Pathogen diagnostic value of metagenomic next-generation sequencing of bronchus alveolus lavage fluid in children with pulmonary infection

doi:10.3877/cma.j.issn.1673-5250.2022.05.010

Objective

To investigate the pathogen diagnostic value of metagenomic next-generation sequencing (mNGS) technology of bronchus alveolus lavage fluid (BALF) in children with pulmonary infections.

Methods

A total of 28 children with suspected pulmonary infections treated at Sun Yat-sen Memorial Hospital from February 2019 to May 2020 were enrolled in this study. According to diagnostic criteria of pulmonary infection used in this study, they were divided into observation group (n=22, children confirmed with pulmonary infection) and control group (n=6, children without pulmonary infection). The clinical data of all children were retrospectively analyzed. Pathogens of BALF in them were detected by mNGS and traditional pathogen detection methods. Rank sum test and chi-square test were used to statistically compare clinical data of children between two groups. Generalized additive model (GAM) was used to analyze the relationship between white blood cell count or serum C-reactive protein (CRP) levels and mNGS pathogen sequencing results of BALF of children. Receiver operating characteristic (ROC) curve was used to assess pathogenic diagnostic values of two methods for pulmonary infections in children. The study was approved by the Medical Ethics Committee of Sun Yat-sen Memorial Hospital (Approval No. SYSKY-2022-300-01). Informed consent forms were obtained by children′s guardians before receiving treatment.

Results

① There were no significant differences between two groups in age, proportion of male and immunocompromised children, body temperature, white blood cell count, absolute neutrophil count, serum CRP and PCT levels, positive rates by BALF mNGS and traditional pathogen testing methods, required-testing-time of BALF mNGS and traditional pathogen testing, BALF mNGS pathogen sequencing results (P>0.05). ② Among 28 children, pathogen positive rate of BALF detected by mNGS was 89.3%(25/28), which was higher than that of 17.9%(5/28) by traditional pathogen detection methods, and the difference was statistically significant (χ²=28.72, P<0.001). ③ GAM analysis results showed a negative linear relationship between white blood cell count and their mNGS pathogen sequencing results of BALF in 28 children (P<0.05). And when serum CRP level was<60 mg/L, the children′s serum CRP level increased with their mNGS pathogen sequencing results. ④ROC-AUC of BALF mNGS for diagnosis of pulmonary infection in children was 0.955 (95%CI: 0.893-1.000, P=0.180), which was higher than that of traditional pathogen detection methods of 0.508 (95%CI: 0.324-0.690, P=0.010), and the difference was statistically significant(Z=4.45, P<0.05).

Conclusions

BALF mNGS can provide a rapid and accurate pathogen diagnosis of pulmonary infections in children. White blood cell count and serum CRP levels in children can be used as indicators to assess the effectiveness of anti-infection treatment.

Key words: Bronchoalveolar lavage fluid, Metagenome, High-throughput nucleotide sequencing, Respiratory tract infections, Child

