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  • Analysis
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Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences

Nature Reviews Microbiology volume 12, pages 635–645 (2014)Cite this article

Subjects

Key Points

  • As the number of environmental small subunit (SSU) ribosomal RNA gene sequences has greatly surpassed the number of cultured microorganisms, reconciliation of the established taxonomy and classification of the uncultured microorganisms are crucial.

  • Rational taxonomic boundaries have been proposed for the high taxa (that is, genus and above) of the Bacteria and the Archaea on the basis of 16S rRNA gene sequence identities. These are : 94.5% for genus, 86.5% for family, 82.0% for order, 78.5% for class and 75.0% for phylum.

  • The application of these thresholds to the clustering of the SILVA database confirms that the current number of formally described taxa at any rank (for example, ∼30 phyla) is negligible compared with the total number of detected taxa (for example, ∼1,300 phyla).

  • In addition, the study of the annual rate of taxa discovery enables a new extrapolation of the total number of species (4 × 105) and high taxa on Earth (for example, 1 × 105 genera), which indicates that most common terrestrial and aquatic habitats will be exhaustively described within the next 5 years.

  • Taxon recovery tests that were carried out using partial 16S rRNA gene sequences show that short reads are not suitable for accurate richness estimations and accurate classifications of high taxa.

  • On the basis of the general taxonomic thresholds and phylogenetic considerations, we suggest a new biodiversity unit known as the candidate taxonomic unit (CTU), which is compatible with the hierarchy that was established in the Bacteriological Code. The ability to specify a taxonomic rank for particular clades is a major advance in understanding tree topologies and goes beyond the classic phylogenetic delineation.

  • The usefulness of CTUs has been intensively tested in the reclassification of the phylum Spirochaetes and the classification of 15 candidate divisions and environmental clades that are presented in this Analysis article, which also provide new insights into the coherence of classes, phyla and superphyla.

  • By providing explicit and well-documented guidelines, it is hoped that this work will facilitate the implementation of the many changes in the current taxonomy that are necessary to develop a common taxonomic classification of high taxa of bacteria and archaea on the basis of SSU rRNA gene sequences.

Abstract

Publicly available sequence databases of the small subunit ribosomal RNA gene, also known as 16S rRNA in bacteria and archaea, are growing rapidly, and the number of entries currently exceeds 4 million. However, a unified classification and nomenclature framework for all bacteria and archaea does not yet exist. In this Analysis article, we propose rational taxonomic boundaries for high taxa of bacteria and archaea on the basis of 16S rRNA gene sequence identities and suggest a rationale for the circumscription of uncultured taxa that is compatible with the taxonomy of cultured bacteria and archaea. Our analyses show that only nearly complete 16S rRNA sequences give accurate measures of taxonomic diversity. In addition, our analyses suggest that most of the 16S rRNA sequences of the high taxa will be discovered in environmental surveys by the end of the current decade.

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Figure 1: Variable regions of the 16S ribosomal RNA.
Figure 2: Predicting taxa richness using partial 16S ribosomal RNA sequences.
Figure 3: Classification of phylum Spirochaetes into candidate taxonomic units.
Figure 4: Phylogenetic reconstruction of the SAR11 group within the Proteobacteria based on 16S ribosomal RNA gene sequences.

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References

  1. Godfray, H. C. J. Challenges for taxonomy. Nature 417, 17–19 (2002).

    Article  CAS  PubMed  Google Scholar 

  2. Mora, C., Tittensor, D. P., Adl, S., Simpson, S. G. B. & Worm, B. How many species are on Earth and in the ocean. PLoS Biol. 9, e1001127 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Quast, C. et al. The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Res. 41, D590–D596 (2013). This paper reports the SILVA project, which is a comprehensive web resource (see Further information) for up-to-date, quality-controlled databases of aligned rRNA gene sequences from the Bacteria, the Archaea and the Eukarya.

    Article  CAS  PubMed  Google Scholar 

  4. Mole, B. Microbiome research goes without a home. Nature 500, 16–17 (2013).

    Article  CAS  PubMed  Google Scholar 

  5. Amann, R. I., Ludwig, W. & Schleifer, K. H. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59, 143–169 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Rosselló-Móra, R. Towards a taxonomy of Bacteria and Archaea based on interactive and cumulative data repositories. Environ. Microbiol. 14, 318–334 (2012).

