Limits...
Dissecting the in vivo metabolic potential of two human gut acetogens.

Rey FE, Faith JJ, Bain J, Muehlbauer MJ, Stevens RD, Newgard CB, Gordon JI - J. Biol. Chem. (2010)

Bottom Line: It increases the efficiency of fermentation by consuming reducing equivalents, thereby maintaining a high NAD(+)/NADH ratio and boosting acetate production.In contrast, M. formatexigens consumes oligosaccharides, does not impact the redox state of the gut, and boosts the yield of succinate.These findings have strategic implications for those who wish to manipulate the hydrogen economy of gut microbial communities in ways that modulate energy harvest.

View Article: PubMed Central - PubMed

Affiliation: Center for Genome Sciences, Washington University School of Medicine, St. Louis, Missouri 63108, USA.

ABSTRACT
Fermenting microbial communities generate hydrogen; its removal through the production of acetate, methane, or hydrogen sulfide modulates the efficiency of energy extraction from available nutrients in many ecosystems. We noted that pathway components for acetogenesis are more abundantly and consistently represented in the gut microbiomes of monozygotic twins and their mothers than components for methanogenesis or sulfate reduction and subsequently analyzed the metabolic potential of two sequenced human gut acetogens, Blautia hydrogenotrophica and Marvinbryantia formatexigens in vitro and in the intestines of gnotobiotic mice harboring a prominent saccharolytic bacterium. To do so, we developed a generally applicable method for multiplex sequencing of expressed microbial mRNAs (microbial RNA-Seq) and, together with mass spectrometry of metabolites, showed that these organisms have distinct patterns of substrate utilization. B. hydrogenotrophica targets aliphatic and aromatic amino acids. It increases the efficiency of fermentation by consuming reducing equivalents, thereby maintaining a high NAD(+)/NADH ratio and boosting acetate production. In contrast, M. formatexigens consumes oligosaccharides, does not impact the redox state of the gut, and boosts the yield of succinate. These findings have strategic implications for those who wish to manipulate the hydrogen economy of gut microbial communities in ways that modulate energy harvest.

Show MeSH

Related in: MedlinePlus

The impact of two human gut acetogens on microbial metabolism in the ceca of gnotobiotic mice. A, multiplex RNA-Seq analysis of the B. hydrogenotrophica transcriptome. The heat map shows selected B. hydrogenotrophica genes, expressed in the ceca of B. thetaiotaomicron (Bt)/B. hydrogenotrophica (Bh) bi-associated mice and in vitro during growth of B. hydrogenotrophica in MA4 medium containing nine different substrates. PPDE > 0.99. Carbon sources were supplied at 1% w/v. The maximal relative expression across a row is red; the minimum is green. stat, stationary phase (note that all other cultures were harvested during mid-log phase). Rumhyd is the locus tag for B. hydrogenotrophica (previously known as R. hydrogenotrophicus). PTS, phosphotransferase system. B, targeted tandem mass spectrometry analysis of amino acid levels. C, nontargeted GC/MS analysis of cecal contents. The data are expressed as unit-less peak areas, normalized to the total ion chromatogram for the cecal sample. 3-IAA, 3-indoleacetic acid; 4-HPA, 4-hydroxyphenylacetic acid. D–H, cecal contents from mice, colonized with either acetogen plus B. thetaiotaomicron or with B. thetaiotaomicron alone, were assayed for NAD+, NADH (D; n = 4–5/group), polyglucose- and polyfructose-containing polysaccharides (E; n = 14–15/group), acetate (F; n = 14–15 animals/group), succinate (G; n = 14–15/group), and propionate (H; n = 14–15/group). The mean values ± S.E. are plotted. *, p < 0.05 based on analysis of variance, except B and C (Student's t test). Quantitative PCR assays revealed the following levels of colonization in the different groups of mice studied: B. thetaiotaomicron mono-association, 7.5 × 1010 ± 1.3 × 1010 genome equivalents/g of wet cecal content, B. thetaiotaomicron/B. hydrogenotrophica bi-association, 6.5 × 1010 ± 9.3 × 109 (B. thetaiotaomicron), 9.4 × 109 ± 2.1 × 109 (B. hydrogenotrophica), and B. thetaiotaomicron/M. formatexigens (Mf) bi-association, 3.1 × 1010 ± 6.3 × 109 (B. thetaiotaomicron), 8.4 ± 109 ± 6.3 × 108 (M. formatexigens); n = 4–5 animals group (three independent experiments).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2903421&req=5

