Cell Host & Microbe

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.

Candidozyma auris is an emerging multi-drug resistant fungal pathogen characterized by high mortality and rapid transmission in healthcare settings, but the genetic drivers of phenotypic variation between strains and the landscape of gene essentiality in this organism remain undercharacterized. Here, we integrate pangenomic analysis with global essentiality screening to establish a foundational understanding of the C. auris genome and identify potential therapeutic targets. We performed pangenome analysis on 695 outbreak strains of C. auris selected to be genetically representative of publicly sequenced genomes. After using BLAST to refine the pangenome, we found that 96.8% of gene families were core, with the remaining high-confidence accessory gene families primarily consisting of gene loss events or clade-specific genes. The high proportion of core genes emphasizes the clonal nature of these outbreak strains, but comparative analysis with the closely related C. haemuli species complex suggested that most of these core genes are functionally dispensible. To examine this hypothesis, we developed a novel insertional mutagenesis approach that leverages the promiscuous integration of linear DNA in the C. auris genome. This global analysis identified 614 high-confidence essential genes. Crucially, nearly one-third of these genes, including the conserved translation initiation factor Sui1, exhibit divergent essentiality patterns compared to the model yeasts Candida albicans and Saccharomyces cerevisiae. These findings highlight organism-specific biology that would be overlooked by orthology alone. By combining pangenomic diversity with functional essentiality, this study provides a comprehensive resource for identifying species-specific determinants of virulence and prioritizing novel targets for antifungal drug development.

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.

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.

WD40 domains are major protein-protein interaction (PPI) scaffolds, yet their contributions to fungal pathogenicity remain poorly defined. We systematically analysed 94 canonical WD40 proteins in Cryptococcus neoformans. Conditional knockdown and sporulation identified 36 essential WD40 proteins, while in vitro and in vivo profiling of 103 signature-tagged deletion strains spanning 52 genes uncovered 31 pathogenicity-related WD40 proteins, including epigenetic and post-transcriptional regulators. We identified Wcp1, a dual-domain protein whose WD40-repeat and cyclophilin domains are required for growth at 37{degrees}C under 5% CO2. Its WD40 scaffold and PPIase domain supported CO2/heat tolerance and virulence. Notably, Wcp1 couples these functions to acidic pH adaptation: wcp1{Delta} failed to grow under elevated temperature and CO2 at acidic pH, exhibited enhanced intracellular acidification, reduced macrophage survival and attenuated virulence in Drosophila and mice. Integrated transcriptomic and proteomic analyses place Wcp1 at the centre of intracellular pH homeostasis, coordinating proton transport, metabolic adaptation and stress-buffering networks.

MDA5 is a cytosolic pattern-recognition receptor (PRR) that binds to double-stranded RNA (dsRNA) and subsequently interacts with the signaling adaptor protein MAVS to initiate the antiviral interferon (IFN) response. Our group previously demonstrated that MDA5 is essential for host resistance against the fungal pathogen, Aspergillus fumigatus. Although fungal dsRNA was sufficient to activate MDA5 signaling, the precise source of A. fumigatus dsRNA responsible for this MDA5-stimulating function remains unknown. Here, we demonstrate that the magnitude of the IFN-dependent antifungal response is A. fumigatus strain dependent. Unexpectedly, we found that A. fumigatus isolates infected with dsRNA mycoviruses triggered a more robust MAVS-dependent inflammatory response within alveolar macrophages. Furthermore, dsRNA mycovirus infection increased fungal susceptibility to antifungal killing without altering other A. fumigatus growth characteristics. Although dsRNA mycovirus infection did not alter virulence in an acute bronchopneumonia model of A. fumigatus infection, it significantly impaired virulence and improved disease parameters in a chronic model of allergic bronchopulmonary aspergillosis (ABPA). Collectively, these findings reveal a novel role for trans-kingdom interactions in driving the host antifungal IFN response and modulating virulence in chronic aspergillosis models.

