宏基因组学在传染病防控中的应用进展

doi:10.13560/j.cnki.biotech.bull.1985.2017-0787

摘要/Abstract

摘要： 新发突发传染病暴发给全球公共卫生防控带来严峻挑战。快速识别致病病原体是应对新发突发传染病的首要问题，传统病原检测方法难以应对已知变异较大病原或未知病原，基于高通量测序的宏基因组学研究给病原识别鉴定带来了新的方法和思路。核酸提取、高通量测序和数据分析等关键技术方法不断发展，使宏基因组学成为新突发传染病防控的重要研究方向。宏基因组学可对传染病防控中的多种类型样本进行直接测序，获得高通量的测序数据，并结合病原核酸数据库，通过序列比对、变异进化分析等生物信息学方法，通过监测可疑样本对疫情暴发进行预测预警；识别传染病患者感染致病病原，为临床诊治提供指导；构建病原系统发育关系，追溯疫情潜在感染来源，最终实现新突发传染病病原的快速识别、分型、耐药及溯源分析。宏基因组学作为一项新兴技术，在传染病防控领域具有巨大潜力和发展空间。通过对宏基因组学在传染病病原监测、检测及溯源等方面的应用进展进行综述，以期为传染病防控提供新的视角。

关键词: 宏基因组学, 传染病防控, 高通量测序

Abstract: The outbreak of new infectious diseases poses a serious challenge to global public health prevention and control.Rapid identification of pathogens is the primary challenge in dealing with outbreaks emerging infectious diseases. It is difficult to detect unknown pathogens or pathogens with large mutations using traditional pathogen detection methods. Metagenomics based on high-throughput sequencing has shed light on the pathogen detection and identification. Nucleic acid extraction，high-throughput sequencing，data analysis and other key technologies continue to develop，making metagenomics become an important research direction for the prevention and control of new infectious diseases. Metagenomics can direct sequence many samples of the prevention and control of infectious diseases to obtain high-throughput sequencing data. It can combine the pathogenic nucleic acid database，through sequence alignment，mutation evolution analysis and other bioinformatics methods in to use. In this way，we can monitor the suspicious samples of the epidemic outbreak forecast and warning，identify pathogens and provide medication guidance，determine the phylogenetic relationship，trace the origin of the pathogen and investigate the epidemic. Ultimately，we can realize the target of rapid identification，typing，drug resistance and traceability analysis of the new outbreak of infectious diseases. As a new technology，metagenomics has great potential and development prospects in the field of prevention and control of infectious diseases. This article reviews the application of metagenomics in the pathogen detection，detection and traceability of infectious diseases in order to provide a new perspective for the prevention and control of infectious diseases.

Key words: metagenomics, prevention and control of infectious diseases, high throughput sequencing

李沛翰, 李鹏, 宋宏彬. 宏基因组学在传染病防控中的应用进展[J]. 生物技术通报, 2018, 34(3): 43-52.

LI Pei-han, LI Peng, SONG Hong-bin. Application of Metagenomics in Prevention and Control of Infectious Diseases[J]. Biotechnology Bulletin, 2018, 34(3): 43-52.

