The 'in vivolifestyle' of bile acid 7α-dehydroxylating bacteria: comparative genomics, metatranscriptomic, and bile acid metabolomics analysis of a defined microbial community in gnotobiotic mice

The formation of secondary bile acids by gut microbes is a current topic of considerable biomedical interest. However, a detailed understanding of the biology of anaerobic bacteria in the genusClostridiumthat are capable of generating secondary bile acids is lacking. We therefore sought to determine the transcriptional responses of two prominent secondary bile acid producing bacteria,Clostridium hylemonaeandClostridium hiranonisto bile salts (in vitro) and the cecal environment of gnotobiotic mice. The genomes ofC. hylemonaeDSM 15053 andC. hiranonisDSM 13275 were closed, and found to encode 3,647 genes (3,584 protein-coding) and 2,363 predicted genes (of which 2,239 are protein-coding), respectively, and 1,035 orthologs were shared betweenC. hylemonaeandC. hiranonis. RNA-Seq analysis was performed in growth medium alone, and in the presence of cholic acid (CA) and deoxycholic acid (DCA). Growth with CA resulted in differential expression (>0.58 log(2)FC; FDR < 0.05) of 197 genes inC. hiranonisand 118 genes inC. hylemonae. The bile acid-inducible operons (bai) from each organism were highly upregulated in the presence of CA but not DCA. We then colonized germ-free mice with human gut bacterial isolates capable of metabolizing taurine-conjugated bile acids. This consortium included bile salt hydrolase-expressingBacteroides uniformisATCC 8492,Bacteroides vulgatusATCC 8482,Parabacteroides distasonisDSM 20701, as well as taurine-respiringBilophila wadsworthiaDSM 11045, and deoxycholic/lithocholic acid generatingClostridium hylemonaeDSM 15053 andClostridium hiranonisDSM 13275. Butyrate and iso-bile acid-formingBlautia productaATCC 27340 was also included. The Bacteroidetes made up 84.71% of 16S rDNA cecal reads,B. wadsworthia, constituted 14.7%, and the clostridia made up <.75% of 16S rDNA cecal reads. Bile acid metabolomics of the cecum, serum, and liver indicate that the synthetic community were capable of functional bile salt deconjugation, oxidation/isomerization, and 7 alpha-dehydroxylation of bile acids. Cecal metatranscriptome analysis revealed expression of genes involved in metabolism of taurine-conjugated bile acids. Thein vivotranscriptomes ofC. hylemonaeandC. hiranonissuggest fermentation of simple sugars and utilization of amino acids glycine and proline as electron acceptors. Genes predicted to be involved in trimethylamine (TMA) formation were also expressed.