The infection of plants by soil-borne fungal pathogens is a complex process and depends on their adhere ability to host surfaces and interactions with the rhizosphere bacteria. In this study, we identify galactosaminogalactan (GAG) as a pivotal virulence determinant in the pathogenesis of Verticillium dahliae. By characterizing the VdGAG biosynthetic gene cluster, we show that the glycosyltransferase VdGtb is essential for GAG synthesis. The knockout of GAG increased fungal sensitivity to the cell wall-perturbing agent calcofluor white (CFW) and reduced mycelial ball formation in liquid culture. The absence of GAG polysaccharides reduced root-binding capacity by 50% and increased cotton immune responses 1 day post fungal infection. The ΔVdGtb mutant exhibited a significant 30.5% decrease in the pathogenicity toward cotton seedlings compared with the wild type V991. Microbiome and bacterial enrichment analysis indicate that the GAG polysaccharides promote the enrichment of soil bacteria and alter the bacterial community structure in the plant rhizosphere. Several bacteria enriched by GAG-contained fungal cells, including Achromobacter animicus, Pseudomonas aeruginosa, and Acinetobacter pittii exhibited strong growth-inhibitory effects against V. dahliae and showed distinct effects on fungal virulence in a GAG-dependent manner. Together, these results reveal that GAG is not merely a cell wall component but a multi-functional molecule that orchestrates fungal protection, host infection, and inter-kingdom microbial communication.

Metabolomic strategies are being increasingly applied in marine systems, leading to unprecedented insights into the ocean and its inhabitants. In particular, exo-metabolites, small molecules produced by marine organisms and released into the environment, have received recognition as factors that influence marine microbial communities, global nutrient cycles, and ecosystem function. Studying exo-metabolites in the marine system is challenging due to the low metabolite concentrations, high salt content, and substantial chemical diversity, all of which can interfere with metabolite detection and therefore necessitate specialized, often tedious sample preparation. Advantageously, new marine metabolomic methods have recently emerged that improve the coverage and detection of key metabolite classes. Here, we offer a practical guide for selecting suitable extraction methods based on specific research needs alongside a systematic review of five recently published methods designed for marine exo-metabolomics. Specifically, these workflows enable quantification of various primary metabolites using liquid and gas chromatography coupled to mass spectrometry. The selection process is guided by several key questions that consider both experimental design and the specific research questions. Additionally, we highlight both the practical constraints and aspects of sample handling for each method. This guide aims to support researchers in effectively choosing a method that aligns with their study goals and logistical capabilities, while also highlighting the opportunities for innovation in the field moving forward.

Phage therapy, the use of viruses that infect bacteria (bacteriophages), is a promising complement to antibiotics during the antimicrobial resistance crisis, but treatment success is very variable. A key variable which likely influences treatment outcomes is how different immune components interact with bacteriophage, with studies finding neutrophils work synergistically while macrophages work antagonistically with bacteriophage. However, many of these studies characterise interactions and outcomes over short timescales, not considering the potential for the evolution of resistance to bacteriophages which can itself greatly affect treatment outcomes. Here, we measure how macrophages and bacteriophages affect densities and resistance evolution of the pathogen Pseudomonas aeruginosa in vitro. Consistent with previous studies, we find macrophages interact antagonistically with bacteriophages in the short term. However, this antagonism was lost following bacterial population recovery associated with rapidly evolved resistance to bacteriophages. Macrophages resulted in greater net levels of resistance and hindered increases in bacteriophage infectivity, but this did not lead to differences in bacteria-phage population dynamics. This work emphasises the importance of characterising the effect of the immune system on phage therapy outcomes over both shorter- and longer- timescales.

Elucidating interaction mechanisms and substrate specialization is central to understanding bacterial adaptation across ecological niches. Specialized mucin-degrading bacteria of the genus Akkermansia are widely recognized for their beneficial roles in the human gut, yet it remains unclear whether this specialization is unique to the gut or reflects a conserved ecological strategy across different hosts and environments. Here, we show that members of the family Akkermansiaceae share a deeply conserved genetic and mechanistic framework, enabling colonization across gut and aquatic ecosystems. Comparative genomics of Akkermansiaceae representatives revealed niche-specific gene repertoires tightly adapted to substrate source and availability. Marine representatives encode distinct combinations of CAZymes and comparatively expanded sulfatase repertoires that enable the degradation of sulfated polysaccharides such as fucoidan, a recalcitrant substrate linked to carbon sequestration. Structural predictions and comparisons identified a conserved molecular system centered on a type IV-like pilus that mediates attachment to complex, fucose-rich glycans. The genes underlying this system are syntenic with the recently described mucin utilization locus in Akkermansia muciniphila, revealing an evolutionary continuity between aquatic and gut lineages. Seawater incubations with fluorescently labeled substrates confirmed fucoidan uptake and degradation by marine Akkermansiaceae. Together, these results reveal a unified glycan-utilization strategy spanning the environmental breadth of Akkermansiaceae and provide a mechanistic framework linking ecological success in marine environments to traits associated with probiotic functions in the human gut.