参考文献

［1］ Fouchier RA, Kuiken T, Schutten M, et al. Aetiology：Koch’s postulates fulfilled for SARS virus［J］. Nature, 2003, 423（6937）：240.
［2］ Smith RD. Responding to global infectious disease outbreaks：lessons from SARS on the role of risk perception, communication and management［J］. Soc Sci Med, 2006, 63（12）：3113-3123.
［3］ Torsvik V, Ovreas L. Microbial diversity and function in soil：from genes to ecosystems［J］. Curr Opin Microbiol, 2002, 5（3）：240-245.
［4］ Roingeard P. Viral detection by electron microscopy：past, present and future［J］. Biol Cell, 2008, 100（8）：491-501.
［5］ Doane FW. Immunoelectron microscopy in diagnostic virology［J］. Ultrastruct Pathol, 1987, 11（5-6）：681-685.
［6］ Rose TM. CODEHOP-mediated PCR - a powerful technique for the identification and characterization of viral genomes［J］. Virol J, 2005, 2：20.
［7］ Ambrose HE, Granerod J, Clewley JP, et al. Diagnostic strategy used to establish etiologies of encephalitis in a prospective cohort of patients in england［J］. Journal of Clinical Microbiology, 2011, 49（10）：3576-3583.
［8］ Finkbeiner SR, Allred AF, Tarr PI, et al. Metagenomic analysis of human diarrhea：viral detection and discovery［J］. PLoS Pathog, 2008, 4（2）：e1000011.
［9］ Handelsman J, Rondon MR, Brady SF, et al. Molecular biological access to the chemistry of unknown soil microbes：a new frontier for natural products［J］. Chem Biol, 1998, 5（10）：R245-249.
［10］ Chen K, Pachter L. Bioinformatics for whole-genome shotgun sequencing of microbial communities［J］. PLoS Comput Biol, 2005, 1（2）：106-112.
［11］ Miller RR, Montoya V, Gardy JL, et al. Metagenomics for pathogen detection in public health［J］. Genome medicine, 2013, 5（9）：81.
［12］ Maukonen J, Simoes C, Saarela M. The currently used commercial DNA-extraction methods give different results of clostridial and actinobacterial populations derived from human fecal samples［J］. FEMS Microbiol Ecol, 2012, 79（3）：697-708.
［13］ Relman DA, Falkow S, Leboit PE, et al. The organism causing bacillary angiomatosis, peliosis hepatis, and fever and bacteremia in immunocompromised patients［J］. New England Journal of Medicine, 1991, 324（21）：1514.
［14］ Weisburg WG, Barns SM, Pelletier DA, et al. 16S ribosomal DNA amplification for phylogenetic study［J］. J Bacteriol, 1991, 173（2）：697-703.
［15］ Marchesi JR, Sato T, Weightman AJ, et al. Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16S rRNA［J］. Appl Environ Microbiol, 1998, 64（2）：795-799.
［16］ Klindworth A, Pruesse E, Schweer T, et al. Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies［J］. Nucleic Acids Res, 2013, 41（1）：e1.
［17］ Claesson MJ, O’sullivan O, Wang Q, et al. Comparative analysis of pyrosequencing and a phylogenetic microarray for exploring microbial community structures in the human distal intestine［J］. PLoS One, 2009, 4（8）：e6669.
［18］ Thurber RV, Haynes M, Breitbart M, et al. Laboratory procedures to generate viral metagenomes. ［J］. Nat Protoc, 2009, 4（4）：470-483.
［19］ Del Sal G, Manfioletti G, Schneider C. The CTAB-DNA precipitation method：a common mini-scale preparation of template DNA from phagemids, phages or plasmids suitable for sequencing. ［J］. Biotechniques, 1989, 7（5）：514-520.
［20］ Tsai Y-L, Olson BH. Rapid method for direct extraction of DNA from soil and sediments. ［J］. Appl Environ Microbiol, 1991, 57（4）：1070-1074.
［21］ Simms D, Cizdziel PE, Chomczynski P. TRIzol：A new reagent for optimal single-step isolation of RNA［J］. Focus, 1993, 15（4）：532-535.
［22］ Grant P, Sims C, Krieg-Schneider F, et al. Automated screening of blood donations for hepatitis C virus RNA using the Qiagen BioRobot 9604 and the Roche COBAS HCV Amplicor assay［J］. Vox Sanguinis, 2002, 82（4）：169-176.
［23］Jonasson J, Olofsson M, Monstein HJ. Classification, identification and subtyping of bacteria based on pyrosequencing and signature matching of 16S rDNA fragments［J］. Apmis, 2002, 110（3）：263-272.
