支气管肺泡灌洗液宏基因组二代测序对儿童肺部感染的病原体诊断价值

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

目的探讨支气管肺泡灌洗液(BALF)宏基因组二代测序(mNGS)技术,对儿童肺部感染的病原体诊断价值。方法选择2019年2月至2020年5月,于中山大学孙逸仙纪念医院诊疗的28例疑似肺部感染患儿为研究对象。根据本研究采用的肺部感染诊断标准,将患儿分为观察组(n=22,确诊为肺部感染)和对照组(n=6,非肺部感染)。采用回顾性分析方法对患儿临床病例资料进行分析。采用mNGS与传统病原体检测方法,对患儿BALF进行病原体检测。采用秩和检验与χ^(2)检验,对2组患儿临床资料进行统计学比较;采用广义相加模型(GAM),对白细胞计数、血清C反应蛋白(CRP)水平分别与患儿BALF中mNGS病原体测序读数的关系进行分析;采用受试者工作特征(ROC)曲线,评估2种方法对儿童肺部感染的病原体诊断价值。本研究通过中山大学孙逸仙纪念医院医学伦理委员会批准(审批文号:SYSKY-2022-300-01)。所有患儿接受治疗前,由监护人签署知情同意书。结果①2组患儿年龄、男性及免疫功能缺陷患儿比例、体温、白细胞计数、中性粒细胞绝对计数、血清CRP及PCT水平、BALF mNGS及传统病原体检测阳性率、BALF mNGS及传统病原体检测报告所需时间、BALF mNGS病原体测序读数比较,差异均无统计学意义(P>0.05)。②28例患儿中,mNGS检测患儿BALF的病原体阳性率为89.3%(25/28),高于传统病原体检测方法的17.9%(5/28),并且差异有统计学意义(χ^(2)=28.72,P<0.001)。③GAM分析显示,28例患儿的血常规白细胞计数与其BALF中mNGS病原体测序读数之间存在负线性关系(P<0.05);当血清CRP水平<60 mg/L时,患儿血清CRP水平随其BALF中mNGS病原体测序读数增加而增加。④BALF mNGS诊断儿童肺部感染的ROC-AUC为0.955(95%CI:0.893~1.000,P=0.180),高于传统病原体检测方法的0.508(95%CI:0.324~0.690,P=0.010),并且差异有统计学意义(Z=4.45,P<0.05)。结论BALF mNGS可为儿童肺部感染提供快速、准确的病原体诊断。患儿血常规白细胞计数和血清CRP水平,可作为评估抗感染效果的指标。

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.Results①There 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).②Among 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(χ^(2)=28.72,P<0.001).③GAM 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.