Genetic specialization of key bifidobacterial phylotypes in multiple mother-infant dyad cohorts from geographically isolated populations

Abstract

Little has been known about symbiotic relationships and host specificity for symbionts in the human gut microbiome so far. Bifidobacteria are a paragon of the symbiotic bacteria biota in the human gut. In this study, we characterized the population genetic structure of three bifidobacteria species from 58 healthy mother-infant pairs of three ethnic groups in China, geographically isolated, by Rep-PCR, multi-locus sequence analysis (MLSA), and in vitro carbohydrate utilization. One hundred strains tested were incorporated into 50 sequence types (STs), of which 29 STs, 17 STs, and 4 STs belong to B. longum subsp. longum, B. breve, and B. animalis subsp. lactis, respectively. The conspecific strains from the same mother-child pair were genetically very similar, supporting the vertical transmission of Bifidobacterium phylotypes from mother to offspring. In particular, results based on allele profiles and phylogeny showed that B. longum subsp. longum and B. breve exhibited considerable intraspecies genetic heterogeneity across three ethnic groups, and strains were clustered into ethnicity-specific lineages. Yet almost all strains of B. animalis subsp. lactis were incorporated into the same phylogenetic clade, regardless of ethnic origin. Our findings support the hypothesis of co-evolution between human gut symbionts and their respective populations, which is closely linked to the lifestyle of specific bacterial lineages. Hence, the natural and evolutionary history of Bifidobacterium species would be an additional consideration when selecting bifidobacterial strains for industrial and therapeutic applications.

Keywords: Bifidobacterium; ethnicity; mother–infant dyad; multilocus sequence typing; probiotic.

Figure 1

Schematic of the sampling data and Bifidobacterial profiling of 58 mother–infant sample sets. (A) Schematic depicts the total number of mother fecal, breast milk, and corresponding infant fecal samples from whom data were available. MF, maternal feces; IF, infant feces; BM, breast milk. (B) The samples were collected in Hoten, Xinjiang (23.19°N, 115.38°E), Changjiang, Hainan (18.53°N, 109.17°E), and Wuwei, Gansu (36.31°N, 103.46°E) Province, P.R. China, respectively. Bar plot of bifidobacterium isolated from each mother–infant pair of three ethnic groups. (C) Uygur group, (D) Li group, and (E) Han group. The number on the horizontal axis represents the family number.

Figure 2

Heat map of bifidobacterial species that shared within mother–infant pairs. I, infant feces; M, mother feces; K, breast milk. I–M–K, bifidobacteria species appeared to be present in all infant feces, mother feces, and breast milk samples, and so on. Singleton, bifidobacteria species from only the mother or infant in each dyad. NA, no bifidobacteria species were obtained. The number on the horizontal axis represents the family number.

Figure 3

Comparison of MLST profiles of 100 bifidobacteria strains. (A) A diagram represents clonal relationships between STs was generated from goeBURST. (B) Minimum spanning tree of 100 Bifidobacterium strains based on MLST profiles according to the isolation source. Each filled circle corresponds to an ST. The color of the circle represents the source of isolation. Pink, Uygur; Green, Li; Yellow, Han; Purple, strains from two or more ethnic groups. Clonal complexes are represented by colored shaded areas.

Figure 4

Maximum-likelihood phylogenetic tree obtained from the concatenated nucleotide sequence of 100 Bifidobacterium strains. For the ML phylogenetic trees, MEGA X software was used with the GTRGAMMAIX model. The number of bootstrap replicates was 1,000 and visualized using ITOL. (A) B. longum subsp. longum, (B) B. breve, and (C) B. animalis subsp. lactis.

Figure 5

Recombination analysis of 100 strains of bifidobacteria. (A) Structure analysis. The plot shows one vertical line for each strain, and the length of the colored segments indicates the proportions of nucleotides from each of the three ancestral populations. (B) Combined split-decomposition analysis based on concatenated sequences of all 7 MLST loci.

Figure 6

Growth of Bifidobacteria isolates on polysaccharides. Isolates were cultured anaerobically at 37°C in an MRS medium supplemented with individual polysaccharides as the sole carbon source. OD₆₀₀ was measured at 0 and 48 h. Data indicate mean ± SD, and each condition was tested in duplicate. The heatmap shows the mean of the highest OD for each condition of each experiment. Bifidobacteria clonal complexes are indicated above the heatmap.