［24］Salipante SJ, Kawashima T, Rosenthal C, et al. Performance comparison of Illumina and ion torrent next-generation sequencing platforms for 16S rRNA-based bacterial community profiling［J］. Appl Environ Microbiol, 2014, 80（24）：7583-7591.
［25］Mosher JJ, Bernberg EL, Shevchenko O, et al. Efficacy of a 3rd generation high-throughput sequencing platform for analyses of 16S rRNA genes from environmental samples［J］. J Microbiol Methods, 2013, 95（2）：175-181.
［26］Mosher JJ, Bowman B, Bernberg EL, et al. Improved performance of the PacBio SMRT technology for 16S rDNA sequencing［J］. J Microbiol Methods, 2014, 10：459-460.
［27］Aagaard K, Riehle K, Ma J, et al. A metagenomic approach to characterization of the vaginal microbiome signature in pregnancy［J］. PLoS One, 2012, 7（6）：e36466.
［28］Lasken RS, Mclean JS. Recent advances in genomic DNA sequencing of microbial species from single cells［J］. Nat Rev Genet, 2014, 15（9）：577-584.
［29］Berger B, Peng J, Singh M. Computational solutions for omics data［J］. Nat Rev Genet, 2013, 14（5）：333-346.
［30］张恩民, 海荣, 俞东征. 基因预测方法的研究进展［J］. 中国媒介生物学及控制杂志, 2009（3）：271-273.
［31］Caporaso JG, Kuczynski J, Stombaugh J, et al. QIIME allows analysis of high-throughput community sequencing data［J］. Nat Methods, 2010, 7（5）：335-336.
［32］Huson DH, Auch AF, Qi J, et al. MEGAN analysis of metagenomic data［J］. Genome Res, 2007, 17（3）：377-386.
［33］Markowitz VM, Ivanova NN, Szeto E, et al. IMG/M：a data management and analysis system for metagenomes［J］. Nucleic Acids Res, 2008, 36（Database issue）：D534-538.
［34］Giardine B, Riemer C, Hardison RC, et al. Galaxy：a platform for interactive large-scale genome analysis［J］. Genome Research, 2005, 15（10）：1451-1455.
［35］Meyer F, Paarmann D, D’souza M, et al. The metagenomics RAST server-a public resource for the automatic phylogenetic and functional analysis of metagenomes［J］. BMC Bioinformatics, 2008, 9（1）：386.
［36］Li W, Wooley JC, Godzik A. Probing metagenomics by rapid cluster analysis of very large datasets［J］. PLoS One, 2008, 3（10）：e3375.
［37］Fischer N, Indenbirken D, Meyer T, et al. Evaluation of unbiased next-generation sequencing of RNA（RNA-seq）as a diagnostic method in influenza virus-positive respiratory samples［J］. J Clin Microbiol, 2015, 53（7）：2238-2250.
［38］Greninger AL, Naccache SN, Federman S, et al. Rapid metagenomic identification of viral pathogens in clinical samples by real-time nanopore sequencing analysis［J］. Genome medicine, 2015, 7：99.
［39］Lysholm F, Wetterbom A, Lindau C, et al. Characterization of the viral microbiome in patients with severe lower respiratory tract infections, using metagenomic sequencing［J］. PLoS One, 2012, 7（2）：e30875.
［40］Temmam S, Davoust B, Chaber AL, et al. Screening for viral pathogens in African simian bushmeat seized at a French airport［J］. Transboundary and Emerging Diseases, 2016, 64（4）：1159-1167.
［41］Ng TF, Alavandi S, Varsani A, et al. Metagenomic identification of a nodavirus and a circular ssDNA virus in semi-purified viral nucleic acids from the hepatopancreas of healthy Farfantepenaeus duorarum shrimp［J］. Dis Aquat Organ, 2013, 105（3）：237-242.
［42］Sachsenroder J, Braun A, Machnowska P, et al. Metagenomic identification of novel enteric viruses in urban wild rats and genome characterization of a group A rotavirus［J］. J Gen Virol, 2014, 95（Pt 12）：2734-2747.
［43］Coffey LL, Page BL, Greninger AL, et al. Enhanced arbovirus surveillance with deep sequencing：Identification of novel rhabdoviruses and bunyaviruses in Australian mosquitoes［J］. Virology, 2014, 448（448）：146.
［44］Ksiazek TG, West CP, Rollin PE, et al. ELISA for the detection of antibodies to Ebola viruses［J］. J Infect Dis, 1999, 179（Suppl1）S192-S198.
［45］Jacobs M, Rodger A, Bell DJ, et al. Late Ebola virus relapse causing meningoencephalitis：a case report. ［J］. Lancet, 2016, 388（10043）：498-503.
