Research ArticleMICROBIAL ECOLOGY

The microbiome of uncontacted Amerindians

Science Advances  17 Apr 2015:
Vol. 1, no. 3, e1500183
DOI: 10.1126/sciadv.1500183
  • Fig. 1 Microbiota diversity in fecal, oral, and skin samples from uncontacted Yanomami in relation to other human groups.

    (A) Faith’s phylogenetic diversity (PD) (average ± SD) of fecal samples from Yanomami and Guahibo Amerindians, Malawians, and U.S. subjects. OTU tables rarefied at 5000 sequences per sample. Interpopulation differences were significant (P < 0.001, ANOVA with Tukey’s HSD) for all but Guahibo-Malawi comparison (P = 0.73). (B) PCoA plot based on UniFrac distances calculated on the OTU table of fecal samples rarefied at 5000 sequences per sample. (C) Top discriminative bacteria among populations in fecal samples as determined by linear discriminant analysis (LDA) effect size (LEfSe) analysis. (D) Normalized prevalence/abundance curves for all OTUs found at 1% abundance or more in fecal samples. (E) Faith’s phylogenetic diversity (average ± SD) of oral samples from Yanomami and U.S. subjects. OTU tables rarefied at 1500 sequences per sample. Interpopulation differences were not significant (P = 0.296, ANOVA with Tukey’s HSD). (F) PCoA plot based on UniFrac distances calculated on OTU tables of oral samples rarefied at 1500 sequences per sample. (G) Top discriminative bacteria among populations in oral samples as determined by LEfSe analysis. (H) Normalized prevalence/abundance curves for all OTUs found at 1% abundance or more in oral samples. (I) Faith’s phylogenetic diversity (average ± SD) of skin samples from Yanomami and U.S. subjects. OTU tables rarefied at 1500 sequences per sample. Interpopulation differences were significant (P < 0.001, ANOVA with Tukey’s HSD). (J) PCoA plot based on UniFrac distances calculated on OTU tables of skin samples rarefied at 1500 sequences per sample. (K) Top discriminative bacteria among populations in skin samples as determined by LEfSe analysis. (L) Normalized prevalence/abundance curves for all OTUs found at 1% abundance or more in skin samples.

  • Fig. 2 Metagenomic diversity of fecal and oral samples from uncontacted Yanomami in relation to other human groups.

    (A) Functional diversity (average ± SD) measured by the observed number of KEGG orthologs in fecal samples of Yanomami and Guahibo Amerindians, Malawians, and U.S. subjects. Metagenomic tables rarefied at 1 million sequences per sample. Interpopulation differences were significant for all comparisons (P < 0.001, ANOVA and Tukey’s HSD). (B) PCoA plot based on Bray-Curtis distances calculated on the metagenomic table of fecal samples rarefied at 1 million sequences per sample. (C) Top discriminative metabolic pathways in Yanomami fecal samples as determined by STAMP; pathways ranked by effect size (η2). (D) Normalized prevalence/abundance curves for all KOs found at 1% abundance or more in fecal samples. (E) Functional diversity (average ± SD) measured by the observed number of KEGG orthologs in oral samples of Yanomami and Guahibo Amerindians, Malawians, and U.S. subjects. Metagenomic tables rarefied at 900,000 sequences per sample. Interpopulation differences were not significant (P = 0.07, t test). (F) PCoA plot based on Bray-Curtis distances calculated on the metagenomic table of oral samples rarefied at 900,000 sequences per sample. (G) Top discriminative metabolic pathways in Yanomami oral samples as determined by STAMP; pathways ranked by effect size (η2). (H) Normalized prevalence/abundance curves for all KOs found at 1% abundance or more in oral samples. (I) Functional diversity (average ± SD) measured by the observed number of KEGG orthologs in skin samples of Yanomami and U.S. subjects. Metagenomic tables rarefied at 1 million sequences per sample. Interpopulation differences were significant for all comparisons (P < 0.001, ANOVA and Tukey’s HSD). (J) PCoA plot based on Bray-Curtis distances calculated on the metagenomic table of skin samples rarefied at 1 million sequences per sample. (K) Top discriminative metabolic pathways in Yanomami skin samples as determined by STAMP; pathways ranked by effect size (η2). (L) Normalized prevalence/abundance curves for all KOs found at 1% abundance or more in skin samples.

  • Fig. 3 Representative AR genes captured through functional metagenomic selection of Amerindian oral and fecal microbiota.

    (A) Maximum likelihood (ML) tree of ceftazidime-resistant PBPs from Amerindian oral microbiota (blue) and their top blastx hits in NCBI nr (black). (B) Identity (%) of the 28 AR genes to their top hits in HMP and MetaHIT (blastn). Fourteen aligned to MetaHIT, all isolated from fecal microbiota. (C) Chloramphenicol acetyltransferase (cat) from O23_CH_21 and homologs in NCBI nt (nucleotide database). Dashed lines indicate 99% identical sequences. Dashes indicate omitted sequence (8487..25716). (D) tetW from F6_TE_1 and homologs in NCBI nt. Dashed lines indicate 98 to 99% identical sequences. Sequences are to scale. Yellow, AR gene; red, other resistance genes; green, mobile genetic elements; purple, bacteriophage genes; blue, other.

  • Fig. 4 Abundance of functionally selected AR genes in Amerindian fecal and oral metagenomes.

