Application and challenge of metagenomic next-generation sequencing in the diagnosis of pulmonary infection_West China Medical Journal

Authors：

ZHAO Zeyu ¹ ,  WANG Hui ²

1. Clinical Medical College, Graduate School of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan 610075, P. R. China;
2. Department of Respiratory and Critical Care Medicine, Chengdu First People’s Hospital, Chengdu, Sichuan 610041, P. R. China;

Corresponding author：

WANG Hui, Email: 13666257233@163.com

Keywords：

Metagenomic next-generation sequencing; pulmonary infection; pathogenic bacteria

DOI：

10.7507/1002-0179.202205127

Video：

Export PDF Favorites Scan Get Citation

Abstract Full text Figures/Tables Video References Cited by

The morbidity and mortality of pulmonary infection are high among infectious diseases worldwide. Rapid and accurate etiological diagnosis is the key to timely and effective treatment. Metagenomic next-generation sequencing (mNGS) technology has brokenthrough the limitations of traditional pathogenic microorganism detection methods and improved the detection rate of pathogens. In this paper, the application and advantages of mNGS technology in the diagnosis of bacteria, fungi, viruses and mixed infections in the lungs are analyzed, and the challenges and breakthroughs in RNA detection, wall breaking of firmicutes and host DNA clearance are described, in order to achieve targeted and accurate etiological diagnosis through mNGS, so as to effectively treat pulmonary infections.

Citation： ZHAO Zeyu, WANG Hui. Application and challenge of metagenomic next-generation sequencing in the diagnosis of pulmonary infection. West China Medical Journal, 2022, 37(8): 1231-1237. doi: 10.7507/1002-0179.202205127 Copy