［46］Loman NJ, Constantinidou C, Christner M, et al. A culture-independent sequence-based metagenomics approach to the investigation of an outbreak of Shiga-toxigenic Escherichia coli O104：H4［J］. JAMA, 2013, 309（14）：1502-1510.
［47］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［J］. PLoS Pathog, 2011, 7（11）：e1002369.
［48］Fischer N, Rohde H, Indenbirken D, et al. Rapid metagenomic diagnostics for suspected outbreak of severe pneumonia［J］. Emerg Infect Dis, 2014, 20（6）：1072-1075.
［49］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.
［50］Yao M, Zhou J, Zhu Y, et al. Detection of Listeria monocytogenes in CSF from three patients with meningoencephalitis by Next-Generation Sequencing［J］. J Clin Neurol, 2016, 12（4）：446-451.
［51］Ye M, Wei W, Yang Z, et al. Rapid diagnosis of Propionibacterium acnes infection in patient with hyperpyrexia after hematopoietic stem cell transplantation by next-generation sequencing：a case report［J］. BMC Infect Dis, 2016, 16：5.
［52］Palacios G, Druce J, Du L, et al. A new arenavirus in a cluster of fatal transplant-associated diseases［J］. N Engl J Med, 2008, 358（10）：991-998.
［53］Wilson MR, Zimmermann LL, Crawford ED, et al. Acute west nile virus meningoencephalitis diagnosed via metagenomic deep sequencing of cerebrospinal fluid in a renal transplant patient［J］. Am J Transplant, 2017, 17（3）：803-808.
［54］Holtz LR, Finkbeiner SR, Kirkwood CD, et al. Identification of a novel picornavirus related to cosaviruses in a child with acute diarrhea［J］. Virol J, 2008, 5：159.
［55］Mlakar J, Korva M, Tul N, et al. Zika virus associated with microcephaly［J］. N Engl J Med, 2016, 374（10）：951-958.
［56］Calvet G, Aguiar RS, Melo AS, et al. Detection and sequencing of Zika virus from amniotic fluid of fetuses with microcephaly in Brazil：a case study［J］. Lancet Infect Dis, 2016, 16（6）：653-660.
［57］Quick J, Grubaugh ND, Pullan ST, et al. Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples［J］. Nat Protoc, 2017, 12（6）：1261-1276.
［58］Guerbois M, Fernandez-Salas I, Azar SR, et al. Outbreak of zika virus infection, chiapas state, mexico, 2015, and first confirmed transmission by Aedes aegypti mosquitoes in the americas［J］. J Infect Dis, 2016, 214（9）：1349-1356.
［59］Snitkin ES, Zelazny AM, Thomas PJ, et al. Tracking a hospital outbreak of carbapenem-resistant Klebsiella pneumoniae with whole-genome sequencing［J］. Sci Transl Med, 2012, 4（148）：148ra116.
［60］Yu X, Jin T, Cui Y, et al. Influenza H7N9 and H9N2 viruses：coexistence in poultry linked to human H7N9 infection and genome characteristics［J］. J Virol, 2014, 88（6）：3423-3431.
［61］Mcmullan LK, Frace M, Sammons SA, et al. Using next generation sequencing to identify yellow fever virus in Uganda［J］. Virology, 2012, 422（1）：1-5.
［62］Keller A, Graefen A, Ball M, et al. New insights into the Tyrolean Iceman’s origin and phenotype as inferred by whole-genome sequencing［J］. Nature Communications, 2012, 3（698）：698.
［63］Schuenemann VJ, Singh P, Mendum TA, et al. Genome-wide comparison of medieval and modern Mycobacterium leprae［J］. Science, 2013, 341（6142）：179-183.
［64］Chan JZ, Sergeant MJ, Lee OY, et al. Metagenomic analysis of tuberculosis in a mummy［J］. N Engl J Med, 2013, 369（3）：289-290.
［65］Human Microbiome Project C. A framework for human microbiome research［J］. Nature, 2012, 486（7402）：215-221.
［66］Reyes A, Haynes M, Hanson N, et al. Viruses in the faecal microbiota of monozygotic twins and their mothers［J］. Nature, 2010, 466（7304）：334-338.
［67］Long Y, Zhang Y, Gong Y, et al. Diagnosis of sepsis with cell-free DNA by next-generation-sequencing technology in ICU patients［J］. Archives of Medical Research, 2016, 47（5）：365-371.
［68］ Callaway E. Microbiome privacy risk［J］. Nature, 2015, 521（7551）：136.
［69］Erlich Y, Narayanan A. Routes for breaching and protecting genetic privacy［J］. Nat Rev Genet, 2014, 15（6）：409-421.
［70］Goswami B, Singh B, Chawla R, et al. Turn around time（TAT）as a benchmark of laboratory performance［J］. Indian Journal of Clinical Biochemistry, 2010, 25（4）：376-379.

