The ISME Journal

As key drivers of nitrification, ammonia-oxidizing archaea (AOA) play a central role in the global nitrogen cycle and contribute significantly to the emissions of the potent greenhouse gas nitrous oxide (N2O). However, the ecological implications of AOA growth as biofilms, remain poorly understood. Since nitrite production can be used to follow cellular activities directly we were able to compare biofilms with planktonic cells of the terrestrial model AOA Nitrososphaera viennensis at ecologically and agriculturally relevant conditions. Biofilms were more resistant across nearly all tested conditions and remained active at lower temperatures, acidic pH, and high ammonium concentrations. Collectively, activities in biofilm help reconcile discrepancies between earlier laboratory and environmental observations of soil AOA. Additionally, biofilms showed a high general resilience and lowered sensitivities to nitrification inhibitors. Although in situ biofilms grown in microrespiratory chambers exhibited activity and ammonia affinity similar to planktonic cells, biofilm cultures produced only half as much N2O. The enhanced fitness of biofilms across all tested conditions vastly expands the potential ecophysiological niche of AOA and supports the hypothesis that biofilm growth represents the in situ phenotype of AOA in soil environments.

Host-associated microbiomes are typically maintained in stable configurations that support host fitness, yet the mechanisms by which metabolic perturbations destabilize these communities remain poorly understood. Using the freshwater cnidarian Hydra vulgaris AEP, we systematically assessed microbiome responses to 326 single-metabolite perturbations. Only 17 metabolites, mostly amino acid-related compounds, induced significant compositional shifts in the microbial community. Most shifts are accompanied by transitions from Curvibacter- to Pseudomonas-dominated or Legionella-dominated states, indicating the existence of three alternative community states which can be induced by metabolic triggers. Integrating 16S sequences with functional genomic information, we found that {beta}-diversity strongly predicted functional shifts, whereas reduced -diversity was associated with loss of metabolic functions. The metabolite perturbations also altered host-microbe interactions, affecting pathogenicity-, glycocalyx-, and nitrogen-related functions. In particular, nitrogen metabolism shifted from ammonia oxidation in Curvibacter-dominated communities to ammonia reduction in Pseudomonas-dominated states. Experimental validation confirmed that Pseudomonas metabolizes L-arginine and drives environmental ammonia accumulation to levels that could impair Hydras fitness and induce disease phenotypes. Conversely, Limnobacter was found to scavenge the environmental ammonia, potentially mitigating the adverse effects. These results demonstrate that metabolite-driven niche reconfiguration can destabilize host-associated microbiomes by coupling compositional shifts to functional change and host pathology, identifying metabolite-driven niche restructuring as a mechanism linking microbial community instability to host dysfunction.

Protistan predators are key regulators of microbial food webs, yet most are considered to occupy relatively narrow trophic niches. Here, we demonstrate that Mayorella spp. (Amoebozoa), isolated from marine and freshwater environments, exhibits exceptional trophic breadth spanning multiple trophic levels. Live-cell imaging revealed predation on bacteria, algae, dinoflagellates, diatoms, flagellates, ciliates, and multicellular prey including rotifers. Large or filamentous prey were engulfed whole or mechanically fragmented during ingestion. Notably, Mayorella consumed both trophozoites and cysts of free-living amoebae (Naegleria and Acanthamoeba), with clear digestion of cyst contents. Dense cultures showed aggregation around large prey and facultative cannibalism. Ingestion of microplastic-like particles occurred without evidence of digestion. Predator cell size and population density increased markedly when feeding on protist or mixed prey relative to bacterial diets, indicating pronounced trophic plasticity. These findings establish Mayorella as a broad-spectrum, cross-trophic predator with the capacity to exert top-down effects across microbial food webs and suggest a previously underappreciated role in the suppression of pathogenic free-living amoebae.

