argeting the trimethylamine-producing bacteria of the human gut microbiota

Background: Trimethylamine (TMA), produced by the gut microbiota from dietary quaternary amines, mainly choline and carnitine, is associated with arteriosclerosis and severe cardiovascular disease. Currently, little information on the composition of TMA producers in the gut is available due to their low abundance and the requirement of specific functional-based detection methods as many taxa show disparate abilities to produce that compound.Results: In order to examine the TMA-forming potential of microbial communities we established databases for the key genes of the main TMA-synthesis pathways, encoding choline TMA-lyase (cutC) and carnitine oxygenase (cntA), using a multilevel screening approach on 67,134 genomes predicting 1,114 and 6,815 bacteria to exhibit cutC and cntA, respectively. Gene–targeted assays enumerating the TMA-producing community by quantitative PCR and characterizing its composition via Illumina sequencing were developed and applied on human fecal samples (n=50) where a majority contained TMA producers (84%; cutC in 76%; cntA in 60%) constituting, however, only a minor part of the total community (below 1% in most samples). Obtained cutC amplicons were associated with various taxa, in particular with several Clostridium XIVa strains and Eubacterium sp. strain AB3007, though a bulk of sequences displayed low nucleotide identities (on average 83% ± 8%) to references indicating that key human TMA producers are yet to be isolated. Co-occurrence analysis revealed three distinct groups governing the community structure of cutC-exhibiting taxa across samples. CntA amplicons displayed high identities (~99%) to Gammaproteobacteria derived references, primarily from E. coli. Metagenomic analysis of samples provided by the Human Microbiome Project (n=154) confirmed the abundance patterns as well as overall taxonomic compositions obtained with our assays, though at much lower resolution, whereas 16S rRNA analysis could not adequately uncover TMA-producing taxa. Conclusions: In this study, we developed a diagnostic framework that enabled the quantification and comprehensive characterization of the TMA-producing potential in human fecal samples. The key players were identified and together with predictions on their environmental niches using functional genomics on most closely related reference strains we provide crucial information towards the development of specific treatment strategies to restrain TMA producers and limit their proliferation.