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Structure-resolved virus-host interactomics by cross-linking mass spectrometry.

Viruses depend on host protein networks to replicate, assemble progeny, and spread between cells and organisms. Defining these virus-host protein interactions is challenging because they are highly dependent on infection stage, cell type, species, and because mechanistic interpretation requires information about structural interfaces and conformational states. Cross-linking mass spectrometry (XL-MS) addresses these challenges by adding a spatial and structural dimension to virus-host interactomics in native systems. In this review, we discuss how XL-MS has advanced from targeted analysis of viral protein complexes to structure-resolved mapping of virion architecture and infected-cell virus-host interactomes. We highlight how XL-MS complements AP-MS, cryo-EM/cryo-ET, quantitative proteomics, genetic perturbation, and structure prediction to connect physical proximity with molecular mechanisms. Finally, we discuss current limitations in sensitivity, chemical coverage, temporal resolution, and model interpretation, and outline how future quantitative and integrative XL-MS workflows may enable systems-level structural virology.

Mass Spectrometry

Unraveling critical role of photosynthetic bacteria in sustaining aquatic microbial community stability and function through large-scale genomic data analyses.

The application of photosynthetic bacteria (PB) in water remediation has demonstrated exceptional advantages in terms of high efficiency and low-carbon benefits. However, the limited understanding of PB across natural aquatic environments has constrained the rational development of this strategy. Here, we analyzed 3198 genomic sequencing samples from seven types of natural aquatic ecosystems to investigate the distribution and functions of 42 PB genera commonly used in water remediation. The results showed that the average abundance of the targeted PB reached 9.83 %, with the highest value of 14.93 % observed in River, while Lake harbored the greatest PB genus diversity. PB genera exhibited high sensitivity to salinity, with Rhodoferax dominating freshwater habitats, whereas Rhodovulum was predominant in marine environments. Notably, co-occurrence network analysis revealed that PB were closely associated with microbial community stability and optimized interspecific interactions. Aquatic microbial communities with high PB abundance were characterized by efficient division-of-labor modules, accompanied by enhanced PB-associated functional potential for carbon fixation, denitrification, and sulfur oxidation. In summary, this study systematically elucidates the regional biogeographical patterns and ecological roles of PB in natural aquatic environments, providing a comprehensive scientific basis and theoretical guidance for the development and practical application of PB-based water remediation technologies.

Bacteria

Type VI secretion system activity at lethal antibiotic concentrations leads to overestimation of weapon potency.

Competition assays are a mainstay of modern microbiology, offering a simple and cost-effective means to quantify microbe-microbe interactions in vitro. Here, we demonstrate a key weakness of this method that arises when competing microbes interact via toxins, such as those secreted via the type VI secretion system (T6SS). Time-lapse microscopy reveals that T6SS-armed Acinetobacter baylyi bacteria can maintain lethal T6SS activity against E. coli target cells, even under selective conditions intended to eliminate A. baylyi. Further, this residual killing creates a density- and T6SS-dependent bias in the apparent recovery of E. coli, leading to a misreporting of competition outcomes where target survival is low. We also show that incubating A. baylyi/E. coli co-cultures in liquid antibiotic prior to selective plating can substantially correct this bias. Our findings demonstrate the need for caution when using selective plating as part of T6SS competition assays, or assays involving other toxin-producing bacteria.

Type VI Secretion Systems

Host life-history strategy is a critical determinant of virulent phage infection propensity.

Bacteriophages shape microbial communities through two major lifestyles: virulent (obligately lytic) and temperate (capable of lysogeny). Prevailing phage ecology frameworks focus on how environmental conditions, host density, and physiological state modulate infection modality. This perspective overlooks how host traits exert selective pressure on the distribution of virulent and temperate lifestyles across bacterial species, which limits understanding of phage ecology. To address this critical knowledge gap, we adopt a host-centric, trait-based perspective and use 5821 complete bacterial genomes to build a host life-history space predominantly defined by genome size, metabolic capacity, and growth rate potential. After mapping phage lifestyle association signals, prophage burden formed a continuous gradient across this space. Also, virulent phage association was positively correlated with prophage burden, revealing a nested structure of lifestyle signals. Functional trait analysis identified enrichment of resource-acquisition modules underlying both temperate and virulent associations. Overall, these findings indicate that phage lifestyle is significantly influenced by host life-history strategies, highlighting fast-growing, metabolically versatile hosts as favorable targets for virulent phage isolation and biocontrol applications.