    Article  PubMed  Google Scholar 

  7. Dykhuizen, D. E. Santa Rosalia revisited: why are there so many species of bacteria? Antonie Van Leeuwenhoek 73, 25–33 (1998).

    Article  CAS  PubMed  Google Scholar 

  8. Richter, M. & Rosselló-Móra, R. Shifting the genomic gold standard for the prokaryotic species definition. Proc. Natl Acad. Sci. USA 106, 19126–19131 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Stackebrandt, E. & Ebers, J. Taxonomic parameters revisited: tarnished gold standards. Microbiol. Today 8, 6–9 (2006).

    Google Scholar 

  10. Tindall, B. J., Rosselló-Móra, R., Busse, H.-J., Ludwig, W. & Kämpfer, P. Notes on the characterization of prokaryote strains for taxonomic purposes. Int. J. Syst. Evol. Microbiol. 60, 249–266 (2010).

    Article  CAS  PubMed  Google Scholar 

  11. Philippot, L. et al. The ecological coherence of high bacterial taxonomic ranks. Nature Rev. Microbiol. 8, 523–529 (2010). This study demonstrates that high bacterial taxa (that is, genus and above) are ecologically meaningful and their coherence is inversely correlated to their taxonomic rank. These observations provide a new perspective for the study of bacterial taxonomy, evolution and ecology.

    Article  CAS  Google Scholar 

  12. Gribaldo, S. & Brochier-Armanet, C. Time for order in microbial systematics. Trends Microbiol. 20, 209–210 (2012).

    Article  CAS  PubMed  Google Scholar 

  13. Garrity, G. M. & Oren, A. Response to Gribaldo and Brochier-Armanet: time for order in microbial systematics. Trends Microbiol. 20, 353–354 (2012). In this paper, the International Committee on Systematics of Prokaryotes (ICSP) supports a call for order in microbial systematics to address the lack of criteria to circumscribe high taxa, which represents a major problem in microbiology today.

    Article  CAS  PubMed  Google Scholar 

  14. Ereshefsky, M. Some problems with the Linnaean hierarchy. Phylos. Sci. 61, 186–205 (1994).

    Article  Google Scholar 

  15. Yarza, P. et al. The All-Species Living Tree Project: a 16S rRNA-based phylogenetic tree of all sequenced type strains. Syst. Appl. Microbiol. 31, 241–250 (2008). This paper reports the All-Species Living Tree Project (LTP), which is an initiative of Systematic and Applied Microbiology for the creation and maintenance of highly curated 16S rRNA and 23S rRNA gene sequence databases, alignments and phylogenetic trees for all the type strains of bacteria and archaea.

    Article  CAS  PubMed  Google Scholar 

  16. Cole, J. R., Konstantinidis, K., Farris, R. J. & Tiedje, J. M. in Environmental molecular microbiology (eds Liu, W.-T. & Jansson, J. K.) 1–19 (Caister Academic Press, 2010).

    Google Scholar 

  17. Ludwig, W., Klenk, H.-P. in Bergey's Manual of Systematic Bacteriology 2nd edn (eds Boone, D. R., Castenholz, R. W. & Garrity, G. M.) 49–65 (Springer, 2001).

    Book  Google Scholar 

  18. Ludwig, W. in Molecular Phylogeny of Microorganisms (eds Oren, A. & Papke, R. T.) 65–83 (Caister Academic Press, 2010). This chapter reports the classification of high ranks of the Bacteria and the Archaea, which is currently based on comparative analyses of rRNA and is supported by other markers and multigene approaches. The high information content and great availability in databases mostly justify the usage of rRNA gene sequences in taxonomy.

    Google Scholar 

  19. Fox, G. E., Pechman, K. R. & Woese, C. R. Comparative cataloging of 16S ribosomal ribonucleic acid: molecular approach to procaryotic systematics. Int. J. Syst. Bacteriol. 27, 44–57 (1977).

