We assessed the subsistence-related variation of the human gut microbiome at a fine resolution for two of the main dimensions of microbiome variation, age and geography. For this, we investigated the fecal microbiome and metabolome in rural Bassa and urbanized individuals from Nigeria, including infants, and compared data with worldwide populations practicing varying subsistence. Our data highlight specific microbiome traits that are progressively lost with urbanization, such as the dominance of pristine fiber degraders and the low inter-individual variation. For the Bassa, this last feature is the result of their subsistence-related practices favoring microbial dispersal, such as their extensive environmental contact and the usage of untreated waters from the Usuma River. The high degree of microbial dispersal observed in the Bassa meta-community nullifies the differences between infant and adult intestinal ecosystems, suggesting that the infant-type microbiome in Western populations could be the result of microbiome-associated neotenic traits favored by urbanization.

In an attempt to bridge these gaps, here we characterized the fecal microbiota and metabolome of two Nigerian communities, the Bassa rural agriculturalists and urban individuals from four state capitals (Ilorin, Abeokuta, Ado Ekiti, and Ibadan) and the Nigerian capital city (Abuja), which also include infants aged <3 years. The Bassa are an agrarian community with limited contact with other populations, who live on a hill about 500 m away from the Chibiri village in Kuje Area Council (Abuja), where they moved from Kogi State (a distance of 160 km) about 100 years ago ( Figure 1 ). Their community comprises about 70–80 people who primarily eat what they grow on their farm, such as tubers, grains, fruit, and other small crops. The Bassa are thus a somewhat isolated group, but nonetheless maintain a self-sufficient rural horticultural subsistence. Microbial communities in the Usuma River, which is a daily feature in Bassa life, both for nourishment and physical exposure, were characterized as well. The urban dwellers recruited in our study were randomly selected from different ethnic groups, including Hausas, Igbos, Yorubas and Ebira, as representative of people geographically close to the Bassa but who are embracing a Western lifestyle. Compositional microbiome data and metabolome profiles from these populations were interpreted across subsistence strategies and age, and integrated with available data from worldwide populations, with varying degrees of traditional or urban lifeways. By exploring, at a finer geographic and age resolution than previous efforts, the variation of the human gut ecosystem along the transition from rural to urbanized communities, this study led to uncovering specific adaptive gradients, at both structural and functional scale.

However, most of the previous studies have focused on the taxonomic variation of the gut microbiota in adult populations living distinct lifestyles, thus leaving a number of unanswered questions, especially about potential subsistence-driven alterations in metabolic networks and the broad-scale application of this body of data toward informing about the infant gut microbiome alterations. Moreover, most studies comparing microbiomes from hunter-gatherers, rural agriculturalists, and urbanized communities have so far dealt with geographically and culturally distant populations, with obvious confounding factors, such as relatedness and local environment.

In recent years, we have witnessed a growing number of studies on the characterization of the human gut microbiome across the globe, in populations adhering to varying subsistence patterns, from more traditional to more urbanized (). In addition to providing valuable information on the specific adaptations of the gut microbiota to diet and other lifestyle factors, such studies have evolutionary relevance, as they recall ways of life that accompanied our history, from the hunting and gathering of our Paleolithic ancestors, to small-scale agriculture and permanent settlements of the Neolithic, to the post-industrial Westernized lifestyle. This body of literature has consistently illustrated distinctive signatures of the urbanization process in intestinal microbial communities, including reduced diversity, loss of bacterial taxa with fibrolytic specializations, and the appearance of microorganisms as a potential adaptive response to the changes in diet, environment, use of antibiotics, and hygiene practices, brought on by the modern lifestyle.

