Advertisement

Microbial Ecology

, Volume 52, Issue 3, pp 408–417 | Cite as

Testing for Differentiation of Microbial Communities Using Phylogenetic Methods: Accounting for Uncertainty of Phylogenetic Inference and Character State Mapping

  • Ryan T. Jones
  • Andrew P. Martin
Article

Abstract

Comparative analyses of microbial communities increasingly involve the assay of 16S rRNA (or other gene) sequences from environmental DNA. Determining whether the composition of two or more communities differ in their phylogenetic composition involves testing for covariation between phylogeny and community type. This approach requires estimating the phylogenetic relationships among all sampled sequences and assessing whether the distribution of sequences among communities differs from the null expectation that sequences are randomly distributed. One method developed for implementing the phylogeny-based test of differentiation, referred to as the Phylogenetic test, relies on a single estimate of the phylogeny. However, for most data sets, many alternative phylogenetic trees provide statistically equivalent descriptions of the data. Because the actual phylogeny is unknown, phylogenetic tests of differentiation among microbial communities must account for phylogenetic uncertainty. In this article, we evaluate bootstrapping and Bayesian phylogenetic methods when implementing the Phylogenetic test using parsimony to map character states, and we investigate the effects of character mapping uncertainty by using a Bayesian approach to stochastically map character states on trees. Our approaches incorporate uncertainty into the tests of two closely related null hypotheses: (1) populations are panmictic, and (2) identical communites existed in both environments over the course of evolutionary history. We use two data sets previously implemented in tests for community differentiation: nitrite reductase genes sampled from marsh and upland soils and 16S rDNA sequences sampled from the human mouth and gut. We show that accounting for phylogenetic and mapping uncertainties can drastically affect results when implementing the Phylogenetic test. Accounting for phylogenetic and character mapping uncertainty provides a more conservative and robust test of covariation between phylogeny and environment when comparing microbial communities using DNA sequences.

Keywords

Microbial Community Character State Null Distribution Terminal Restriction Fragment Length Polymorphism Posterior Probability Distribution 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

This work was supported by an NSF Microbial Observatory grant to S. Schmidt and A. Martin. Thanks are due to Steve Schmidt, Sasha Reed, and Elizabeth Costello for editorial comments. We thank J. Bollback for help with SIMMAP. We would also like to thank an anonymous reviewer who helped us immensely with the direction of this article.