Bacteriophages

Concurrent stimulation of diflufenican biodegradation and changes in the active microbiome in gravel revealed by Total RNA.

The use of slowly degraded pesticides poses a particular problem when these are applied to urban areas such as gravel paths. The urban gravel provides an environment very different from agricultural soils; i.e., it is both lower in carbon and microbial activity. We, therefore, endeavored to stimulate the degradation of the pesticide diflufenican added to urban gravel microcosms amended with dry alfalfa to increase microbial activity. In the present study, alfalfa addition significantly increased the formation of diflufenican's primary metabolite, 2-[3-(trifluoromethyl)phenoxy]nicotinic acid (AE-B), indicating stimulated biotransformation. The concurrent changes of the active microbial communities within the gravel were explored using shotgun metatranscriptomic sequencing of ribosomal RNA and messenger RNA. Although bacterial taxa remained dominant (87.0%-98.5% relative abundance), the alfalfa treatment led to a 4-5-fold increase in eukaryotic groups, including fungi and microbial grazers. Several microbial taxa potentially involved in the degradation of complex carbon compounds and aromatic pollutants-including Bacteroidetes, Verrucomicrobia, Sordariomycetes, Mortierellales, Tremellales, Sphingopyxis, and Phenylobacterium-increased in relative abundance following alfalfa amendment. Functional gene profiling revealed elevated expression of genes related to microbial activity and biomass production. Genes with potential roles in the breakdown of complex carbon structures (e.g., xylanases/chitin deacetylases) and in the transformation of aromatic compounds (e.g., ring-cleaving dioxygenases) were revealed. We conclude that complex carbon amendments can enhance the microbial activity, promoting the biotransformation of diflufenican in urban gravel environments. These findings provide new insights into the interactions between microbial community dynamics, gene expression profiles, and pesticide biotransformation in non-agricultural matrices.IMPORTANCEPesticides used on urban areas, e.g., gravel paths, are likely to have different effects and fates than when these are used on agricultural soils. Hence, studies into the degradation of pesticides applied to urban matrices are needed. We have previously shown that metabolites of the persistent pesticide diflufenican are even more persistent in urban soils, and it has also previously been shown that these metabolites leach from gravel surfaces. The reasons behind this are that the urban gravel provides an environment very different from agricultural soils; i.e., it is both lower in carbon and microbial activity. In the present study, we, therefore, endeavored to stimulate the degradation of the pesticide diflufenican added to urban gravel microcosms amended with dry alfalfa to increase microbial activity, concurrently studying the changes in the active microbiome by Total RNA-metatranscriptomics.

Biodegradation, Environmental

Impacts of host genetics on gut microbiome composition in Alzheimer's disease.

BACKGROUND: Host-microbiome interactions play essential roles in the development of Alzheimer's disease (AD), yet the host genetic impacts on gut microbial alterations in AD remain poorly understood. RESULTS: Here, we simultaneously profiled host genotype and gut microbiome in 252 Chinese individuals with varying degrees of cognitive disability. Using the latent Dirichlet allocation topic model, we identified the Anaerostipes-enriched enterosignature (ES-Ana) at the microbial subgroup level as significantly negatively associated with cognitive disability, which could be recapitulated in external cohorts. With the whole-genome sequencing data, we performed microbiome genome-wide association studies for the ES-Ana relative abundance. We prioritized 41 lead genetic variants and confirmed that the high ES-Ana relative abundance showed a negative correlation with the polygenic risk score of AD, indicating its protective effect against AD. Furthermore, we identified 174 ES-Ana-associated genes, which are enriched in AD-related biological functions and phenotypes, and exhibite pervasive underexpression in glial cells during brain aging. CONCLUSIONS: In summary, our study reveals the complex genetic effects on the gut microbiota in AD, and provides novel evidence for the roles of the gut-brain axis in AD. Video Abstract.