With regard to metabolite data, the Bassa and urban Nigerians cluster in distinct groups alongside either rural (hunter-gatherer) or urban individuals for which metabolomics data produced through the same analytical method are available (). Additionally, these cross-study populations cluster into distinct groups according to subsistence strategy and age (adonis: p = 1 × 10, R= 0.56; ANOSIM: p = 1 × 10, R = 0.30) ( Figure 7 D). In particular, the Bassa population separates from all the others, whereas urban Nigerians overlap with urban Italians and are similarly enriched in phenylalanine and tyrosine. Despite this, Nigerian urban individuals are still distinguishable from Italians by higher proportions of glycerophospholipids (Nigerians versus Italians, 1.73% versus 0.60%), sphingolipids (0.23% versus 0.07%), and acylcarnitines (0.50% versus 0.26%) (p ≤ 0.02, Wilcoxon rank-sum test). The same signatures emerge from the comparison between Bassa and Italians ( Figure S5 ). On the other hand, differences between the traditional subsistence groups, the Bassa and the Hadza, include enrichment in biogenic amines (Bassa versus Hadza, 2.75% versus 2.50%) and amino acids (73.61% versus 45.12%) for Bassa, and significant enrichment in hexoses (21.13% versus 50.54%) for Hadza (p ≤ 0.005). The same differences are seen by comparing the Hadza with Bassa adults, but not with infants, whose metabolic profiles are overall more similar to those of the Hadza, with hexoses as main discriminant metabolites ( Figure 7 D; Figure S5 ).

To further explore the variation of the gut microbiome across populations with different geographic origin and lifeway, the genus-level abundance profiles of the Bassa and urbanite Nigerians were compared with those from two previous works, dealing with a hunter-gatherer community (Hadza from Tanzania), rural agricultural communities from Malawi and Amazonas State of Venezuela, and urban-industrialized societies (from Italy and the USA) (). As expected, the Bray-Curtis distances between the Bassa and the other traditionally living rural populations are lower than those between Bassa and urban groups ( Figure 7 A). Alternatively, the Bray-Curtis distances between urban Nigerians and the other groups are overall similar. A source tracking analysis using the previously published datasets as source populations confirmed that rural sources account for the vast majority (mean, 81%) of the Bassa microbiome, with the hunter-gatherer source making a substantial contribution (up to 59%) in eight individuals ( Figures 7 B and 7C). Compared with adults, Bassa infants have a greater contribution from rural (84% versus adults, 78%) as well as hunter-gatherer (15% versus 11%) sources, with a corresponding decrease in unknown sources (0.8% versus 11%). Supporting the inference that the urban Nigerian population represents a middle ground between a rural-traditional and fully urban-industrial lifestyle, the Nigerian urban profiles generally have a higher contribution from rural (mean, 57%) than urban sources (32%) ( Figures 7 B and 7C). It is worth noting that the rural source contributions decrease from infants (66%) to adults (50%), parallel to the increase in the urban source proportion (infants, 29%; adults, 34%). The hunter-gatherer source contributes 12%–88% in six urban individuals (five adults and one infant).

(D) PCA of Euclidean distances between the metabolic profiles of the study populations (same color code as the horizontal bars in C), as well as Hadza (yellow) and urban Italians (blue) (), as assessed using a semi-untargeted metabolomics approach. Ellipses include 99% confidence area based on the SE of the weighted average of sample coordinates and are colored by population (salmon, Bassa; green, urbanite Nigerians; yellow, Hadza; light blue, Italians). p = 1 × 10, adonis. The main discriminant metabolites are mapped on the plot.

(B and C) Bayesian source-tracking analysis. Source contributions, estimated using SourceTracker (), are shown averaged within Bassa or urban Nigerians (B), and for individual samples (C). Bars below the histograms are colored by population and age (orange, Bassa infants; red, Bassa adults; green, urban infants; olive green, urban adults). Hunter-gatherer, rural agricultural, urban, and unknown sources are colored in dark blue, light blue, brick red, and gray, respectively (as in).

(A) Bray-Curtis distances (means ± SD) between genus-level microbiota profiles. Publicly available sequences from Hadza hunter-gatherers and urban Italian adults (), rural agriculturalists from Malawi and Amazonas State of Venezuela, and urban US adults () were used.