Hospital outbreaks caused by the fungal pathogen Candida parapsilosis are of growing concern due to their increased drug resistance and high mortality rates. However, the genetic bases of clinically-relevant traits in this species remain poorly explored. Here, we mapped genotype-phenotype relationships across 189 isolates from a multi-hospital Candida parapsilosis outbreak, for which we measured 61 diverse clinical phenotypes and generated complete genome sequences. As variation in previously-known genes explained little of the observed phenotypic diversity, we leveraged convergence genome-wide association studies and interpretable machine-learning models that predict phenotypes from genetic variants. These approaches identified candidate drivers of virulence and antifungal resistance, confirming expected mechanisms while uncovering novel ones. Predictive models were accurate for key traits, including azole resistance and clinical features of infected patients. Our results shed light on the genetic bases of clinically-relevant traits in a major fungal pathogen, and pave the way towards sequence-based diagnostics for improved patient outcomes.

The tuft cell-ILC2 amplification circuit has emerged as a central paradigm of anti-helminth immunity in laboratory animals, driving the so-called weep and sweep response, required for parasite expulsion. Yet soil-transmitted helminths (STHs) commonly establish chronic infections in humans. Whether tuft cell expansion is required for parasite clearance under naturalistic conditions remains unknown. Here, using wildlings, a naturalized mouse model exposed from birth to complex microbial communities and pathogens, we re-investigated anti-STH immunity in the context of ecological realism. Following infection with Nippostrongylus brasiliensis, specific pathogen-free (SPF) mice mounted markedly amplified type 2 responses in both lung and intestine compared to wildlings. In the intestine, SPF mice mounted robust tuft cell expansion, IL-25 production, ILC2 accumulation, and goblet cell hyperplasia. In contrast, infected wildlings exhibited delayed parasite expulsion, limited tuft cell expansion, reduced IL-25 and ILC2 responses, and attenuated goblet cell expansion. Wildling tuft cells, but not goblet cells, displayed markedly reduced expansion in response to succinate or exogenous IL-13, indicating selective hypo-responsiveness of the epithelial sensory compartment. Microbial transfer into adult SPF mice selectively conferred tuft cell hypo-responsiveness without impairing ILC2 accumulation or goblet cell expansion. Tuft cell hypo-responsiveness in wildlings and FMT recipients was associated with enrichment of fermentative bacteria and increased levels of the short chain fatty acids acetate and propionate. Together, these findings indicate that ecological microbial exposure imprints systemic type 2 immunity during early-life, whereas epithelial responsiveness remains plastic and microbiome-dependent, thereby revealing regulatory constraints not evident under SPF conditions. One Sentence SummaryOur findings reveal that in a naturalized immune-microbiome context, intestinal tuft cells are surprisingly hypo-responsive, highlighting how environmental microbial exposure can calibrate type 2 immunity and helminth resistance.

Clostridioides difficile infection (CDI) susceptibility and severity are strongly associated with preexisting colonic inflammation. However, chronic inflammatory conditions such as cystic fibrosis rarely progress to symptomatic CDI despite high rates of C. difficile colonization, suggesting that inflammation alone is insufficient to explain disease vulnerability. Notably, populations relatively protected from symptomatic CDI exhibit impaired regenerative capacity within the colon epithelium. Here, we used single cell RNA sequencing of human colonoid monolayers to map markers of CDI susceptibility and severity to cell populations associated with inflammation and epithelial repair. We identified an inducible microfold-like (M-like) population that is largely absent from the healthy colon but emerges during inflammation and regeneration. These cells were enriched for markers of severe CDI, C. difficile toxin interaction genes, and elevated CCL20 and CFTR expression. Spatial imaging localized CCL20-producing cells to wound-like gaps in mock and CDI-treated colonoids, identifying a repair-associated niche active independent of infection. Following exposure to C. difficile, wound-healing transcription within the M-like lineage declined while tuft-like populations expanded and upregulated genes associated with immune cell recruitment. These findings demonstrate that epithelial regeneration shapes host CDI vulnerability. IMPORTANCEClostridioides difficile infection can lead to severe illness and death in vulnerable populations despite available treatments. Clinical signs of inflammation during active Clostridioides difficile infection are strongly associated with disease outcome, yet these responses primarily reflect tissue damage already underway, limiting opportunities to prevent progression. In contrast, conditions linked to severe disease, including inflammatory bowel disease and antibiotic exposure, are associated with colonic inflammation before infection or at the time of diagnosis, highlighting an opportunity for earlier identification of high-risk individuals. Using human colonoid single cell transcriptomics and spatial imaging, we identified a microfold-like cell population enriched for inflammatory mediators and Clostridioides difficile toxin interaction genes linked to severe disease. This population was active even in the absence of infection, suggesting that repair-associated populations within the inflamed colon may help identify susceptibility to severe CDI before clinical progression occurs.