1.	Xie G, Zhao B, Wang X, et al. Exploring the clinical utility of metagenomic next-generation sequencing in the diagnosis of pulmonary infection. Infect Dis Ther, 2021, 10(3): 1419-1435.
2.	Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A, 1977, 74(12): 5463-5467.
3.	Margulies M, Egholm M, Altman WE, et al. Genome sequencing in microfabricated high-density picolitre reactors. Nature, 2005, 437(7057): 376-380.
4.	Palacios G, Druce J, Du L, et al. A new arenavirus in a cluster of fatal transplant-associated diseases. N Engl J Med, 2008, 358(10): 991-998.
5.	Xu B, Liu L, Huang X, et al. Metagenomic analysis of fever, thrombocytopenia and leukopenia syndrome (FTLS) in Henan Province, China: discovery of a new bunyavirus. PLoS Pathog, 2011, 7(11): e1002369.
6.	Wilson MR, Naccache SN, Samayoa E, et al. Actionable diagnosis of neuroleptospirosis by next-generation sequencing. N Engl J Med, 2014, 370(25): 2408-2417.
7.	Miao Q, Ma Y, Wang Q, et al. Microbiological diagnostic performance of metagenomic next-generation sequencing when applied to clinical practice. Clin Infect Dis, 2018, 67(Suppl 2): S231-S240.
8.	Li H, Gao H, Meng H, et al. Detection of pulmonary infectious pathogens from lung biopsy tissues by metagenomic next-generation sequencing. Front Cell Infect Microbiol, 2018, 8: 205.
9.	郑一惠, 林威, 张天蕾, 等. 宏基因组二代测序在儿童重症感染中的应用价值. 中国当代儿科杂志, 2022, 24(3): 273-278.
10.	Liu H, Zhang Y, Yang J, et al. Application of mNGS in the etiological analysis of lower respiratory tract infections and the prediction of drug resistance. Microbiol Spectr, 2022, 10(1): e0250221.
11.	Qasem A, Shaw AM, Elkamel E, et al. Coronavirus disease 2019 (COVID-19) diagnostic tools: a focus on detection technologies and limitations. Curr Issues Mol Biol, 2021, 43(2): 728-748.
12.	van Boheemen S, van Rijn AL, Pappas N, et al. Retrospective validation of a metagenomic sequencing protocol for combined detection of RNA and DNA viruses using respiratory samples from pediatric patients. J Mol Diagn, 2020, 22(2): 196-207.
13.	Liu H, Zhang Y, Chen G, et al. Diagnostic significance of metagenomic next-generation sequencing for community-acquired pneumonia in southern China. Front Med (Lausanne), 2022, 9: 807174.
14.	胡继红, 高振祥. 应规范下呼吸道感染微生物检验的标本采集及报告方式. 中华医学杂志, 2015, 95(40): 3248-3250.
15.	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. BMC Infect Dis, 2022, 22(1): 416.
16.	Lei C, Zhou X, Ding S, et al. Case report: community-acquired Legionella gormanii pneumonia in an immunocompetent patient detected by metagenomic next-generation sequencing. Front Med (Lausanne), 2022, 9: 819425.
17.	Chen X, Ding S, Lei C, et al. Blood and bronchoalveolar lavage fluid metagenomic next-generation sequencing in pneumonia. Can J Infect Dis Med Microbiol, 2020, 2020: 6839103.
18.	Wang Q, Wu B, Yang D, et al. Optimal specimen type for accurate diagnosis of infectious peripheral pulmonary lesions by mNGS. BMC Pulm Med, 2020, 20(1): 268.
19.	Long Y, Zhang Y, Gong Y, et al. Diagnosis of sepsis with cell-free DNA by next-generation sequencing technology in ICU patients. Arch Med Res, 2016, 47(5): 365-371.
20.	Grumaz S, Grumaz C, Vainshtein Y, et al. Enhanced performance of next-generation sequencing diagnostics compared with standard of care microbiological diagnostics in patients suffering from septic shock. Crit Care Med, 2019, 47(5): e394-e402.
21.	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 Med Microbiol, 2021, 70(1): 001259.
22.	Qian YY, Wang HY, Zhou Y, et al. Improving pulmonary infection diagnosis with metagenomic next generation sequencing. Front Cell Infect Microbiol, 2021, 10: 567615.
23.	Ma N, Chen M, Ding J, et al. Recurrent pneumonia with tuberculosis and candida co-infection diagnosed by metagenomic next-generation sequencing: a case report and literature review. Front Med (Lausanne), 2022, 9: 755308.
24.	Shi CL, Han P, Tang PJ, et al. Clinical metagenomic sequencing for diagnosis of pulmonary tuberculosis. J Infect, 2020, 81(4): 567-574.
25.	Ullmann AJ, Aguado JM, Arikan-Akdagli S, et al. Diagnosis and management of aspergillus diseases: executive summary of the 2017 ESCMID-ECMM-ERS guideline. Clin Microbiol Infect, 2018, 24(Suppl 1): e1-e38.