Nitrous oxide (N2O) is a potent greenhouse gas, and soil is its largest terrestrial source. Microbial N2O reductase (NosZ) is the only known enzyme capable of reducing N2O to N2, making nosZ-harboring prokaryotes important sinks in terrestrial ecosystems. Despite being among the most abundant and ubiquitous bacterial phyla in soil, the potential role of Acidobacteriota in N2O reduction remains largely unexplored. In this study, we addressed this gap using genomic, metagenomic, and physiological analyses. We first analyzed 199,602 prokaryotic genomes, including genomes from both isolated strains and metagenome-assembled genomes (MAGs). We found that 491 Acidobacteriota genomes harbored nosZ, predominantly the Sec-dependent NosZ gene (nosZII). Global metagenomic analysis of 321 soil samples revealed that Acidobacteriota nosZII is one of the most abundant groups of nosZ and distributed across different continents. Among Acidobacteriota, nosZII from the class Vicinamibacteria was the most prevalent in the soils. Finally, we provide the physiological evidence of N2O-reducing activity in Acidobacteriota by demonstrating that the Vicinamibacteria type strain, Luteitalea pratensis KCTC52215T, can reduce N2O. Taken together, these findings highlight the previously overlooked potential role of Acidobacteriota as a global N2O sink and underscore the need to include them in future studies on soil N2O dynamics.

Associations with microbial symbionts shape the ecology and evolution of almost all eukaryotes. One of their defining features is their specificity, but despite this, many symbioses show a degree of flexibility, with some symbiont species capable of colonizing multiple (often closely related) host species. Although widespread, the functional and evolutionary consequences of flexibility in host-symbiont pairings are poorly understood. Bivalves from the diverse, globally distributed, and ecologically important family Lucinidae are ideal for investigating this, as multiple host species can associate with the same symbiont species, often at the same location. We used metatranscriptomics to investigate the molecular responses of one symbiont species, Candidatus Thiodiazotropha endolucinida, in association with three different host species that co-occur in seagrass meadows in the Caribbean Sea. In replicated experiments, we identified host species-specific patterns of symbiont gene expression including those for key functions such as carbon fixation, cell division, and sulfide oxidation. Our work shows that the symbiont consistently responds in different ways to association with different host species. Because all samples were collected at the same site on the same day, and were thus exposed to the same environmental conditions, these differences are likely driven by host rather than environmental factors. In addition, host species had significantly different carbon isotope signatures, which were consistent with distinct modes of host-microbe interaction indicated by transcriptomics. Our results show that not only symbiont genotype, but also symbiont phenotype may enable coexistence of closely related host species, demonstrating the power of symbiosis in promoting and maintaining biodiversity.

Toxic cyanobacterial harmful algal blooms (cyanoHABs) threaten freshwater resources globally and are intensifying with increasing eutrophication. Bloom toxicity is strongly influenced by intraspecific variation in the biosynthetic repertoires of toxic cyanobacteria, yet few studies examine the diversity of cyanobacterial cyanopeptides beyond hepatotoxic microcystins. To understand the dynamics and drivers of cyanopeptide diversity in cyanoHABs, we analyzed temporal patterns of cyanobacteria, metabolites, and their biosynthetic gene clusters (BGCs) in western Lake Erie using a seven-year time series (2016-2022) of metagenomic and metabolomic data. Our findings demonstrate that shifts from Microcystis to Dolichospermum occur later in the bloom season, coinciding with lower temperatures. Modules of co-varying BGCs (biosynthesis modules) from these genera were identified with hierarchical clustering, with uncharacterized BGCs among the most abundant. Biosynthesis modules rich in nonribosomal peptide synthetases (NRPS) peaked in early August, coinciding with elevated levels of inorganic nitrogen, warmer temperatures, and high Microcystis abundance. In contrast, modules rich in polyketide synthases (PKS) and ribosomally synthesized and post-translationally modified peptides (RiPPs) peaked following the Microcystis maximum in mid-August. Metabolomic analyses confirmed that metabolites followed shared seasonal patterns with their associated biosynthesis modules, forming three phases characterized by (1) microcystins, (2) anabaenopeptins and aeruginosins, and (3) aerucyclamides. These phases co-varied with bottom-up and top-down pressures, with later phases coinciding with increased microbially processed organic nitrogen and reduced detection of grazers. This study demonstrates consistent seasonal patterns of cyanobacterial metabolite succession and co-occurrence beyond microcystins, suggesting tradeoffs between biosynthetic resource demands and ecological controls.

