Cell Host & Microbe

Using a longitudinal cohort of 633 Gambian children (IHAT-GUT, NCT02941081), we resolve two mechanistically distinct ecological pathways linking Prevotella stercorea to infection risk. Its abundance positively predicts gut microbiome richness, consistent with community-level colonisation resistance for enteric outcomes. However, its association with reduced acute respiratory infection (ARI) persists unchanged after richness adjustment, identifying a species-autonomous pathway independent of community diversity. Weight-for-age z-score (WAZ) is uncorrelated with microbiome richness within strata, supporting WAZ as a proxy for host immune-metabolic reserve rather than a determinant of microbiome composition. In Low-WAZ children, P. stercorea at Day 1 associates with suppressed CRP, whereas in higher-WAZ children, elevated Day 1 inflammation predicts subsequent P. stercorea colonisation at Day 85, consistent with host-context-dependent immune selection. ARI and fever protection is richness-independent and concentrated in Low-WAZ children. P. copri does not retain an independent protective association when modelled jointly. These findings have direct implications for microbiome-directed interventions.

The mechanisms by which maturation of the infant gut microbiota promotes resistance to pathogen colonization remain poorly understood. Infant botulism, a potentially fatal disease caused by intestinal colonization by Clostridium botulinum, provides a striking example of age-dependent susceptibility to infection1. Although the gut microbiota has long been implicated in protection against C. botulinum2-6, the responsible bacterial species and underlying mechanisms have yet to be elucidated. Here, we show that acquisition of Stickland-metabolizing Clostridia during infant gut microbiota maturation confers resistance to C. botulinum colonization through competition for shared amino-acid-dependent nutritional niches. In human fecal microbiota-transplanted mice, longitudinally collected infant microbiotas exhibited a clear transition from susceptibility to resistance. Intestinal metabolomic analysis identified 5-aminovalerate as a hallmark metabolite of the resistant microbiota, implicating Stickland metabolism, an amino acid metabolic pathway also utilized by C. botulinum. Guided by this finding, metagenomic analysis revealed enrichment of Stickland-metabolizing Clostridia in resistant microbiotas, including Clostridioides difficile, a bacterium frequently carried by healthy infants. Intestinal metabolic signatures of C. botulinum and infant-derived Stickland-metabolizing Clostridia suggested competition for shared amino-acid-dependent nutritional niches. Consistent with this model, C. difficile suppressed C. botulinum expansion through nutrient competition. Together, these findings identify nutritional niche competition as a mechanism by which microbiota maturation promotes resistance to C. botulinum colonization. This work demonstrates how acquisition of specific microbial metabolic functions during early life can shift the gut microbiota from a susceptible to a resistant state.

Antibiotic exposure disrupts enteric pathogen colonization resistance, yet how antibiotics reshape pathogen population dynamics, infection bottlenecks, and strain-level heterogeneity in the gut remains poorly understood. Here, we combine high-resolution pathogen barcoding, transcriptomic, and metabolomic analyses to quantify how short-term vancomycin perturbation alters infection ecology in vivo. We use Citrobacter rodentium as a model for human infection by pathogenic Escherichia coli-an antimicrobial resistance priority group-to demonstrate that just two days of vancomycin pre-treatment profoundly reshapes infection trajectories, driving rapid, global gut colonization, a dramatic increase in pathogen founding population size, and preservation of strain diversity across intestinal sites. Notably, vancomycin eliminated the hallmark heterogeneity of C. rodentium infection, resulting in fully reproducible colonization across hosts. Population-level analysis revealed that antibiotic treatment relaxes competitive constraints both with the resident microbiota and among clonal pathogen lineages, allowing early-established founders to persist and expand. Despite accelerated pathogen engraftment and tissue pathology, transcriptomic analysis revealed reduced virulence gene expression. Instead, antibiotic-induced metabolic restructuring of the gut created permissive conditions for pathogen expansion. Interactions with a vancomycin-altered microbiota, dominated by Akkermansia and Bacteroides, further promoted nutrient cross-feeding and influenced epithelial attachment. Together, we illustrate how short-term antibiotic exposure reshapes enteric infection by removing ecological bottlenecks that normally constrain strain diversity and infection outcomes. These findings have implications for antibiotic use, antimicrobial resistance transmission, and therapeutic strategies that rely on competition-driven dynamics, such as strain replacement.