Microbial habitats that receive repeated external input may not remain shaped by that input forever if local retention allows resident communities to build up over time. Here, we used a controlled bench-scale sink p-trap system to examine how community assembly unfolded during initial establishment in new, bleach-treated p-traps. Two p-traps received repeated handwashing-water input, while one received tap water as baseline. The treated p-traps, but not the control, showed clear successional change toward later resident-like states. Nested-model comparisons further showed that recent external input had its greatest influence early in succession, but the p-traps own prior state remained the stronger predictor throughout. Final-day post-flush trajectories indicated short-term displacement from pre-flush positions, with later time points tending to move back toward late-stage resident centroids. Together, these results show that repeated inoculation does not necessarily keep communities under continued outside influence. Instead, retentive microbial habitats can shift over time from early sensitivity to external input toward persistence shaped more by local history.

1Consortia of archaea and partner bacteria couple the anaerobic oxidation of alkanes to sulfate reduction. While catabolic pathways in anaerobic alkane-oxidizing archaea (ANKA) are increasingly understood, their anabolic capacities remain poorly characterized. Here, we examined nine enrichment cultures dominated by ANKA and their partner bacteria for small-molecular compounds using solvent extraction and gas chromatographic analysis of derivatized extracts. All hydrocarbon-degrading cultures contained substantial amounts of disaccharides in their metabolite pools. Cold-adapted methane-oxidizing cultures dominated by ANME-2c and Seep-SRB2 contained up to 1.5 mg of trehalose per mg soluble protein. Trehalose was also abundant in ethane-oxidizing cultures of Candidatus Ethanoperedens and its distinct partner SRBs, accounting for up to 75 % of the extracted metabolites. In contrast, thermophilic ANKA cultures dominated by ANME-1 or Ca. Syntropharchaeum and Ca. Desulfofervidus contained an abundant as-yet-unidentified glucose-containing disaccharide. Metagenomic analysis revealed widespread trehalose metabolism genes among partner Desulfobacterota and in ANME-2c and Ca. Ethanoperedens, but a lower potential in ANME-1 and Syntropharchaeum, consistent with metabolite profiles. If exogenous trehalose was added to the Ethane50 culture, we observed rapid metabolization by heterotrophic microorganisms, but poor assimilation by the Ca. Ethanoperedens/ Ca. Desulfofervidus core community, indicating that ANKA/SRB consortia do not consume externally supplied trehalose. Instead, Ca. Ethanoperedens/ Ca. Desulfofervidus, as well as other ANKA/SRB consortia, may use the disaccharides as energy-storage molecules, osmolytes, or components of the extracellular matrix. Notably, the disaccharides produced by the consortia also sustain ancillary heterotrophs, thereby linking alkane oxidation to broader sedimentary carbon cycling.

Patescibacteria are an elusive linage of "microbial dark matter" bacteria predicted to represent [~]25% of total bacterial diversity. Despite this abundance and ubiquity, these organisms are challenging to cultivate, resulting from their specialized episymbiotic lifestyle. All cultivated representatives to date, predominantly composed of Saccharibacteria from the oral microbiome, depend on cognate prokaryotic hosts for growth and reproduction. Studying the growth dynamics of episymbiotic bacteria and their hosts in batch cultures has suggested that many episymbionts initially reduce host populations, and that hosts eventually adapt to episymbiont stress after serial passaging. However, discontinuous batch cultures do not reflect natural interactions between these organisms due to their drastically different growth rates. An episymbiont requires several ([~]2-4) serial passages alongside its host to reach the high cell densities needed to impact host growth, which complicates investigation of host inhibition and adaptation to episymbiont stress. To describe these dynamics accurately, we utilized continuous culture via small-scale Raspberry Pi powered bioreactors, called Pioreactors. Within a bioreactor, host bacteria can be cultivated at a consistent growth rate indefinitely, providing the perfect substrate for cultivation of model Saccharibacteria. Quantification of time until host crash, crash severity, time until recovery, and stable co-culture density provides mechanistic ways to describe episymbiont-host interactions. First, we used these techniques to compare episymbiont infection by three different episymbionts, revealing distinct infection patterns ranging from mild inhibition with rapid host adaptation, to rapid host collapse followed by "arms race" oscillation dynamics. Then, bioreactors were used to quantify the episymbiotic role played by a known host-binding type 4 pili (T4P-2), demonstrating that loss of long-distance host binding significantly delayed the host crash without altering general crash dynamics. These experiments reveal that episymbionts can have drastically different effects on bacterial communities and provide the tools necessary to describe strain/species differences and molecular interactions. ImportanceEpisymbiotic Patescibacteria represent one of the largest branches of life on Earth, as well as one of the least understood. Furthermore, because Patescibacteria can manipulate their hosts growth and morphology they have immense ecological potential to be shaping the communities they occupy, both environmental and microbiome-associated. Our study highlights for the first time the potential of small-scale continuous cultivation for studying episymbiotic interactions that cannot be captured in discontinuous cultures. Herein we used these techniques to interrogate inter-species variation in host inhibition potential and to determine how loss of a long-distance episymbiosis factor mechanistically alters the cycle of episymbiont infection; however, this cultivation platform will enable researchers to answer many new questions about these ubiquitous host-episymbiont interactions.

