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Supplemental Information

Environmental distribution of the Lachnospiraceae.

A total of 25 16S rRNA gene surveys containing a total of 1,697 samples covering 17 different habitat classes were taxonomically profiled to identify the overall percentage of Lachnospiraceae. Boxplots outline the 25th, 50th and 75th percentiles of the data. The minimum, maximum and average (red box) percent abundance per sample of this family are also indicated. The number of samples per environment is listed beside habitat type and in Supplementary Table 1. Each GI tract-associated habitat is highlighted in bold.

DOI: 10.7287/peerj.preprints.168v1/supp-1

Grouping of genomes based upon counts of shared gene clusters.

Heatmap showing the number of gene clusters shared between genomes, inversely weighted by genome size. Genomes are clustered with intersecting cells between two genomes colored based on similarity ranging from low (red) to high (blue). The hierarchy of clustering is displayed along the side and top of the heat map with branches colored according to habitat (yellow = oral; red = sediment; green = rumen; blue = human GI tract). Names of gut-associated members predicted to be lacking butyric acid production are highlighted by an asterisk.

DOI: 10.7287/peerj.preprints.168v1/supp-2

Distribution of sporulation-associated genes within Lachnospiraceae genomes.

A range of sporulation genes was examined for each genome to assess the capabilities of producing endospores within each strain. Each gene is displayed as present (green) or absent (white) from each Lachnospiraceae genome. Organisms are clustered based upon their distribution of sporulation genes. Hierarchical clustering of genomes is displayed at the top of the grid with branches colored according to habitat (yellow = oral; red = sediment; green = rumen; blue = human GI tract). Gray lines separate sporulation genes into the broad categories listed on the right-hand side

DOI: 10.7287/peerj.preprints.168v1/supp-3

Relationships of 30 Lachnospiraceae genomes based on marker-gene and concatenated alignments.

Phylogenetic trees based upon the 16S ribosomal RNA gene (A) and the family-wide shared orthologs (B). Trees are rooted using two Ruminococcaceae as outgroup. Branches are colored based upon listed habitat (yellow = oral; red = sediment; green = rumen; blue = human GI tract). Bootstrap support values greater than 0.5 are displayed. Locations of putative gain and loss of functions are also shown on the trees. Stars mark the gain of butyric acid production capabilities (pink = butyrate kinase; orange = butyryl-CoA:acetate CoA-transferase). An alternative gain of butyrate kinase is marked with a pink X on the 16S tree (part A). Putative loss of sporulation capabilities is marked with a black bar. Strains classified as gut-restricted based upon shared gene clusters are underlined.

DOI: 10.7287/peerj.preprints.168v1/supp-4

Phylogenetic analysis of Lachnospiraceae-associated genes involved in the production of butyric acid and their associated 16S phylogenies.

The genes for butyryl-CoA:acetate CoA-transferase (A) and butyrate kinase (B) within Lachnospiraceae genomes were compared to 3,500 other prokaryotic genomes to find sources of potential LGT of these functions. Individual phylogenies were built using 16S sequences from genomes found to have the relevant butyrate-related gene and are displayed beside the BCoAT (A) and butyrate kinase (B) phylogenies. Lachnospiraceae members are highlighted in red.

DOI: 10.7287/peerj.preprints.168v1/supp-5

Sporulation-related sequences that differ in abundance between the human GI tract microbiome, and the cow rumen and human oral cavity.

The abundance of reads assigned to Lachnospiraceae-associated sporulation genes within metagenomic samples from the human gut microbiome were compared to those within the cow rumen and the human oral cavity using STAMP. This revealed several genes that were more abundant in the human GI tract (blue) compared to the rumen (green) or oral cavity (yellow). The mean proportions of assigned reads within each dataset are shown in addition to the difference of these proportions between datasets. The p-value from the Bonferroni-corrected two-sided Welch’s t-test is shown for each comparison.

DOI: 10.7287/peerj.preprints.168v1/supp-6

Maximum agreement forest between the 16S and shared gene cluster phylogenetic trees.

SPR operations were used to assess the congruence of phylogenetic trees based upon the 16S gene and the shared gene clusters of all analyzed genomes. The maximum agreement forest displays components that are in present in both trees. Branches are colored based upon listed habitat (yellow = oral; red = sediment; green = rumen; blue = human GI tract).

DOI: 10.7287/peerj.preprints.168v1/supp-7

Phylogenetic network of shared gene clusters based upon individual gene tree topologies.

The gene trees of 91 family-wide shared gene clusters were input to SplitsTree4 to construct an unrooted phylogenetic network that best represented all the individual relationships. Most gene trees were found to disagree, resulting in a star-like topology. Branch coloring is based upon listed habitat (yellow = oral; red = sediment; green = rumen; blue = human GI tract).

DOI: 10.7287/peerj.preprints.168v1/supp-8

Metagenomic samples utilized for environmental distribution analysis.

Multiple habitat types were tested for the presence of Lachno- spiraceae. Each habitat type is listed along with the MG-RAST ID of the project samples were retrieved from and the number of sample obtained from each project.

DOI: 10.7287/peerj.preprints.168v1/supp-9

Lachnospiraceae genomes

The designation, abbreviation used in this manuscript, NCBI taxon identification number, associated habitat according to IMG and source of for each genome utilized in this study is listed.

DOI: 10.7287/peerj.preprints.168v1/supp-10

Functions characterizing sub-groups of Lachnospiraceae.

Gene clusters present in over 90% of one group of Lachnospiraceae genomes and absent in over 90% of another were analyzed using Interproscan to determine their functions, as was the reverse. The general InterPro functional categories and GO terms, if any, for each gene cluster are listed.

DOI: 10.7287/peerj.preprints.168v1/supp-11

Shared features between members of the human gut Lachnospiraceae listed as gut-restricted/non-gut-restricted and those either possessing or lacking the capability to produce butyric acid.

Lachnospiraceae residing in the human GI tract were classified in 2 ways: those classed as gut-restricted or not based upon shared gene clusters (Fig. 2) and those classed based upon their capability to produce butyric acid or not (Table 1). Overlap of species assigned as one or the other within each classification was identified, as were functions associated with each classification.

DOI: 10.7287/peerj.preprints.168v1/supp-12

The distribution of butyric acid production genes.

The final stage of butyric acid production can be undertaken by 2 gene groups: butyrate kinase or butyryl-CoA:acetate CoA-transferase. The presence of each gene within a Lachnospiraceae genome is marked with a +.

DOI: 10.7287/peerj.preprints.168v1/supp-13

Additional Information

Competing Interests

We have no competing interests.

Author Contributions

Conor J. Meehan conceived and designed the experiments, performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper.

Robert G Beiko conceived and designed the experiments, analyzed the data, wrote the paper.

Grant Disclosures

The following grant information was disclosed by the authors:

CMF-108026

Funding

This work is supported by the Canadian Institute for Health Research (grant number CMF-108026). RGB acknowledges support from Genome Atlantic and the Canada Research Chairs program. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.


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