The bile acid pool, which is synthesized collaboratively by the host and its microbiome, impacts metabolism, immunity, and disease risk. Targeted microbiome interventions could in principle reshape the bile acid pool for therapeutic benefit, but practical strategies remain elusive. In the course of screening a complex defined community for metabolic phenotypes by dropping out individual strains, we observed that several of the single-strain dropout communities had markedly increased deoxycholic and lithocholic acid levels and a larger bile acid pool. In each of these communities, a second strain--Lactobacillus plantarum--had bloomed. The bile salt hydrolase activity of L. plantarum was necessary and sufficient to expand the size of the bile acid pool. An engineered community in which the bsh gene is overexpressed in multiple Lactobacillus strains confers on mice increased levels of secondary bile acid levels and a larger pool size. By overexpressing a different pair of bile acid metabolic genes in multiple strains of Lactobacillus--7- and 7{beta}-hydroxysteroid dehydrogenase--we changed the composition of the bile acid pool, enlarging it and redirecting it toward ursodeoxycholic acid. Together, these results demonstrate that fine details of the microbiomes strain composition can have a substantial effect on bile acid metabolism, and that rational manipulation of the microbiome can alter the size and composition of the bile acid pool.

The complexity of the gut microbiome has made it challenging to define the role of individual species in community-level function. Here, we constructed 56 single-strain dropout variants of a defined 118-member community and used each one to colonize a group of germ-free mice. In many cases, removing a single strain triggered a large reordering of a small group of species, which in turn altered the communitys metabolic output. En bloc removal of the eight-strain acetogen compartment markedly reduced acetate production and caused intestinal H2 accumulation and bloating; a specific subset of four acetogens was sufficient to relieve bloating and restore acetate production. Together, these data show that small disturbances in community composition can trigger a confined ecological reorganization with a large chemical phenotype, and they reveal novel strategies for engineering communities with altered metabolic output.

Candida glabrata is a leading cause of invasive candidiasis. The gut serves as its primary reservoir, yet factors governing colonization and pathogenic potential remain poorly defined. Here, we identify immunoglobulin A (IgA) as a key regulator of C. glabrata within the intestinal microbiome. We found that C. glabrata induces an IgA response in a strain-specific manner. Comparative transcriptional and proteomic analyses of IgA-inducing and non-inducing strains identified a putative adhesin, Awp11, whose expression correlated with IgA induction. Awp11 is directly targeted by IgA and is required for inducing C. glabrata-specific IgA and Th17 responses in vivo. Functionally, Awp11 promotes colonization of a complex intestinal microbiome, and intestinal IgA limits this advantage. In most strains, AWP11 transcription is dynamic and limited by IgA in the gut. This identifies Awp11 as a key determinant of strain-dependent immunogenicity and gut colonization that C. glabrata may dynamically regulate to balance colonization and immune evasion.

Recovery of the gut microbiome after antibiotic exposure is often incomplete and variable, and the processes underlying this variation remain unclear. We performed longitudinal shotgun metagenomic sequencing of 2876 daily fecal samples from replicated humanized and conventional mouse cohorts exposed to controlled antibiotic perturbations. Metagenomic profiling recapitulated ecological trajectories previously observed by 16S sequencing, while revealing extensive strain-level dynamics, including reproducible sweeps of standing variants and de novo mutations in antibiotic target sites and regulatory loci. We also identified genetic changes whose effects depended on community composition, competitive release, and perturbation history. Cross-housing experiments revealed bidirectional strain transfer, with antibiotic-induced niche clearance enabling replacement of resident strains. In parallel, phage dynamics were heterogeneous and clustered by cage. Together, these findings show that post-antibiotic microbiome recovery is a path-dependent process shaped by selection, transmission, and phage activity, producing divergent outcomes even among closely matched communities exposed to the same perturbations.