BackgroundLeafhoppers are among the most important insect vectors of plant pathogens worldwide and depend on microbial symbionts to exploit nutrient-poor phloem diets. However, most studies of leafhopper-associated microbiota have focused on a limited number of taxa or marker-gene surveys, leaving the genomic diversity, ecological organization, and functional potential of these microbial communities poorly understood. Here, we generated the Global Leafhopper Microbiome Catalog by integrating genome-resolved metagenomics from 171 leafhopper species across 11 subfamilies and 13 countries, including the first microbiomes characterized from Arctic leafhoppers. ResultsDe novo assembly and genome reconstruction generated 337 high-quality non-redundant microbial genomes and 18.6 million non-redundant genes, substantially expanding the known microbial diversity associated with Cicadellidae, including several previously undescribed bacterial lineages. Comparative analyses revealed a recurrent modular microbiome architecture composed of: (i) a conserved core of obligate nutritional symbionts, dominated by Candidatus Karelsulcia and Candidatus Nasuia; (ii) a heterogeneous layer of secondary symbionts, including Wolbachia, Arsenophonus, Rickettsia, and Diplorickettsia; and (iii) a dynamic pool of environmentally acquired bacteria. While obligate symbionts remained highly conserved across divergent hosts, secondary and environmental taxa varied substantially among species and regions, suggesting repeated acquisition shaped by ecological filtering rather than host phylogeny alone. Comparative analyses between the specialist corn leafhopper Dalbulus maidis and the more polyphagous aster leafhopper Macrosteles quadrilineatus further showed that closely related vectors can maintain conserved ancestral symbionts while harboring markedly distinct accessory microbiomes. Arctic populations contained unique microbial assemblages enriched in functions associated with cold tolerance, oxidative stress, and reproductive manipulation. In addition, we identified numerous plant-associated bacteria, including phytoplasmas, spiroplasmas, Pantoea, and Erwinia, alongside taxa with predicted nutritional and plant growth-promoting functions. ConclusionsOur findings reveal that leafhopper microbiomes are structured through the interaction of ancient obligate symbioses and flexible environmentally responsive microbial layers. This work establishes a genome-resolved framework for understanding microbiome evolution in insect vectors and highlights the potential role of microbial community structure in host adaptation, pathogen ecology, and sustainable pest management.

Long term chronic infections of plants by bacterial pathogens are largely unknown. Understanding how pathogens adapt during chronic infection provides a key insight to pathogen evolutionary strategy both for persistence and survival, but also for potential future outbreaks. Pseudomonas savastanoi pv. fraxini (Psf), a member of phylogroup 3 within the Pseudomonas syringae species complex, causes canker disease in European ash (Fraxinus excelsior). Infections persist for years within the bark parenchyma, where bacteria are enclosed in cavities that contribute to the gradual expansion of host periderm. This pathosystem therefore provides an opportunity to examine pathogen evolution in a long-lived, largely unmanaged host. We combined population genomics and phenotypic analysis of 124 Psf strains collected from six sites across the UK. Phylogenetic analysis revealed a highly clonal population, with only 833 core genome SNPs across a 5.3 Mb genome, and a relatively small accessory genome largely shaped by gain and loss of large mobile genetic elements. Despite this limited genomic diversity, mutations were enriched in regulatory genes, including two-component systems, chemotaxis proteins, and cell envelope-associated loci. Notably, the global regulator gacA/S was independently mutated multiple times within the same clonal lineage. These mutations, typically small deletions, were associated with changes in motility, nutrient utilisation, stress tolerance, and virulence across genetic backgrounds. As a result, phenotypic heterogeneity was observed within otherwise clonal populations, including within individual lesions. These findings indicate that repeated mutation of regulatory systems represents a key mechanism of adaptation in this chronic plant-pathogen interaction, enabling phenotypic diversification despite limited sequence divergence. This study provides a microevolutionary perspective on P. syringae populations in the phyllosphere and highlights the role of regulatory variation in the evolution of low-virulence, ecologically restricted pathogens.

