FEMS Microbiology Ecology

Bacterial interactions-whether positive or negative - are crucial for the functioning of microbial communities. Though bacterial interactions are mainly expected to be negative, the sign and strength of interactions are predicted to be context dependent, with interactions typically being more positive in more stressful and nutrient-poor conditions. However, systematic studies investigating how the environment affects interactions between multiple taxa are lacking. Here, we determine if interactions between a panel of natural soil isolates change in response to the environment in which they are grown, with two different artificial media used (one simple and one complex) and a more ecologically relevant soil wash. To maximise natural variation in interactions, we collected multiple isolates from multiple sites: co-occurring (sympatric) isolates were predicted to show more negative interactions than allopatric isolates because of greater overlap in resource use. Pairwise interactions were in general negative, but more negative when grown in a complex lab-derived medium (Tryptic Soy Broth). Mutually beneficial interactions were most common in a simple resource medium (M9 minimal media) and exploitative interactions were most frequent in a soil broth. These patterns were independent of whether species originated from the same or a different site. The study supports the prediction that nutrient rich environments promote more negative interactions, and that measuring interactions of soil isolates in standard lab media is likely to misrepresent interactions occurring in natural environments.

BackgroundThe role of the gut microbiome and specific enteric bacteria in influencing the development of colorectal cancer (CRC) remains incompletely understood. Recently, it was shown that human CRC-derived strains of Clostridioides difficile were capable of inducing colonic tumorigenesis in a susceptible mouse model. We hypothesized that C. difficile contributes to the pathogenesis of human CRC and would be enriched in CRC tumors compared to paired normal tissues from the same individual. MethodsWe analyzed matched tumor/normal tissue samples from a cohort of 108 individuals presenting to a tertiary care hospital in Kuala Lumpur, Malaysia for CRC resection between 2013-2014. We assessed the prevalence of C. difficile detection using 16S rRNA amplicon sequencing with high-resolution taxonomic assignment as well as culture and PCR. ResultsWe found that detection of C. difficile was prevalent (38% of individuals), but of low abundance (tumor median relative abundance 0.01%, paired normal 0.006% [p=0.4]). Detection of C. difficile was more prevalent in individuals with biofilm-positive tumor tissues than biofilm-negative (i.e., 81% of C. difficile-positive individuals were biofilm-positive vs. 63% of C. difficile-negative individuals [p=0.04]). Additionally, in exploratory analyses, we describe patterns of taxonomic and inferred functional pathway differences between C. difficile-positive and C. difficile-negative groups. ConclusionThese findings suggest that C. difficile is frequently present in low abundance in the tumor microbiome with a potentially significant impact on community composition and function.

Propionic acid (HPr) accumulation is a major indicator of anaerobic digestion (AD) dysfunction, yet the relative contributions of acidity, undissociated HPr, and propionate ions (Pr-) to process inhibition remain poorly understood. We investigated these effects in mesophilic batch AD microcosms fed with municipal sewage sludge, using a comparative design involving HPr, sodium propionate (NaPr), NaCl, and HCl treatments across two series of experiments. While 20 mM HPr caused a 22% reduction in the maximal methane production rate, 81 mM HPr led to complete inhibition, with the initial pH dropping to 5.1. By contrast, 81 mM NaPr reduced methane production rate by only 40%, and 81 mM NaCl caused no inhibition, demonstrating that acidity is the dominant inhibitory factor, with Pr- exerting a secondary concentration-dependent effect. 16S rRNA gene amplicon sequencing revealed strong, compound-specific shifts in microbial community composition, affecting key functional groups including syntrophs and methanogenic archaea. The proportion of methanogens dropped from 2-3% in control reactors to less than 0.2% under 81 mM HPr, consistent with the observed methane production inhibition. Under HPr81, over 100 ASVs were differentially abundant compared to controls, a pattern largely shared with HCl-treated reactors, further confirming the predominant role of acidity. The number of differentially abundant ASVs was negatively correlated with methane production rates (R{superscript 2} = 0.97), underscoring the link between community reshaping and process impairment. These results provide a unifying framework for propionate inhibition in AD and suggest that microbial community profiling could serve as an early warning tool for process imbalance detection.

