鸟枪法宏基因组测序分析作为下呼吸道感染患者的诊断策略

Shotgun Metagenomic Sequencing Analysis as a Diagnostic Strategy for Patients with Lower Respiratory Tract Infections.

作者信息

Cho Ha-Eun, Kim Min Jin, Choi Jongmun, Sohn Yong-Hak, Lee Jae Joon, Park Kyung Sun, Cho Sun Young, Park Ki-Ho, Kim Young Jin

机构信息

Department of Laboratory Medicine, Graduate School, Kyung Hee University, Kyung Hee University Hospital, Seoul 02447, Republic of Korea.

Department of Laboratory Medicine, Seegene Medical Foundation, Seoul 04805, Republic of Korea.

出版信息

Microorganisms. 2025 Jun 9;13(6):1338. doi: 10.3390/microorganisms13061338.

Abstract

Conventional diagnostic methods (CDMs) for lower respiratory infections (LRIs) have limitations in detecting causative pathogens. This study evaluates the utility of shotgun metagenomic sequencing (SMS) as a complementary diagnostic tool using bronchoalveolar lavage (BAL) fluid. Sixteen BAL fluid samples from pneumonia patients with positive CDM results-including bacterial/fungal cultures; PCR for or cytomegalovirus; and the BioFire FilmArray Pneumonia Panel (BioFire Diagnostics LLC, Salt Lake City, UT, USA)-underwent 10 Gb SMS on the Illumina NovaSeq 6000 platform (Illumina, San Diego, CA, USA). Reads were aligned to the NCBI RefSeq database; with fungal identification further supported by internal transcribed spacer (ITS) analysis. Antibiotic resistance genes (ARGs) were annotated using the Comprehensive Antibiotic Resistance Database. Microbial reads accounted for 0.00002-0.04971% per sample. SMS detected corresponding bacteria in 63% of cases, increasing to 69% when subdominant taxa were included. Fungal reads were low; however, species were identified in four samples via ITS. No viral reads were detected. ARGs meeting perfect match criteria were found in two cases. This is the first real-world study comparing SMS with CDMs, including semiquantitative PCR, in BAL fluid for LRI. SMS shows promise as a supplementary diagnostic method, with further research needed to optimize its performance and cost-effectiveness.

摘要

下呼吸道感染(LRI)的传统诊断方法(CDM)在检测致病病原体方面存在局限性。本研究评估了鸟枪法宏基因组测序(SMS)作为一种使用支气管肺泡灌洗(BAL)液的补充诊断工具的效用。对16份来自肺炎患者且CDM结果呈阳性的BAL液样本进行了研究,这些样本包括细菌/真菌培养;针对 或巨细胞病毒的PCR检测;以及BioFire FilmArray肺炎检测板(美国犹他州盐湖城的BioFire Diagnostics LLC公司),在Illumina NovaSeq 6000平台(美国加利福尼亚州圣地亚哥的Illumina公司)上对其进行了10 Gb的SMS分析。读取序列与NCBI RefSeq数据库进行比对;通过内部转录间隔区(ITS)分析进一步支持真菌鉴定。使用综合抗生素抗性数据库注释抗生素抗性基因(ARG)。每个样本中微生物读取序列占0.00002 - 0.04971%。SMS在63%的病例中检测到了相应细菌,若包括次要分类群,这一比例增至69%。真菌读取序列较少;然而,通过ITS在4个样本中鉴定出了 种真菌。未检测到病毒读取序列。在2例病例中发现了符合完全匹配标准的ARG。这是第一项在BAL液中对LRI将SMS与CDM(包括半定量PCR)进行比较的实际研究。SMS显示出作为一种补充诊断方法的前景,需要进一步研究以优化其性能和成本效益。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e53c/12196101/43e942b2f4b4/microorganisms-13-01338-g001.jpg

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