Bacteriophages are key drivers of microbial ecology and evolution, and the rapid expansion of phage sequencing has created sustained demand for curated reference genome databases. We released the INfrastructure for a PHAge REference Database (INPHARED) in January 2021 to provide quality-controlled metadata for complete phage genomes from cultured isolates. Here, we compare the 2021 and 2026 snapshots, spanning a five-year period that included a substantial overhaul of bacterial virus taxonomy by the ICTV. The database has approximately doubled, from 14,244 to 28,777 genomes, yet the proportion representing novel species-level diversity has declined, indicating that redundant sequencing is outpacing new discovery. Host bias persists despite the addition of 97 new host genera. We have incorporated genome quality assessments, lifestyle predictions, and defence and anti-defence system annotations, providing an updated resource and a snapshot of the current state of phage genomics.

C. elegans, a bacterivore living in microbially-complex environments, harbors a characteristic community of gut bacteria that contribute to its health and fitness. What determines which environmental bacteria end up as commensals is largely unknown in C. elegans, as in other animals. Previous work found that gut Pantoea isolates supported rapid worm development and infection resistance, while environmental congenerics were inferior. Notably, worms were preferentially attracted to the more beneficial gut isolates. Using bioactivity-guided fractionation and gas chromatography-mass spectrometry analysis, we identified bacterially derived isoamyl alcohol (IAA) as a secreted volatile attractant underlying this preference. Screening of worm mutants implicated AWC sensory neuron-associated genes in preferential attraction to beneficial Pantoea and established a causal link between IAA sensing and colonization by beneficial strains. While IAA sensing was important for initial colonization, gut-associated Pantoea strains ultimately outcompeted environmental congenerics over time, indicating that microbiome assembly is shaped by two complementary processes: host behavioral preference for high-IAA producers and bacterial competitive fitness within the gut. While IAA is a product of leucine metabolism and may function as a nutritional cue, we found that it could also directly enhance host infection resistance, suggesting an additional role in modulating host physiology. Finally, knockout analysis identified a bacterial branched-chain amino acid aminotransferase homolog as important for IAA production. Together, these findings identify bacterial volatile sensing as an important and underexplored mechanism shaping microbiome composition and its contributions to host fitness.

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.

Bacteria can outcompete rivals by using specialized secretion systems to deliver toxic effectors into prey cells. How these systems distinguish prey from kin remains a fundamental unanswered question. In many Xanthomonadales (Lysobacterales) species, the antibacterial type IV secretion system (X-T4SS) mediates cell-cell contact-dependent killing of competing bacteria. Here, we identified XAC2611, a chromosomally encoded, cysteine-rich DUF4189 protein, as essential for preventing X-T4SS-mediated fratricide among sibling cells in Xanthomonas citri. XAC2611 homologs are found within or proximal to the genomic loci coding almost all identified X-T4SS. Live-cell microscopy and biochemical data reveal that XAC2611 is abundantly produced and widely distributed throughout the periplasm of recipient cells and is required to prevent X. citri cells from intoxicating each other in a contact-dependent manner. We also show that XAC2611 homologs from Stenotrophomonas maltophilia protect X. citri cells from attack by S. maltophilia. Protein-protein interaction assays show that XAC2611 interacts directly with the VirB5 subunit. Since VirB5 is predicted to be localized at the tip of the X-T4SS pilus, its interaction with XAC2611 in a neighboring sister cell could block X-T4SS-mediated effector delivery. We therefore name this family of proteins trans-intoxication protection factors (Tpfs).