    The heatmap displays the abundances in RPKM (reads per 1-kb gene sequence per million reads mapped) of functionally selected AR genes in the fecal and oral metagenomes of Amerindians. The AR genes were functionally selected from the oral and fecal microbiota of the Yanomami individuals and matched Puerto Rican controls. The shotgun-sequenced metagenomes are from the Yanomami villagers, including individuals not investigated with functional metagenomic selections. Shotgun-sequenced metagenomes were clustered by UPGMA (unweighted pair group method with arithmetic mean) on the AR gene abundance profiles. Fecal and oral microbiota cluster apart.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/1/3/e1500183/DC1

    Text

    Table S1. Samples obtained from the 34 Amerindian subjects included in this study: skin from right (BD) or left (BI) forearm (n = 28), oral (n = 28), and fecal (n = 12) specimens.

    Table S2. Composition of Amerindian E. coli isolate genomic libraries.

    Table S3. Functional capture of AR genes from Amerindian E. coli isolate genomic libraries.

    Table S4. AR genes identified from functional selections of Amerindian E. coli isolate genomic libraries and Amerindian and Puerto Rican metagenomic libraries.

    Table S5. Composition of metagenomic libraries from Amerindian and Puerto Rican fecal and oral microbiota.

    Table S6. Antibiotics used in genomic and metagenomic library selections for AR genes.

    Table S7. List of all taxa significantly associated with each population in fecal samples as determined by LEfSe analysis.

    Table S8. List of all taxa significantly associated with each population in oral samples as determined by LEfSe analysis.

    Table S9. List of all taxa significantly associated with each population in skin samples as determined by LEfSe analysis.

    Table S10. List of clusters at different k-mer and minimum contig length and their associated taxonomy.

    Table S11. Taxa of 329 bacterial strains cultured from feces of 12 isolated Amerindians.

    Table S12. Typing of 24 E. coli strains from 11 isolated Amerindians.

    Fig. S1. Meta-analysis of 16S V4 region fecal data.

    Fig. S2. Microbiome diversity between different human groups.

    Fig. S3. Microbiota diversity in fecal, oral, and skin samples from uncontacted Yanomami in relation to U.S. subjects using the V2 region of the 16S rRNA gene.

    Fig. S4. Taxonomic distribution across gut samples sorted by the dominant genera within each population.

    Fig. S5. Prevalence/abundance curves of representative bacterial OTUs in feces (A), oral (B), and skin (C).

    Fig. S6. ClonalFrame analysis of Escherichia coli strains based on the sequence of seven housekeeping genes.

    Fig. S7. PCoA plot of Yanomami, Malawian, and Guahibo fecal samples.

    Fig. S8. Fecal microbiome of non-Bacteroides OTUs in Yanomami, Malawian,Guahibo, and U.S. samples.

    Fig. S9. Bacterial diversity as a function of age and body site.

    Fig. S10. Procrustes analyses of shotgun metagenomic and 16S rRNA data for different subsets of samples.

    Fig. S11. Comparison of PICRUSt versus sequenced shotgun metagenomes of fecal and oral samples with at least 10,000 mapped reads.

    Fig. S12. Bacterial genome assembly.

    References (7276)

  • Supplementary Materials

    This PDF file includes:

    • Text
    • Table S1. Samples obtained from the 34 Amerindian subjects included in this study: skin from right (BD) or left (BI) forearm (n = 28), oral (n = 28), and fecal (n = 12) specimens.
    • Table S2. Composition of Amerindian E. coli isolate genomic libraries.
    • Table S3. Functional capture of AR genes from Amerindian E. coli isolate genomic libraries.
    • Table S4. AR genes identified from functional selections of Amerindian E. coli isolate genomic libraries and Amerindian and Puerto Rican metagenomic libraries.
    • Table S5. Composition of metagenomic libraries from Amerindian and Puerto Rican fecal and oral microbiota.
    • Table S6. Antibiotics used in genomic and metagenomic library selections for AR genes.
    • Table S10. List of clusters at different k-mer and minimum contig length and their associated taxonomy.
    • Table S11. Taxa of 329 bacterial strains cultured from feces of 12 isolated Amerindians.
    • Table S12. Typing of 24 E. coli strains from 11 isolated Amerindians.
    • Fig. S1. Meta-analysis of 16S V4 region fecal data.
    • Fig. S2. Microbiome diversity between different human groups.
    • Fig. S3. Microbiota diversity in fecal, oral, and skin samples from uncontacted Yanomami in relation to U.S. subjects using the V2 region of the 16S rRNA gene.
    • Fig. S4. Taxonomic distribution across gut samples sorted by the dominant genera within each population.
    • Fig. S5. Prevalence/abundance curves of representative bacterial OTUs in feces (A), oral (B), and skin (C).
    • Fig. S6. ClonalFrame analysis of Escherichia coli strains based on the sequence of seven housekeeping genes.
    • Fig. S7. PCoA plot of Yanomami, Malawian, and Guahibo fecal samples.
    • Fig. S8. Fecal microbiome of non-Bacteroides OTUs in Yhotgun metagenomes of fecal and oral samples with at least 10,000 mapped reads.
    • anomami, Malawian, Guahibo, and U.S. samples.
    • Fig. S9. Bacterial diversity as a function of age and body site.
    • Fig. S10. Procrustes analyses of shotgun metagenomic and 16S rRNA data for different subsets of samples.
    • Fig. S11. Comparison of PICRUSt versus sequenced s
    • Fig. S12. Bacterial genome assembly.
    • References (72–76)

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    Other Supplementary Material for this manuscript includes the following:

    • Table S7 (Microsoft Excel format). List of all taxa significantly associated with each population in fecal samples as determined by LEfSe analysis.
    • Table S8 (Microsoft Excel format). List of all taxa significantly associated with each population in oral samples as determined by LEfSe analysis.
    • Table S9 (Microsoft Excel format). List of all taxa significantly associated with each population in skin samples as determined by LEfSe analysis.

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