Figure 2: The impact of two human gut acetogens on microbial metabolism in the ceca of gnotobiotic mice. A, multiplex RNA-Seq analysis of the B. hydrogenotrophica transcriptome. The heat map shows selected B. hydrogenotrophica genes, expressed in the ceca of B. thetaiotaomicron (Bt)/B. hydrogenotrophica (Bh) bi-associated mice and in vitro during growth of B. hydrogenotrophica in MA4 medium containing nine different substrates. PPDE > 0.99. Carbon sources were supplied at 1% w/v. The maximal relative expression across a row is red; the minimum is green. stat, stationary phase (note that all other cultures were harvested during mid-log phase). Rumhyd is the locus tag for B. hydrogenotrophica (previously known as R. hydrogenotrophicus). PTS, phosphotransferase system. B, targeted tandem mass spectrometry analysis of amino acid levels. C, nontargeted GC/MS analysis of cecal contents. The data are expressed as unit-less peak areas, normalized to the total ion chromatogram for the cecal sample. 3-IAA, 3-indoleacetic acid; 4-HPA, 4-hydroxyphenylacetic acid. D–H, cecal contents from mice, colonized with either acetogen plus B. thetaiotaomicron or with B. thetaiotaomicron alone, were assayed for NAD+, NADH (D; n = 4–5/group), polyglucose- and polyfructose-containing polysaccharides (E; n = 14–15/group), acetate (F; n = 14–15 animals/group), succinate (G; n = 14–15/group), and propionate (H; n = 14–15/group). The mean values ± S.E. are plotted. *, p < 0.05 based on analysis of variance, except B and C (Student's t test). Quantitative PCR assays revealed the following levels of colonization in the different groups of mice studied: B. thetaiotaomicron mono-association, 7.5 × 1010 ± 1.3 × 1010 genome equivalents/g of wet cecal content, B. thetaiotaomicron/B. hydrogenotrophica bi-association, 6.5 × 1010 ± 9.3 × 109 (B. thetaiotaomicron), 9.4 × 109 ± 2.1 × 109 (B. hydrogenotrophica), and B. thetaiotaomicron/M. formatexigens (Mf) bi-association, 3.1 × 1010 ± 6.3 × 109 (B. thetaiotaomicron), 8.4 ± 109 ± 6.3 × 108 (M. formatexigens); n = 4–5 animals group (three independent experiments).

Mentions: Groups of 8-week-old germ-free mice were colonized with B. thetaiotaomicron alone or with B. thetaiotaomicron for 2–3 days followed by B. hydrogenotrophica or M. formatexigens for 14 days (n = 4–5 mice/experiment; n = 3 independent experiments). In the presence of B. thetaiotaomicron, both acetogens robustly colonized the distal gut where they represented 13 ± 3% (B. hydrogenotrophica) and 22 ± 2% (M. formatexigens) of the two-member model communities (see Fig. 2 legend for absolute numbers). These levels of colonization are sufficient to detect expression changes in all community members with RNA-Seq.


Dissecting the in vivo metabolic potential of two human gut acetogens.

Rey FE, Faith JJ, Bain J, Muehlbauer MJ, Stevens RD, Newgard CB, Gordon JI - J. Biol. Chem. (2010)

The impact of two human gut acetogens on microbial metabolism in the ceca of gnotobiotic mice. A, multiplex RNA-Seq analysis of the B. hydrogenotrophica transcriptome. The heat map shows selected B. hydrogenotrophica genes, expressed in the ceca of B. thetaiotaomicron (Bt)/B. hydrogenotrophica (Bh) bi-associated mice and in vitro during growth of B. hydrogenotrophica in MA4 medium containing nine different substrates. PPDE > 0.99. Carbon sources were supplied at 1% w/v. The maximal relative expression across a row is red; the minimum is green. stat, stationary phase (note that all other cultures were harvested during mid-log phase). Rumhyd is the locus tag for B. hydrogenotrophica (previously known as R. hydrogenotrophicus). PTS, phosphotransferase system. B, targeted tandem mass spectrometry analysis of amino acid levels. C, nontargeted GC/MS analysis of cecal contents. The data are expressed as unit-less peak areas, normalized to the total ion chromatogram for the cecal sample. 3-IAA, 3-indoleacetic acid; 4-HPA, 4-hydroxyphenylacetic acid. D–H, cecal contents from mice, colonized with either acetogen plus B. thetaiotaomicron or with B. thetaiotaomicron alone, were assayed for NAD+, NADH (D; n = 4–5/group), polyglucose- and polyfructose-containing polysaccharides (E; n = 14–15/group), acetate (F; n = 14–15 animals/group), succinate (G; n = 14–15/group), and propionate (H; n = 14–15/group). The mean values ± S.E. are plotted. *, p < 0.05 based on analysis of variance, except B and C (Student's t test). Quantitative PCR assays revealed the following levels of colonization in the different groups of mice studied: B. thetaiotaomicron mono-association, 7.5 × 1010 ± 1.3 × 1010 genome equivalents/g of wet cecal content, B. thetaiotaomicron/B. hydrogenotrophica bi-association, 6.5 × 1010 ± 9.3 × 109 (B. thetaiotaomicron), 9.4 × 109 ± 2.1 × 109 (B. hydrogenotrophica), and B. thetaiotaomicron/M. formatexigens (Mf) bi-association, 3.1 × 1010 ± 6.3 × 109 (B. thetaiotaomicron), 8.4 ± 109 ± 6.3 × 108 (M. formatexigens); n = 4–5 animals group (three independent experiments).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2903421&req=5