Streptococcus pneumoniae (Spn) asymptomatically colonizes the upper respiratory tract (URT), a niche from which it can transmit to another host or cause invasive disease in the same host. The in vivo transcriptional adaptations that Spn undergoes during nasopharyngeal colonization, particularly during influenza A virus (IAV) coinfection, are poorly understood. Here, we leveraged an established infant mouse model of colonization, shedding, and transmission to perform genome-wide transcriptomic profiling of Spn during mono- and during IAV coinfection. Compared with broth-grown controls, pneumococci isolated from the URT exhibited distinct transcriptional programs, with over 200 genes differentially expressed across time points. Genes involved in carbohydrate uptake and metabolism, glycan degradation, amino sugar and nucleotide sugar metabolism, and amino acid biosynthesis were consistently enriched during colonization, highlighting metabolic adaptation to the nasopharyngeal niche. In contrast, IAV coinfection induced a markedly distinct transcriptional signature, including upregulation of branched-chain amino acid biosynthesis, bacteriocin production, and phosphate acquisition systems. Notably, the pilus islet-1 locus was upregulated during Spn-IAV coinfection. Functional studies demonstrated that while the pilus was dispensable for colonization under mono- and coinfection conditions, it promoted high-shedding events and enhanced inflammatory responses during IAV coinfection. However, reduced inflammation and reduced high shedding events from pups inoculated with a pilus-deficient mutant did not alter transmission frequency in the infant mouse model. Collectively, our findings define the in vivo transcriptional landscape of Spn during URT colonization and reveal distinct bacterial adaptations during viral coinfection, providing insight into mechanisms that influence pneumococcal persistence, inflammation, and transmission.

Cryptococcus neoformans is a global fungal pathogen that causes fatal cryptococcosis, underscoring the urgent need for novel therapeutics given the limitations of current antifungals. Here, we systematically investigate essential transcription factors (TFs) in C. neoformans, focusing on their roles in growth and their potential as drug targets. We developed experimental pipelines to assess growth requirement, essentiality, and function using conditional gene expression, constitutive overexpression, and meiotic spore analysis. Through these systematic analyses, we identify one quasi-essential (growth-required but non-essential) TF, Fhl1, and 13 essential TFs, including three (Ezt1, Ezt2 and Cbf1) that are highly divergent from counterparts in other eukaryotes. Notably, Ezt1 emerged as a central TF regulating over 1,200 genes, controlling growth, antifungal drug and stress responses, sexual development, and virulence. Collectively, our findings define the essential TF landscape of C. neoformans and provide a framework for leveraging these regulators, particularly Ezt1, to develop targeted therapies for cryptococcosis.

