This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Blackwood, C. B.
Right arrow Articles by Paul, E. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Blackwood, C. B.
Right arrow Articles by Paul, E. A.
Agricola
Right arrow Articles by Blackwood, C. B.
Right arrow Articles by Paul, E. A.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, February 2003, p. 926-932, Vol. 69, No. 2
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.2.926-932.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Terminal Restriction Fragment Length Polymorphism Data Analysis for Quantitative Comparison of Microbial Communities

Christopher B. Blackwood,1,2* Terry Marsh,1 Sang-Hoon Kim,1 and Eldor A. Paul1,2,{dagger}

Center for Microbial Ecology,1 Department of Crop and Soil Sciences, Michigan State University, East Lansing, Michigan 488242

Received 22 May 2002/ Accepted 12 November 2002

Terminal restriction fragment length polymorphism (T-RFLP) is a culture-independent method of obtaining a genetic fingerprint of the composition of a microbial community. Comparisons of the utility of different methods of (i) including peaks, (ii) computing the difference (or distance) between profiles, and (iii) performing statistical analysis were made by using replicated profiles of eubacterial communities. These samples included soil collected from three regions of the United States, soil fractions derived from three agronomic field treatments, soil samples taken from within one meter of each other in an alfalfa field, and replicate laboratory bioreactors. Cluster analysis by Ward's method and by the unweighted-pair group method using arithmetic averages (UPGMA) were compared. Ward's method was more effective at differentiating major groups within sets of profiles; UPGMA had a slightly reduced error rate in clustering of replicate profiles and was more sensitive to outliers. Most replicate profiles were clustered together when relative peak height or Hellinger-transformed peak height was used, in contrast to raw peak height. Redundancy analysis was more effective than cluster analysis at detecting differences between similar samples. Redundancy analysis using Hellinger distance was more sensitive than that using Euclidean distance between relative peak height profiles. Analysis of Jaccard distance between profiles, which considers only the presence or absence of a terminal restriction fragment, was the most sensitive in redundancy analysis, and was equally sensitive in cluster analysis, if all profiles had cumulative peak heights greater than 10,000 fluorescence units. It is concluded that T-RFLP is a sensitive method of differentiating between microbial communities when the optimal statistical method is used for the situation at hand. It is recommended that hypothesis testing be performed by redundancy analysis of Hellinger-transformed data and that exploratory data analysis be performed by cluster analysis using Ward's method to find natural groups or by UPGMA to identify potential outliers. Analyses can also be based on Jaccard distance if all profiles have cumulative peak heights greater than 10,000 fluorescence units.

* Corresponding author. Present address: Sustainable Agricultural Systems Laboratory, Beltsville Agricultural Research Center, Bldg. 001 Rm. 140, USDA-ARS, Beltsville, MD 20705. Phone: (301) 504-8022. Fax: (301) 504-8370. E-mail: blackwoc{at}ba.ars.usda.gov.

{dagger} Present address: Natural Resource Ecology Laboratory, Colorado State University, Fort Collins, CO 80523-1965.