Alzheimer Disease

Benchmarking with synthetic communities provides a baseline for virus-host inferences from Hi-C proximity linking.

Microbiomes influence diverse ecosystems, and viruses increasingly appear to impose key constraints. While viromics has expanded genomic catalogs, host identification for these viruses remains challenging due to the limitations in scaling cultivation-based approaches and the uncertain reliability and relative low resolution of in silico predictions - particularly for understudied viral taxa. Towards this, Hi-C proximity ligation uses sequenced, cross-linked virus and host genomic fragments to infer virus-host linkages and has now been applied in at least 10 studies. However, its accuracy remains unknown. Here we assess Hi-C performance in recovering virus-host interactions using synthetic communities (SynComs) composed of four marine bacterial strains and nine phages with known interactions and then apply optimized bioinformatic protocols to natural soil samples. In SynComs, standard Hi-C sample preparations and analyses showed poor normalized contact score performance (26% specificity, 100% sensitivity, incorrect matches up to class level) that could be dramatically improved by Z-score filtering (Z ≥ 0.5, 99% specificity), though at reduced sensitivity (62% down from 100%). Detection limits were established as reproducibility was poor below minimal phage abundances of 105 PFU/mL. Applying optimized bioinformatic protocols to natural soil samples, we compared virus-host linkages inferred from proximity-ligated Hi-C sequencing with predictions generated by in silico homology-based and machine learning-based bioinformatic approaches. Prior to Z-score thresholding, agreement was relatively high at the phylum to family levels (72%), but not at the genus (43%) or species (15%) levels. Z-score thresholding reduced sensitivity (only 34% of predictions were retained), with only modest improvements in congruence with bioinformatic methods (48% or 18% at genus or species levels, respectively). Regardless, this led to 79 genus-level-congruent virus-host linkages and 293 new ones revealed by Hi-C alone, i.e., providing many new virus-host interactions to explore in already well-studied climate-critical soils. Overall, these findings provide empirical benchmarks and methodological guidelines to improve the accuracy and reliability of Hi-C for virus-host linkage studies in complex microbial communities.

Benchmarking

Function-based selection of synthetic communities enables mechanistic microbiome studies.

Understanding the complex interactions between microbes and their environment requires robust model systems such as synthetic communities (SynComs). We developed a functionally directed approach to generate SynComs by selecting strains that encode key functions identified in metagenomes. This approach enables the rapid construction of SynComs tailored to any ecosystem. To optimize community design, we implemented genome-scale metabolic models, providing in silico evidence for cooperative strain coexistence prior to experimental validation. Using this strategy, we designed multiple host-specific SynComs, including those for the rumen, mouse, and human microbiomes. By weighting functions differentially enriched in diseased versus healthy individuals, we constructed SynComs that capture complex host-microbe interactions. We designed an inflammatory bowel disease SynCom of 10 members that successfully induced colitis in gnotobiotic IL10-/- mice, demonstrating the potential of this method to model disease-associated microbiomes. Our study establishes a framework for designing functionally representative SynComs of any microbial ecosystem, facilitating mechanistic study.

Animals

A deep insight into the sialome of the house fly, Musca domestica, infected with the salivary gland hypertrophy virus (MdSGHV).

The house fly, Musca domestica, serves as a mechanical vector for numerous pathogens, posing a significant risk to human and animal health. More than two decades ago, the Musca domestica salivary gland hypertrophy virus (MdSGHV) was discovered, infecting both males and females flies and disrupting mating and the reproductive process. While MdSGHV can infect various tissues, its primary replication site is the house fly salivary gland. It is well established that arthropod salivary glands play an important role not only in acquiring food but also in transmitting pathogens. Therefore, understanding the composition of vector salivary glands and the interactions between vector and pathogen components is essential for developing future control strategies. To this end, we conducted a comprehensive RNA-sequencing of salivary glands from both infected and non-infected house flies. Our analysis identified a total of 6,410 putative sequences, with 6,309 originating from M. domestica and 101 from the MdSGHV, categorized into 25 functional groups. Furthermore, differential expression analysis between infected and non-infected salivary glands revealed 2,852 significantly modulated transcripts, highlighting profound transcriptional changes triggered by MdSGHV infection. Overall, these findings not only deepen our understanding of the composition of M. domestica salivary glands but also provide valuable insight into the virus-vector interaction, which could serve as a model to understand other medically relevant interactions.