26.	Yang L, Song J, Wang Y, et al. Metagenomic next-generation sequencing for pulmonary fungal infection diagnosis: lung biopsy versus bronchoalveolar lavage fluid. Infect Drug Resist, 2021, 14: 4333-4359.
27.	Li Z, Tang J, Zhu J, et al. The convoluted process of diagnosing pulmonary mycosis caused by Exophiala dermatitidis: a case report. BMC Infect Dis, 2022, 22(1): 433.
28.	Lu X, Zhang J, Ma W, et al. Pneumocystis Jirovecii pneumonia diagnosis via metagenomic next-generation sequencing. Front Med (Lausanne), 2022, 9: 812005.
29.	Sun H, Wang F, Zhang M, et al. Diagnostic value of bronchoalveolar lavage fluid metagenomic next-generation sequencing in Pneumocystis jirovecii pneumonia in non-HIV immunosuppressed patients. Front Cell Infect Microbiol, 2022, 12: 872813.
30.	Xu J, Yu Y, Lv J, et al. Application of metagenomic next-generation sequencing to diagnose Pneumocystis jirovecii pneumonia in kidney transplantation recipients. Ann Transplant, 2021, 26: e931059.
31.	Bai GH, Lin SC, Hsu YH, et al. The human virome: viral metagenomics, relations with human diseases, and therapeutic applications. Viruses, 2022, 14(2): 278.
32.	Shi L, Xia H, Moore MD, et al. Metagenomic next-generation sequencing in the diagnosis of HHV-1 reactivation in a critically ill COVID-19 patient: a case report. Front Med (Lausanne), 2021, 8: 715519.
33.	Wang J, Han Y, Feng J. Metagenomic next-generation sequencing for mixed pulmonary infection diagnosis. BMC Pulm Med, 2019, 19(1): 252.
34.	Chen P, Sun W, He Y. Comparison of the next-generation sequencing (NGS) technology with culture methods in the diagnosis of bacterial and fungal infections. J Thorac Dis, 2020, 12(9): 4924-4929.
35.	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. 中华急诊医学杂志, 2019, 28(2): 151-155.
36.	宏基因组学测序技术在中重症感染中的临床应用共识专家组, 中国研究型医院学会脓毒症与休克专业委员会, 中国微生物学会微生物毒素专业委员会, 等. 宏基因组学测序技术在中重症感染中的临床应用专家共识(第一版). 中华危重病急救医学, 2020, 32(5): 531-536.
37.	黄辉, 沈亦平, 顾卫红, 等. 临床基因检测报告规范与基因检测行业共识探讨. 中华医学遗传学杂志, 2018, 35(1): 1-8.
38.	中华医学会检验医学分会. 宏基因组测序病原微生物检测生物信息学分析规范化管理专家共识. 中华检验医学杂志, 2021, 44(9): 799-807.
39.	Langelier C, Kalantar KL, Moazed F, et al. Integrating host response and unbiased microbe detection for lower respiratory tract infection diagnosis in critically ill adults. Proc Natl Acad Sci U S A, 2018, 115(52): E12353-E12362.
40.	Deng X, Achari A, Federman S, et al. Metagenomic sequencing with spiked primer enrichment for viral diagnostics and genomic surveillance. Nat Microbiol, 2020, 5(3): 443-454.
41.	He Y, Fang K, Shi X, et al. Enhanced DNA and RNA pathogen detection via metagenomic sequencing in patients with pneumonia. J Transl Med, 2022, 20(1): 195.
42.	陆泓雨, 刘波, 秦超勇, 等. 基于模板转换的微量 RNA 测序建库方案探索. 生物技术通讯, 2018, 29(6): 796-801.
43.	Dupuy AK, David MS, Li L, et al. Redefining the human oral mycobiome with improved practices in amplicon-based taxonomy: discovery of malassezia as a prominent commensal. PLoS One, 2014, 9(3): e90899.
44.	Angebault C, Payen M, Woerther PL, et al. Combined bacterial and fungal targeted amplicon sequencing of respiratory samples: does the DNA extraction method matter?. PLoS One, 2020, 15(4): e0232215.
45.	Vesty A, Biswas K, Taylor MW, et al. Evaluating the impact of DNA extraction method on the representation of human oral bacterial and fungal communities. PLoS One, 2017, 12(1): e0169877.
46.	Huseyin CE, Rubio RC, O’Sullivan O, et al. The fungal frontier: a comparative analysis of methods used in the study of the human gut mycobiome. Front Microbiol, 2017, 8: 1432.
47.	Yang J, Yang F, Ren L, et al. Unbiased parallel detection of viral pathogens in clinical samples by use of a metagenomic approach. J Clin Microbiol, 2011, 49(10): 3463-3469.
48.	余汉忠, 牛璐璐, 孔倩倩, 等. 不同核酸提取方法和新型冠状病毒核酸检测试剂室间质评结果分析. 临床检验杂志, 2021, 39(2): 146-147.
49.	陈晓东, 陶志华, 周武. 核酸提取方法在聚合酶链反应测定乙肝病毒核酸中的评价. 中华检验医学杂志, 2002, 25(4): 212-214.
50.	林祖锐, 周耀武, 孙晓东, 等. 捕获和连接探针 PCR 与巢氏 PCR 检测疟原虫效果的比较. 中国病原生物学杂志, 2020, 15(6): 682-684, 707.
51.	Charalampous T, Kay GL, Richardson H, et al. Nanopore metagenomics enables rapid clinical diagnosis of bacterial lower respiratory infection. Nat Biotechnol, 2019, 37(7): 783-792.