Viruses are abundant and ecologically important in soils, yet the persistence and production dynamics of extracellular virions remain poorly understood. We applied a genome-resolved stable isotope probing viromics (SIP-viromics) approach, combining H 18O labeling with viral metagenomics, to track virion turnover in seasonally dry grassland soils following rewetting. We identified 354 viral populations (vOTUs) using individual-sample and combined metagenome assemblies. Only 22% of vOTUs exhibited significant 18O enrichment, indicating active replication and new virion production during the 1-week incubation; the majority (78%) persisted without detectable replication, consistent with a viral seed bank. Active vOTUs accounted for 4.76-5.15% of total virions per gram of soil, with viral loads ranging from 3.15 x 1010 to 6.59 x 1010 virions per gram. Probabilistic and deterministic sensitivity analyses spanning viral DNA fraction and genome length reinforced that persistent virions represented the majority of the extracellular viral pool post-wet-up, regardless of parameter assumptions. Host predictions linked both active and persistent vOTUs primarily to Actinomycetota and Pseudomonadota--bacterial groups known to rapidly resuscitate following rewetting--suggesting that some viruses exhibit rapid turnover while others persist over longer timescales, forming a stable viral pool capable of reinitiating infections during favorable conditions. These results demonstrate that SIP-viromics can distinguish newly produced from persistent virions and reveal host-associated patterns of lytic infection and virion production. Our findings advance understanding of soil virus-host interactions and highlight the ecological role of persistent virions as a genetic reservoir contributing to microbial turnover and biogeochemical cycling following environmental disturbance. ImportanceUnderstanding the persistence and production dynamics of soil viruses is critical for elucidating their roles in microbial community dynamics and nutrient cycling, yet these processes have remained largely uncharacterized due to methodological limitations. By integrating stable isotope probing with viromics, this study provides a robust framework for directly distinguishing newly produced from persistent virions in situ. Unlike conventional viromics, which only catalogs viral diversity, SIP-viromics enables quantification of active viral replication and persistence under natural soil conditions. Our results demonstrate that most virions in a seasonally dry soil persisted through a rewetting event, with active replication limited to a minority of viral populations. Persistent virions were primarily linked to dominant bacterial groups, indicating that host ecophysiology and environmental stability strongly influence lytic infection. Collectively, these findings highlight viruses as long-term reservoirs of genetic material, capable of shaping microbial dynamics and ecosystem processes over time. This work establishes SIP-viromics as a powerful approach for studying virus-host interactions and their ecological significance in terrestrial environments.

BackgroundCloud water harbors diverse microbial communities despite its extreme oligotrophic conditions. However, the ecological and evolutionary dynamics of viruses in these transient atmospheric habitats remain poorly understood. Clouds have traditionally been regarded primarily as passive carriers of microorganisms rather than as active ecological environments supporting microbial interactions. In this study, cloud water was sampled at Mount Verde, Cape Verde Islands (744 m a.s.l.). We performed metagenomic analyses of iron-flocculated cloud water alongside genome analyses of a bacterial isolate and metagenome-assembled genomes using established bioinformatic approaches. Viral diversity, virus-host interactions, metabolic functions, genetic adaptations, and viral population dynamics across cloud events were investigated. In addition, UV-B resistance experiments were conducted for a novel cloud-water isolate. ResultsWe isolated 24 cloud water bacteria, including four novel species lineages, and recovered 62 high-quality metagenome-assembled genomes, including 10 novel species lineages. We identified 458 viral operational taxonomic units and 237 virus-host linkages across diverse prokaryotic hosts, revealing active viral predation across diverse bacterial taxa. In addition, CRISPR spacer matches from isolates of novel bacterial lineages such as Deinococcus nubigenus MPC36 were found. Viruses carried genes involved in host adaptation to environmental stressors, including cold-shock response, UV radiation resistance, and osmotic stress. In addition, viral populations exhibited SNP-level microdiversity and shifts in single-nucleotide variant composition across temporally proximate cloud events, indicating rapid population turnover. Experimental characterization of the cloud isolate Curtobacterium nubigenum MPC39 further revealed pronounced resistance to UV-B radiation and the presence of an inducible prophage, Curtobacterium phage vB_CnuS_Cirrus1 assigned to the new viral family Nebulaviridae, which could be validated in transmission electron microscopy. Reconstructed genomes from cloud-associated bacteria encoded carbon monoxide dehydrogenase genes and UV resistance genes, suggesting trace gas metabolism and enhanced UV protection as survival strategies in oligotrophic cloud droplets. In silico replication rates estimated using iRep were consistent with active bacterial replication at the time of sampling. ConclusionsTogether, these findings demonstrate that clouds are not merely passive carriers of microorganisms, but dynamic atmospheric ecosystems in which virus-host interactions shape microbial diversity and contribute to microbial turnover, atmospheric dispersal, and cloud-water biogeochemistry.