[1]	孙海航, 官会林, 王旭, 王童, 李泓霖, 彭文洁, 刘柏桢, 樊芳玲. 生物炭对三七连作土壤性质及真菌群落的影响[J]. 生物技术通报, 2023, 39(2): 221-231.
[2]	鲁兆祥, 王夕冉, 连新磊, 廖晓萍, 刘雅红, 孙坚. 基于功能宏基因组学挖掘抗生素耐药基因研究进展[J]. 生物技术通报, 2022, 38(9): 17-27.
[3]	王子夜, 王志刚, 阎爱华. 不同树龄桑根际土壤原生生物群落组成多样性[J]. 生物技术通报, 2022, 38(8): 206-215.
[4]	陈天赐, 武少兰, 杨国辉, 江丹霞, 江玉姬, 陈炳智. 无柄灵芝醇提物对小鼠睡眠及肠道菌群的影响[J]. 生物技术通报, 2022, 38(8): 225-232.
[5]	钟辉, 刘亚军, 王滨花, 和梦洁, 吴兰. 分析方法对细菌群落16S rRNA基因扩增测序分析结果的影响[J]. 生物技术通报, 2022, 38(6): 81-92.
[6]	赵林艳, 官会林, 向萍, 李泽诚, 柏雨龙, 宋洪川, 孙世中, 徐武美. 白及根腐病植株根际土壤微生物群落组成特征分析[J]. 生物技术通报, 2022, 38(2): 67-74.
[7]	陈宇捷, 郑华宝, 周昕彦. 改良高通量测序技术揭示除藻剂对藻类群落的影响[J]. 生物技术通报, 2022, 38(11): 70-79.
[8]	曹修凯, 王珊, 葛玲, 张卫博, 孙伟. 染色体外环形DNA研究进展及其在畜禽育种中的应用[J]. 生物技术通报, 2022, 38(1): 247-257.
[9]	毛婷, 牛永艳, 郑群, 杨涛, 穆永松, 祝英, 季彬, 王治业. 菌剂对苜蓿青贮发酵品质及微生物群落的影响[J]. 生物技术通报, 2021, 37(9): 86-94.
[10]	唐蝶, 周倩. 植物基因组组装技术研究进展[J]. 生物技术通报, 2021, 37(6): 1-12.
[11]	吕燕, 刘建利, 李靖宇, 候琳琳, 孙敏, 苟琪. 不同品种和产区宁夏枸杞根系AMF多样性[J]. 生物技术通报, 2021, 37(6): 36-48.
[12]	朱斌, 甘晨晨, 王洪程. 球花石斛（Dendrobium thyrsiflorum）叶绿体基因组特征及亲缘关系解析[J]. 生物技术通报, 2021, 37(5): 38-47.
[13]	张秫华, 方千, 贾红梅, 韩桂琪, 严铸云, 何冬梅. 川芎非根际、根际及根茎内生真菌群落差异分析[J]. 生物技术通报, 2021, 37(4): 56-69.
[14]	郭艳萍, 张浩, 赵新钢, 罗海玲, 张英俊. DNA宏条形码技术在食草动物食性研究中的应用[J]. 生物技术通报, 2021, 37(3): 252-260.
[15]	郑芳芳, 林俊生. 增殖诱导配体蛋白的核酸适配体筛选与特异性研究[J]. 生物技术通报, 2021, 37(10): 196-202.