The final clinical diagnosis was used as the reference standard to compare the efficacy of the three methods in the diagnosis of osteoarticular tuberculosis, and to analyze the positivity of the three methods in the detection of pus, granulation tissue and bone tissue specimens. Results: Of the 185 patients with suspected bony joint tuberculosis, the positive rate of NGS for detecting osteoarticular tuberculosis (63.24%, 117/185) was significantly higher than that of Xpert (54.05%, 100/185) and culture (35.68%, 66/185), and the differences were statistically significant (χ²=24.982, P<0.001; χ²=37.934, P<0.001). Using the final clinical diagnosis as the reference standard, the sensitivity of NGS, Xpert and culture methods for the diagnosis of osteoarticular tuberculosis were 74.84% (116/155), 64.52% (100/155) and 42.58% (66/155), respectively, and the specificity were 96.67% (29/30), 100.00% (30/30) and 100.00% (30/30), the diagnostic compliance rates were 78.38% (145/185), 70.27% (130/185) and 51.89% (96/185), respectively, and the Kappa values were 0.799, 0.590 and 0.504, respectively, and the AUC (95%CI) values were 0.867 (0.693-0.941), 0.703 (0.612-0.784) and 0.623 (0.529-0.717), respectively. The positive rates of pus detected by NGS was 80.39% (41/51), respectively, which were significantly higher than that granulation (62.92% (56/89)) and bone tissue specimens (44.44% (20/45)), and the differences were all statistically significant (χ²=4.560, P=0.031; χ²=13.335, P<0.001). Conclusion: NGS technology can significantly increase the pathogenetically positive detection rate in patients with osteoarticular tuberculosis with high detection efficacy, and has the highest diagnostic value with pus specimens.

Key words: Tuberculosis, osteoarticular, Metagenomic next-generation sequencing, Diagnostic techniques and procedures, Comparative study

中图分类号:

R529.2
R8

姚晓伟, 刘树仁, 景艳色, 贾晨光. 二代测序技术在骨关节结核临床诊断中的应用价值[J]. 结核与肺部疾病杂志, 2024, 5(1): 37-43. doi: 10.19983/j.issn.2096-8493.20230094

Yao Xiaowei, Liu Shuren, Jing Yanse, Jia Chenguang. Evaluation of next generation sequencing for the diagnosis of osteoarticular tuberculosis infection[J]. Journal of Tuberculosis and Lung Disease, 2024, 5(1): 37-43. doi: 10.19983/j.issn.2096-8493.20230094

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临床资料	合计(185例)	结核组(155例)	非结核组(30例)	统计检验值	P值
年龄范围(岁)	12~70	12~66	13~65	-	-
年龄[岁,M(Q₁,Q₃)]	35(17,71)	31(13,67)	43(25,78)	Z=6.961	0.117
性别[例(构成比,%)]				χ²=4.398	0.036
男性	110(59.46)	87(56.13)	23(76.67)
女性	75(40.54)	68(43.87)	7(23.33)
发生部位[例(发生率,%)]				χ²=20.199	0.003
脊柱	135(72.97)	129(83.22)	6(20.00)	χ²=50.946	<0.001
膝关节	13(7.03)	7(4.52)	6(20.00)	χ²=9.224	0.002
肩关节	11(5.95)	5(3.23)	6(20.00)	χ²=12.646	0.000
髋关节	9(4.86)	4(2.58)	5(16.67)	χ²=10.776	0.001
肘关节	8(4.32)	5(3.23)	3(10.00)	χ²=2.788	0.095
踝关节	5(2.70)	2(1.29)	3(10.00)	χ²=7.251	0.007
腕关节	4(2.16)	3(1.94)	1(3.33)	χ²=0.232	0.630
合并基础疾病[例(发生率,%)]
糖尿病	13(7.03)	9(5.81)	4(13.33)	χ²=2.180	0.140
高血压	16(8.65)	12(7.74)	4(13.33)	χ²=0.995	0.319
冠心病	6(3.24)	4(2.58)	2(6.67)	χ²=1.337	0.248
风湿免疫性疾病	11(5.95)	9(5.81)	2(6.67)	χ²=0.033	0.855

标本	标本份数	NGS		Xpert		培养法		χ²值^c	P值	χ²值^d	P值
标本	标本份数	阳性	阴性	阳性	阴性	阳性	阴性	χ²值^c	P值	χ²值^d	P值
脓液	51	41(80.39)	10(19.61)	34(66.67)	17(33.33)	21(41.18)	30(58.82)	2.468	0.016	16.452	<0.001
肉芽组织	89	56(62.92)	33(37.08)	48(53.93)	41(46.07)	30(33.71)	59(66.29)	1.480	0.024	15.208	<0.001
骨组织	45	20(44.44)	25(55.56)	18(40.00)	27(60.00)	15(33.33)	30(66.67)	1.182	0.067	1.169	0.028
合计	185	117(63.24)	68(36.76)	100(54.05)	85(45.95)	66(35.68)	119(64.32)
χ²值^a		4.560		2.167		0.781
P值		0.031		0.041		0.077
χ²值^b		13.335		6.848		0.627
P值		<0.001		0.009		0.028