References

  1. 1.
    Acinas, SG, Klepac-Ceraj, V, Hunt, DE, Pharino, C, Ceraj, I, Distel, MF, Polz, MF (2004) Fine-scale phylogenetic arachitecture of a complex bacterial community. Nature 430(6999): 551–554PubMedCrossRefGoogle Scholar
  2. 2.
    Alfaro, ME, Zoller, S, Lutzoni, F (2003) Bayes or bootstrap? A simulation study comparing the performance of Bayesian Markov chain Monte Carlo sampling and bootstrapping in assessing phylogenetic confidence. Mol Biol Evol 20: 255–266PubMedCrossRefGoogle Scholar
  3. 3.
    Brauer, MJ, Holder, MT, Dries, LA, Zwickl, DJ, Lewis, PO, Hillis, DM (2002) Genetic algorithms and parallel processing in maximum-likelihood phylogeny inference. Mol Biol Evol 19(10): 1717–1726PubMedGoogle Scholar
  4. 4.
    Cummings, MP, Handley, SA, Myers, DS, Reed, DL, Rokas, A, Winka, K (2003) Comparing bootstrap and posterior probability values in the four-taxon case. Syst Biol 52: 477–487PubMedCrossRefGoogle Scholar
  5. 5.
    Donachie, SP, Hou, S, Lee, KS, Riley, CW, Pikina, A, Belisle, C, Kempe, S, Gregory, TS, Bossuyt, A, Boerema, J, Liu, J, Frietas, TA, Malahoff, A, Alam, M (2004) The Hawaiian Archipelago: a microbial diversity hotspot. Microb Ecol 48: 509–520PubMedCrossRefGoogle Scholar
  6. 6.
    Eckburg, PB, Bik, EM, Bernstein, CN, Purdom, E, Dethlefsen, L, Sargent, M, Gill, SR, Nelson, KE, Relman, DA (2005) Diversity of the human intestinal microbial flora. Science 308: 1635–1638PubMedCrossRefGoogle Scholar
  7. 7.
    Felsenstein, J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39: 783–791CrossRefGoogle Scholar
  8. 8.
    Giribet, G, Edgecombe, GD, Wheeler, WC (2001) Arthropod phylogeny based on eight molecular loci and morphology. Nature 413(6852): 157–161PubMedCrossRefGoogle Scholar
  9. 9.
    Hackl, E, Zechmeister-Boltenstern, S, Bodrossy, L, Sessitsch, A (2004) Comparison of diversities and compositions of bacterial populations inhabiting natural forest soils. Appl Environ Microbiol 70: 5057–5065PubMedCrossRefGoogle Scholar
  10. 10.
    Harvey, PH, Pagel, MD (1991) The Comparative Method in Evolutionary Biology. Oxford University Press. Oxford, UKGoogle Scholar
  11. 11.
    Heijs, SK, Damste, JSS, Forney, LJ (2005) Characterization of a deep-sea microbial mat from an active cold seep at the Milano mud volcano in the Eastern Mediterranean Sea. FEMS Microb Ecol 54: 47–56CrossRefGoogle Scholar
  12. 12.
    Huelsenbeck, JP, Ronquist, FR (2001) MRBAYES: Bayesian inference of phylogeny. Biometrics 17: 754–756Google Scholar
  13. 13.
    Huelsenbeck, JP, Rannala, B, Masly, JP (2000) Accommodating phylogenetic uncertainty in evolutionary studies. Science 288: 2349–2350PubMedCrossRefGoogle Scholar
  14. 14.
    Huelsenbeck, JP, Ronquist, F, Nielsen, R, Bollback, JP (2001) Bayesian inference of phylogeny and its impact on evolutionary biology. Science 2944: 2310–2314CrossRefGoogle Scholar
  15. 15.
    Huelsenbeck, JP, Nielsen, R, Bollback, JP (2003) Stochastic mapping of morphological characters. Syst Biol 52(2): 131–158PubMedCrossRefGoogle Scholar
  16. 16.
    Hughes, JB, Hellmann, JJ, Ricketts, TH, Bohannan, BJ (2001) Counting the uncountable: statistical approaches to estimating microbial diversity. Appl Environ Microbiol 67: 4399–4406PubMedCrossRefGoogle Scholar
  17. 17.
    Keane, TM, Naughton, TJ, Travers, SA, McInerney, JO, McCormack, GP (2004) DPRml: distributed phylogeny reconstruction by maximum likelihood. Bioinformatics. 2004 Oct. 28Google Scholar
  18. 18.
    Kroes, R, Lepp, PW, Relman, D (1999) Bacterial diversity within the human subgingival crevice. Proc Natl Acad Sci USA 96: 14547–14552PubMedCrossRefGoogle Scholar
  19. 19.
    Kumar, PS, Griffen, AL, Moeschberger, ML, Leys, EJ (2005) Identification of candidate periodontal pathogens and beneficial species by quantitative 16S clonal analysis. J Clin Microb 43: 3944–3955CrossRefGoogle Scholar