Phyllosphere microbiomes are increasingly recognized as key regulators of plant health and stress responses, although they are also known to change considerably over both space and time. In the phyllosphere, members of the genus Methylobacterium are often abundant and ecologically important as plant growth promoting bacteria. However, knowledge about the temporal abundances and community dynamics of Methylobacterium in agricultural systems remains limited. To address this gap, we characterized seasonal shifts in Methylobacterium-specific and total phyllosphere bacterial loads and community structure on two common summer crops and one overwintering cover crop. Leaf samples of Zea mays (corn), Glycine max (soybean), and Thlaspi arvense L. (pennycress) plants were collected over one year in Minnesota, USA and analyzed with host-associated microbial PCR (hamPCR). Microbial loads and community composition varied strongly among hosts and across growing seasons. Corn supported the highest Methylobacterium and total bacterial loads, increasing towards senescence, while pennycress exhibited the lowest loads and the most distinct communities. While there were strong host-specific patterns, a group of most abundant genera were shared across all crops (Methylobacterium, Sphingomonas, Pseudomonas, and Massilia) and the most abundant Methylobacterium amplicon sequence variants were present on all three hosts. Our findings highlight how microbial loads and community composition change during phyllosphere assembly across diverse summer and overwintering crops, with a small core of versatile taxa dominating multiple agricultural hosts. Understanding these host and season-linked patterns provides a foundation of harnessing Methylobacterium strains to enhance crop productivity and resilience.

Corals are complex holobionts, encompassing numerous prokaryotes, viruses, and protists. This associated microbial community strongly influences coral health and its resilience to global ocean warming. Corallicolid apicomplexans are widespread coral-infecting parasites, yet their impact on the coral host remains poorly understood. This knowledge gap largely stems from the low abundance of these parasites in coral tissues, which makes them difficult to isolate and access to their genetic material. Here we analyzed nearly 1,000 Pocillopora coral colonies collected from 32 islands during the Tara Pacific expedition to identify the drivers of corallicolid prevalence and abundance. Corallicolids were detected in almost all Pocillopora colonies with variable relative abundances between islands. The high abundance of specific corallicolid populations correlates with seawater temperature and levels of host protein carbonylation. We used a large collection of 297 metatranscriptomes to assemble a corallicolid transcriptome and we identified apicomplexan parasite signature genes, including the GRA9 and PV2 confirming the close phylogenetic relationship with the family of Eimeriidae. Gene expression patterns indicate that the high abundance of corallicolids correlates with a high transcription of genes encoding apical complex proteins and genes involved in the control of host immune defenses. Overall, this study provides new insights into corallicolid biology and its interaction with the coral host by combining a newly generated transcriptome with a large-scale sampling of Pocillopora corals across the Pacific Ocean.