表1 2组疑似肺部感染患儿临床资料比较

组别	例数	年龄[岁，M(Q₁，Q₃)]	男性[例数(%)]	免疫功能缺陷[例数(%)]	体温[℃，M(Q₁，Q₃)]	白细胞计数[×10⁹/L，M(Q₁，Q₃)]	中性粒细胞绝对计数[×10⁹/L，M(Q₁，Q₃)]	血清CRP水平[mg/L，M(Q₁，Q₃)]
观察组	22	4.0(3.0，7.8)	9(40.9)	14(63.6)	37.9(35.4，39.6)	3.1(0.5，18.9)	1.9(0.0，14.6)	12.6(0.0，133.4)
对照组	6	3.0(1.2，4.8)	5(83.3)	5(83.3)	37.5(36.6，39.0)	6.0(2.2，17.2)	3.3(0.6，11.0)	7.0(3.0，65.0)
统计量		Z＝1.45	χ²＝3.39	χ²＝0.84	Z<0.01	Z＝0.91	Z＝0.52	Z＝0.74
P值		0.153	0.065	0.360	0.997	0.364	0.615	0.463
组别	例数	血清PCT水平[mg/L，M(Q₁，Q₃)]	BALF mNGS阳性率[例数(%)]	BALF传统病原体检测阳性率[例数(%)]	BALF mNGS检测报告所需时间[h，M(Q₁，Q₃)]	BALF传统病原体检测报告所需时间[h，M(Q₁，Q₃)]	BALF mNGS病原体测序读数[读数，M(Q₁，Q₃)]
观察组	22	0.2(0.0，8.4)	20(90.9)	4(18.2)	48.0(24.0，56.0)	75(72，150)	670.5(0.0，1 469 565.0)
对照组	6	0.1(0.0，0.8)	5(83.3)	1(16.7)	50.5(35.1，55.0)	80(75，168)	212.0(0.0，12 350.0)
统计量		Z＝0.72	χ²＝0.28	χ²＝0.01	Z<0.01	Z＝1.58	Z＝1.67
P值		0.447	0.594	0.932	0.925	0.126	0.107

表1 2组疑似肺部感染患儿临床资料比较

组别	例数	年龄[岁，M(Q₁，Q₃)]	男性[例数(%)]	免疫功能缺陷[例数(%)]	体温[℃，M(Q₁，Q₃)]	白细胞计数[×10⁹/L，M(Q₁，Q₃)]	中性粒细胞绝对计数[×10⁹/L，M(Q₁，Q₃)]	血清CRP水平[mg/L，M(Q₁，Q₃)]
观察组	22	4.0(3.0，7.8)	9(40.9)	14(63.6)	37.9(35.4，39.6)	3.1(0.5，18.9)	1.9(0.0，14.6)	12.6(0.0，133.4)
对照组	6	3.0(1.2，4.8)	5(83.3)	5(83.3)	37.5(36.6，39.0)	6.0(2.2，17.2)	3.3(0.6，11.0)	7.0(3.0，65.0)
统计量		Z＝1.45	χ²＝3.39	χ²＝0.84	Z<0.01	Z＝0.91	Z＝0.52	Z＝0.74
P值		0.153	0.065	0.360	0.997	0.364	0.615	0.463
组别	例数	血清PCT水平[mg/L，M(Q₁，Q₃)]	BALF mNGS阳性率[例数(%)]	BALF传统病原体检测阳性率[例数(%)]	BALF mNGS检测报告所需时间[h，M(Q₁，Q₃)]	BALF传统病原体检测报告所需时间[h，M(Q₁，Q₃)]	BALF mNGS病原体测序读数[读数，M(Q₁，Q₃)]
观察组	22	0.2(0.0，8.4)	20(90.9)	4(18.2)	48.0(24.0，56.0)	75(72，150)	670.5(0.0，1 469 565.0)
对照组	6	0.1(0.0，0.8)	5(83.3)	1(16.7)	50.5(35.1，55.0)	80(75，168)	212.0(0.0，12 350.0)
统计量		Z＝0.72	χ²＝0.28	χ²＝0.01	Z<0.01	Z＝1.58	Z＝1.67
P值		0.447	0.594	0.932	0.925	0.126	0.107

表2 对28例疑似肺部感染患儿BALF采取2种方法检出病原体构成比[例数(%)]

病原体		传统病原体检测	mNGS	病原体		传统病原体检测	mNGS
支原体				病毒
	肺炎支原体	-	5(20.0)		巨细胞病毒	-	3(12.0)
细菌					人类腺病毒7型	-	2(8.0)
	耶氏肺孢子菌	-	2(8.0)		人类疱疹病毒4型	-	1(4.0)
	铜绿假单胞菌	1(20.0)	-		人类细小病毒B19	-	1(4.0)
	副血链球菌	-	2(8.0)		EB病毒	-	1(4.0)
	鲍曼不动杆菌	-	2(8.0)		人类多瘤病毒3型	-	1(4.0)
	嗜酸乳球菌	-	1(4.0)	真菌
	肺炎克雷伯菌	-	1(4.0)		热带念珠菌	1(20.0)
	儿童链球菌	-	1(4.0)		曲霉菌	-	2(8.0)
	口腔链球菌	1(20.0)	-		白色念珠菌	1(20.0)
	人类葡萄球菌	1(20.0)	-	合计		5(100.0)	25(100.0)