The pattern of metabolome variation between populations was further explored through a CAG analysis, leading to the identification of three CAGs of correlated metabolites ( Figure 6 C; Figure S6 ). According to our findings, the alanine CAG (brownish), including most of the amino acids, is exclusively present in the fecal metabolome of adults, but far more represented in urban compared with rural individuals. Conversely, the hexoses CAG (ivory), also including several glycerophospholipids, some acylcarnitines, along with most of the biogenic amines, is more represented in the infant metabolome, but far more in rural than urban. The threonine CAG (purple) is variously represented in all study groups except in Bassa infants. It is worth noting the Bassa-specific age-independent co-abundance of asparagine, histamine, and acetylornithine, with the first being widely distributed in the plant kingdom, the second derived by enzymatic decarboxylation of histidine, and the last available in several “rural-like” foods used by Bassa ( http://www.hmdb.ca/ http://foodb.ca/ ). At the same time, it is interesting to note the absence of histamine (with the abundance of histidine), as well as that of acetylornithine and ornithine (with the abundance of putrescine, a product of ornithine decarboxylation), in the CAGs of urban infants, suggesting the establishment of alternate co-metabolisms in early life. Contrarily to Bassa, no overlap between infant and adult CAGs is observed for the urban population, consistent with the Western-type age-related differentiation of microbiota profiles.

Along PC1, which accounted for nearly all of the variation (81.4%), the infant groups segregate from the respective adult groups, with the metabolomics profiles of the latter located at positive values of the axis (p ≤ 0.04, Wilcoxon rank-sum test). Microbial taxa that positively correlate to PC1 (i.e., to the intra-group segregation by metabolites) include Oscillospira and unclassified Christensenellaceae (p ≤ 3 × 10, Kendall tau correlation test) ( Figure 6 B; Figure S4 ). Regardless of population, adult individuals are characterized by a greater presence of amino acids (infants versus adults: Bassa, 54.75% versus 83.03%; urban, 73.67% versus 85.82%; p ≤ 0.04, Wilcoxon rank-sum test) ( Figure S5 ). When looking at individual metabolites, alanine is the main discriminant amino acid of Bassa adults, while phenylalanine, tyrosine, and branched-chain amino acids (BCAAs; leucine, isoleucine, valine) are far more represented in urban adults. With respect to infants, hexoses are largely dominant in the Bassa, while taurine is greatly overrepresented in the urban infant metabolomes ( Figure 6 B). Compared with infants, urban adults also have different proportions of glycerophospholipids (infants versus adults, 1.84% versus 1.67%), sphingolipids (0.25% versus 0.22%), biogenic amines (5.34% versus 2.53%), and hexoses (18.36% versus 9.28%) (p ≤ 0.04). Aside from the previously described amino acid category, the Bassa adult metabolome significantly differs from that of infants by a slightly diminished abundance of acylcarnitines (infants versus adults, 0.88% versus 0.35%; p = 0.02) ( Figure S5 ).

Principal component analysis (PCA) of the relative abundance profiles of 185 key metabolites from core metabolic pathways, as detected through a semi-untargeted metabolomics approach ( Table S3 ), shows separation between Bassa (3 infants, 6 adults) and urban populations (7 infants, 14 adults), which is particularly evident along PC2 (adonis: p = 0.04, R= 0.11; ANOSIM: p = 0.03, R = 0.21) ( Figure 6 B). Procrustes analysis of Euclidean distances between metabolomes and weighted or unweighted UniFrac distances highlights significant association between the microbiota taxonomic and metabolic profiles across the entire cohort (for weighted UniFrac: m12 squared = 0.78, correlation value = 0.47; for unweighted UniFrac: m12 squared = 0.55, correlation value = 0.67; p ≤ 0.002, PROTEST) ( Figure S3 ). As expected, PC2 coordinate-related genera are among those previously identified as discriminatory for the Bassa from the above taxonomic findings. In particular, Prevotella, [Prevotella], and Succinivibrio are positively correlated to the axis, whereas Bifidobacterium, Bacteroides, Blautia, and unclassified Lachnospiraceae and Erysipelotrichaceae correlate with negative values of PC2 coordinates (p ≤ 0.007, Kendall tau correlation test) ( Figure S4 ). According to a Random Forests analysis (), seven metabolites are highly discriminatory between Bassa and urban individuals, of which five are over-represented in the former (histamine, alanine, lysoPC a C16:1, lysoPC a C18:1, and C10:2) and two in the latter (serotonin and tyrosine).

Gas chromatograph-mass spectrometry (GC-MS) analysis of SCFAs revealed a different profile for Bassa (3 infants and 8 adults) compared with urban individuals (8 infants and 15 adults). Bassa are significantly enriched in propionate (relative abundance, Bassa versus urban settlers, 21.86% versus 12.53%) and proportionally depleted in acetate (65.22% versus 78.07%) (p ≤ 0.005, Wilcoxon rank-sum test). These signatures are shared by both infant and adult groups within the urban population, while the Bassa exhibit varying SCFA profiles depending on the age group. Specifically, compared with adults, Bassa infants show a greater abundance of acetate and valerate (p = 0.04) ( Figures 6 A).