    Article  CAS  Google Scholar 

  20. Ludwig, W. & Schleifer, K. H. Bacterial phylogeny based on 16S and 23S rRNA sequence analysis. FEMS Microbiol. Rev. 15, 155–173 (1994).

    Article  CAS  PubMed  Google Scholar 

  21. Van de Peer, Y., Chapelle, S. & Wachter, R. D. A quantitative map of nucleotide substitution rates in bacterial rRNA. Nucleic Acids Res. 24, 3381–3391 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Fuchs, B. M. et al. Flow cytometric analysis of the in situ accessibility of Escherichia coli 16S rRNA for fluorescently labeled oligonucleotide probes. Appl. Environ. Microbiol. 64, 4973–4982 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Ludwig, W. et al. ARB: a software environment for sequence data. Nucleic Acids Res. 32, 1363–1371 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Yarza, P. et al. Update of the all-species living tree project based on 16S and 23S rRNA sequence analyses. Syst. Appl. Microbiol. 33, 291–299 (2010).

    Article  CAS  PubMed  Google Scholar 

  25. Shida, O., Takagi, H., Kadowaki, K. & Komagata, K. Proposal for two new genera, Brevibacillus gen. nov. and Aneurinibacillus gen. nov. Int. J. Syst. Bacteriol. 46, 939–946 (1996).

    Article  CAS  PubMed  Google Scholar 

  26. Kang, S.-J. et al. Brevundimonas naejangsanensis sp. nov., a proteolytic bacterium isolated from soil, and reclassification of Mycoplana bullata into the genus Brevundimonas as Brevundimonas bullata comb. nov. Int. J. Syst. Evol. Microbiol. 59, 3155–3160 (2009).

    Article  CAS  PubMed  Google Scholar 

  27. Keswani, J. & Whitman, W. B. Relationship of 16S rRNA sequence similarity to DNA hybridization in prokaryotes. Int. J. Syst. Evol. Microbiol. 51, 667–678 (2001).

    Article  CAS  PubMed  Google Scholar 

  28. Chakravorty, S., Helb, D., Burday, M., Connell, N. & Alland, D. A detailed analysis of 16S ribosomal RNA gene segments for the diagnosis of pathogenic bacteria. J. Microbiol. Methods 69, 330–339 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Mizrahi-Man, O., Davenport, E. R. & Gilad, Y. Taxonomic classification of bacterial 16S rRNA genes using short sequencing reads: evaluation of effective study designs. PLoS ONE 8, e53608 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Ashelford, K. E., Chuzhanova, N. A., Fry, J. C., Jones, A. J. & Weightman, A. J. At least 1 in 20 16S rRNA sequence records currently held in public repositories is estimated to contain substantial anomalies. Appl. Environ. Microbiol. 71, 7724–7736 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Saddler, G. S. International Committee on Systematics of Prokaryotes. Xth International (IUMS) Congress of Bacteriology and Applied Microbiology. Minutes of the meetings, 28 and 30 July 2002, Paris, France. Int. J. Syst. Evol. Microbiol. 55, 533–537 (2005).

    Article  Google Scholar 

  32. Ritalahti, K. M. et al. Sphaerochaeta globosa gen. nov., sp. nov. and Sphaerochaeta pleomorpha sp. nov., free-living, spherical spirochaetes. Int. J. Syst. Evol. Microbiol. 62, 210–216 (2012).

    Article  CAS  PubMed  Google Scholar 

  33. Meier-Kolthoff, J. P., Göker, M., Spröer, C. & Klenk, H. P. When should a DDH experiment be mandatory in microbial taxonomy? Arch. Microbiol. 195, 413–418 (2013).

    Article  CAS  PubMed  Google Scholar 

  34. Curtis, T. P., Sloan, W. T., & Scannell, J. W. Estimating prokaryotic diversity and its limits. Proc. Natl Acad. Sci. USA 99, 10494–10499 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Salman, V., Amann, R., Shub, D. A. & Schulz-Vogt, H. N. Multiple self-splicing introns in the 16S rRNA genes of giant sulfur bacteria. Proc. Natl Acad. Sci. USA 109, 4203–4208 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Klindworth, A. et al. Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies. Nucleic Acids Res. 41, e1 (2013).

