MALDI-TOF MS在结核分枝杆菌检测及耐药性分析中的应用价值

doi:10.20199/j.issn.1672-2302.2025.04.012

摘要/Abstract

摘要：

目的分析基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）技术在结核分枝杆菌鉴定及相关耐药基因检测中的临床应用价值，为结核病精准诊断提供技术支撑。方法收集2020年11月—2023年5月安徽省胸科医院结核病住院患者的临床资料，所有患者均接受涂片显微镜检查、分枝杆菌生长指示管（MGIT）960培养及药敏试验、Xpert MTB/RIF检测和MALDI-TOF MS检测。以MGIT 960培养及药敏试验结果作为金标准，评估MALDI-TOF MS在结核分枝杆菌菌种鉴定和耐药性分析中的检测效能。结果 MALDI-TOF MS、Xpert MTB/RIF、MGIT 960和涂片镜检的结核分枝杆菌检出率分别为94.70%（125/132）、84.09%（111/132）、67.42%（89/132）和29.55%（39/132），差异有统计学意义（Q=172.87，P<0.001）。MALDI-TOF MS、Xpert MTB/RIF和MGIT 960对利福平耐药的检测阳性率分别为53.03%（70/132）、45.45%（60/132）和44.70%（59/132），差异有统计学意义（Q=9.920，P=0.004）。MALDI-TOF MS对利福平耐药检测的一致性最好，为几乎完全一致（Kappa=0.879，P<0.001）；其次是对氟喹诺酮类药物和异烟肼，一致性为高度一致（Kappa=0.698、0.667，P均<0.001）；对氯法齐明检测的一致性较差，为中度一致（Kappa=0.476，P<0.001）。结论 MALDI-TOF MS技术作为结核病分子诊断及耐药筛查检测方法，具有灵敏度高、可靠性强的优点，值得专科医院进一步推广应用。

关键词: 结核分枝杆菌, 质谱分析法, 耐药, 灵敏度, 特异度

Abstract:

Objective To evaluate the clinical utility of the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) technology for identification of Mycobacterium tuberculosis and detection of associated drug-resistance genes, so as to provide technical suport for precision diagnosis of tuberculosis (TB). Methods Clinical data were collected from tuberculosis patients treated in Anhui Provincial Chest Hospital between November 2020 and May 2023. All patients underwent smear microscopy, mycobacterial growth indicator tube (MGIT) 960 culture and drug susceptibility testing (DST), Xpert MTB/RIF assay, and MALDI-TOF MS. MGIT 960 culture and DST results were used as the reference standard to evaluate the performance of MALDI-TOF MS for Mycobacterium tuberculosis identification and drug-resistance detection. Results The detection rate of Mycobacterium tuberculosis was 94.70% (125/132) by MALDI-TOF MS, 84.09% (111/132) by Xpert MTB/RIF, 67.42% (89/132) by MGIT 960, and 29.55% (39/132) by smear microscopy. The difference was significant (Q=172.87, P<0.001). The positive detection rate for rifampicin resistance was 53.03% (70/132) by MALDI-TOF MS, 45.45% (60/132) by Xpert MTB/RIF, and 44.70% (59/132) by MGIT 960 (Q=9.920, P=0.004). MALDI-TOF MS demonstrated the highest consistency for rifampicin resistance, which showed almost perfect agreement (Kappa=0.879, P<0.001), followed by high agreement with the results for fluoroquinolones and isoniazid (Kappa=0.698 and 0.667, both P<0.001). The consistency in clofazimine detection was relatively poor, showing moderate agreement (Kappa=0.476, P<0.001). Conclusion MALDI-TOF MS is worthy of recommendation in specialized hospitals, because this technique is highly sensitive and reliable for molecular diagnosis and drug-resistance screening of TB.

Key words: Mycobacterium tuberculosis, Mass spectrometry, Drug resistance, Sensitivity, Specificity

中图分类号:

R521

熊延军, 肖雨, 方雪晖, 黄鹏飞, 王华. MALDI-TOF MS在结核分枝杆菌检测及耐药性分析中的应用价值[J]. 热带病与寄生虫学, 2025, 23(4): 254-258.

XIONG Yanjun, XIAO Yu, FANG Xuehui, HUANG Pengfei, WANG Hua. Clinical value of MALDI-TOF MS for detection of Mycobacterium tuberculosis and drug resistance[J]. Journal of Tropical Diseases and Parasitology, 2025, 23(4): 254-258.

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药物	基因	位点	参考结果	突变结果
利福平	rpoB	511	T	CGA
		513	A/A/C	G/TC/TA
		516	G/A/C	TA/TCG/TG
		522	T/C/G	CAG/TC/TC
		526	C/A	TAG/TG
		531	T/C	C/T
		533	T	C
		572	A	T
异烟肼	katG	315	A/G/C	G/CAT/G
		316	G	A
	inhA	-15	C	T
氟喹诺酮类药物	gyrA/gyrB	80	G	AT
		90	C	T
		91	T	CG
		94	G/A	CTA/GC
		538	A	G
		543	C	T
		551	G	A
氯法齐明	Rv0678	193	G	AC
		466	C	T

抗结核药物	灵敏度（%）	特异度（%）	阳性预测值（%）	阴性预测值（%）	Kappa值（95%CI）	P值
利福平	91.53	100.00	100.00	85.71	0.879（0.777~0.981）	<0.001
异烟肼	85.71	81.82	88.89	77.14	0.667（0.508~0.826）	<0.001
氟喹诺酮类药物	77.27	92.54	77.27	92.54	0.698（0.530~0.866）	<0.001
氯法齐明	50.00	97.65	50.00	97.65	0.476（0.039~0.913）	<0.001

抗结核药物	MGIT 960药敏结果	MassARRAY法结果	耐药基因及相关位点突变	发生次数
异烟肼	敏感	耐药	inhA-15突变（C→T）	2
	敏感	耐药	katG 315突变（AGC→AAC）	1
	敏感	耐药	katG 315突变（AGC→ACC）	4
	耐药	敏感	未检测到突变	8
利福平	耐药	敏感	rpoB 513（CAA→CAG）同义突变Q513Q，与耐药无关	1
	耐药	敏感	rpoB 522（TCG→TC?）不确定	1
	耐药	敏感	rpoB 531（TCG→T?G）不确定	1
	耐药	敏感	rpoB 513（CAA→CAG）同义突变Q513Q，与耐药无关	2
	耐药	敏感	未检测到突变	3
氟喹诺酮类药物	敏感	耐药	gyrA 90（GCG→GTG）	2
	敏感	耐药	gyrA 94（GAC→GGC）	4
	耐药	敏感	gyrA 90（GCG→G?G）不确定	2
	耐药	敏感	未检测到突变	3
氯法齐明	敏感	耐药	Rv0678 193突变（G→DEL）	2
	耐药	敏感	未检测到突变	2