Inferring bacterial growth rates is fundamental to understanding microbial interactions and community dynamics, but remains difficult in natural settings where timepoints are limited or organisms are unculturable. In these cases, a widely used method is the origin-to-terminus ratio, or peak-to-trough ratio (PTR), which estimates DNA replication activity by comparing the copy number of DNA at the replication origin and terminus. While PTR correlates well with cellular growth in uniform, idealized environments, it measures replication rather than net growth rate, and thus reflects growth only when there is no cell death. Despite this, PTR is widely applied across a range of laboratory and environmental contexts, where microbial populations frequently experience fluctuating stress, mortality, and subpopulation heterogeneity. Given its widespread use in such settings, we developed a stochastic, cell-based model that explicitly tracks DNA replication and cell death to quantify how different patterns and levels of mortality affect the relationship between PTR and net growth rate. We found that PTR and net growth rate are tightly correlated in idealized conditions; however, systematic deviations emerge when death rates vary over time or across subpopulations. We experimentally validated these predictions by exposing Escherichia coli to osmotic shock or antibiotics, and measuring net growth rate (by spot plating and observing the change in colony counts over time) and DNA replication activity (from qPCR with primers for the origin and terminus). Consistent with the predictions from our model, PTR correlated strongly with net growth rate in standard rich media, but not under stress. Together, these results provide a mechanistic and quantitative framework that clarifies the biological conditions under which PTR can be interpreted as a proxy for net growth rate.

The Canadian Arctic Archipelago (CAA) is warming at an unprecedented rate, leading to sea ice loss and glacial retreat. Marine-terminating (tidewater) glaciers can fuel summertime marine productivity by delivering nutrient-rich deep waters via upwelling to the surface ocean. While the impact of glacier-induced upwelling has been well-studied in the context of phytoplankton and primary productivity, its effects on broader marine microbial communities remain poorly understood. We investigated how glacier-driven upwelling shapes marine microbial (bacterial and archaeal) communities across a series of sites in the CAA. At upwelling sites, the upper 50 m of the water column exhibited elevated nutrient concentrations and physical characteristics that resembled deeper waters, which were associated with differences in microbial community composition relative to non-upwelling sites. Our results indicate that upwelling influences microbial communities in surface waters in two ways. It directly introduces typically deeper-water-associated taxa into surface waters and reshapes ecological niches by enhancing nutrient supply and stimulating primary production, indirectly driving changes in microbial communities. The enrichment of Candidatus Nitrosopumilus, a deep water nitrifier, likely affects nitrogen cycling and raises the possibility of active nitrification in surface waters. Likewise, the increased abundance of taxa known to be associated with phytoplankton-derived organic matter in upwelling regions suggests an enhanced capacity to process organic matter generated from elevated primary productivity. Ultimately, as tidewater glaciers continue to retreat, the resulting changes in the glacially-driven upwelling regime will likely shift marine microbial communities towards assemblages adapted to less productive ecosystems, with implications for nutrient cycling in these systems. ImportanceClimate change has a disproportionate impact on the Arctic, with rising temperatures causing increased marine-terminating glacier retreat and changes in the marine water column structure. The consequent loss of the ability of these glaciers to upwell deep water to the surface ocean results in a reduction of nutrient delivery and mixing in these ecosystems. Previous work has highlighted the importance of marine-terminating glaciers in sustaining phytoplankton productivity during the summer season through this delivery of deep-water nutrients to the surface ocean. The impact of glacially-induced upwelling on marine bacterial and archaeal communities, however, remains underexplored. We found that in regions with glacially-driven upwelling, the surface ocean showed enrichment of phytoplankton-associated taxa and nitrifiers commonly associated with deep waters. This work underscores the role of glacially-driven upwelling in structuring both microbial communities and nutrient cycling, suggesting that glacier loss could reshape community composition and biogeochemical processes in a rapidly changing Arctic.

The role of chemodiversity in plant-insect interactions is widely recognised. However, our understanding of the extent to which chemodiversity connects both partners remains limited. Here, we investigated how aphid chemistry is linked to their plant diet and whether aphids capture plant inter- and intraspecific chemodiversity. Up to 93% of aphid chemical features were detected in plants. Untargeted metabolomics of aphids feeding on diets composed of distinct species or chemotypes within species unveiled the aphid capacity to capture inter- and intraspecific chemodiversity. Multiple chemodiversity indices and metabolic features significantly tracked diet variation and plant chemotypes were reflected in aphid metabolites. These features included phenolics and amino acids, likely ingested with the phloem sap, and fatty acids and terpenoids, potentially captured from the leaf surface. Overall, these findings expand our knowledge of the aphid plant-derived chemical repertoire and highlight that plant chemodiversity can be transmitted, supporting the need for chemodiversity preservation programs.