(C) Wiggum plots indicate pattern of variation of the three identified metabolic co-abundance groups (CAGs) in Bassa (top) and urban Nigerians (bottom), both in infants (left) and in adults (right). Each node represents a metabolite, and its dimension is proportional to the over-abundance relative to background. Connections between nodes indicate positive and significant Kendall correlations between metabolites (p < 0.05). Only metabolites with ≥0.1% relative abundance in at least two subjects were considered.

(B) PCA of Euclidean distances between the metabolic profiles of the study populations, assessed using a semi-untargeted metabolomics approach (). p = 0.04, adonis. The main discriminant metabolites are mapped on the plot. Genera of the gut microbiota significantly correlated to PC1 and PC2 (p < 0.05, Kendall tau correlation test) are displayed at the bottom and on the right, respectively. BA, Bassa adults (red); BI, Bassa infants (orange); UA, urban adults (olive green); UI, urban infants (green).

We also examined the microbial community in water samples from the Usuma River, which is a daily feature, both in nourishment and physical exposure, in Bassa life. Consistent with previous findings on other African rivers (), the dominant phyla in the Usuma are Proteobacteria (relative abundance, 37.13%) and Firmicutes (15.11%), with Acidobacteria (13.54%), Chloroflexi (5.26%), Bacteroidetes (5.04%), Planctomycetes (4.86%), and Actinobacteria (4.51%) as other major groups ( Figure 5 A). Within Proteobacteria, the Betaproteobacteria members (mainly Comamonadaceae and Oxalobacteraceae families, and unknown taxa in the SC-I-84 and Ellin6067 orders) dominate the ecosystem, followed by Alphaproteobacteria (mainly Sphingomonadaceae, Rhodospirillaceae, and Hyphomicrobiaceae), Gammaproteobacteria (with Coxiellaceae as the most abundant family), and Deltaproteobacteria (with an unknown Myxococcales family as the most represented) members ( Figure 5 B). Interestingly, among the most abundant (>1%) genera, we could recognize typical commensals of the human gut microbiota belonging to the Clostridiales order and specifically to the Lachnospiraceae and Ruminococcaceae families, such as Oscillospira and Blautia (cumulative relative abundance, 10.62%) ( Figure 5 C). It should be pointed out that during the wet season, when sampling took place, the rain waters wash down the surrounding hill-lands into the river, including the bush and scrub land near the Bassa settlement where Bassa usually defecate. Therefore, the high presence of human-associated commensal microorganisms in river waters could be the consequence of fecal pollution from anthropogenic sources, as well as other wild fauna. These observations are supported by recent work demonstrating that Ruminococcaceae and especially Lachnospiraceae bacteria could serve as human fecal signatures, as an alternative to traditional indicators based on enterococci and enterobacteria ().

The intestinal microbiota structure of Bassa and urban settlers was further analyzed through the determination of co-abundance groups (CAGs) (). Four CAGs were identified based on pairwise Kendall correlation of taxa from the genus-level summarized taxa table, and each one was named according to the most abundant taxon ( Figure 4 Figure S2 ). Interestingly, the Faecalibacterium CAG (yellow), showing the co-abundance of several Western-like short-chain fatty acid (SCFA) producers (including the butyrate-producing Faecalibacterium and Coprococcus, and the acetate-producing Blautia, Bacteroides, Parabacteroides, Bifidobacterium, and Lachnospira) (), is absent in the Bassa microbiota, regardless of age, suggesting a different ecological layout supporting SCFA production in the intestinal ecosystem of study populations. On the other hand, the Prevotella CAG (light blue) is far more represented in Bassa rather than urban individuals and shows the Bassa-specific co-abundance of bacteria with xylan- and/or starch-degrading and succinogenic capabilities, such as Prevotella and Succinivibrio (), and Phascolarctobacterium, an asaccharolytic succinate-utilizing and propionate-producing bacterium (), suggesting the establishment of cross-feeding mechanisms leading to propionate production. With regard to the transition from infants to adults, population-specific inter- and intra-CAG rearrangements are observed. In particular, for the urban community, changes in the proportions of common Western SCFA producers and other commensals accompany the shift from the infant to the adult group, along with an increasing representation of the Roseburia CAG (dark blue). Alternatively, the Roseburia CAG decreases in representation among the Bassa from infants to adults. Furthermore, in the transition to Bassa adults, a shuffling of the Dialister CAG (pink) is observed, along with increased proportions of Turicibacter, Streptococcus, and [Eubacterium] in the Prevotella CAG.