Antibiotic exposure is a significant risk factor for Crohns disease, yet the tissue-specific consequences of antibiotic-driven dysbiosis remain poorly defined. Adherent-invasive Escherichia coli (AIEC), a pathobiont enriched in Crohns disease, expands following antibiotic treatment, but whether discrete mucosal niches support this expansion is unknown. Peyers patches are specialized lymphoid structures that coordinate mucosal immunity and are frequently associated with early disease lesions, suggesting they may represent a vulnerable site for pathobiont colonization. Here, we show that vancomycin disrupts the Peyers patch-associated microbiome, creating a permissive niche that is selectively exploited by AIEC and associated with focal inflammation. Antibiotic treatment markedly increased AIEC burden within Peyers patches. AIEC localized within the lymphoid follicle was accompanied by focal tissue pathology and a distinct cytokine signature. In contrast, expansion of resident E. coli in the absence of AIEC did not elicit comparable inflammation, indicating that the pathogenic traits of AIEC are required to trigger disease-relevant responses in this niche. Supporting this, genetic disruption of flagellin, long polar fimbriae, or antimicrobial peptide resistance in AIEC attenuated Peyers patch colonization or inflammation, revealing separable mechanisms governing niche access and immunopathology. Together, these findings identify Peyers patches as a previously unrecognized reservoir for antibiotic-driven AIEC expansion and define a localized host-microbe interaction that links dysbiosis to focal intestinal inflammation. These results provide a mechanistic framework for understanding how antibiotic exposure may precipitate site-specific pathology in Crohns disease. Further, these findings highlight that mucosal lymphoid tissues should be considered when evaluating microbiome-targeted therapeutic interventions in Crohns disease.

Host genetics shapes gut microbiome composition, yet the physiological mechanisms underlying this relationship remain poorly understood. Characterizing associations between host gene expression and the mucosal microbiome offers a promising route to identifying the host pathways and microbial taxa most likely to interact physiologically. However, existing investigations have been conducted primarily in acute disease contexts and within the colon, leaving host-microbiome associations outside of acute inflammatory contexts and those in undersampled regions such as the terminal ileum poorly characterized. To address these gaps, we profiled paired host gene expression from full-thickness resections and mucosal microbiome data, both from macroscopically non-inflamed tissue from Crohns disease patients undergoing surgery across three intestinal sites: terminal ileum (n = 32), cecum (n = 35), and right colon (n = 30). Using a multi-level analytical framework including Procrustes analysis, sparse canonical correlation analysis, and elastic net regression, we identified significant associations between the mucosal transcriptome and microbiome. Intestine-wide, genes enriched in immune and intestinal barrier integrity pathways were associated with heritable taxa including Fusicatenibacter, consistent with patterns observed in microbiome genome-wide association studies. Region-specific analysis identified the terminal ileum as a distinct site of host-microbiome interaction, with associations involving metabolic and barrier-related pathways not observed in the large intestine. Notable terminal ileum-specific associations included PCDH20 with Faecalitalea and ACAT1 with Lactococcus, implicating epithelial barrier maintenance and host-microbiome metabolic interactions, respectively. These findings advance our understanding of the physiological basis of host-microbiome interactions across the intestine. ImportanceThe human gut is home to trillions of microorganisms that interact with the intestinal lining, yet we have a limited understanding of the specific biological processes involved in these interactions. Most studies characterizing the relationships between host gene expression and the gut microbiome have focused on the colon and on active disease contexts, leaving it unclear whether the associations observed reflect fundamental host-microbiome biology or disease-specific responses. By examining mucosal tissue, where host cells and microbes are in direct contact, across three sites in non-acutely inflamed tissue, we show that expression of immune defense and barrier maintenance genes is broadly associated with the microbiome across the intestine. We also identify distinct classes of associations in the terminal ileum, including host genes involved in metabolic processes. These findings provide a foundation for understanding how host biology and the gut microbiome are linked outside of acute disease.

The kynurenine pathway (KP) of tryptophan metabolism shapes host immunity and is implicated in inflammatory, malignant, and neurodegenerative diseases. Although it is well known that inflammation influences KP activity, whether and how gut microbiota regulates KP in the intestine remains unclear. Here, we find that in allogeneic hematopoietic cell transplant patients, the kynurenine-to-tryptophan ratio tracks plasma neopterin, whereas intestinal levels do not, suggesting that local KP regulation is not explained by inflammation alone. In mice, targeted microbiota perturbation downregulates intestinal expression of indoleamine 2,3-dioxygenase 1 (IDO1), the rate-limiting enzyme of the KP, while microbiota restoration reversed this effect. Computational analyses associate Lachnospiraceae and Oscillospiraceae with colonic IDO1 in mice and inflammatory bowel disease cohorts. In vitro experiments support a mechanism whereby microbiota perturbation alters host absorption of the vitamin E isoform, {gamma}-tocotrienol, which impacts IDO1 expression. Our findings support that the gut microbiota controls colonic KP through micronutrient absorption.