Figure 2: The impact of two human gut acetogens on microbial metabolism in the ceca of gnotobiotic mice. A, multiplex RNA-Seq analysis of the B. hydrogenotrophica transcriptome. The heat map shows selected B. hydrogenotrophica genes, expressed in the ceca of B. thetaiotaomicron (Bt)/B. hydrogenotrophica (Bh) bi-associated mice and in vitro during growth of B. hydrogenotrophica in MA4 medium containing nine different substrates. PPDE > 0.99. Carbon sources were supplied at 1% w/v. The maximal relative expression across a row is red; the minimum is green. stat, stationary phase (note that all other cultures were harvested during mid-log phase). Rumhyd is the locus tag for B. hydrogenotrophica (previously known as R. hydrogenotrophicus). PTS, phosphotransferase system. B, targeted tandem mass spectrometry analysis of amino acid levels. C, nontargeted GC/MS analysis of cecal contents. The data are expressed as unit-less peak areas, normalized to the total ion chromatogram for the cecal sample. 3-IAA, 3-indoleacetic acid; 4-HPA, 4-hydroxyphenylacetic acid. D–H, cecal contents from mice, colonized with either acetogen plus B. thetaiotaomicron or with B. thetaiotaomicron alone, were assayed for NAD+, NADH (D; n = 4–5/group), polyglucose- and polyfructose-containing polysaccharides (E; n = 14–15/group), acetate (F; n = 14–15 animals/group), succinate (G; n = 14–15/group), and propionate (H; n = 14–15/group). The mean values ± S.E. are plotted. *, p < 0.05 based on analysis of variance, except B and C (Student's t test). Quantitative PCR assays revealed the following levels of colonization in the different groups of mice studied: B. thetaiotaomicron mono-association, 7.5 × 1010 ± 1.3 × 1010 genome equivalents/g of wet cecal content, B. thetaiotaomicron/B. hydrogenotrophica bi-association, 6.5 × 1010 ± 9.3 × 109 (B. thetaiotaomicron), 9.4 × 109 ± 2.1 × 109 (B. hydrogenotrophica), and B. thetaiotaomicron/M. formatexigens (Mf) bi-association, 3.1 × 1010 ± 6.3 × 109 (B. thetaiotaomicron), 8.4 ± 109 ± 6.3 × 108 (M. formatexigens); n = 4–5 animals group (three independent experiments).
Mentions: Groups of 8-week-old germ-free mice were colonized with B. thetaiotaomicron alone or with B. thetaiotaomicron for 2–3 days followed by B. hydrogenotrophica or M. formatexigens for 14 days (n = 4–5 mice/experiment; n = 3 independent experiments). In the presence of B. thetaiotaomicron, both acetogens robustly colonized the distal gut where they represented 13 ± 3% (B. hydrogenotrophica) and 22 ± 2% (M. formatexigens) of the two-member model communities (see Fig. 2 legend for absolute numbers). These levels of colonization are sufficient to detect expression changes in all community members with RNA-Seq.

Bottom Line: It increases the efficiency of fermentation by consuming reducing equivalents, thereby maintaining a high NAD(+)/NADH ratio and boosting acetate production.In contrast, M. formatexigens consumes oligosaccharides, does not impact the redox state of the gut, and boosts the yield of succinate.These findings have strategic implications for those who wish to manipulate the hydrogen economy of gut microbial communities in ways that modulate energy harvest.

View Article: PubMed Central - PubMed

Affiliation: Center for Genome Sciences, Washington University School of Medicine, St. Louis, Missouri 63108, USA.

ABSTRACT
Fermenting microbial communities generate hydrogen; its removal through the production of acetate, methane, or hydrogen sulfide modulates the efficiency of energy extraction from available nutrients in many ecosystems. We noted that pathway components for acetogenesis are more abundantly and consistently represented in the gut microbiomes of monozygotic twins and their mothers than components for methanogenesis or sulfate reduction and subsequently analyzed the metabolic potential of two sequenced human gut acetogens, Blautia hydrogenotrophica and Marvinbryantia formatexigens in vitro and in the intestines of gnotobiotic mice harboring a prominent saccharolytic bacterium. To do so, we developed a generally applicable method for multiplex sequencing of expressed microbial mRNAs (microbial RNA-Seq) and, together with mass spectrometry of metabolites, showed that these organisms have distinct patterns of substrate utilization. B. hydrogenotrophica targets aliphatic and aromatic amino acids. It increases the efficiency of fermentation by consuming reducing equivalents, thereby maintaining a high NAD(+)/NADH ratio and boosting acetate production. In contrast, M. formatexigens consumes oligosaccharides, does not impact the redox state of the gut, and boosts the yield of succinate. These findings have strategic implications for those who wish to manipulate the hydrogen economy of gut microbial communities in ways that modulate energy harvest.

Show MeSH
Related in: MedlinePlus