The gut microbiome is essential for human health. Although the gut microbiota is largely stable at the species level in healthy individuals, strain-level variation remains less understood. Many bacterial strains encode toxin delivery systems that may shape competition within the gut. Here, we investigate how contact-dependent growth inhibition (CDI) and colicins influence intestinal colonization by a competitive murine Escherichia coli isolate, R12. We show that R12 can colonize an intact mouse gut microbiota by displacing resident Enterobacteriaceae, but success depends on multiple interacting factors. CDI systems and colicins provide a competitive advantage against resident E. coli, particularly during early colonization, while metabolic flexibility and access to alternative carbon sources support long-term persistence. Colonization outcomes vary between hosts and are shaped by resident microbiota composition, strain-level competition, and the initial invader-to-resident ratio. Overall, successful gut invasion is determined by the combined effects of bacterial antagonistic systems, metabolic capacity, and ecological context.

Sialic acids are abundant components of host- and diet-derived glycans in the human gut and serve as important nutrients that shape microbial fitness and interspecies competition. Excess free sialic acids are also linked to inflammation and pathogen susceptibility. While well-studied gut bacteria such as E. coli and Bacteroides spp. catabolize sialic acids via the NanAKE or NanLE-RokA pathways, the metabolic capacity of many microbiome members remains undefined. To identify sialic acid catabolizing bacteria, we cultured fecal samples from healthy human donors. The gut anaerobe Hungatella hathewayi was selected under sialic acid-supplemented conditions. H. hathewayi is a poorly characterized gram-positive Lachnospiraceae associated with long-lived individuals and purine metabolism. Here we establish that H. hathewayi grows robustly on sialic acids as a sole carbon source using a pathway homologous to the canonical NanAKE system of E. coli, despite the species phylogenetic distance. We functionally validated these orthologs through growth assays and heterologous complementation in E. coli knockout strains. Comparative analyses further showed that key catalytic residues in H. hathewayi NanA are conserved despite overall sequence divergence from E. coli. Additionally, we find that colocalized sialic acid transporters and regulatory proteins are not orthologous to E. coli proteins and instead are related to proteins from other gut anaerobes. Together, these findings expand our understanding of sialic acid utilization within the human gut microbiome. We identify H. hathewayi as an overlooked but capable sialic acid degrader that can contribute to modulation of gut sialic acid levels and related inflammation. ImportanceSialic acids play an important role in mammalian and microbial signalling. Excess free sialic acids increase susceptibility to gut pathogens and induce inflammation. Gut bacteria can both generate and consume free sialic acids, and these pathways are conserved across diverse bacteria. E. coli and B. fragilis consume sialic acids as a carbon source, decreasing free sialic acid levels. We identify H. hathewayi as another bacteria capable of sialic acid consumption and define the enzymes responsible. H. hathewayi is a prevalent member of the human gut microbiome, but it is not genetically tractable, limiting enzymatic characterization. H. hathewayi is enriched in the gut microbiomes of long-lived individuals and expected to be an important contributor to purine degradation to limit gout risk. Defining sialic acid catabolism in non-model species is essential to understanding the evolution and conservation of this pathway as well as how nutrient competition shapes gut microbiome composition.

Bacteriophage therapy is needed to treat antibiotic resistant infections; however, when a clinical isolate resists a given phage, it is often unclear why. It is therefore currently unknown how to rationally fortify phage therapies to circumvent a priori resistance. Using a family of broad host range therapeutic Pseudomonas aeruginosa phages (Pbunaviruses), we show that cell surface receptor masking and intracellular defenses are both common barriers in distinct clinical isolates. In some cases these barriers can be bypassed by intrafamily phage engineering. Using unbiased genome-wide CRISPRi screens, we reveal that the broadly conserved L-Rhamnose in the core polysaccharide is the receptor for Pbunavirus family. This molecule is often masked by diverse O-antigen structures. In other isolates with the L-Rha receptor accessible, internal defense mechanisms commonly prevent Pbunavirus DNA replication. A single anti-defense locus often encoding 8-11 different genes within the Pbunavirus family is required for optimal host range, providing anti-defense genes that enable replication of both Pbunavirus phages and phages of other families. Our work demonstrates the importance of both internal and surface defense mechanisms in clinical isolates causally antagonizing a commonly used phage therapeutic and presents phage engineering strategies to circumvent a priori resistance.