1. Xie G, Zhao B, Wang X, et al. Exploring the clinical utility of metagenomic next-generation sequencing in the diagnosis of pulmonary infection. Infect Dis Ther, 2021, 10(3): 1419-1435.
2. Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A, 1977, 74(12): 5463-5467.
3. Margulies M, Egholm M, Altman WE, et al. Genome sequencing in microfabricated high-density picolitre reactors. Nature, 2005, 437(7057): 376-380.
4. Palacios G, Druce J, Du L, et al. A new arenavirus in a cluster of fatal transplant-associated diseases. N Engl J Med, 2008, 358(10): 991-998.
5. Xu B, Liu L, Huang X, et al. Metagenomic analysis of fever, thrombocytopenia and leukopenia syndrome (FTLS) in Henan Province, China: discovery of a new bunyavirus. PLoS Pathog, 2011, 7(11): e1002369.
6. Wilson MR, Naccache SN, Samayoa E, et al. Actionable diagnosis of neuroleptospirosis by next-generation sequencing. N Engl J Med, 2014, 370(25): 2408-2417.
7. Miao Q, Ma Y, Wang Q, et al. Microbiological diagnostic performance of metagenomic next-generation sequencing when applied to clinical practice. Clin Infect Dis, 2018, 67(Suppl 2): S231-S240.
8. Li H, Gao H, Meng H, et al. Detection of pulmonary infectious pathogens from lung biopsy tissues by metagenomic next-generation sequencing. Front Cell Infect Microbiol, 2018, 8: 205.
9. 郑一惠, 林威, 张天蕾, 等. 宏基因组二代测序在儿童重症感染中的应用价值. 中国当代儿科杂志, 2022, 24(3): 273-278.
10. Liu H, Zhang Y, Yang J, et al. Application of mNGS in the etiological analysis of lower respiratory tract infections and the prediction of drug resistance. Microbiol Spectr, 2022, 10(1): e0250221.
11. Qasem A, Shaw AM, Elkamel E, et al. Coronavirus disease 2019 (COVID-19) diagnostic tools: a focus on detection technologies and limitations. Curr Issues Mol Biol, 2021, 43(2): 728-748.
12. van Boheemen S, van Rijn AL, Pappas N, et al. Retrospective validation of a metagenomic sequencing protocol for combined detection of RNA and DNA viruses using respiratory samples from pediatric patients. J Mol Diagn, 2020, 22(2): 196-207.
13. Liu H, Zhang Y, Chen G, et al. Diagnostic significance of metagenomic next-generation sequencing for community-acquired pneumonia in southern China. Front Med (Lausanne), 2022, 9: 807174.
14. 胡继红, 高振祥. 应规范下呼吸道感染微生物检验的标本采集及报告方式. 中华医学杂志, 2015, 95(40): 3248-3250.
15. 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. BMC Infect Dis, 2022, 22(1): 416.
16. Lei C, Zhou X, Ding S, et al. Case report: community-acquired Legionella gormanii pneumonia in an immunocompetent patient detected by metagenomic next-generation sequencing. Front Med (Lausanne), 2022, 9: 819425.
17. Chen X, Ding S, Lei C, et al. Blood and bronchoalveolar lavage fluid metagenomic next-generation sequencing in pneumonia. Can J Infect Dis Med Microbiol, 2020, 2020: 6839103.
18. Wang Q, Wu B, Yang D, et al. Optimal specimen type for accurate diagnosis of infectious peripheral pulmonary lesions by mNGS. BMC Pulm Med, 2020, 20(1): 268.