表2 对28例疑似肺部感染患儿BALF采取2种方法检出病原体构成比[例数(%)]

病原体		传统病原体检测	mNGS	病原体		传统病原体检测	mNGS
支原体				病毒
	肺炎支原体	-	5(20.0)		巨细胞病毒	-	3(12.0)
细菌					人类腺病毒7型	-	2(8.0)
	耶氏肺孢子菌	-	2(8.0)		人类疱疹病毒4型	-	1(4.0)
	铜绿假单胞菌	1(20.0)	-		人类细小病毒B19	-	1(4.0)
	副血链球菌	-	2(8.0)		EB病毒	-	1(4.0)
	鲍曼不动杆菌	-	2(8.0)		人类多瘤病毒3型	-	1(4.0)
	嗜酸乳球菌	-	1(4.0)	真菌
	肺炎克雷伯菌	-	1(4.0)		热带念珠菌	1(20.0)
	儿童链球菌	-	1(4.0)		曲霉菌	-	2(8.0)
	口腔链球菌	1(20.0)	-		白色念珠菌	1(20.0)
	人类葡萄球菌	1(20.0)	-	合计		5(100.0)	25(100.0)

图2 对本组疑似肺部感染患儿BALF样本采取传统病原体检测和mNGS技术进行儿童肺部感染病原体诊断的ROC曲线比较

[1]	Bhuiyan MU, Blyth CC, West R, et al. Combination of clinical symptoms and blood biomarkers can improve discrimination between bacterial or viral community-acquired pneumonia in children[J]. BMC Pulm Med, 2019, 19(1): 71. DOI: 10.1186/s12890-019-0835-5.
[2]	Zhang S, Sammon PM, King I, et al. Cost of management of severe pneumonia in young children: systematic analysis[J]. J Glob Health, 2016, 6(1): 010408. DOI: 10.7189/jogh.06.010408.
[3]	Xie Y, Du J, Jin W, et al. Comparison the pathogen diagnosis of severe pneumonia by using next generation sequencing and traditional detection methods, China, 2010-2018[J]. J Infect, 2018, 78(2): S0163445318302779. DOI: 10.1016/j.jinf.2018.09.004.
[4]	中华医学会呼吸病学分会感染学组．中国成人医院获得性肺炎与呼吸机相关性肺炎诊断和治疗指南(2018年版)[J]. 中华结核和呼吸杂志，2018, 41(4): 255-280. DOI: 10.3760/cma.j.issn.1001-0939.2018.04.006.
[5]	宏基因组分析和诊断技术在急危重症感染应用专家共识组．宏基因组分析和诊断技术在急危重症感染应用的专家共识[J]. 中华急诊医学杂志，2019, 28(2): 151-155. DOI: 10.3760/cma.j.issn.1671-0282.2019.02.005.
[6]	Miao Q, Ma Y, Wang Q, et al. Microbiological diagnostic performance of metagenomic next-generation sequencing when applied to clinical practice[J]. Clin Infect Dis, 2018, 67(suppl_2): S231-S240. DOI: 10.1093/cid/ciy693.
[7]	Li Y, Sun B, Tang X, et al. Application of metagenomic next-generation sequencing for bronchoalveolar lavage diagnostics in critically ill patients[J]. Eur J Clin Microbiol Infect Dis, 2020, 39(2): 369-374. DOI: 10.1007/s10096-019-03734-5.
[8]	Huang J, Jiang E, Yang D, et al. Metagenomic next-generation sequencing versus traditional pathogen detection in the diagnosis of peripheral pulmonary infectious lesions[J]. Infect Drug Resist, 2020, 13: 567-576. DOI: 10.2147/IDR.S235182.
[9]	Guan H, Shen A, Lv X, et al. Detection of virus in CSF from the cases with meningoencephalitis by next-generation sequencing[J]. J Neurovirol, 2016, 22(2): 240-245. DOI: 10.1007/s13365-015-0390-7.
[10]	Patterson TF, Thompson GR 3rd, Denning DW, et al. Executive summary: practice guidelines for the diagnosis and management of aspergillosis: 2016 update by the Infectious Diseases Society of America[J]. Clin Infect Dis, 2016, 63(4): 433-442. DOI: 10.1093/cid/ciw444.
[11]	Chen X, Ding S, Lei C, et al. Blood and bronchoalveolar lavage fluid metagenomic next-generation sequencing in pneumonia[J]. Can J Infect Dis Med Microbiol, 2020, 2020: 6839103. DOI: 10.1155/2020/6839103.