(A–D) Wiggum plots indicate pattern of variation of the four identified CAGs in Bassa (top) and urban Nigerians (bottom), both in infants (A and C) and in adults (B and D). CAGs were named according to the most abundant genera, as follows: Faecalibacterium (yellow), Dialister (pink), Roseburia (dark blue), and Prevotella (light blue). Each node represents a genus, and its dimension is proportional to the over-abundance relative to background. Connections between nodes indicate positive and significant Kendall correlations between genera (p < 0.05). Line thickness is proportional to correlation strength. Only genera with ≥0.1% relative abundance in at least 30% of subjects were considered. Asterisk ( ∗ ) indicates unclassified OTU reported at higher taxonomic level.

At phylum level, Firmicutes and Bacteroidetes dominate the gut microbiota of rural and urban populations ( Figure 3 A), but with different ratios (Firmicutes-to-Bacteroidetes ratio: Bassa versus urban, 1.37 versus 2.85; p = 3 × 10). Moreover, the Bassa microbial communities are comparatively enriched in Spirochaetes (relative abundance, Bassa versus urban, 1.54% versus 0.18%) and Fusobacteria (0.94% versus 0.01%), while depleted in Actinobacteria (1.13% versus 3.87%) (p ≤ 0.005). The main discriminant genera are members of the Lachnospiraceae and Ruminococcaceae families, such as Blautia, Coprococcus, Lachnospira, Faecalibacterium, and Oscillospira, which are less abundant in the microbiota of Bassa compared with urban individuals (p ≤ 0.03) ( Figure 3 B). The Bassa are also depleted in other common commensal inhabitants of the Western gut, including Bacteroides and Bifidobacterium (p ≤ 4 × 10). As expected based on previous reports for non-Western populations (), the Bassa microbiome is enriched in Prevotella and other Bacteroidales members (including [Prevotella] and an unknown S24-7 genus), as well as in Bulleidia, [Eubacterium], Cetobacterium, Succinivibrio, and unclassified Peptostreptococcaceae. Furthermore, in accordance with available data on traditional populations (), Phascolarctobacterium and Treponema are more abundant in the microbial ecosystem of Bassa compared with the urban community (p ≤ 0.03). Notably, Ruminobacter and Butyrivibrio are exclusively present in the Bassa microbiota, whereas Megamonas is only detected in urban individuals ( Figure 3 C). The relative abundance of several taxa increase with age within the urban cohort, including Oscillospira, Clostridium, Odoribacter, and unknown genera in the Clostridiales order and in the families Christensenellaceae, Rikenellaceae, and [Barnesiellaceae] (p < 0.05, Spearman’s correlation test). Otherwise, no differences are found between the two Bassa age groups at the various taxonomic levels. However, the exact age of Bassa is uncertain, which prevented us from assessing the precise contribution of age to microbial variation. Likewise, sex was not recorded for participating Bassa individuals, making us blind to possible sex-related differences.

(B) Log2 fold changes of the main discriminant genera between Bassa and urban Nigerians (p < 0.05, Wilcoxon rank-sum test). Genera with ≥0.5% of mean relative abundance in at least one population were considered. When the difference was significant only between adults or infants, the bar was colored according to the population with the highest relative abundance of that genus (same color code as in A). Asterisk ( ∗ ) indicates unclassified OTU reported at higher taxonomic level.