  20. 20.
    Leache, AD, Reeder, TW (2002) Molecular systematics of the eastern fence lizard (Sceloporus undulatus): a comparison of parsimony, likelihood and Bayesian approaches. Syst Biol 51: 44–68PubMedCrossRefGoogle Scholar
  21. 21.
    Ley, RE, Backhed, F, Turnbaugh, P, Lozupone, CA, Knight, RD, Gordon, JI (2005) Obesity alters gut microbial community. Proc Natl Acad Sci USA 102: 11070–11075PubMedCrossRefGoogle Scholar
  22. 22.
    Maddison, WP, Maddison, DR (1993) MacClade, v. 3. Sinauer Press. Sunderland, MAGoogle Scholar
  23. 23.
    Maddison, WP, Slatkin, M (1991) Null models for the number of evolutionary steps in a character on a phylogenetic tree. Evolution 45: 1184–1197CrossRefGoogle Scholar
  24. 24.
    Martin, AP (2002) Phylogenetic approaches for describing and comparing the diversity of microbial communities. Appl Environ Microbiol 68: 3673–3682PubMedCrossRefGoogle Scholar
  25. 25.
    McGarvey, JA, Miller, WG, Sanchez, S, Stanker, L (2004) Identification of bacterial populations in dairy wastewaters by use of 16S rRNA gene sequences and other genetic markers. Appl Environ Microbiol 70: 4267–4275PubMedCrossRefGoogle Scholar
  26. 26.
    Nielsen, R (2002) Mapping mutations on phylogenies. Syst Biol 51(5): 729–739PubMedCrossRefGoogle Scholar
  27. 27.
    Pace, N (1997) A molecular view of microbial diversity and the biosphere. Science 276: 734–740PubMedCrossRefGoogle Scholar
  28. 28.
    Posada, D, Crandall, KA (1998) Modeltest: testing the model of DNA substitution. Bioinformatics 14: 817–818PubMedCrossRefGoogle Scholar
  29. 29.
    Prieme, A, Braker, G, Tiedje, JM (2002) Diversity of nitrite reductase (nirK and nirS) gene fragments in forested upland and wetland soils. Appl Environ Microbiol 68: 1893–1900PubMedCrossRefGoogle Scholar
  30. 30.
    Ronquist, F (2004) Bayesian inference of character evolution. Trends Ecol Evol 19(9): 475–481PubMedCrossRefGoogle Scholar
  31. 31.
    Schadt, CW, Martin, AP, Lipson, DA, Schmidt, SK (2003) Seasonal dynamics of previously unknown fungal lineages in tundra soils. Science 301: 1359–1361PubMedCrossRefGoogle Scholar
  32. 32.
    Schloss, PD, Larget, BR, Handelsman, J (2004) Integration of microbial ecology and statistics: a test to compare gene libraries. Appl Environ Microbiol 70: 5485–5492PubMedCrossRefGoogle Scholar
  33. 33.
    Schultz, TR, Churchill, GA (1999) The role of subjectivity in reconstructing ancestral character states: a Bayesian approach to unknown rates, states, and transformation asymmetries. Syst Biol 48(3): 651–664CrossRefGoogle Scholar
  34. 34.
    Suau, A, Bonnet, R, Sutren, M, Godon, JJ, Gibson, G, Collins, MD, Dore, J (1999) Direct analysis of genes encoding 16S rRNA from complex communities reveals many novel molecular species within the human gut. Appl Environ Microbiol 65: 4799–4807PubMedGoogle Scholar
  35. 35.
    Suzuki, Y, Glazko, GV, Nei, M (2002) Overcredibility of molecular phylogenies obtained by Bayesian phylogenetics. Proc Natl Acad Sci USA 99: 16138–16143PubMedCrossRefGoogle Scholar
  36. 36.
    Swofford, DL, Olsen, GJ, Waddell, PJ, Hillis, DM (1996) Phylogenetic inference. In: Hillis DM, Moritz C, Mable BK (Eds.) Molecular Systematics2Sinauer Press. Sunderland, MA, pp 407–514Google Scholar
  37. 37.
    Swofford, DS (2002) PAUP* 4.0. Sinauer Press. Sunderland, MA, USAGoogle Scholar
  38. 38.
    Theron, J, Cloete, TE (2000) Molecular techniques for determining microbial diversity and community structure in natural environments. Crit Rev Microbiol 26: 37–57PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, Inc. 2006

Authors and Affiliations

  1. 1.Department of Ecology and EvolutionUniversity of ColoradoBoulderUSA

Personalised recommendations