The seed is a habitat with limited resources and space. Although it is widely accepted that microbial competition is a key driver of the assembly of seed-associated microbial communities, the underlying mechanisms of this competition are not well understood. The initial objective of this work was to assess the importance of contact-independent microbial competition between the phytopathogenic bacterium Xanthomonas campestris pv. campestris 8004 (Xcc8004) and 30 strains representative of the bacterial populations most commonly associated with radish (Raphanus sativus) seeds. We identified Pseudomonas lactucae CFBP 13502 as a potent inhibitor of Xcc8004, mediated by exometabolites, specifically induced by certain seed-borne strains. Transcriptomic analysis linked this inducible activity to the upregulation of a gene cluster encoding a lipopeptide siderophore. Targeted gene deletion in P. lactucae CFBP 13502 confirmed that this cluster is essential for antagonism against Xcc8004. Furthermore, iron supplementation abolished this inhibitory effect, strongly supporting iron chelation as the underlying mechanism. Through comparative metabolomics, we elucidated the structure of a family of lipopeptide siderophores, produced by P. lactucae CFBP 13502, which we named lactuchelins. Our findings provide molecular evidence of competitive exclusion mechanisms at the seed microbiome interface, highlighting lactuchelins as a promising avenue for the development of seed-based biocontrol strategies against seed-borne phytopathogens.

Marine anammox bacteria of the genus Scalindua play a pivotal role in nitrogen loss within oceanic oxygen minimum zones (OMZs), yet their physiological resilience to oxygen exposure remains poorly characterized. Here, we show that Scalindua sp. possesses an intrinsic capacity to reduce ambient oxygen under microoxic conditions (headspace O₂ ≈ 3%), exhibiting specific oxygen reduction rates of ~10nmol-O₂ min-1 mg- protein-1. Anammox activity, evidenced by 29N₂ production, resumed once dissolved oxygen (DO) declined below ~5μM, defining the DO threshold. This oxygen-scavenging ability is likely mediated by direct oxygen reduction to water via cbb₃-type cytochrome c oxidase (CcO) and A-type flavodiiron proteins, with minimal reactive oxygen species (ROS) generation. CcO activity increased rapidly and markedly upon nitrite addition, consistent with the immediate onset of oxygen consumption and the transcriptional induction of CcO subunit genes. Nitrite was essential for sustaining oxygen consumption, and the rapid CcO activation suggests that nitrite stimulates CcO function through an unresolved regulatory mechanism. Nitrite oxidation also supplies supplementary electrons for oxygen reduction independently of core anammox metabolism, underscoring a flexible substrate-driven detoxification mechanism that enables Scalindua sp. to cope with transient microoxia. Together, these results show that Scalindua sp. employs a multi-layered defense-moderate oxygen-reduction capacity coupled with efficient ROS detoxification-conferring high and reversible oxygen tolerance. Although laboratory conditions cannot fully replicate natural OMZ complexity, our findings indicate that this physiological flexibility reflects the potential ecological resilience of Scalindua sp. in dynamic ocean environments.

Marine microbial communities are fundamental to nutrient and biogeochemical cycling, with intricate networks of metabolic interdependencies influencing their structure and dynamics. Among these, vitamins B1 (thiamin) and B12 (cobalamin) play crucial roles as enzymatic cofactors in central metabolic pathways. Despite their importance, the temporal dynamics of vitamin production, bioavailability, and associated microbial interactions remain poorly understood. Using a 7-year monthly metagenomic time series from the NW Mediterranean Sea (SOLA station), we found that vitamin B1/B12 auxotrophs (need for an exogenous vitamin source) were present throughout the year. Among B1 auxotrophs, those requiring the thiamin precursor pyrimidine were the most prevalent, with peak abundances in summer. Distinct metagenome-assembled genome co-abundance patterns between B1 and B12 producers/auxotrophs across seasons suggested mutualistic relationships. Double B1/B12 vitamin complementarities were more common in summer, and single vitamin complementarity was dominant in winter. As previously shown for vitamin B12, which is limiting during winter, bioassays revealed variable availability of vitamin B1 in winter seawater despite the abundance of its producers, suggesting potential transfer of vitamin B1 among microorganisms. Finally, microcosm experiments showed that B1 and B12 amendments significantly influenced the composition of microbial communities, with temporal variations in their impact. In some cases, B12 and B1 amendments favored both vitamin auxotrophs and producers, highlighting complex interdependencies between B1 and B12 producers and consumers. Our findings highlight the complexity of B vitamin–mediated metabolic interactions that shape microbial community dynamics and underscore the need for long-term, high-resolution studies to better understand vitamin-driven ecological processes in marine systems.