THE INTRA-FAMILY MICROBIAL DYNAMICS
By Naseer Sangwan, PhD
The gut microbiome (total microbial gene content existing in the gastrointestinal tract) has been established as a significant factor in the development of human health (1). Interaction between co-habitating individuals and their physical environment, e.g. shared physical space, has significant impact on their associated microbiome (2). Recent studies have shown that ‘nuclear families’ provides a unique framework to analyze how perturbations to one family member’s microbiome would impact the microbiome of others in the family. Here we present a brief summary of the microbiome analysis of a nuclear family with two children (juvenile female=12 years, juvenile male=8 years, adult female=42 years, adult male=44 years) who have distinct interpersonal health profiles. As an example, metadata highlighted that both juveniles suffered from seasonal allergies, while the juvenile male also suffered from eczema and food allergies to peanuts, lentils, soy and wheat. Neither adult family member suffered from these conditions. We performed whole genome shot gun metagenomic sequencing of fecal samples (one sample per individual) of all family members. Metagenome sequences were analyzed to characterize the taxonomic and functional potential of the gut microbiota using MetaPhlAn (3) and HUMAnN, respectively (4).
Figure 1. Intra-family microbial dynamics. (A) Meta data and total microbial gene content assembled across all family members. (B) Bargraph representing relative abundance of the signature genera in all family members. (C) Table presenting pairwise simialrity of microbiome samples measured by pairwise distance based on quantitative taxa abundance data. (D) Whole genome based synteny (order of similar genes) comparison of complete genome of A. muciniphila ATCC BAA-835 and Akkermansia Meta assembled across adult male microbiome.
Genus level analysis revealed quite a few bacteria that were common to all members, such as the genera Escherichia, Bacteroides, Eubacterium and Faecalibacterium (Figure 1B). High abundance of Bacteroides was clearly evident across all members (i.e. Adult male=40.8%, Adult female=24.5%, juvenile female=36.0%, juvenile male=34.6%). However, upon closer inspection it was revealed that a certain species of the genus Bacteroides, called Bacteroides intestinalis, was enriched in the adult male and female and juvenile female samples, where it comprised >5% of the community. However, in the juvenile male, this species was mostly absent, instead Bacteroides vulgatus was highly abundant (>5% of the community). Overall, the juvenile male’s microbiome was the most different (Figure 1C), and contained bacterial genomic sequences that are related to organisms that can produce butyrate (a short chain fatty acid; SCFA), such as the species Roseburia intestinalis, Bacteroides vulgatus and Ruminococcus bromii. This species specific SCFA production helps in developing tolerance acquisition, e.g. (i) Expansion of intraepithelial lymphocytes, (ii) lowers the pH, favoring the colonization of other commonsel anaerobes. It is possible that the juvenile male’s microbiome is responding to his allergies, as we have shown in previous studies (5) by increasing the abundance of taxa that can produce SCFAs. The adult male and female had genomic DNA related to Akkermansia municiphilia, which has been identified as being a negatively correlated with obesity (6). A. muciniphila has been shown to regulate human metabolism through increasing intestinal levels of endocannabinoids that control inflammation and gut peptide secretion. Using metagenome assembly and binning methods we compared the genome of Akkermansia municiphilia found in the microbiome of the adult male and female against genome sequenced strains in the public databases (specifically A. muciniphila ATCC BAA-835; (7); Fig. 1D). Th A. muciniphila species found in the adults had a different gene content and a different order for the same genes when compared to the known strains. This points to the individuality of each person’s microbiome; A. municiphilia in one person may be different to A. municiphilia in another!
We further analyzed the functional potential of each person’s microbiome, and determined how well conserved microbial function was between family members. We randomly picked and annotated 76,000 genes from the microbiome of each member, and mapped these genes to metabolic pathways. Most metabolic pathways were found in similar abundance in each family member. However, the juvenile male had a significant enrichment for lipid metabolism, which may be related to his diet. To achieve higher resolution we focused our analysis on determining the differences in enzyme potential between individuals, ignoring core metabolic functions such as energy metabolism, and DNA/RNA transcription and translation). Enzymes involved in Acetyl-CoA-based butyrate production were at very different abundances in each family member. The enzyme 3-hydroxy-3-methylglutaryl-CoA reductase and butyrate kinase were significant enriched (average mean proportion > 5%) in the microbiome of the juvenile male compared to the other family members. This could be related, as shown previously (Canini et al., 2015) to the juvenile male’s allergies (Figure 1A). Although, in the present study it is not possible to determine the mechanism by which seasonal and food allergy influence microbial composition.
We further quantified the ‘equivalent’ genes, i.e. orthologous genes, across all family members. Orthologs are defined as similar genes that are related through speciation from a single ancestral gene, not through gene duplication. Pairwise analysis revealed 12,512 genes as orthologous across all family members, and 19,069 genes in parents versus progeny analysis. In addition, 30,668 and 35,621 genes were characterized as orthologous across the pairwise analysis of parents and progeny samples, respectively. Our results indicate that the parents share similar gene content with each other more than they do to the children, and vice versa. Finally, we analyzed the microbial antibiotic resistance potential across all family members. The juvenile male showed the highest number of microbial antibiotic resistance genes (n = 170) in comparison to other family members, i.e. (Adult male = 117, adult female = 142, juvenile female=110). However, the importance of this observation is hard to define.
In summary, we demonstrated clear differences in the microbiome recovered from the members of a nuclear family with various aged children. Also, the juvenile male’s microbiome showed signatures that relate to his food allergies. We expect that further inclusion of data from more families in future studies would help to understand the role of microbiome in tolerance acquisition against food and seasonal allergies.
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Naseer Sangwan, PhD, is a post-doc in the Gilbert lab at the University of Chicago. His research is focused on understanding the food allergies-induced dysbiosis of intestinal microbiota using comparative population genomics of microbial gene-complements re-assembled across deeply sequenced metagenome datasets.