Salivary Glands

Rgg144/SHP144-controlled streptolancidin D mediates intra-species competition in Streptococcus pneumoniae with cumulative effect from other bacteriocins and fratricide.

UNLABELLED: Streptococcus pneumoniae is a major colonizer of the human nasopharynx, where inter- and intra-strain competition plays a critical role in shaping population structure and influencing vaccine outcomes. Bacteriocins are key mediators of intra-species competition, yet many of their functions and regulatory mechanisms remain poorly understood. Here, we identify and characterize streptolancidin D, a previously uncharacterized bacteriocin encoded by the sldA-T locus, and demonstrate its contribution to pneumococcal competition. Using isogenic streptolancidin-producing and non-producing variants of a naturally colonizing strain, we show that sldA-T contributes to the inhibition of competitor strains in in vitro biofilms and during murine co-colonization. Importantly, streptolancidin D also inhibited in vitro a subset of genetically diverse pneumococcal isolates representing multiple serotypes, whereas non-producing variants showed no activity. This indicates that its effect is broad and not restricted to isogenic interactions. Genomic analysis of over 7,500 pneumococcal genomes revealed that sldA-T is present in ~12% of isolates, with lineage-associated distribution patterns, and is consistently encoded downstream of the Rgg144/SHP144 quorum sensing system. We further demonstrate that sldA-T is regulated by this system, with sldA-T promoter activity abolished in a SHP-deficient background and partially restored by exogenous peptide stimulation. Finally, we show that streptolancidin D acts in concert with other bacteriocin systems and competence-mediated fratricide, highlighting a multifactorial antimicrobial strategy that enhances pneumococcal competitiveness. Overall, our findings identify a quorum sensing-regulated bacteriocin that contributes to pneumococcal competition and helps shape population dynamics. IMPORTANCE: Bacteriocins are central to bacterial competition and niche occupation, particularly in structured environments like the human nasopharynx. While several pneumococcal bacteriocins have been characterized, the functions of many remain unknown, limiting our understanding of how these systems shape strain fitness and population dynamics. We characterize streptolancidin D, a bacteriocin that enhances intraspecies competitiveness in vitro and in vivo and contributes to the inhibition of genetically diverse pneumococcal strains. We demonstrate that its expression is tightly regulated by the conserved Rgg144/SHP144 quorum sensing system and that the locus is distributed and shows synteny across multiple pneumococcal lineages. Our findings reveal that streptolancidin D operates within a broader network of bacteriocins and competence-associated mechanisms that collectively shape competitive interactions. By integrating genomic, functional, and regulatory analyses, this work expands the known repertoire of pneumococcal antimicrobial systems and provides new insights into the mechanisms underpinning competition and population structure in S. pneumoniae.

Bacteriocins

Serine: From Metabolic Intermediate to Signaling Entity.

Serine, a nonessential amino acid classically defined as a precursor for protein synthesis and one-carbon metabolism, is increasingly recognized as a signaling metabolite that links the cellular metabolic status to regulatory decision-making. Intracellular serine availability is shaped by nutrient conditions, glycolytic flux, and activity of the serine synthesis pathway, and these fluctuations are sensed to elicit coordinated metabolic and signaling responses. This review discusses mechanisms by which serine modulates cell growth and stress responses, with particular emphasis on its interaction with central nutrient-sensing pathways, including mTORC1 and the integrated stress response. In parallel, serine-driven one-carbon metabolism is examined for its role in supporting nucleotide biosynthesis, methylation reactions, and redox homeostasis through folate-dependent pathways and NADPH generation, thereby coupling anabolic processes to the maintenance of redox balance and genome integrity. In addition to intracellular functions, serine contributes to intercellular signaling. Conversion of l-serine to d-serine mediates neuromodulatory activity via N-methyl-d-aspartate receptors, while serine availability also influences immune cell function, inflammatory signaling, and host-microbe interactions. Dysregulation of serine metabolism and signaling is further considered in the context of disease states, including cancer, neurodegeneration, and metabolic disorders. Together, these observations support a framework in which serine functions as an information-bearing metabolic signal that coordinates the biosynthetic capacity with cellular adaptation and intercellular communication.

