Single-cell metagenomics: challenges and applications
Yuan Xu, Fangqing Zhao
Single-cell metagenomics: challenges and applications
With the development of high throughput sequencing and single-cell genomics technologies, many uncultured bacterial communities have been dissected by combining these two techniques. Especially, by simultaneously leveraging of single-cell genomics and metagenomics, researchers can greatly improve the efficiency and accuracy of obtaining whole genome information from complex microbial communities, which not only allow us to identify microbes but also link function to species, identify subspecies variations, study host-virus interactions and etc. Here, we review recent developments and the challenges need to be addressed in single-cell metagenomics, including potential contamination, uneven sequence coverage, sequence chimera, genome assembly and annotation. With the development of sequencing and computational methods, single-cell metagenomics will undoubtedly broaden its application in various microbiome studies.
metagenomics / bioinformatics / single-cell genomics
[1] |
Albanese D, Donati C (2017) Strain profiling and epidemiology of bacterial species from metagenomic sequencing. Nat Commun 8:2260
CrossRef
Google scholar
|
[2] |
Avital G, Avraham R, Fan A, Hashimshony T, Hung DT, Yanai I (2017) scDual-Seq: mapping the gene regulatory program of Salmonella infection by host and pathogen single-cell RNAsequencing. Genome Biol 18:200
CrossRef
Google scholar
|
[3] |
Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD
CrossRef
Google scholar
|
[4] |
Becraft ED, Dodsworth JA, Murugapiran SK, Ohlsson JI, Briggs BR, Kanbar J, De Vlaminck I, Quake SR, Dong H, Hedlund BP
CrossRef
Google scholar
|
[5] |
Blainey PC (2013) The future is now: single-cell genomics of bacteria and archaea. FEMS Microbiol Rev 37:407–427
CrossRef
Google scholar
|
[6] |
Blanco L, Bernad A, Lazaro JM, Martin G, Garmendia C, Salas M (1989) Highly efficient DNA synthesis by the phage phi 29 DNA polymerase. Symmetrical mode of DNA replication. J Biol Chem 264:8935–8940
|
[7] |
Boisvert S, Raymond F, Godzaridis E, Laviolette F, Corbeil J (2012) Ray Meta: scalable de novo metagenome assembly and profiling. Genome Biol 13:R122
CrossRef
Google scholar
|
[8] |
Brown CT (2015) Strain recovery from metagenomes. Nat Biotechnol 33:1041–1043
CrossRef
Google scholar
|
[9] |
Chaisson MJ, Pevzner PA (2008) Short read fragment assembly of bacterial genomes. Genome Res 18:324–330
CrossRef
Google scholar
|
[10] |
Champlot S, Berthelot C, Pruvost M, Bennett EA, Grange T, Geigl EM (2010) An efficient multistrategy DNA decontamination procedure of PCR reagents for hypersensitive PCR applications. PLoS ONE 5:e13042
CrossRef
Google scholar
|
[11] |
Chen M, Song P, Zou D, Hu X, Zhao S, Gao S, Ling F (2014) Comparison of multiple displacement amplification (MDA) and multiple annealing and looping-based amplification cycles (MALBAC) in single-cell sequencing. PLoS ONE 9:e114520
CrossRef
Google scholar
|
[12] |
Chitsaz H, Yee-Greenbaum JL, Tesler G, Lombardo MJ, Dupont CL, Badger JH, Novotny M, Rusch DB, Fraser LJ, Gormley NA
CrossRef
Google scholar