UniFrac (weighted and unweighted) and Bray-Curtis distance ordination show separation between Bassa and urban individuals ( Figures 2 C and 2D; Figure S1 Table S2 ). No clustering is observed within the populations between infants and adults, except for the unweighted UniFrac analysis, which shows barely separable clusters by age group among urban individuals (adonis: p = 0.03, R= 0.03; ANOSIM: p = 0.02, R = 0.14). Regardless of the metrics used, lower interpersonal variation is observed in the microbiota structure of Bassa compared with urban individuals (p ≤ 0.001, Wilcoxon rank-sum test). With regard to age, greater inter-individual variation is found among urban infants compared with urban adults, according to unweighted UniFrac metrics (p = 3 × 10) ( Figure 2 E). The opposite is true for the Bassa based on weighted UniFrac distances, with a lower inter-individual variation among infants than among adults (p = 0.009), as well as compared with urban groups (p ≤ 3 × 10) ( Figure 2 F). The analysis based on Bray-Curtis dissimilarities confirms the lower beta diversity for Bassa infants compared with urban groups ( Figure S1 ).

According to common diversity indices, similar alpha diversity is seen regardless of community membership and age (p > 0.05, Wilcoxon rank-sum test) ( Figures 2 A and 2B ; Figure S1 ). However, it should be noted that the observed number of operational taxonomic units (OTUs) in the microbiota of Bassa infants (mean, 758) is significantly higher compared with urban adults (651; p = 0.03) and tends to be greater than Bassa adults (636; p = 0.08) and urban infants (615; p = 0.2).

(C–F) Beta diversity, based on unweighted (C and E) and weighted (D and F) UniFrac distances, and PCoA plots. A significant separation between Bassa and urbanized Nigerians was found (p < 0.05, adonis). Different letters in the bar plots (means ± SD) indicate significant differences (p < 0.05, Kruskal-Wallis test). Same color code as in (A).

The gut microbiota structure from 18 Bassa (9 infants aged <3 years and 9 individuals aged 3–60 years, hereafter referred to as adults, according to previous evidence on the establishment of the typical adult-like microbiota profile;) and 30 urban Nigerians (12 infants aged <3 years; 18 adults aged 5–75 years) was profiled through 16S rRNA gene sequencing of stool samples. A total of 3,346,376 high-quality reads (mean, 69,716; range, 8,480–133,340) were produced and analyzed.

The diet of the urban dwellers was a mixture of traditional Nigerian foods and Westernized dietary items, with only 9 subjects (out of 30) declaring to follow a purely Nigerian diet. Breakfast was generally based on cereals, tea, bread, and pap, while lunch consisted of cooked rice, beans, pastries, and fruit, and dinner usually included foods made from common African tuber crops (e.g., iyan, eba, fufu, tuwo), eaten with spicy soup. In contrast to the Bassa, the urban infants were on average exclusively breastfed for the first 3 months of life (during the working mother national maternity leave) before the introduction of cereals, both traditional (e.g., koko or ogi) and more processed (e.g., flakes, oats, and commercial breakfast cereals). Enrolled urban Nigerians mostly (73%) belonged to the middle class, were educated, had a high level of hygiene and access to potable water, and made a moderate use of antibiotics. Only 5 infants out of the 12 enrolled (and none of the adults) came from the lower class, were exposed to low standards of hygiene, and were usually given natural remedies rather than antibiotics. See Table S1 for a short description of local ingredients and foods.

The Bassa diet mostly consists of foods derived from their farming activities, such as tubers (yams and cassava), grains (guinea corn, millet, and maize), fruit (banana and mango), and soup condiments (okra and melon). Grains are processed into a flour used to make the traditional stiff porridge “tuwo,” and tubers are used to make solid foods (e.g., sokora). The Bassa regularly eat soups (e.g., ayoyo from Corchorus leaves and kuka from leaves of Adansonia digitata) and fish, given their proximity to the river Usuma. They rear goats and chickens that they sometimes sell to farmers who cross over to their land, but they rarely eat meat, except during festival periods. Breakfast usually consists of koko (or pap), made from the fermentation of maize, millet, and sorghum into a thick paste with hot water. The Usuma River serves as their main water source for drinking and cooking because they have no access to potable water. The river water is not heated or subjected to any form of treatment. The Bassa also bathe in the Usuma and cross it on foot, when it is not flooded, to get to neighboring communities and to the nearest school (attended by very few children). The Bassa breastfeed their infants up to 7–12 months, but they do not practice exclusive breastfeeding. As soon as the baby is able to handle other foods, they supplement breast milk with liquid and semi-liquid foods (e.g., koko, kunu, a popular drink consumed in Northern Nigeria, made from cereal fermentation, and dame, a paste obtained by dissolving in water 24-hour rested tuwo); then solid food is gradually introduced. Antibiotic use is infrequent, and they still rely on local remedies for health needs, favoring herbs to modern medicines, because there is no health care center nearby.