Uric acid, the final product of purine metabolism in humans, accumulates in blood and tissues at relatively high concentrations1 as humans lack the enzyme uricase2,3. Under inflammatory conditions, uric acid can be oxidised to yield reactive intermediates4. In activated neutrophils, myeloperoxidase (MPO) catalyses the oxidation of uric acid by hydrogen peroxide, leading to the formation of urate hydroperoxide (UH)5,6. While recent studies have shown that UH is toxic to bacteria lacking peroxiredoxins7, its precise mechanism of toxicity and the existence of dedicated bacterial defence systems remain unknown. Here, we identify HiuH as a periplasmic enzyme, conserved across E. coli strains, that specifically degrades UH. Our findings reveal that UH selectively induces the expression of hiuH and that HiuH efficiently detoxifies UH both in vitro and in bacterial cells. HiuH cooperates with MsrP, a periplasmic methionine sulfoxide reductase that repairs UH-induced protein-bound methionine oxidation. This combined defence offering both direct detoxification and damage repair, is essential for bacterial survival under UH stress, and confers a competitive fitness advantage in a DSS-induced mouse model of colitis. Although UH is chemically transient, our work shows that it imposes durable biological consequences and a sufficient fitness cost in the in vivo niches occupied by E. coli to favour the evolution of a dedicated detoxification pathway beyond general oxidative-stress responses, defining a key adaptation to periods of gut inflammation.

Using a longitudinal cohort of 633 Gambian children (IHAT-GUT, NCT02941081), we resolve two mechanistically distinct ecological pathways linking Prevotella stercorea to infection risk. Its abundance positively predicts gut microbiome richness, consistent with community-level colonisation resistance for enteric outcomes. However, its association with reduced acute respiratory infection (ARI) persists unchanged after richness adjustment, identifying a species-autonomous pathway independent of community diversity. Weight-for-age z-score (WAZ) is uncorrelated with microbiome richness within strata, supporting WAZ as a proxy for host immune-metabolic reserve rather than a determinant of microbiome composition. In Low-WAZ children, P. stercorea at Day 1 associates with suppressed CRP, whereas in higher-WAZ children, elevated Day 1 inflammation predicts subsequent P. stercorea colonisation at Day 85, consistent with host-context-dependent immune selection. ARI and fever protection is richness-independent and concentrated in Low-WAZ children. P. copri does not retain an independent protective association when modelled jointly. These findings have direct implications for microbiome-directed interventions. Impact statementGut Prevotella stercorea protects young children against respiratory infection through a species-specific, community-independent immune pathway that is most active in immunologically vulnerable hosts, defining a new mechanistic target for microbiome-directed respiratory therapeutics.

The skin microbiome regulates key skin processes, yet the functional diversity of a dominant genus, Staphylococcus, remains poorly resolved at the strain level for multiple species across its pathogenic and commensal continuum. It is likely that Staphylococcus effects on skin are diverse at these finest taxonomic resolutions, but current skin models lack the physiological relevance and scalability needed to profile this diversity. Using an organotypic 3D human skin model (reconstructed human epidermis, RHE), we profiled skin responses to 187 Staphylococcus strains across seven dominant species. Canonically pathogenic species (e.g., S. aureus) induced broad inflammatory responses, whereas prototypical commensal species (e.g., S. hominis) elicited more nuanced effects on innate immune and skin barrier responses. Strikingly, S. epidermidis displayed pronounced strain-level heterogeneity, with subsets inducing either commensal or pathogen-like responses despite lacking canonical virulence factors, suggesting pleiotropic effects. Comparative genomics, dual-transcriptomics, untargeted metabolomics, and growth phenotyping revealed species- and strain-specific traits underlying these differential effects on RHE, including the presence of select cell surface proteins and differential arginine metabolism. Together, our study provides the first high-throughput, species- and strain-resolved analysis of skin-Staphylococcus interactions, offering mechanistic insights and a platform for microbiome-based strategies to modulate skin inflammation and diseases. One-line summaryHigh-throughput profiling of Staphylococcus in a human skin model shows that species- and strain-level diversity underlies a continuum of host barrier and immune responses.