Aquaculture is an essential food production sector for meeting the global demand for high-quality protein. However, the sector faces significant challenges from bacterial pathogens, particularly Vibrio anguillarum, which causes vibriosis in numerous commercially important fish species. Current disease management strategies rely heavily on antibiotics, leading to antimicrobial resistance and environmental concerns. Microalgal microbiomes represent promising alternatives for sustainable pathogen control, yet the molecular mechanisms underlying their inhibitory activity remain poorly understood. Here, we employed an integrated multi-omics approach to elucidate the mechanisms by which the microbiome of microalga Isochrysis galbana inhibits the highly virulent fish pathogen V. anguillarum strain 90-11-286. Using a GFP-based inhibition assay, we confirmed potent pathogen suppression by the algal microbiome, achieving complete inhibition at a 1:1000 ratio of pathogen to microbiome. Through 16S rRNA gene amplicon sequencing, metagenomics, metatranscriptomics, and metabolomics, we characterized community composition, genomic potential, gene expression patterns, and metabolite production during pathogen challenge. The inhibitory microbiome was dominated by Alteromonas macleodii and Vreelandella alkaliphila, with high-quality metagenome-assembled genomes revealing substantial secondary metabolite biosynthetic potential. Metatranscriptomic analysis revealed active expression of biosynthetic gene clusters encoding, for example, non-ribosomal peptide synthetases, particularly a siderophore gene cluster in V. alkaliphila. Metabolomic profiling confirmed the production of hydroxamate siderophores in the microbiome, including desferrioxamine analogues, proferrioxamine G1t, and tenacibactin D, which accumulated during pathogen inhibition, as well as 10 putative new compounds. Notably, siderophore production was constitutive rather than pathogen-induced, suggesting iron competition as the primary inhibitory mechanism. Our findings demonstrate that iron sequestration through siderophore production represents a key strategy for pathogen suppression in marine microbial communities. This work provides molecular evidence for microbiome-mediated disease control and establishes a foundation for developing rationally designed multi-strain probiotic consortia for sustainable aquaculture applications, offering an environmentally friendly alternative to antibiotic-based pathogen management strategies.

Stability is a desirable property for agricultural microbiomes, but there is a poor understanding of the mechanisms that mediate microbial community stability. A representative bacterial synthetic community from maize roots has been proposed by Niu et al. (2017, PNAS, 114:E2450) as a model system to study microbiome stability. This SynCom assembles stably when all seven members are present, but community diversity collapses without the keystone E. ludwigii strain. In this study, we used complementary in vitro experiments and in silico metabolic modelling to assess the role of metabolites for the stability of this SynCom, by defining the metabolic niches occupied by each strain, as well as their cross-feeding phenotypes and B-vitamin dependencies. We show that the individual member strains occupy complementary metabolic niches, measured by the depletion of distinct metabolites in exometabolomic experiments, as well as contrasting growth phenotypes on diverse carbon substrates, patterns which are largely recapitulated by computational simulations. Minimal medium experiments show that the established seven-member community comprises a mixture of prototrophic and auxotrophic strains. Correspondingly, experimental and in silico cross-feeding phenotypes showed that spent media harvested from the prototrophic strains can sustain growth of two auxotrophs and let to the identification of B-vitamin dependencies. Altogether, this study highlights the complementary power of in vitro and in silico approaches and suggests that the metabolic mechanisms of this SynCom can serve as design principles to inform the rational assembly of stable plant-associated microbial communities.

