Metagenomic analysis of the human distal gut microbiome

Abstract

The human intestinal microbiota is composed of 10(13) to 10(14) microorganisms whose collective genome ("microbiome") contains at least 100 times as many genes as our own genome. We analyzed approximately 78 million base pairs of unique DNA sequence and 2062 polymerase chain reaction-amplified 16S ribosomal DNA sequences obtained from the fecal DNAs of two healthy adults. Using metabolic function analyses of identified genes, we compared our human genome with the average content of previously sequenced microbial genomes. Our microbiome has significantly enriched metabolism of glycans, amino acids, and xenobiotics; methanogenesis; and 2-methyl-d-erythritol 4-phosphate pathway-mediated biosynthesis of vitamins and isoprenoids. Thus, humans are superorganisms whose metabolism represents an amalgamation of microbial and human attributes.

Fig. 1

Comparison of random metagenome reads with completed genome of Bifidobacterium longum and Methanobrevibacter smithii. (A) Percent identity plot (PIP) of alignments of shotgun reads along the genome of B. longum strain NCC2705. The x axis represents the coordinate along the genome, and the y axis represents the percent identity of the match. (B) Percent identity plot (PIP) of the alignment of shotgun reads along the draft genome of M. smithii. The x axis represents the coordinate along a pseudomolecule formed by concatenating all contigs in the M. smithii draft assembly. The y axis represents the percent identity of the match. The variation in the percent identity of the matches between the shotgun reads from subjects 7 and 8 as compared with the genome sequences of B. longum NCC2705 suggests considerable diversity among Bifidobacterium-like organisms within our samples. Alignments of the reads to the draft genome of M. smithii exhibit a much narrower range of percent identity (89% of alignments were at 95% or better identity as compared with 48% for B. longum), consistent with lower levels of diversity among archaeal members of the gastrointestinal tract.

Fig. 2

COG analysis reveals metabolic functions that are enriched or underrepresented in the human distal gut microbiome (relative to all sequenced microbes). Color code: black, subject 7; gray, subject 8. Bars above both dashed lines indicate enrichment, and bars below both lines indicate underrepresentation (P < 0.05). Asterisks indicate categories that are significantly different between the two subjects (P < 0.05). Secondary metabolites biosynthesis includes antibiotics, pigments, and nonribosomal peptides. Inorganic ion transport and metabolism includes phosphate, sulfate, and various cation transporters.

Fig. 3

KEGG pathway reconstructions reveal metabolic functions that are enriched or underrepresented in the human distal gut microbiome as follows: both samples compared with all sequenced bacterial genomes in KEGG (blue), the human genome (red), and all sequenced archaeal genomes in KEGG (yellow). Asterisks indicate enrichment (odds ratio > 1, P < 0.05) or underrepresentation (odds ratio < 1, P < 0.05). The KEGG category, “metabolism of other amino acids,” includes amino acids that are not incorporated into proteins, such as β-alanine, taurine, and glutathione. Odds ratios are a measure of relative gene content based on the number of independent hits to enzymes present in a given KEGG category.

Fig. 4

Isoprenoid biosynthesis via the MEP pathway and methanogenesis are highly enriched in the distal gut microbiome. (A) MEP pathway for isoprenoid biosynthesis. (B) Odds ratio for each COG in the MEP pathway. All enzymes necessary to convert DXP to IPP and thiamine are enriched (P < 0.0001 relative to all sequenced microbes). (C) Location and role of key enzymes in methanogenesis. (D) Odds ratio for each COG highlighted in (C).