Candidozyma auris is an emerging, multidrug-resistant (MDR) fungal pathogen that persistently colonizes human skin and disproportionately causes invasive infections in patients with metabolic or immune dysfunction. Despite strong epidemiological links to diabetes and immunosuppression, how these host conditions shape skin colonization, immune defense, and dissemination remain poorly defined. Here, we establish complementary murine model of C. auris skin colonization under immunocompetent, immunosuppressed, and diabetic ketoacidosis (DKA) conditions. DKA mice exhibited significantly increased skin fungal burden, impaired clearance, and frequent systemic dissemination. Notably, DKA permitted dissemination despite preserved granulocyte and neutrophil recruitment to the skin, indicating a functional rather than quantitative defect in innate immunity. Consistent with this, phagocytes from DKA mice displayed impaired antifungal activity characterized by reduced phagocytosis and killing despite elevated reactive oxygen species production. Hyperglycemic and ketone-rich conditions (BHB) remodel the C. auris cell wall, reducing mannan, increasing chitin, and upregulating adhesins, thereby enhancing adhesion and inflammatory activation while impairing neutrophil killing. Together, these findings reveal host metabolic dysfunction as a primary driver of persistent C. auris skin colonization and dissemination, identify qualitative defects in innate antifungal immunity as a key determinant of invasive risk and highlight metabolic condition as a critical target for infection prevention strategies. Short SummaryThis study establishes the first physiologically relevant murine model of Candidozyma auris skin colonization under diabetic ketoacidosis and immunosuppression, revealing distinct immune dysfunction and systemic dissemination that can inform targeted antifungal strategies.

Microbial dysbiosis is a hallmark of inflammatory bowel diseases (IBD); however, its drivers and impact on disease pathophysiology are poorly understood. Applying neural network-based feature attribution to metabolomics and metagenomics datasets from >5000 individuals, we identified epimerized host derived bile acids (BAs) produced by microbial hydroxysteroid dehydrogenases (HSDHs) as a novel hallmark of IBD-associated dysbiosis. Epimerized BAs reduce FXR activity in intestinal epithelial cells and dampen their production of FGF19, a negative feedback regulator of host-derived bile acid (HBA) production in the liver. Increased HBA levels drive colonic epithelial remodeling by impacting goblet cell maturation and select for HSDH-carrying bacteria that transform bactericidal HBA into less toxic, epimerized forms. Confirming the translational relevance of these findings, we demonstrated that high HBA levels limit fecal microbiota transplant engraftment and show that BA sequestering drugs support microbiome recovery in patients with high HBA levels. Together, we discover that elevated HBAs deplete BA-sensitive commensals and favor the growth of HSDH-encoding pathobionts that disrupt host BA feedback signaling, establishing a causal link between changes in microbial ecology and IBD pathophysiology.

Clostridioides difficile is an urgent threat to human health. Current treatments for C. difficile infections (CDIs) are antibiotics and microbiome restoration therapy (MRT) for recurrent cases. However, antibiotics contribute to antibiotic resistance and recurrent CDIs and the long-term sustainability and accessibility of MRTs remains to be determined. Since a dysbiotic gut microbiome is the primary risk factor for CDI, a better understanding of the interactions between C. difficile, the microbiome, and the host will aid development of treatments with improved precision. Emerging evidence supports that butyrate, a prominent end product of gut microbiome metabolism, is a key determinant of C. difficile pathogenesis. Notably, C. difficile releases more of its toxins TcdA and TcdB in butyrate-rich environments. Here, we demonstrate that butyrate-dependent toxin release is not driven by two previously characterized modes of toxin release (e.g., TcdE-dependent secretion or Cwp19-dependent autolysis). Instead, butyrate enhances the expression of a broadly conserved endolysin (EndD), which is responsible for butyrate-dependent toxin release. We additionally demonstrate that endD-dependent toxin release does not universally occur under all growth conditions and that its expression is dependent on the late-stage sporulation sigma factor SigK. Overall, our findings provide deeper insight into butyrate-dependent effects on C. difficile pathogenesis and set the stage for future work to better understand the molecular and genetic underpinnings of endD regulation.

Bacteriophages are a promising tool for microbiome editing, yet their development has been constrained by limited insights into bacteriophage-host interactions within their shared mammalian body habitat. We isolated a lytic phage {Phi}KL11 that efficiently targets a disease-associated member of the human gut microbiota, Eggerthella lenta, during in vitro growth. However, {Phi}KL11 selects for a pre-existing and reversible bacteriophage-resistant sub-population in mice. Long-read sequencing revealed a massive genomic inversion event, representing >50% of the E. lenta genome, enriched in response to bacteriophage infection. Transcriptomics linked this inversion to the altered expression of three capsular polysaccharide synthesis (CPS) gene clusters and transmission electron microscopy confirmed differential capsule production. Finally, we show that {Phi}KL11 has a broad host range attributable to CPS and other strain-variable genes. These findings suggest a previously unrecognized strategy for phage evasion in the gut, involving megabase-scale genomic inversions and reversible capsule variation driving phage resistance.