Discussion

In this study, we demonstrate that two human communities living in a geographically proximate region in Nigeria follow a predictive pattern of dissimilarity in taxonomic and metabolic traits of the gut microbiome that mirror the traditional and/or rural versus urban and/or industrialized subsistence dichotomy. Importantly, these results have allowed us to witness specific traits that indicate a progressive adaption of the intestinal microbial ecosystem toward urbanization.

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Sonnenburg J.L. Starving our microbial self: the deleterious consequences of a diet deficient in microbiota-accessible carbohydrates. According to the metabolomics results, the Bassa, especially infants, show an overall healthy profile with greater proportions of hexoses and fewer amounts of amino acids and biogenic amines compared with urban individuals. As previously discussed (), the abundance of hexoses may be indicative of a diet high in microbiota-accessible carbohydrates (), as that of the Bassa, heavily based on tubers, grains, and derived processed foods, as well as a variety of leafy soups, with the microbiota-dependent release of monosaccharides probably exceeding the enteric nutritional demands and thus excreted in the feces. With specific regard to Bassa infants, hexoses may also result from the digestion of the sugary and starchy liquid or semisolid foods they are fed with during weaning. On the other hand, the smaller amounts of amino acids and derivatives in Bassa feces may reflect less protein consumption compared with urban Nigerians and/or altered metabolisms or absorption.

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Glew R.H. The amino acid and mineral content of Baobab (Adansonia digitata L.) leaves. When focusing on age, our compositional data on urban infants corroborated what is well-known in developed countries, i.e., that the gut microbiota of infants aged <3 years is unstable, with high inter-individual diversity and a taxonomic structure progressively approaching the more complex and stable adult-type microbiota (). Conversely, different and at times opposite features were observed for the intestinal microbial ecosystem of Bassa infants that, compared with the adult counterparts, showed high biodiversity, lower inter-individual variability, and no difference at the various taxonomic levels. This in turn brought about a finding that lends clues to a lingering question from the work of, which is whether bifidobacteria are indeed absent in the kinetics of assembly and development of traditional microbiotas. The data we present here of pre- and peri-weaned Bassa infants confirms that bifidobacteria, which are undoubtedly beneficial for Western human populations, are missing from certain traditional population infant guts. Collectively, these data are in contrast with the work of, which identified distinctive microbiome features in early childhood in rural populations, including greater inter-individual variation among children than adults, an increasing biodiversity with age, and the dominance of Bifidobacterium. While we are aware that the much smaller sample size of our study could have led us to overestimate our results, we believe that our data merit continued consideration of the hypothesis that in a non-Westernized context with vastly different environmental conditions, socio-economic structures, subsistence practices, and external contact with other populations, assumptions about microbiome traits derived from studies on Western populations lead interpretations about human-microbe associations astray. From an ecological standpoint, it is possible to speculate that the extensive sharing of life within the Bassa community (in terms of lifestyle habits, contact with the environment, and usage of untreated river water, which indeed shows traces of microbiota components) results in a high degree of microbial dispersal, thus allowing the human microbiome to behave as a meta-community (). In turn, the establishment of a meta-community, a feature probably common to traditional populations, has the potential to nullify the differences between infant and adult microbiomes, as mainly observed in Western populations, significantly shortening the microbiota assembly process. On the contrary, along with Westernization (involving sanitization, water treatment and other hygienic practices, and reduced life sharing with dispersal limitation), the human microbiome has lost its meta-community feature, resulting in increased individuality, and consequently driving the differentiation of the infant-type microbiome, as well as the trajectory of microbiome assembly, typical of Western populations. Indeed, the acquisition or, more likely, the extension of an infant-type microbiota in modern populations could be the result of neoteny in human evolution, favored by the establishment of profound differences in diet and lifestyle between infants and adults in modern societies. Further supporting this, also metabolic data suggest opposing processes of maturation of the intestinal ecosystem from infancy to adulthood, with the urban metabolome rearranging with increasing age, counter to what is seen among Bassa adults, which remain comparable with the overall Bassa infant profile. In particular, in urban adults, we witnessed a reduction in abundance of several metabolite categories, including hexoses, glycerophospholipids, sphingolipids, and biogenic amines, and a corresponding increase in amino acids. As shown elsewhere, sphingolipids play a crucial, lasting role in the modulation of the intestinal natural killer T cell homeostasis in early life (), but their presence, as well as that of glycerophospholipids, may be of some relevance even in adults by triggering anti-inflammatory cascades (). It is interesting to note that although the age-related increase in fecal amino acids occurred in both populations, the discriminant amino metabolites were yet distinct by population; alanine enriched in Bassa adults, and phenylalanine, tyrosine, and BCAAs in urban Nigerians. Alanine is among the most abundant amino acids in leaves of Corchorus olitorius (jute) ( https://phytochem.nal.usda.gov/ ), a native plant of tropical Africa, whose leafy vegetable is popularly used in folk medicine and the preparation of soups (), such as ayoyo, which is an integral part of Bassa cuisine. Furthermore, alanine is gluconeogenic, and thus may relate to the enrichment in hexoses, as observed in rural subjects. Similarly, the increased phenylalanine and tyrosine amounts in urban adult metabolomes could be related to the diet, because these amino acids are particularly abundant in the leaves of the baobab tree (A. digitata) (), which serve as the main ingredient of the kuka soup, another staple food in Nigeria.