The assembly and maturation of the infant gut microbiome is a critical developmental process. Yet the dynamics of the viral community, particularly in the context of stunting (chronic malnutrition) remain underexplored. Leveraging longitudinal fecal metagenomes from Zimbabwean infants with normal and stunted growth trajectories, we characterized the development of the gut bacterial and temperate phage communities from birth to 18 moths old. We found that infant gut temperate phages target hallmark early-life bacterial taxa, such as Bifidobacteriaceae, and exhibit an age-dependent maturation that parallels bacterial succession. Notably, both bacterial and temperate phage alpha diversity increased with age. This contrasts with previous studies focused on the extracellular viral fraction and highlights a strong coupling between prophage early-life dynamics and during bacterial gut colonization. Using abundance-based maturation models, we identified successional phases of colonization for both bacteria and their associated temperate viral clusters. Importantly, a viral microdiversity maturation model provided a stronger prediction of chronological age than viral abundance-based model, revealing within-phage genomic variation as a key signal of virome assembly, particularly around weaning. Contrary to findings in wasting (or severe acute malnutrition), stunted growth trajectories were not associated with a significant delay in either bacterial or temperate phage maturation. These results demonstrate that viral genomic variation is a new, informative dimension of early-life gut microbial assembly and that stunting may not impair infants gut maturation process. ImportanceThe early-life period represents a critical window for the establishment of the gut microbial communities, a process that is often affected by environmental factors such as diet. While severe acute malnutrition (SAM) is known to delay bacterial maturation, the impact of chronic, moderate undernutrition, such as stunting is poorly understood. Stunting is a highly prevalent global health condition with irreversible consequences on long-term host health, yet its implications on gut microbiome assembly remain unclear. Our study provides novel insights into the maturation of temperate phages, which to prime the infant gut by colonizing alongside their bacterial hosts and acting as drivers of bacterial evolution via lysogeny. By demonstrating that viral strain-level (genomic) variation captures a stronger age-related signal than viral abundance, we identified an underexplored dimension of microbial assembly. The finding that stunting, in contrast to SAM, does not impact microbial maturation provides essential context for public health interventions and future studies addressing this condition.

Clinically relevant infections commonly develop within polymicrobial environments where interkingdom interactions shape host responses and disease trajectories. Candida albicans and Klebsiella pneumoniae are critical pathogens that can co-exist in the respiratory tract, yet the consequences of their interaction in terms of fungal physiology, pathogenicity and impact on disease outcomes remain poorly understood. Here, we show that K. pneumoniae enhances C. albicans virulence traits suggesting that co-infections could exacerbate lung disease. Mechanistically, bacterial presence induces fungal hyphal morphogenesis via MAPK-CEK signaling, coupled to metabolic rewiring and alterations in cell wall remodeling and septation, resulting in highly elongated hyphae that escape faster from macrophages. At the host level, co-infection reprograms macrophages into a non-canonical state characterized by overlapping pro- and anti-inflammatory modules, integrating type I interferon and IL-10 signaling. This response contributes to tissue damage and facilitates fungal persistence. Our findings reveal that the bacterial-fungal interactions coordinately reprogram pathogen behavior and host immunity, promoting pathogenic synergy and potentially conferring a negative impact on disease outcomes. HIGHLIGHTSCandida-Klebsiella interactions modulate hyphal morphogenesis. Ectopic morphogenesis encompasses septation, cell wall remodeling and carbon metabolism. Candida-Klebsiella co-infections trigger tissue hyperinflammation and compensatory regulation. Candida-Klebsiella co-infection establishes a host environment facilitating microbial dissemination and tissue pathology. Candida-Klebsiella interactions enhance fungal virulence potentially impacting the severity of co-infections