|
[13] |
Cole JR, Wang Q, Fish JA, Chai B, McGarrell DM, Sun Y, Brown CT, Porras-Alfaro A, Kuske CR, Tiedje JM (2014) Ribosomal Database Project: data and tools for high throughput rRNA analysis. Nucleic Acids Res 42:D633–D642
CrossRef
Google scholar
|
[14] |
de la Cruz Peña MJ, Martinez-Hernandez F, Garcia-Heredia I, Lluesma Gomez M, Fornas Ò, Martinez-Garcia M (2018) Deciphering the human virome with single-virus genomics and metagenomics. Viruses 10:113
CrossRef
Google scholar
|
[15] |
De Smet J, Hendrix H, Blasdel BG, Danis-Wlodarczyk K, Lavigne R (2017) Pseudomonas predators: understanding and exploiting phage-host interactions. Nat Rev Microbiol 15:517–530
CrossRef
Google scholar
|
[16] |
Dean FB, Nelson JR, Giesler TL, Lasken RS (2001) Rapid amplification of plasmid and phage DNA using Phi 29 DNA polymerase and multiply-primed rolling circle amplification. Genome Res 11:1095–1099
CrossRef
Google scholar
|
[17] |
Delcher AL, Bratke KA, Powers EC, Salzberg SL (2007) Identifying bacterial genes and endosymbiont DNA with Glimmer. Bioinformatics 23:673–679
CrossRef
Google scholar
|
[18] |
Delcher AL, Harmon D, Kasif S, White O, Salzberg SL (1999) Improved microbial gene identification with GLIMMER. Nucleic Acids Res 27:4636–4641
CrossRef
Google scholar
|
[19] |
DeSantis TZ, Hugenholtz P, Larsen N, Rojas M, Brodie EL, Keller K, Huber T, Dalevi D, Hu P, Andersen GL (2006) Greengenes, a chimera-checked 16S rRNA gene database and workbench compatible with ARB. Appl Environ Microbiol 72:5069–5072
CrossRef
Google scholar
|
[20] |
Dodsworth JA, Blainey PC, Murugapiran SK, Swingley WD, Ross CA, Tringe SG, Chain PS, Scholz MB, Lo CC, Raymond J
CrossRef
Google scholar
|
[21] |
Dupont CL, Rusch DB, Yooseph S, Lombardo MJ, Richter RA, Valas R, Novotny M, Yee-Greenbaum J, Selengut JD, Haft DH
CrossRef
Google scholar
|
[22] |
Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R (2011) UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27:2194–2200
CrossRef
Google scholar
|
[23] |
Erkel C, Kube M, Reinhardt R, Liesack W (2006) Genome of Rice Cluster I archaea—the key methane producers in the rice rhizosphere. Science 313:370–372
CrossRef
Google scholar
|
[24] |
Garcia Martin H, Ivanova N, Kunin V, Warnecke F, Barry KW, McHardy AC, Yeates C, He S, Salamov AA, Szeto E
CrossRef
Google scholar
|
[25] |
Hasegawa M, Hashimoto T (1993) Ribosomal RNA trees misleading? Nature 361:23
CrossRef
Google scholar
|
[26] |
Hosono S, Faruqi AF, Dean FB, Du Y, Sun Z, Wu X, Du J, Kingsmore SF, Egholm M, Lasken RS (2003) Unbiased whole-genome amplification directly from clinical samples. Genome Res 13:954–964
CrossRef
Google scholar
|
[27] |
Huerta-Cepas J, Szklarczyk D, Forslund K, Cook H, Heller D, Walter MC, Rattei T, Mende DR, Sunagawa S, Kuhn M
CrossRef
Google scholar
|
[28] |
Ji P, Zhang Y, Wang J, Zhao F (2017) MetaSort untangles metagenome assembly by reducing microbial community complexity. Nat Commun 8:14306
CrossRef
Google scholar
|
[29] |
Kanehisa M, Araki M, Goto S, Hattori M, Hirakawa M, Itoh M, Katayama T, Kawashima S, Okuda S, Tokimatsu T
CrossRef
Google scholar
|
[30] |