19. Long Y, Zhang Y, Gong Y, et al. Diagnosis of sepsis with cell-free DNA by next-generation sequencing technology in ICU patients. Arch Med Res, 2016, 47(5): 365-371.
20. Grumaz S, Grumaz C, Vainshtein Y, et al. Enhanced performance of next-generation sequencing diagnostics compared with standard of care microbiological diagnostics in patients suffering from septic shock. Crit Care Med, 2019, 47(5): e394-e402.
21. 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 Med Microbiol, 2021, 70(1): 001259.
22. Qian YY, Wang HY, Zhou Y, et al. Improving pulmonary infection diagnosis with metagenomic next generation sequencing. Front Cell Infect Microbiol, 2021, 10: 567615.
23. Ma N, Chen M, Ding J, et al. Recurrent pneumonia with tuberculosis and candida co-infection diagnosed by metagenomic next-generation sequencing: a case report and literature review. Front Med (Lausanne), 2022, 9: 755308.
24. Shi CL, Han P, Tang PJ, et al. Clinical metagenomic sequencing for diagnosis of pulmonary tuberculosis. J Infect, 2020, 81(4): 567-574.
25. Ullmann AJ, Aguado JM, Arikan-Akdagli S, et al. Diagnosis and management of aspergillus diseases: executive summary of the 2017 ESCMID-ECMM-ERS guideline. Clin Microbiol Infect, 2018, 24(Suppl 1): e1-e38.
26. Yang L, Song J, Wang Y, et al. Metagenomic next-generation sequencing for pulmonary fungal infection diagnosis: lung biopsy versus bronchoalveolar lavage fluid. Infect Drug Resist, 2021, 14: 4333-4359.
27. Li Z, Tang J, Zhu J, et al. The convoluted process of diagnosing pulmonary mycosis caused by Exophiala dermatitidis: a case report. BMC Infect Dis, 2022, 22(1): 433.
28. Lu X, Zhang J, Ma W, et al. Pneumocystis Jirovecii pneumonia diagnosis via metagenomic next-generation sequencing. Front Med (Lausanne), 2022, 9: 812005.
29. Sun H, Wang F, Zhang M, et al. Diagnostic value of bronchoalveolar lavage fluid metagenomic next-generation sequencing in Pneumocystis jirovecii pneumonia in non-HIV immunosuppressed patients. Front Cell Infect Microbiol, 2022, 12: 872813.
30. Xu J, Yu Y, Lv J, et al. Application of metagenomic next-generation sequencing to diagnose Pneumocystis jirovecii pneumonia in kidney transplantation recipients. Ann Transplant, 2021, 26: e931059.
31. Bai GH, Lin SC, Hsu YH, et al. The human virome: viral metagenomics, relations with human diseases, and therapeutic applications. Viruses, 2022, 14(2): 278.
32. Shi L, Xia H, Moore MD, et al. Metagenomic next-generation sequencing in the diagnosis of HHV-1 reactivation in a critically ill COVID-19 patient: a case report. Front Med (Lausanne), 2021, 8: 715519.
33. Wang J, Han Y, Feng J. Metagenomic next-generation sequencing for mixed pulmonary infection diagnosis. BMC Pulm Med, 2019, 19(1): 252.
34. Chen P, Sun W, He Y. Comparison of the next-generation sequencing (NGS) technology with culture methods in the diagnosis of bacterial and fungal infections. J Thorac Dis, 2020, 12(9): 4924-4929.