    Article  CAS  PubMed  Google Scholar 

  37. Wang, T. et al. Structural segregation of gut microbiota between colorectal cancer patients and healthy volunteers. ISME J. 6, 320–329 (2012).

    Article  CAS  PubMed  Google Scholar 

  38. Guazzaroni, M.-E. et al. Metaproteogenomic insights beyond bacterial response to naphthalene exposure and bio-stimulation. ISME J. 7, 122–136 (2013).

    Article  CAS  PubMed  Google Scholar 

  39. Stackebrandt, E., Rainey, F. A. & Ward-Rainey, N. L. Proposal for a new hierarchic classification system, Actinobacteria classis nov. Int. J. Syst. Bacteriol. 47, 479–491 (1997).

    Article  Google Scholar 

  40. Lee, K. C. Y. et al. Phylogenetic delineation of the novel phylum Armatimonadetes (former candidate division OP10) and definition of two novel candidate divisions. Appl. Environ. Microbiol. 79, 2484–2487 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Cavalier-Smith, T. The neomuran origin of archaebacteria, the negibacterial root of the universal tree and bacterial megaclassification. Int. J. Syst. Evol. Microbiol. 52, 7–76 (2002).

    Article  CAS  PubMed  Google Scholar 

  42. Buchanan, R. E. Studies in the nomenclature and classification of the bacteria: II. The primary subdivisions of the Schizomycetes. J. Bacteriol. 2, 155–164 (1917).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Hovind-Hougen, K. Leptospiraceae, a new family to include Leptospira Noguchi 1917 and Leptonema gen. nov. Int. J. Syst. Bacteriol. 29, 245–251 (1979).

    Article  Google Scholar 

  44. Swellengrebel, N. H. Sur la cytologie comparée des spirochètes et des spirilles. Ann. Inst. Pasteur. 21, 562–586 (in French) (1907).

    Google Scholar 

  45. Gupta, R. S., Mahmood, S. & Adeolu, M. A phylogenomic and molecular signature based approach for characterization of the phylum Spirochaetes and its major clades: proposal for a taxonomic revision of the phylum. Front. Microbiol. 4, 217 (2013).

    PubMed  PubMed Central  Google Scholar 

  46. Harris, J. K., Kelley, S. T. & Pace, N. R. New perspective on uncultured bacterial phylogenetic division OP11. Appl. Environ. Microbiol. 70, 845–849 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Giebel, H.-A. et al. Distribution of Roseobacter RCA and SAR11 lineages in the North Sea and characteristics of an abundant RCA isolate. ISME J. 5, 8–19 (2011).

    Article  PubMed  Google Scholar 

  48. Wagner, M. & Horn, M. The Planctomycetes, Verrucomicrobia, Chlamydiae and sister phyla comprise a superphylum with biotechnological and medical relevance. Curr. Opin. Biotechnol. 17, 241–249 (2006).

    Article  CAS  PubMed  Google Scholar 

  49. Guy, L. & Ettema, T. J. G. The archaeal 'TACK' superphylum and the origin of eukaryotes. Trends Microbiol. 19, 580–587 (2011).

    Article  CAS  PubMed  Google Scholar 

  50. Hugenholtz, P., Goebel, B. M. & Pace, N. R. Impact of culture-independent studies on the emerging phylogenetic view of bacterial diversity. J. Bacteriol. 180, 4765–4774 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Yilmaz, P. et al. The SILVA and 'All-species Living Tree Project (LTP)' taxonomic frameworks. Nucl. Acids Res. 42, D643–D648 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  52. Geissinger, O., Herlemann, D. P. R., Mörschel, E., Maier, U. G. & Brune, A. The ultramicrobacterium 'Elusimicrobium minutum' gen. nov., sp. nov., the first cultivated representative of the Termite Group 1 phylum. Appl. Environ. Microbiol. 75, 2831–2840 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. R Core Team. R: A Language and Environment for Statistical Computing. (R Foundation for Statistical Computing, 2014) [online]