Applied and Environmental Microbiology, February 2003, p. 926-932, Vol. 69, No. 2
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.2.926-932.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Rossi, P., Gillet, F., Rohrbach, E., Diaby, N., Holliger, C. (2009). Statistical Assessment of Variability of Terminal Restriction Fragment Length Polymorphism Analysis Applied to Complex Microbial Communities. Appl. Environ. Microbiol. 75: 7268-7270 [Abstract] [Full Text]  
  • Berthrong, S. T., Schadt, C. W., Pineiro, G., Jackson, R. B. (2009). Afforestation Alters the Composition of Functional Genes in Soil and Biogeochemical Processes in South American Grasslands. Appl. Environ. Microbiol. 75: 6240-6248 [Abstract] [Full Text]  
  • Feinstein, L. M., Sul, W. J., Blackwood, C. B. (2009). Assessment of Bias Associated with Incomplete Extraction of Microbial DNA from Soil. Appl. Environ. Microbiol. 75: 5428-5433 [Abstract] [Full Text]  
  • Engelbrektson, A., Korzenik, J. R., Pittler, A., Sanders, M. E., Klaenhammer, T. R., Leyer, G., Kitts, C. L. (2009). Probiotics to minimize the disruption of faecal microbiota in healthy subjects undergoing antibiotic therapy. J Med Microbiol 58: 663-670 [Abstract] [Full Text]  
  • Ramette, A. (2009). Quantitative Community Fingerprinting Methods for Estimating the Abundance of Operational Taxonomic Units in Natural Microbial Communities. Appl. Environ. Microbiol. 75: 2495-2505 [Abstract] [Full Text]  
  • Flores-Mireles, A. L., Winans, S. C., Holguin, G. (2007). Molecular Characterization of Diazotrophic and Denitrifying Bacteria Associated with Mangrove Roots. Appl. Environ. Microbiol. 73: 7308-7321 [Abstract] [Full Text]  
  • Bordenave, S., Goni-Urriza, M. S., Caumette, P., Duran, R. (2007). Effects of Heavy Fuel Oil on the Bacterial Community Structure of a Pristine Microbial Mat. Appl. Environ. Microbiol. 73: 6089-6097 [Abstract] [Full Text]  
  • Blackwood, C. B., Hudleston, D., Zak, D. R., Buyer, J. S. (2007). Interpreting Ecological Diversity Indices Applied to Terminal Restriction Fragment Length Polymorphism Data: Insights from Simulated Microbial Communities. Appl. Environ. Microbiol. 73: 5276-5283 [Abstract] [Full Text]  
  • Collins, R. E., Rocap, G. (2007). REPK: an analytical web server to select restriction endonucleases for terminal restriction fragment length polymorphism analysis. Nucleic Acids Res 35: W58-W62 [Abstract] [Full Text]  
  • Bent, S. J., Pierson, J. D., Forney, L. J., Danovaro, R., Luna, G. M., Dell'Anno, A., Pietrangeli, B. (2007). Measuring Species Richness Based on Microbial Community Fingerprints: the Emperor Has No Clothes. Appl. Environ. Microbiol. 73: 2399-2401 [Full Text]  
  • Thies, J. E. (2007). Soil Microbial Community Analysis using Terminal Restriction Fragment Length Polymorphisms. Soil Sci. 71: 579-591 [Abstract] [Full Text]  
  • Das, M., Royer, T. V., Leff, L. G. (2007). Diversity of Fungi, Bacteria, and Actinomycetes on Leaves Decomposing in a Stream. Appl. Environ. Microbiol. 73: 756-767 [Abstract] [Full Text]  
  • Singh, B. K., Nazaries, L., Munro, S., Anderson, I. C., Campbell, C. D. (2006). Use of Multiplex Terminal Restriction Fragment Length Polymorphism for Rapid and Simultaneous Analysis of Different Components of the Soil Microbial Community{triangledown}. Appl. Environ. Microbiol. 72: 7278-7285 [Abstract] [Full Text]  
  • Bhatia, M., Sharp, M., Foght, J. (2006). Distinct Bacterial Communities Exist beneath a High Arctic Polythermal Glacier. Appl. Environ. Microbiol. 72: 5838-5845 [Abstract] [Full Text]  
  • Danovaro, R., Luna, G. M., Dell'Anno, A., Pietrangeli, B. (2006). Comparison of Two Fingerprinting Techniques, Terminal Restriction Fragment Length Polymorphism and Automated Ribosomal Intergenic Spacer Analysis, for Determination of Bacterial Diversity in Aquatic Environments. Appl. Environ. Microbiol. 72: 5982-5989 [Abstract] [Full Text]  
  • Souza, V., Espinosa-Asuar, L., Escalante, A. E., Eguiarte, L. E., Farmer, J., Forney, L., Lloret, L., Rodriguez-Martinez, J. M., Soberon, X., Dirzo, R., Elser, J. J. (2006). From the Cover: An endangered oasis of aquatic microbial biodiversity in the Chihuahuan desert. Proc. Natl. Acad. Sci. USA 103: 6565-6570 [Abstract] [Full Text]  
  • Osborne, C. A., Rees, G. N., Bernstein, Y., Janssen, P. H. (2006). New Threshold and Confidence Estimates for Terminal Restriction Fragment Length Polymorphism Analysis of Complex Bacterial Communities. Appl. Environ. Microbiol. 72: 1270-1278 [Abstract] [Full Text]  
  • Pett-Ridge, J., Firestone, M. K. (2005). Redox Fluctuation Structures Microbial Communities in a Wet Tropical Soil. Appl. Environ. Microbiol. 71: 6998-7007 [Abstract] [Full Text]  
  • Rhee, H., Holditch-Davis, D., Miles, M. S. (2005). Patterns of Physical Symptoms and Relationships With Psychosocial Factors in Adolescents. Psychosom. Med. 67: 1006-1012 [Abstract] [Full Text]  
  • Fogarty, L. R., Voytek, M. A. (2005). Comparison of Bacteroides-Prevotella 16S rRNA Genetic Markers for Fecal Samples from Different Animal Species. Appl. Environ. Microbiol. 71: 5999-6007 [Abstract] [Full Text]  
  • Kuehl, C. J., Wood, H. D., Marsh, T. L., Schmidt, T. M., Young, V. B. (2005). Colonization of the Cecal Mucosa by Helicobacter hepaticus Impacts the Diversity of the Indigenous Microbiota. Infect. Immun. 73: 6952-6961 [Abstract] [Full Text]  
  • Sanchez, M. A., Vasquez, M., Gonzalez, B. (2004). A Previously Unexposed Forest Soil Microbial Community Degrades High Levels of the Pollutant 2,4,6-Trichlorophenol. Appl. Environ. Microbiol. 70: 7567-7570 [Abstract] [Full Text]  
  • Cardinale, M., Brusetti, L., Quatrini, P., Borin, S., Puglia, A. M., Rizzi, A., Zanardini, E., Sorlini, C., Corselli, C., Daffonchio, D. (2004). Comparison of Different Primer Sets for Use in Automated Ribosomal Intergenic Spacer Analysis of Complex Bacterial Communities. Appl. Environ. Microbiol. 70: 6147-6156 [Abstract] [Full Text]  
  • Ayala-del-Rio, H. L., Callister, S. J., Criddle, C. S., Tiedje, J. M. (2004). Correspondence between Community Structure and Function during Succession in Phenol- and Phenol-plus-Trichloroethene-Fed Sequencing Batch Reactors. Appl. Environ. Microbiol. 70: 4950-4960 [Abstract] [Full Text]  
  • Devare, M. H., Jones, C. M., Thies, J. E. (2004). Effect of Cry3Bb Transgenic Corn and Tefluthrin on the Soil Microbial Community: Biomass, Activity, and Diversity. J. Environ. Qual. 33: 837-843 [Abstract] [Full Text]  
  • Grant, A., Ogilvie, L. A., Blackwood, C. B., Marsh, T., Kim, S.-H., Paul, E. A. (2003). Terminal Restriction Fragment Length Polymorphism Data Analysis. Appl. Environ. Microbiol. 69: 6342-6343 [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»