35. 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. 中华急诊医学杂志, 2019, 28(2): 151-155.
36. 宏基因组学测序技术在中重症感染中的临床应用共识专家组, 中国研究型医院学会脓毒症与休克专业委员会, 中国微生物学会微生物毒素专业委员会, 等. 宏基因组学测序技术在中重症感染中的临床应用专家共识(第一版). 中华危重病急救医学, 2020, 32(5): 531-536.
37. 黄辉, 沈亦平, 顾卫红, 等. 临床基因检测报告规范与基因检测行业共识探讨. 中华医学遗传学杂志, 2018, 35(1): 1-8.
38. 中华医学会检验医学分会. 宏基因组测序病原微生物检测生物信息学分析规范化管理专家共识. 中华检验医学杂志, 2021, 44(9): 799-807.
39. Langelier C, Kalantar KL, Moazed F, et al. Integrating host response and unbiased microbe detection for lower respiratory tract infection diagnosis in critically ill adults. Proc Natl Acad Sci U S A, 2018, 115(52): E12353-E12362.
40. Deng X, Achari A, Federman S, et al. Metagenomic sequencing with spiked primer enrichment for viral diagnostics and genomic surveillance. Nat Microbiol, 2020, 5(3): 443-454.
41. He Y, Fang K, Shi X, et al. Enhanced DNA and RNA pathogen detection via metagenomic sequencing in patients with pneumonia. J Transl Med, 2022, 20(1): 195.
42. 陆泓雨, 刘波, 秦超勇, 等. 基于模板转换的微量 RNA 测序建库方案探索. 生物技术通讯, 2018, 29(6): 796-801.
43. Dupuy AK, David MS, Li L, et al. Redefining the human oral mycobiome with improved practices in amplicon-based taxonomy: discovery of malassezia as a prominent commensal. PLoS One, 2014, 9(3): e90899.
44. Angebault C, Payen M, Woerther PL, et al. Combined bacterial and fungal targeted amplicon sequencing of respiratory samples: does the DNA extraction method matter?. PLoS One, 2020, 15(4): e0232215.
45. Vesty A, Biswas K, Taylor MW, et al. Evaluating the impact of DNA extraction method on the representation of human oral bacterial and fungal communities. PLoS One, 2017, 12(1): e0169877.
46. Huseyin CE, Rubio RC, O’Sullivan O, et al. The fungal frontier: a comparative analysis of methods used in the study of the human gut mycobiome. Front Microbiol, 2017, 8: 1432.
47. Yang J, Yang F, Ren L, et al. Unbiased parallel detection of viral pathogens in clinical samples by use of a metagenomic approach. J Clin Microbiol, 2011, 49(10): 3463-3469.
48. 余汉忠, 牛璐璐, 孔倩倩, 等. 不同核酸提取方法和新型冠状病毒核酸检测试剂室间质评结果分析. 临床检验杂志, 2021, 39(2): 146-147.
49. 陈晓东, 陶志华, 周武. 核酸提取方法在聚合酶链反应测定乙肝病毒核酸中的评价. 中华检验医学杂志, 2002, 25(4): 212-214.
50. 林祖锐, 周耀武, 孙晓东, 等. 捕获和连接探针 PCR 与巢氏 PCR 检测疟原虫效果的比较. 中国病原生物学杂志, 2020, 15(6): 682-684, 707.
51. Charalampous T, Kay GL, Richardson H, et al. Nanopore metagenomics enables rapid clinical diagnosis of bacterial lower respiratory infection. Nat Biotechnol, 2019, 37(7): 783-792.

Previous Article
A meta-analysis of the incidence of resorption of lumbar disc herniation
Next Article
Research progress on detection technology of human immunodeficiency virus-1 p24 antigen

West China Medical Journal

Application and challenge of metagenomic next-generation sequencing in the diagnosis of pulmonary infection

Abstract Full text Figures/Tables Video References Cited by

Previous Article

Next Article

Format

Content