Gene-centric metagenomics of the fiber-adherent bovine rumen microbiome reveals forage specific glycoside hydrolases

The complex microbiome of the rumen functions as an effective system for the conversion of plant cell wall biomass to microbial protein, short chain fatty acids, and gases. As such, it provides a unique genetic resource for plant cell wall degrading microbial enzymes that could be used in the production of biofuels. The rumen and gastrointestinal tract harbor a dense and complex microbiome. To gain a greater understanding of the ecology and metabolic potential of this microbiome, we used comparative metagenomics (phylotype analysis and SEED subsystems-based annotations) to examine randomly sampled pyrosequence data from 3 fiber-adherent microbiomes and 1 pooled liquid sample (a mixture of the liquid microbiome fractions from the same bovine rumens). Even though the 3 animals were fed the same diet, the community structure, predicted phylotype, and metabolic potentials in the rumen were markedly different with respect to nutrient utilization. A comparison of the glycoside hydrolase and cellulosome functional genes revealed that in the rumen microbiome, initial colonization of fiber appears to be by organisms possessing enzymes that attack the easily available side chains of complex plant polysaccharides and not the more recalcitrant main chains, especially cellulose. Furthermore, when compared with the termite hindgut microbiome, there are fundamental differences in the glycoside hydrolase content that appear to be diet driven for either the bovine rumen (forages and legumes) or the termite hindgut (wood).

[1]  J. Kruskal Multidimensional scaling by optimizing goodness of fit to a nonmetric hypothesis , 1964 .

[2]  B. A. Dehority,et al.  Effect of Short-Term Chilling of Rumen Contents on Viable Bacterial Numbers , 1980, Applied and environmental microbiology.

[3]  A. Klieve,et al.  Morphological diversity of ruminal bacteriophages from sheep and cattle , 1988, Applied and environmental microbiology.

[4]  B Flesher,et al.  Use of phylogenetically based hybridization probes for studies of ruminal microbial ecology , 1988, Applied and environmental microbiology.

[5]  R. Anthony,et al.  Use of DNA probes to monitor nutritional effects on ruminal prokaryotes and Fibrobacter succinogenes S85. , 1992, Journal of animal science.

[6]  Keith Gull,et al.  Anaerobic fungi in herbivorous animals , 1994 .

[7]  R. Amann,et al.  Taxon Specific Hybridization Probes for Fiber-digesting Bacteria Suggest Novel Gut-associated Fibrobacter , 1994 .

[8]  D. Stahl,et al.  Taxon-specific probes for the cellulolytic genus Fibrobacter reveal abundant and novel equine-associated populations , 1995, Applied and environmental microbiology.

[9]  D. Stahl,et al.  Microbial community structure in gastrointestinal tracts of domestic animals: comparative analyses using rRNA‐targeted oligonucleotide probes , 1997 .

[10]  B. White,et al.  Polysaccharide Degradation in the Rumen and Large Intestine , 1997 .

[11]  D. E. Akin,et al.  Bacteria, Fungi, and Protozoa of the Rumen , 1997 .

[12]  M. Cotta,et al.  Digestion of Nitrogen in the Rumen: A Model for Metabolism of Nitrogen Compounds in Gastrointestinal Environments , 1997 .

[13]  N. Pace A molecular view of microbial diversity and the biosphere. , 1997, Science.

[14]  R. Forster,et al.  Phylogenetic analysis of rumen bacteria by comparative sequence analysis of cloned 16S rRNA genes. , 1998, Anaerobe.

[15]  J. Handelsman,et al.  Molecular biological access to the chemistry of unknown soil microbes: a new frontier for natural products. , 1998, Chemistry & biology.

[16]  G. Gottschalk,et al.  Construction of Environmental DNA Libraries inEscherichia coli and Screening for the Presence of Genes Conferring Utilization of 4-Hydroxybutyrate , 1999, Applied and Environmental Microbiology.

[17]  B. Dalrymple,et al.  16S rDNA sequencing of Ruminococcus albus and Ruminococcus flavefaciens: design of a signature probe and its application in adult sheep. , 1999, Microbiology.

[18]  T. E. Cloete,et al.  Molecular Techniques for Determining Microbial Diversity and Community Structure in Natural Environments , 2000, Critical reviews in microbiology.

[19]  D. Stahl,et al.  Comparison of microbial populations in model and natural rumens using 16S ribosomal RNA-targeted probes. , 2000, Environmental microbiology.

[20]  Y. Benno,et al.  Rumen Bacterial Community Transition During Adaptation to High-grain Diet , 2000 .

[21]  R. Aminov,et al.  Phylogenetic analysis of archaeal 16S rRNA libraries from the rumen suggests the existence of a novel group of archaea not associated with known methanogens. , 2001, FEMS microbiology letters.

[22]  B. White,et al.  Analysis of the rumen bacterial diversity under two different diet conditions using denaturing gradient gel electrophoresis, random sequencing, and statistical ecology approaches , 2001 .