Mechanistic understanding of gut ecology is limited by the availability of tools for precise manipulation of microbiome composition. Here, we isolate lytic phages to enable targeted removal of gut commensal Escherichia fergusonii (Ef) from complex, undefined stool-derived in vitro communities. A single phage drove resistance without fitness cost in monoculture, but resistant Ef exhibited reduced fitness in communities, enabling expansion of closely related Proteobacteria. Resistance arose via reversible promoter inversion linked to outer-membrane function. A phage cocktail overcame resistance to achieve Ef knockout across communities with minimal collateral effects. Using knockout communities, we show that Ef is necessary and sufficient for preventing Salmonella invasion. Replacement with an Ef transposon-mutant library revealed that community-specific fitness defects are enriched in genes involved in outer-membrane assembly. Disruption of these genes sensitized Ef to antagonistic community members, highlighting interspecies warfare as a key driver of microbiome ecology. These results establish phage-mediated perturbation as a framework for linking species to community-level function and for enabling precision microbiome engineering.

Infancy is a critical developmental window during which the gut ecosystem assembles and helps train the immune system, thereby setting trajectories for lifelong health. Bacteria and viruses are equally numerous in this early ecosystem, yet the gut viromes composition, dynamics, and health relevance remain poorly understood. Here, we show that the infant gut virome is diverse, dynamic, and linked to health outcomes. We performed comprehensive virome profiling of 1,110 longitudinal fecal samples from 314 mother-infant pairs from the Dutch birth cohort Lifelines NEXT using both virus-like particle enrichment (VLP) and total metagenomic sequencing (MGS). We find only 18.9% compositional overlap between the VLP- and MGS-metaviromes, with VLP recovering the active virome and most novel species and MGS predominantly capturing temperate phages. By combining both methods, we identified 8,348 novel virus species spanning diverse hosts, from bacteria to humans, and all major viral genome types (dsDNA, ssDNA, and RNA). We find that bacteriophages frequently encode metabolic functions, including genes related to B vitamin metabolism. We further observe that the development of the infant gut virome is shaped by both host factors, including delivery mode and feeding practices, and continuous switching of temperate phage lifecycles. Notably, the relative abundance of induced temperate phages is also associated with eczema development within the first year of life. Together, these findings establish the infant gut virome as a dynamic and clinically relevant component of early-life microbial development and highlight how comprehensive dual-method profiling is a necessary framework for future virome research.

Whole genome sequences of microbial isolates are essential to link microbial diversity to population and community ecology, because metagenomics alone cannot resolve strain-level variation, particularly in complex and mobile genetic elements that confer strain-specific traits such as virulence. We cultured and whole-genome sequenced 2,920 Staphylococcus isolates from healthy subjects and patients with Dariers disease and congenital ichthyosis. Our results show that disease imposes selection at the species level, while strain populations within those species diversify across individuals in a manner consistent with ecological drift, resulting in increased interpersonal heterogeneity of strain composition. Despite this variability, strain populations converged on shared functional traits, suggesting constraints imposed by the disease environment. We further examined key functional elements of clinical and ecological relevance, including the agr quorum sensing system, the mobile genetic element SCCmec that carries methicillin resistance, and CRISPR/Cas systems, contextualizing these features both within isolate population structure and the surrounding microbial community using metagenomic data. We found disease-dependent enrichment of virulence genes, striking species-specific phylogenetic sorting of agr variants, unexpected modularity of previously undocumented SCCmec elements in diseased skin, and links between phages in the microbiota and CRISPR spacers in the isolate genomes. Taken together, our integrated, large-scale analysis of isolate genomics and metagenomics reveals that skin disease restructures staphylococcal strain populations through combined effects of selection, drift, and functional constraint. One line summaryWhole-genome sequencing of 2,920 skin Staphylococcus isolates shows that disease selects at the species level while reshaping strain populations and their functional diversity within hosts.