Restricted Distribution of the Butyrate Kinase Pathway among Butyrate-Producing Bacteria from the Human Colon

Abstract

The final steps in butyrate synthesis by anaerobic bacteria can occur via butyrate kinase and phosphotransbutyrylase or via butyryl-coenzyme A (CoA):acetate CoA-transferase. Degenerate PCR and enzymatic assays were used to assess the presence of butyrate kinase among 38 anaerobic butyrate-producing bacterial isolates from human feces that represent three different clostridial clusters (IV, XIVa, and XVI). Only four strains were found to possess detectable butyrate kinase activity. These were also the only strains to give PCR products (verifiable by sequencing) with degenerate primer pairs designed within the butyrate kinase gene or between the linked butyrate kinase/phosphotransbutyrylase genes. Further analysis of the butyrate kinase/phosphotransbutyrylase genes of one isolate, L2-50, revealed similar organization to that described previously from different groups of clostridia, along with differences in flanking sequences and phylogenetic relationships. Butyryl-CoA:acetate CoA-transferase activity was detected in all 38 strains examined, suggesting that it, rather than butyrate kinase, provides the dominant route for butyrate formation in the human colonic ecosystem that contains a constantly high concentration of acetate.The human colon harbors a diverse range of bacteria that form a complex anaerobic ecosystem. Apart from interacting with each other, for example by cross-feeding, colonic bacteria also interact with the human host and play a vital role in maintaining human health through mechanisms such as protecting against pathogenic bacteria, influencing the host's immune system, and supplying nutrients (23).The recent development of molecular techniques for studying phylogenetic diversity of the colonic microflora is leading to a better understanding of its composition (9, 21). One group of bacteria, which has received relatively little previous attention because of their fastidious growth requirements, are the strictly anaerobic gram-positive bacteria of low mol% GϩC-content clostridial groups. Several studies employing a variety of molecular approaches indicate that members of this group related to clostridial clusters IV and XIVa (14) might make up about 30 to 60% of the total bacterial population within the colon (22,25,38,46,50).Bacteria have been identified within these groups that carry out a fermentative metabolism leading to the buildup of butyrate; lactate and formate are often being produced, while acetate can be either produced or consumed (18). Butyrate is considered to exert health-promoting effects on the colon. It serves as a major energy source for the colonocytes and has also been claimed to be protective against colon cancer and inflammatory bowel diseases via effects on gene expression and cellular development (reviewed in references 36 and 51).The pathway for butyrate formation and its respective genes have been examined in the solventogenic bacterium Clostridium acetobutylicum. Two molecules of acetyl coenzyme A (acetyl-CoA) are condensed and subsequently reduced to butyryl-CoA in a ...

Phylogenetic distribution of three pathways for propionate production within the human gut microbiota

“…The 16S rRNA sequence of GD/7 was determined (accession number EU266552) as described before (Louis et al, 2004) and blastn analysis revealed that it was 97% identical to Coprococcus catus VPI-C6-61. A fluorescent in situ hybridisation (FISH) probe was designed for C. catus and was used to determine the abundance of this species in human volunteers (for details see Supplementary Information S1).…”

“…Bacterial isolation and cultivation Strain GD/7 was isolated on yeast extract-casitonefatty acid (YCFA) medium (Lopez-Siles et al, 2012) containing 25 mM DL lactate from a faecal sample of a 42-year-old man consuming an omnivorous diet as described previously (Louis et al, 2004). The 16S rRNA sequence of GD/7 was determined (accession number EU266552) as described before (Louis et al, 2004) and blastn analysis revealed that it was 97% identical to Coprococcus catus VPI-C6-61.…”

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Voluntary Running Exercise Alters Microbiota Composition and Increases n-Butyrate Concentration in the Rat Cecum

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Proposal of Roseburia faecis sp. nov., Roseburia hominis sp. nov. and Roseburia inulinivorans sp. nov., based on isolates from human faeces

“…Ethical approval was obtained from Grampian Research Ethics Committee (project number 00/00133). The isolations were made from roll tubes of anaerobic M2GSC medium (Miyazaki et al, 1997) or a medium designed to select for Selenomonas-like bacteria, as described previously (Louis et al, 2004). All media were prepared and maintained anaerobically using oxygen-free carbon dioxide.…”