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一种使用16S核糖体RNA准确诊断肺炎病原体的系统算法。

A systematic algorithm using 16S ribosomal RNA for accurate diagnosis of pneumonia pathogens.

作者信息

Kurniawan Ferry Dwi, Alia Dina, Shiraishi Mamoru, Higo Megumi, Inoue Yoshiaki, Hagiwara Koichi

机构信息

Comprehensive Medicine I, Saitama Medical Center, Jichi Medical University, 1-847 Amanuma-cho, Omiya-ku, Saitama-shi, Saitama, 330-8503, Japan.

Department of Pulmonology and Respiratory Medicine, Faculty of Medicine, University of Syiah Kuala, Aceh, Indonesia.

出版信息

Sci Rep. 2025 Aug 10;15(1):29253. doi: 10.1038/s41598-025-14841-z.

DOI:10.1038/s41598-025-14841-z
PMID:40784906
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12336332/
Abstract

Over half of community-acquired pneumonia cases are caused by a few dozen bacterial species, and accurate identification of these pathogens is essential for effective treatment. In this study, we developed a reliable diagnostic method using 16S ribosomal RNA (16S rRNA) sequencing, considering intra-species variation, the need to differentiate Streptococcus pneumoniae from oral α-hemolytic streptococci, and applicability to the battlefield hypothesis, which helps distinguish true pathogens from commensal organisms that are not causative pathogens. We designed specific primers and a BLAST wrapper program, Cheryblast + ob, to classify 37 pneumonia-causing bacteria and 4 α-hemolytic streptococci. In simulation experiments involving a total of 20,309 copies of the 16S rRNA from 41 species of bacteria deposited in Genbank, the algorithm achieved a sensitivity greater than 0.996 and a specificity of 1.000. It was robust against sequencing errors and successfully distinguished S. pneumoniae from closely related species. In an experiment using next-generation sequencing on artificial mixtures containing genomic DNA from 10 bacterial species and human DNA at varying two-fold ratios, the species with the highest copy number was correctly identified in 8 out of 11 samples, and the top two species by copy number were identified in all 11 samples. This high-performance method offers a promising tool for accurate pneumonia diagnosis and could also be applied to other infections in which a limited number of bacterial species must be reliably identified.

摘要

超过一半的社区获得性肺炎病例是由几十种细菌引起的,准确识别这些病原体对于有效治疗至关重要。在本研究中,考虑到种内变异、区分肺炎链球菌与口腔α溶血性链球菌的需要以及适用于战场假说(该假说有助于区分真正的病原体与非致病共生生物),我们开发了一种使用16S核糖体RNA(16S rRNA)测序的可靠诊断方法。我们设计了特异性引物和一个BLAST包装程序Cheryblast + ob,以对37种引起肺炎的细菌和4种α溶血性链球菌进行分类。在涉及Genbank中41种细菌的16S rRNA共20309个拷贝的模拟实验中,该算法的灵敏度大于0.996,特异性为1.000。它对测序错误具有鲁棒性,并成功地将肺炎链球菌与密切相关的物种区分开来。在一项对含有10种细菌基因组DNA和人类DNA且比例呈两倍变化的人工混合物进行下一代测序的实验中,11个样本中有8个正确识别出拷贝数最高的物种,所有11个样本中都识别出了拷贝数排名前两位的物种。这种高性能方法为准确的肺炎诊断提供了一种有前景的工具,也可应用于必须可靠识别有限数量细菌物种的其他感染。

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本文引用的文献

1
Etiology of community-acquired pneumonia in adults: a systematic review.成人社区获得性肺炎的病因:一项系统评价
Pneumonia (Nathan). 2020 Oct 5;12:11. doi: 10.1186/s41479-020-00074-3. eCollection 2020.
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HIRA-TAN detects pathogens of pneumonia with a progressive course despite antibiotic treatment.HIRA-TAN可检测出即便经过抗生素治疗但病情仍呈进展性的肺炎病原体。
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Evolving Understanding of the Causes of Pneumonia in Adults, With Special Attention to the Role of Pneumococcus.
成人肺炎病因的演变认识,特别关注肺炎球菌的作用。
Clin Infect Dis. 2017 Oct 30;65(10):1736-1744. doi: 10.1093/cid/cix549.
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PCR-Based Rapid Identification System Using Bridged Nucleic Acids for Detection of Clarithromycin-Resistant Mycobacterium avium-M. intracellulare Complex Isolates.基于聚合酶链式反应的快速鉴定系统,使用桥接核酸检测克拉霉素耐药鸟分枝杆菌-胞内分枝杆菌复合群分离株
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Insight into biases and sequencing errors for amplicon sequencing with the Illumina MiSeq platform.深入了解Illumina MiSeq平台进行扩增子测序时的偏差和测序错误。
Nucleic Acids Res. 2015 Mar 31;43(6):e37. doi: 10.1093/nar/gku1341. Epub 2015 Jan 13.
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HIRA-TAN: a real-time PCR-based system for the rapid identification of causative agents in pneumonia.HIRA-TAN:一种基于实时PCR的系统,用于快速鉴定肺炎的病原体。
Respir Med. 2014 Feb;108(2):395-404. doi: 10.1016/j.rmed.2013.11.018. Epub 2013 Dec 10.
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Prediction of the pathogens that are the cause of pneumonia by the battlefield hypothesis.根据战场假说预测导致肺炎的病原体。
PLoS One. 2011;6(9):e24474. doi: 10.1371/journal.pone.0024474. Epub 2011 Sep 1.
8
Guidelines for the management of adults with hospital-acquired, ventilator-associated, and healthcare-associated pneumonia.成人医院获得性肺炎、呼吸机相关性肺炎和医疗保健相关性肺炎管理指南。
Am J Respir Crit Care Med. 2005 Feb 15;171(4):388-416. doi: 10.1164/rccm.200405-644ST.
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Community-acquired pneumonia.社区获得性肺炎
Lancet. 2003 Dec 13;362(9400):1991-2001. doi: 10.1016/S0140-6736(03)15021-0.