Kang DD, Froula J, Egan R, Wang Z (2015) MetaBAT, an efficient tool for accurately reconstructing single genomes from complex microbial communities. PeerJ 3:e1165
CrossRef
Google scholar
|
[31] |
Kashtan N, Roggensack SE, Rodrigue S, Thompson JW, Biller SJ, Coe A, Ding H, Marttinen P, Malmstrom RR, Stocker R
CrossRef
Google scholar
|
[32] |
Koren S, Treangen TJ, Pop M (2011) Bambus 2: scaffolding metagenomes. Bioinformatics 27:2964–2971
CrossRef
Google scholar
|
[33] |
Kvist T, Ahring BK, Lasken RS, Westermann P (2007) Specific single-cell isolation and genomic amplification of uncultured microorganisms. Appl Microbiol Biotechnol 74:926–935
CrossRef
Google scholar
|
[34] |
Labonte JM, Swan BK, Poulos B, Luo H, Koren S, Hallam SJ, Sullivan MB, Woyke T, Wommack KE, Stepanauskas R (2015) Single-cell genomics-based analysis of virus-host interactions in marine surface bacterioplankton. ISME J 9:2386–2399
CrossRef
Google scholar
|
[35] |
Lasken RS, Stockwell TB (2007) Mechanism of chimera formation during the multiple displacement amplification reaction. BMC Biotechnol 7:19
CrossRef
Google scholar
|
[36] |
Lin H-H, Liao Y-C (2016) Accurate binning of metagenomic contigs via automated clustering sequences using information of genomic signatures and marker genes. Sci Reports 6:24175
CrossRef
Google scholar
|
[37] |
Liu J, Wang H, Yang H, Zhang Y, Wang J, Zhao F, Qi J (2013) Composition-based classification of short metagenomic sequences elucidates the landscapes of taxonomic and functional enrichment of microorganisms. Nucleic Acids Res 41:e3
CrossRef
Google scholar
|
[38] |
Marcy Y, Ishoey T, Lasken RS, Stockwell TB, Walenz BP, Halpern AL, Beeson KY, Goldberg SM, Quake SR (2007) Nanoliter reactors improve multiple displacement amplification of genomes from single cells. PLoS Genet 3:1702–1708
CrossRef
Google scholar
|
[39] |
Marshall IP, Blainey PC, Spormann AM, Quake SR (2012) A singlecell genome for Thiovulum sp. Appl Environ Microbiol 78:8555–8563
CrossRef
Google scholar
|
[40] |
Martinez-Garcia M, Santos F, Moreno-Paz M, Parro V, Anton J (2014) Unveiling viral-host interactions within the ‘microbial dark matter’. Nat Commun 5:4542
CrossRef
Google scholar
|
[41] |
Munson-McGee JH, Peng S, Dewerff S, Stepanauskas R, Whitaker RJ, Weitz JS, Young MJ (2018) A virus or more in (nearly) every cell: ubiquitous networks of virus–host interactions in extreme environments. ISME J. https://doi.org/10.1038/s41396-018-0071-7
CrossRef
Google scholar
|
[42] |
Namiki T, Hachiya T, Tanaka H, Sakakibara Y (2012) MetaVelvet: an extension of Velvet assembler to de novo metagenome assembly from short sequence reads. Nucleic Acids Res 40:e155
CrossRef
Google scholar
|
[43] |
Nobu MK, Narihiro T, Rinke C, Kamagata Y, Tringe SG, Woyke T, Liu WT (2015) Microbial dark matter ecogenomics reveals complex synergistic networks in a methanogenic bioreactor. ISME J 9:1710–1722
CrossRef
Google scholar
|
[44] |
Nurk S, Meleshko D, Korobeynikov A, Pevzner PA (2017) metaSPAdes: a new versatile metagenomic assembler. Genome Res 27:824–834
CrossRef
Google scholar
|
[45] |
Ochman H, Lawrence JG, Groisman EA (2000) Lateral gene transfer and the nature of bacterial innovation. Nature 405:299–304
CrossRef