Yatsunenko et al., 2012 Yatsunenko T.

Rey F.E.

Manary M.J.

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Contreras M.

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et al. Human gut microbiome viewed across age and geography. Schnorr et al., 2014 Schnorr S.L.

Candela M.

Rampelli S.

Centanni M.

Consolandi C.

Basaglia G.

Turroni S.

Biagi E.

Peano C.

Severgnini M.

et al. Gut microbiome of the Hadza hunter-gatherers. When placed in an international scenario with compositional data from other human populations with varying subsistence patterns (), our data confirm that the way of life (traditional, including hunter-gathering and rural agricultural, versus urban-industrialized), rather than geography, drives structural convergence in gut ecosystem profiles, but with specific gradients in relation to the degree of transition from pre-agricultural to modern lifestyles. Despite the apparent presence of several Western-type taxonomic signatures, urban Nigerians are indeed partly similar to traditional populations and partly to typical urban-industrial communities, indicating an incomplete transition to microbial Westernization. In support of this, the retrieved diet information showed that some urban participants still follow a highly traditional Nigerian diet with limited consumption of processed store-bought foods. In parallel, the degree of schooling was found to vary among the urban subjects enrolled, as well as their hygiene practices, their social class, and the use of antibiotics.

Turroni et al. (2016) Turroni S.

Fiori J.

Rampelli S.

Schnorr S.L.

Consolandi C.

Barone M.

Biagi E.

Fanelli F.

Mezzullo M.

Crittenden A.N.

et al. Fecal metabolome of the Hadza hunter-gatherers: a host-microbiome integrative view. The comparison of metabolite profiles from the Bassa with those from the Hadza hunter-gatherers reveals a shared pattern of enrichment in hexoses and reduction in amino acids and biogenic amines relative to urbanized counterparts. The differences between the populations of the present study are, however, less pronounced as those between Hadza and Italians in, consistent with the less, and more recently, divergent lifestyle and environmental contexts of Bassa and urban Nigerians.

In summary, the microbial and metabolic characterization of the intestinal ecosystem of rural Bassa and urbanized individuals in Nigeria provided insights into the complex host-microbiome relationships across subsistence strategies, advancing our understanding of the changes in gut microbial communities and metabolic networks that probably accompanied human evolutionary history but, above all, stressing the relevance of the progressive adoption of a Western lifestyle as a major driver selecting for the loss of ancient signatures. Moreover, our findings support the existence of distinct trajectories of development of the intestinal ecosystem in early life, depending on human ecological context.

Further studies on larger worldwide cohorts, possibly with exact age information, are needed to evaluate the scope of these results, in the perspective of establishing the actual impacts on health of the lifestyle contribution to the compositional and functional structure of our gut, and its trajectories across lifespan.