[12]	Peng JM, Du B, Qin HY, et al. Metagenomic next-generation sequencing for the diagnosis of suspected pneumonia in immunocompromised patients[J]. J Infect, 2021, 82(4): 22-27. DOI: 10.1016/j.jinf.2021.01.029.
[13]	Chen J, Zhao Y, Shang Y, et al. The clinical significance of simultaneous detection of pathogens from bronchoalveolar lavage fluid and blood samples by metagenomic next-generation sequencing in patients with severe pneumonia[J]. J Med Microbiol, 2021, 70(1): 10.1099/jmm.0.001259. DOI: 10.1099/jmm.0.001259.
[14]	Fang X, Mei Q, Fan X, et al. Diagnostic value of metagenomic next-generation sequencing for the detection of pathogens in bronchoalveolar lavage fluid in ventilator-associated pneumonia patients[J]. Front Microbiol, 2020, 11: 599756. DOI: 10.3389/fmicb.2020.599756.
[15]	Wang H, Lu Z, Bao Y, et al. Clinical diagnostic application of metagenomic next-generation sequencing in children with severe nonresponding pneumonia[J]. PLoS One, 2020, 15(6): e0232610. DOI: 10.1371/journal.pone.0232610.
[16]	Shi W, Zhu S. The application of metagenomic next-generation sequencing in detection of pathogen in bronchoalveolar lavage fluid and sputum samples of patients with pulmonary infection[J]. Comput Math Methods Med, 2021, 2021: 7238495. DOI: 10.1155/2021/7238495.
[17]	Lin P, Chen Y, Su S, et al. Diagnostic value of metagenomic next-generation sequencing of bronchoalveolar lavage fluid for the diagnosis of suspected pneumonia in immunocompromised patients[J]. BMC Infect Dis, 2022, 22(1): 416. DOI: 10.1186/s12879-022-07381-8.
[18]	Shi Y, Peng JM, Qin HY, et al. Metagenomic next-generation sequencing: a promising tool for diagnosis and treatment of suspected pneumonia in rheumatic patients with acute respiratory failure: retrospective cohort study[J]. Front Cell Infect Microbiol, 2022, 12: 941930. DOI: 10.3389/fcimb.2022.941930.
[19]	Liu Y, Zhu H, Zheng Y. Detection of pneumocystis jirovecii pneumonia in infants with non-human immunodeficiency virus admitted to pediatric intensive care using metagenomics next-generation sequencing[J]. Infect Drug Resist, 2022, 15: 1889-1902. DOI: 10.2147/IDR.S358483.
[20]	Qu Y, Ding W, Liu S, et al. Metagenomic next-generation sequencing vs. traditional pathogen detection in the diagnosis of infection after allogeneic hematopoietic stem cell transplantation in children[J]. Front Microbiol, 2022, 13: 868160. DOI: 10.3389/fmicb.2022.868160.
[21]	Zhang D, Yang X, Wang J, et al. Application of metagenomic next-generation sequencing for bronchoalveolar lavage diagnostics in patients with lower respiratory tract infections[J]. J Int Med Res, 2022, 50(4): 3000605221089795. DOI: 10.1177/03000605221089795.

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