Google scholar
|
[46] |
Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M
CrossRef
Google scholar
|
[47] |
Page AJ, Cummins CA, Hunt M, Wong VK, Reuter S, Holden MT, Fookes M, Falush D, Keane JA, Parkhill J (2015) Roary: rapid large-scale prokaryote pan genome analysis. Bioinformatics 31:3691–3693
CrossRef
Google scholar
|
[48] |
Peng Y, Leung HC, Yiu SM, Chin FY (2011) Meta-IDBA: a de Novo assembler for metagenomic data. Bioinformatics 27:i94–i101
CrossRef
Google scholar
|
[49] |
Peng Y, Leung HC, Yiu SM, Chin FY (2012) IDBA-UD: a de novo assembler for single-cell and metagenomic sequencing data with highly uneven depth. Bioinformatics 28:1420–1428
CrossRef
Google scholar
|
[50] |
Quast C, Pruesse E, Yilmaz P, Gerken J, Schweer T, Yarza P, Peplies J, Glockner FO (2013) The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Res 41:D590–D596
CrossRef
Google scholar
|
[51] |
Quince C, Delmont TO, Raguideau S, Alneberg J, Darling AE, Collins G, Eren AM (2017) DESMAN: a new tool for de novo extraction of strains from metagenomes. Genome Biol 18:181
CrossRef
Google scholar
|
[52] |
Raghunathan A, Ferguson HR Jr, Bornarth CJ, Song W, Driscoll M, Lasken RS (2005) Genomic DNA amplification from a single bacterium. Appl Environ Microbiol 71:3342–3347
CrossRef
Google scholar
|
[53] |
Rinke C, Lee J, Nath N, Goudeau D, Thompson B, Poulton N, Dmitrieff E, Malmstrom R, Stepanauskas R, Woyke T (2014) Obtaining genomes from uncultivated environmental microorganisms using FACS-based single-cell genomics. Nat Protoc 9:1038–1048
CrossRef
Google scholar
|
[54] |
Rodrigue S, Malmstrom RR, Berlin AM, Birren BW, Henn MR, Chisholm SW (2009) Whole genome amplification and de novo assembly of single bacterial cells. PLoS ONE 4:e6864
CrossRef
Google scholar
|
[55] |
Roux S, Hawley AK, Torres Beltran M, Scofield M, Schwientek P, Stepanauskas R, Woyke T, Hallam SJ, Sullivan MB (2014) Ecology and evolution of viruses infecting uncultivated SUP05 bacteria as revealed by single-cell- and meta-genomics. Elife 3: e03125
CrossRef
Google scholar
|
[56] |
Seemann T (2014) Prokka: rapid prokaryotic genome annotation. Bioinformatics 30:2068–2069
CrossRef
Google scholar
|
[57] |
Shaw K, Sesardic I, Bristol N, Ames C, Dagnall K, Ellis C, Whittaker F, Daniel B (2008) Comparison of the effects of sterilisation techniques on subsequent DNA profiling. Int J Legal Med 122:29–33
CrossRef
Google scholar
|
[58] |
Shi W, Ji P, Zhao F (2017) The combination of direct and paired link graphs can boost repetitive genome assembly. Nucleic Acids Res 45:e43
CrossRef
Google scholar
|
[59] |
Spencer SJ, Tamminen MV, Preheim SP, Guo MT, Briggs AW, Brito IL, Weitz A, PitkänenVigneault LK, Virta MP
CrossRef
Google scholar
|
[60] |
Stepanauskas R, Fergusson EA, Brown J, Poulton NJ, Tupper B, Labonte JM, Becraft ED, Brown JM, Pachiadaki MG, Povilaitis T
CrossRef
Google scholar
|
[61] |
Swan BK, Martinez-Garcia M, Preston CM, Sczyrba A, Woyke T, Lamy D, Reinthaler T, Poulton NJ, Masland ED, Gomez ML
CrossRef
Google scholar
|
[62] |
Szollosi GJ, Boussau B, Abby SS, Tannier E, Daubin V (2012) Phylogenetic modeling of lateral gene transfer reconstructs the pattern and relative timing of speciations. Proc Natl Acad Sci USA 109:17513–17518