While Antarctic terrestrial ecosystems support low metazoan diversity, the surrounding marine macrobenthos is rich. However, marine meiofauna remains historically neglected, leaving its diversity patterns unclear. In this study, we used 18S rRNA gene metabarcoding alongside an enhanced taxonomic annotation pipeline to characterize marine meiofauna diversity in the Ross Sea, comparing it to global datasets. We evaluated how depth, habitat type, and mesh size influence community structures to test if habitat heterogeneity drives diversity despite the harsh Southern Ocean conditions. Our results revealed exceptionally high diversity, with metazoans richness comparable to or higher than temperate regions. Although environmental variables had limited effects on taxonomic richness, they significantly shaped community composition, with habitat type explaining the highest proportion of variance. Interestingly, we detected several ASVs 100% identical to North Sea and North Atlantic sequences, likely reflecting the limited taxonomic resolution of the 18S marker rather than global dispersal (the "meiofaunal paradox"). Overall, these findings demonstrate that Antarctic marine sediments host rich meiofaunal communities where ecological processes operate similarly to other global regions, contrasting sharply with depauperate continental Antarctic ecosystems.

Methane is a powerful greenhouse gas. Typically, a large fraction of the methane formed in coastal sediments is removed via anaerobic methane oxidation (AOM). Here, we demonstrate the potential for a range of AOM pathways in brackish coastal sediments by ANME-2a archaea. At our study site, geochemical profiles indicate that AOM is primarily restricted to a shallow, metal-oxide-rich sulfate-methane transition zone (SMTZ). ANME-2a were the sole methanotrophs detected, and metatranscriptomics showed the highest expression levels of the ANME-2a genes in the SMTZ. AOM activity was observed in sediment incubations with various electron acceptors, including sulfate, metal oxides, and the organic matter analogue graphene oxide. Highest potential rates were observed in sediments from below the SMTZ, pointing towards fast stimulation of the deeper methanotrophic community when alleviating the electron acceptor limitation. The variety of AOM pathways and persistence of methanotrophs below the SMTZ likely contribute to the resilience of the microbial methane filter in brackish coastal sediments.

Brown rot wood-degrading fungi release carbon (C) from deadwood but leave behind a large fraction of C sequestered in lignin residues or as fungal metabolites. The strength of sequestration in these C residuals remains unclear, but proteobacteria-dominated bacterial communities have been implicated in metabolizing C from decay residues, possibly erasing the C sequestration potential assumed for brown rot. Here, we paired a model brown rot fungus (Rhodonia placenta) with a model Alphaproteobacterium (Rhodopseudomonas palustris) to track fungal release and bacterial utilization of C derived from decaying wood. We found that fungal decay products generated by R. placenta could be used by R. palustris for growth, and later decay stages contained more usable substrates than early stages. High performance liquid chromatography with mass spectrometry identified a range of aromatic and non-aromatic compounds in the fungal-decayed wood, but after 95 days of bacterial growth, R. palustris preferentially consumed non-aromatic acids over aromatic lignin monomers. Genes involved with aromatic compound degradation were unimportant for bacterial growth, and RNA sequencing revealed that aromatic compound degradation genes were repressed on decayed wood extract. Randomly barcoded transposon sequencing failed to identify a solitary catabolic pathway used by R. palustris, suggestive of substrate co-utilization, and surprisingly, showed that genes involved with copper toxicity were essential. Finally, we found that genes involved with biosynthesis of certain cofactors and amino acids were no longer essential on decayed wood extract, suggesting these nutrients were readily accessible. This study helps lay the foundation to understand potential bacterial-fungal interactions in decayed wood. Graphical abstractTo explore how brown rot fungi support and compete with bacterial partners in the wood decay environment, the model brown rot fungus Rhodonia placenta was used to degrade aspen wafers which were then infused into bacterial growth medium. By leveraging the range of molecular biology tools available for the model Alphaproteobacterium Rhodopseudomonas palustris, we discovered that R. palustris preferentially consumes short organic acids instead of aromatic lignin monomers which it would otherwise consume if provided in isolation. Additionally, R. palustris scavenged certain amino acids (AAs) and enzyme cofactors including methionine, biotin, and PLP from the decayed wood extract, highlighting these as key shared resources for bacterial-fungal partnerships. We found that R. placenta increased the concentration of certain metals (Cu and Al) inducing a metal stress response in R. palustris, indicating that metal toxicity could be an important mode of competition between fungi and bacteria in the wood decay environment. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/723453v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@16f31fcorg.highwire.dtl.DTLVardef@13a9b34org.highwire.dtl.DTLVardef@a37dcforg.highwire.dtl.DTLVardef@198bf1c_HPS_FORMAT_FIGEXP M_FIG C_FIG