  54. Fu, L., Niu, B., Zhu, Z., Wu, S. & Li, W. CD-HIT: accelerated for clustering the next-generation sequencing data. Bioinformatics 28, 3150–3152 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Stamatakis, A. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics 22, 2688–2690 (2006).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This work has been co-funded by the Max Planck Society and the European Union (EU) project SYMBIOMICS (grant number 264774). R.R.M. acknowledges the scientific support given by the Spanish Ministry of Economy with the projects CE-CSD2007-0005 and CGL2012-39627-C03-03, which are both also supported with European Regional Development Fund (FEDER) funds, and the preparatory phase of Microbial Resource Research Infrastructure (MIRRI) funded by the EU (grant number 312251). W.B.W. acknowledges support of the Dimensions in Biodiversity program at the US National Science Foundation (NSF). P.Y. acknowledges support of the EU's Seventh Framework Program funds BioVeL, grant no. 283359.

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Authors and Affiliations

  1. Department of Ecology and Marine Resources, Marine Microbiology Group, Mediterranean Institute for Advanced Studies (Spanish National Research Council (CSIC)-University of the Balearic Islands (UIB)), Esporles, E-07190, Balearic Islands, Spain

    Pablo Yarza & Ramon Rosselló-Móra

  2. Max Planck Institute for Marine Microbiology, Celsiusstrasse 1, D-28359, Bremen, Germany

    Pablo Yarza, Pelin Yilmaz, Elmar Pruesse, Frank Oliver Glöckner & Rudolf Amann

  3. Ribocon GmbH, Fahrenheitstrasse 1, D-28359, Bremen, Germany

    Pablo Yarza

  4. Jacobs University Bremen, Campus Ring 1, D-28759, Bremen, Germany

    Frank Oliver Glöckner

  5. Lehrstuhl für Mikrobiologie, Technische Universität München, D-85350, Freising, Germany

    Wolfgang Ludwig & Karl-Heinz Schleifer

  6. Department of Microbiology, University of Georgia, 527 Biological Sciences Building, Athens, 30605–2605, Georgia, USA

    William B. Whitman

  7. Société de Bactériologie Systématique et Vétérinaire (SBSV) and École Nationale Vétérinaire de Toulouse (ENVT), F-31076, Toulouse cedex 03, France

    Jean Euzéby

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Supplementary information

Supplementary information S1 (table)

Intra-taxon sequence identity measures used to calculate the taxonomic thresholds. (XLS 114 kb)

Supplementary information S2 (Box)

Additional tables and figures. (PDF 776 kb)

Supplementary information S3 (figure)

Phylogenetic reconstruction of 15 candidate divisions and environmental clades (PDF 11624 kb)

Supplementary information S4 (table)

Sequence associated meta-data including CTU classification. Fields: acc,start,stop,tax_CTU,tax_xref_embl (XLS 1080 kb)

Glossary

Diversity

A term used to describe the effective number of taxa (that is, the richness) of a particular rank and their respective abundances (that is, the evenness).

SSU

(Small subunit). The small subunit of the ribosome, which is 16S ribosomal RNA for the Bacteria and the Archaea and 18S rRNA for the Eukarya.

Bacterial and archaeal species

A monophyletic group of organisms with a high degree of coherence in their genetic and phenotypic traits, which differentiate it from its close relatives.

High taxonomic ranks

The taxonomic categories of genus and above.

OTUs

(Operational taxonomic units). Groups of sequences that are meaningfully separated from other sequences by hierarchical clustering techniques (independent of phylogenetic inferences) and using strict sequence identity thresholds.

CTU

(Candidate taxonomic unit). A biological entity that is delineated by a monophyletic set of sequences with a sequence identity that stays within, or very close to, the taxonomic threshold that is proposed for a given rank.

OPU

(Operational phylogenetic unit). A group of sequences that appear as a monophyletic clade that is meaningfully separated from the remaining sequences in a genealogical reconstruction.

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Yarza, P., Yilmaz, P., Pruesse, E. et al. Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences. Nat Rev Microbiol 12, 635–645 (2014). https://doi.org/10.1038/nrmicro3330

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