[23]  Y. Benno,et al.  Diet-Dependent Shifts in the Bacterial Population of the Rumen Revealed with Real-Time PCR , 2001, Applied and Environmental Microbiology.

[24]  L. Øvreås,et al.  Microbial diversity and function in soil: from genes to ecosystems. , 2002, Current opinion in microbiology.

[25]  J. Guinea,et al.  Characterization of several Psychrobacter strains isolated from Antarctic environments and description of Psychrobacter luti sp. nov. and Psychrobacter fozii sp. nov. , 2003, International journal of systematic and evolutionary microbiology.

[26]  S. Koike,et al.  Phylogenetic analysis of fiber-associated rumen bacterial community and PCR detection of uncultured bacteria. , 2003, FEMS microbiology letters.

[27]  K. Nelson,et al.  Phylogenetic analysis of the microbial populations in the wild herbivore gastrointestinal tract: insights into an unexplored niche. , 2003, Environmental microbiology.

[28]  A. Travis,et al.  16S rDNA library-based analysis of ruminal bacterial diversity , 2004, Antonie van Leeuwenhoek.

[29]  Bryan A White,et al.  Suppressive subtractive hybridization as a tool for identifying genetic diversity in an environmental metagenome: the rumen as a model. , 2004, Environmental microbiology.

[30]  Zhongtang Yu,et al.  Improved extraction of PCR-quality community DNA from digesta and fecal samples. , 2004, BioTechniques.

[31]  J. Handelsman,et al.  Introducing DOTUR, a Computer Program for Defining Operational Taxonomic Units and Estimating Species Richness , 2005, Applied and Environmental Microbiology.

[32]  S. Tringe,et al.  Comparative Metagenomics of Microbial Communities , 2004, Science.

[33]  Naryttza N. Diaz,et al.  The Subsystems Approach to Genome Annotation and its Use in the Project to Annotate 1000 Genomes , 2005, Nucleic acids research.

[34]  James R. Cole,et al.  The Ribosomal Database Project (RDP-II): sequences and tools for high-throughput rRNA analysis , 2004, Nucleic Acids Res..

[35]  James R. Knight,et al.  Genome sequencing in microfabricated high-density picolitre reactors , 2005, Nature.

[36]  M. Breitbart,et al.  Using pyrosequencing to shed light on deep mine microbial ecology , 2006, BMC Genomics.

[37]  Zhongtang Yu,et al.  Novel microbial diversity adherent to plant biomass in the herbivore gastrointestinal tract, as revealed by ribosomal intergenic spacer analysis and rrs gene sequencing. , 2005, Environmental microbiology.

[38]  K. Timmis,et al.  Novel hydrolase diversity retrieved from a metagenome library of bovine rumen microflora. , 2005, Environmental microbiology.

[39]  Forest Rohwer,et al.  An application of statistics to comparative metagenomics , 2006, BMC Bioinformatics.

[40]  S. Denman,et al.  Development of a real-time PCR assay for monitoring anaerobic fungal and cellulolytic bacterial populations within the rumen. , 2006, FEMS microbiology ecology.

[41]  E. Mardis,et al.  An obesity-associated gut microbiome with increased capacity for energy harvest , 2006, Nature.

[42]  Susan M. Huse,et al.  Microbial diversity in the deep sea and the underexplored “rare biosphere” , 2006, Proceedings of the National Academy of Sciences.

[43]  M. Pop,et al.  Metagenomic Analysis of the Human Distal Gut Microbiome , 2006, Science.

[44]  K. Timmis,et al.  Novel Polyphenol Oxidase Mined from a Metagenome Expression Library of Bovine Rumen , 2006, Journal of Biological Chemistry.

[45]  Natalia N. Ivanova,et al.  Metagenomic and functional analysis of hindgut microbiota of a wood-feeding higher termite , 2007, Nature.

[46]  D. Relman,et al.  An ecological and evolutionary perspective on human–microbe mutualism and disease , 2007, Nature.

[47]  G. Casella,et al.  Pyrosequencing enumerates and contrasts soil microbial diversity , 2007, The ISME Journal.

[48]  Florent E. Angly,et al.  Comparative Metagenomics Reveals Host Specific Metavirulomes and Horizontal Gene Transfer Elements in the Chicken Cecum Microbiome , 2008, PloS one.

[49]  Naryttza N. Diaz,et al.  Phylogenetic classification of short environmental DNA fragments , 2008, Nucleic acids research.

[50]  R. Knight,et al.  Evolution of Mammals and Their Gut Microbes , 2008, Science.

[51]  P. Hugenholtz,et al.  Why the ‘ meta ’ in metagenomics ? , 2022 .

[52]  R. Knight,et al.  Worlds within worlds: evolution of the vertebrate gut microbiota , 2008, Nature Reviews Microbiology.

[53]  Rick L. Stevens,et al.  Functional metagenomic profiling of nine biomes , 2008, Nature.