Serine

Dissecting host-microbe interactions with modern functional genomics.

Interrogation of host-microbe interactions has long been a source of both basic discoveries and benefits to human health. Here, we review the role that functional genomics approaches have played in such efforts, with an emphasis on recent examples that have harnessed technological advances to provide mechanistic insight at increased scale and resolution. Finally, we discuss how concurrent innovations in model systems and genetic tools have afforded opportunities to interrogate additional types of host-microbe relationships, such as those in the mammalian gut. Bringing these innovations together promises many exciting discoveries ahead.

Genomics

Autoimmune disease-associated pathobionts: mechanisms and therapeutic potential of phage-based approaches.

The gut microbiota is a critical regulator of systemic immune homeostasis; accumulating evidence implicates specific commensal bacteria, termed "pathobionts," in autoimmune disease pathogenesis. However, the definition of pathobionts remains context-dependent, as their effects are influenced by host genetics and host-microbe interactions. In this review, we summarize representative pathobionts supported by functional evidence in selected extraintestinal autoimmune diseases and discuss how these mechanisms may inform phage-based microbiome-targeted interventions. Mechanistically, pathobionts contribute to autoimmune disease through multiple pathways, including molecular mimicry, induction of intestinal T helper 17 and T follicular helper cell responses, disruption of regulatory T cell homeostasis, intestinal barrier dysfunction, and bacterial translocation from the gut to extraintestinal sites. These processes highlight the central role of gut-associated lymphoid tissue in initiating systemic autoimmunity, and targeting disease-associated microbes represents a promising therapeutic strategy. Whole-phage therapy, which enables highly specific bacterial elimination, has shown efficacy in preclinical immune-mediated disease models, but may be affected by variable in vivo replication, bacterial receptor-mediated resistance, anti-phage immune responses, and ecological effects on the resident microbiome. Phage-derived enzymes that lyse bacterial cell walls, such as endolysins, represent a complementary therapeutic modality that specifically targets bacterial peptidoglycan through cell wall-binding and catalytic domains. Collectively, these findings support the concept that pathobiont-targeted interventions, particularly phage-based strategies, may provide microbiome-directed, immunosuppression-sparing therapeutic approaches for selected patient subsets.

Humans

Desert-derived Ensifer sp. SA403 enhances potato salt tolerance by reshaping rhizosphere microbiome functions and host responses.

Soil salinization increasingly threatens global food security, and potato (Solanum tuberosum L.), a moderately salt-sensitive crop, is particularly vulnerable to saline soils. Plant growth-promoting rhizobacteria (PGPR) offer a promising strategy to improve crop performance, yet how PGPR interact with native microorganisms to enhance potato salt tolerance remains poorly understood. In this study, we identified a desert-derived PGPR strain, Ensifer sp. SA403, which substantially enhanced potato performance under high salinity across sterile, non-sterile and field conditions. Physiologically, inoculation with SA403 reduced shoot Na⁺ accumulation and increased the K⁺/Na⁺ ratio; notably, these effects were markedly stronger in non-sterile substrates than under sterile conditions, indicating that SA403-mediated ion homeostasis relies on cooperation with the resident microbiota rather than on the strain acting alone. Metagenomic profiling indicated that SA403 strain reshaped rhizosphere communities, significantly enriching beneficial taxa such as Priestia and Bradyrhizobium, and upregulated functional pathways involved in glutathione and sulfur metabolism. Furthermore, host transcriptomic analyses showed that SA403 modulated plant responses to salt stress, with differentially expressed genes enriched in jasmonic acid signaling, ethanolamine metabolism and amino-acid biosynthesis pathways. Field trials on saline soils confirmed that SA403 significantly increased seedling emergence and tuber weight. Together, our results demonstrate that SA403 functions as a biological mediator that optimizes rhizosphere microecology and coordinates ion balance and host signaling to enhance potato salt tolerance. These findings support the potential of SA403 as a robust PGPR-based tool for sustainable potato production on saline soils.