CrossRef
Google scholar
|
[63] |
Truong DT, Tett A, Pasolli E, Huttenhower C, Segata N (2017) Microbial strain-level population structure and genetic diversity from metagenomes. Genome Res 27:626–638
CrossRef
Google scholar
|
[64] |
Wang J, Gao Y, Zhao F (2016) Phage-bacteria interaction network in human oral microbiome. Environ Microbiol 18:2143–2158
CrossRef
Google scholar
|
[65] |
Wang Y, Leung HC, Yiu SM, Chin FY (2012) MetaCluster 5.0: a tworound binning approach for metagenomic data for low-abundance species in a noisy sample. Bioinformatics 28:i356–i362
CrossRef
Google scholar
|
[66] |
Woese CR, Achenbach L, Rouviere P, Mandelco L (1991) Archaeal phylogeny: reexamination of the phylogenetic position of Archaeoglobus fulgidus in light of certain composition-induced artifacts. Syst Appl Microbiol 14:364–371
CrossRef
Google scholar
|
[67] |
Woyke T, Sczyrba A, Lee J, Rinke C, Tighe D, Clingenpeel S, Malmstrom R, Stepanauskas R, Cheng JF (2011) Decontamination of MDA reagents for single cell whole genome amplification. PLoS ONE 6:e26161
CrossRef
Google scholar
|
[68] |
Woyke T, Xie G, Copeland A, Gonzalez JM, Han C, Kiss H, Saw JH, Senin P, Yang C, Chatterji S
CrossRef
Google scholar
|
[69] |
Wright ES, Yilmaz LS, Noguera DR (2012) DECIPHER, a searchbased approach to chimera identification for 16S rRNA sequences. Appl Environ Microbiol 78:717–725
CrossRef
Google scholar
|
[70] |
Wu D, Hugenholtz P, Mavromatis K, Pukall R, Dalin E, Ivanova NN, Kunin V, Goodwin L, Wu M, Tindall BJ
CrossRef
Google scholar
|
[71] |
Wu Y-W, Tang Y-H, Tringe SG, Simmons BA, Singer SW (2014) MaxBin: an automated binning method to recover individual genomes from metagenomes using an expectation-maximization algorithm. Microbiome 2:26
CrossRef
Google scholar
|
[72] |
Yilmaz S, Singh AK (2012) Single cell genome sequencing. Curr Opin Biotechnol 23:437–443
CrossRef
Google scholar
|
[73] |
Yoon HS, Price DC, Stepanauskas R, Rajah VD, Sieracki ME, Wilson WH, Yang EC, Duffy S, Bhattacharya D (2011) Single-cell genomics reveals organismal interactions in uncultivated marine protists. Science 332:714–717
CrossRef
Google scholar
|
[74] |
Yu FB, Blainey PC, Schulz F, Woyke T, Horowitz MA, Quake SR (2017) Microfluidic-based mini-metagenomics enables discovery of novel microbial lineages from complex environmental samples. Elife. https://doi.org/10.7554/eLife.26580
CrossRef
Google scholar
|
[75] |
Zaneveld JR, Lozupone C, Gordon JI, Knight R (2010) Ribosomal RNA diversity predicts genome diversity in gut bacteria and their relatives. Nucleic Acids Res 38:3869–3879
CrossRef
Google scholar
|
[76] |
Zerbino DR, Birney E (2008) Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821–829
CrossRef
Google scholar
|
[77] |
Zhang K, Martiny AC, Reppas NB, Barry KW, Malek J, Chisholm SW, Church GM (2006) Sequencing genomes from single cells by polymerase cloning. Nat Biotechnol 24:680–686
CrossRef
Google scholar
|
[78] |
Zhang Y, Ji P, Wang J, Zhao F (2016) RiboFR-Seq: a novel approach to linking 16S rRNA amplicon profiles to metagenomes. Nucleic Acids Res 44:e99
CrossRef
Google scholar
|
/
〈 | 〉 |