Rhizosphere

Lack of in vitro antiviral activity of fluoroquinolones against herpes simplex virus type 2.

The antiviral activity against herpes simplex virus type 2 (HSV-2) of five fluoroquinolones (ciprofloxacin, lomefloxacin, ofloxacin, pefloxacin, rufloxacin) was tested in vitro. Their efficacy was evaluated as reduction of the cytopathic effect (CPER) exerted by HSV-2 on Vero cells in comparison with novobiocin and acycloguanosine. Our results show a very poor antiviral effect of five quinolones (CPER50 = 200 mg/l) that was comparable with their cytotoxicity (TCIC50 less than 200 mg/l). Novobiocin shows a lower toxicity (TCIC50 = 400 mg/l) and a slight antiviral activity (CPER50 = 120 mg/l). Acycloguanosine shows a TCIC50 greater than 400 mg/l and a CPER50 of 3.125 mg/l. The therapeutic indices gave values ranging from 0.12 to 2 for quinolones, of 3.3 for novobiocin, and greater than 128 for acycloguanosine. The antiviral efficacy of acycloguanosine was not affected by concentrations of quinolones active against bacteria (1-10 mg/l) whereas it was drastically reduced by higher doses of quinolones (greater than 50 mg/l). Our data suggest that fluoroquinolones cannot be considered drugs able to inhibit HSV-2 replication in vitro.

Acyclovir

Advancing the Deciphering of Host-Microbe Crosstalk with Spatial Omics: A Mini-Review.

Host-microbe crosstalk refers to the reciprocal influences between a host and its resident or invading microorganisms. This crosstalk plays important roles in maintaining host health, regulating physiological functions, and coordinating responses to infection. The rapid rise of spatial omics is transforming how this crosstalk is studied in both animals and plants. Unlike traditional bulk omics, which homogenize tissues and erase spatial context, spatial methods preserve in situ organization and can simultaneously capture molecular information from hosts and microbes. As a result, researchers can characterize the spatial organization of colonization and infection, identify spatial associations between microbial niches and host cell states, and visualize local host response gradients across intact tissues. Current spatial omics technologies encompass sequencing-based, imaging-based, and hybrid platforms. Spatial multi-omics approaches enable the joint measurement or integration of gene expression, protein abundance, and metabolite distributions. Although spatial association alone does not establish causality, spatial omics provides a high-resolution framework for characterizing host-microbe relationships within intact tissues and generating spatially constrained, testable hypotheses. When combined with perturbation experiments and complementary experimental evidence, these hypotheses can contribute to mechanistic interpretation of host-microbe crosstalk. Here, we review spatial omics technologies, compare their suitability and major trade-offs for host-microbe studies, and discuss computational strategies, analytical challenges, and future prospects.

Multiomics

A global survey of taxa-metabolic associations across mouse microbiome communities.

Host-microbiota mutualism is rooted in the exchange of dietary and metabolic molecules. Microbial diversity broadens the metabolite pool, with each taxon contributing distinct compounds in varying proportions. In the human microbiome, high variability in consortial composition is largely compensated by similar metabolic functions across different taxa. However, the extent of compensation in lower diversity mouse models, and whether vivaria are metabolically equivalent, is unknown. We provide a searchable resource of microbiome composition variability across 51 murine vivaria and 12 wild mouse colonies worldwide, with vivarium-specific variants mapped according to predicted 3D structures for each microbial species. Our matched metabolomics data show that realized metabolic potential has relatively low variability, providing functional evidence for metabolic compensation. Additionally, variability is related to taxonomic composition rather than vivarium, revealing taxa-metabolite associations that are potentially relevant to phenotypic differences between vivaria. Collectively, this resource offers tools to strengthen microbiome studies and collaborative science.

Animals