BackgroundThe root microbiome plays a critical role in nutrient acquisition, stress tolerance and overall plant health. Rice, a staple food for more than half of the worlds population, is commonly cultivated under flooded conditions. Despite its agronomical importance, our current understanding of root-associated microbiomes in rice grown under flooded conditions is limited. On the other hand, nitrogen (N) and phosphorus (P) fertilizers are routinely applied to maximize rice yield. It is also well known that root colonization by arbuscular mycorrhizal (AM) fungi enhances mineral nutrition in plants, but whether mycorrhizal associations influence the composition of the rice root microbiome remains poorly understood. In this study, shotgun metagenomic sequencing was used to characterize the root endosphere and rhizosphere microbiomes in two temperate japonica rice varieties (cv. Bomba and JSendra) grown under flooded conditions. The impact of colonization by the AM fungus Rhizophagus irregularis on the root microbiome was investigated. ResultsRoot-associated compartments harbour distinct microbial communities in rice with bacterial taxa comprising approximately 95% of the total microbia in rice roots. At the Phylum level, the root bacteriome was primarily composed of Pseudomonadota (Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria) followed by Actinomycetota. The fungal microbiome was dominated by Ascomycota (Sordariomycetes, Eurotiomycetes and Dothideomycetes) and Basidiomycota. Not only the root compartment, but also the host genotype can shape the root microbiome. Recruitment of specific microorganism mainly occurs at the species level. Genotype-specific and compartment-specific associations of microbial species in mycorrhizal rice roots were also observed supporting that root colonization by an AM fungus contributes to variations in the root microbiome. Further, key microbial species primarily associated to methane production and nutrient cycling (e.g. Phosphate Solubilizing Bacteria and Nitrogen cycling bacteria) colonizing root compartments in each rice genotype and mycorrhizal condition are described. ConclusionsThe rice genotype, root compartment and mycorrhizal condition markedly influence the microbiome in roots of rice plants growing in flooded rice fields. These findings illustrate the potential of the plant to shape its associated root microbiome, thus, offering valuable insights for the development of microbiome-based strategies to improve growth and performance in rice plants under flooded conditions.

Controlled environment agriculture (CEA), including soilless farming systems, is rapidly expanding as a strategy to improve food security and resource efficiency. However, limited information is available on how different soilless farming system designs influence microbial populations relevant to plant health and food safety. This study investigated the effects of soilless growing systems and growing season on aerobic plate counts (APC) and bacterial community composition in nutrient solution and on bok choy (Brassica rapa subsp. chinensis) leaves. Five soilless systems, deep water culture (DWC), Kratky (KR), nutrient film technique (NFT), ebb and flow (EF), and drip irrigation (DI), were evaluated across fall and spring growing seasons. Soilless system type significantly influenced APC in nutrient solution, with the DI system consistently exhibiting the highest counts across both seasons. Increased nutrient solution pH was negatively associated with APC, whereas temperature did not significantly affect bacterial concentrations. In contrast, APC on bok choy leaves were not significantly influenced by system type, season, pH, or temperature. Bacterial community composition in nutrient solution was strongly shaped by season, soilless system type, sampling day, and temperature, as determined by 16S rRNA V4 amplicon sequencing. Microbial diversity varied primarily by system type, with limited influence of pH or temperature. Core microbiota analysis identified a small subset of taxa that persisted across systems and seasons, with Acidovorax detected in all samples. We found that soilless system design and seasonal conditions strongly influence microbial load and community structure in nutrient solution, providing a foundation for developing system-specific microbial management strategies. ImportanceUnderstanding factors that shape microbial community composition in soilless farming systems is critical for optimizing plant health, system productivity, and food safety. Microbial communities influence nutrient cycling, biofilm formation, and pathogen survival, all of which affect the ecological stability and performance of these systems. By identifying how system design, seasonal variation, and environmental conditions influence shifts in microbial populations, targeted strategies can be developed to promote beneficial microorganisms and mitigate risks associated with pathogens. This knowledge contributes to advancing safe and sustainable soilless farming practices that can meet the growing demand for fresh produce grown in controlled environments.

O_LIPhyllosphere microbiomes are subject to microbial import from various sources and undergo substantial changes during phenological changes of plants. However, these processes are still poorly understood for forest canopies. We propose that phenology-driven changes in host properties, and rainwater-mediated, within-canopy transport shape the phyllosphere microbiome in temperate forests. Leaves and throughfall samples were collected from oak, ash and linden trees at top, mid, and bottom canopy positions at the Leipzig canopy crane facility (Germany) at time points representing early, mid and late phenological stages. Bacterial community composition was assessed by 16S rRNA gene amplicon sequencing. C_LIO_LIPhenological stages explained 19% of phyllosphere bacterial community variation, followed by tree species identity (12%) and canopy position (2%). Later phenological stages exhibited more homogeneous and functionally redundant phyllosphere communities along with a strong decline of plant pathogens and increasing potential for microbially mediated biocontrol mechanisms. Throughfall transported up to 1011 bacterial cells per litre with maximum bacterial fluxes at the canopy top. C_LIO_LIOur findings demonstrate that in temperate forests, phenology-driven effects on the phyllosphere microbiome are far more important than tree species specific effects. Extent and selectivity of throughfall-mediated mobilization may play a crucial role for the spatial heterogeneity of microbial communities in tree crowns. C_LI

AbstractSoil microbes have sophisticated mechanisms to detect and respond to short pulses of C inputs, often involving changes in gene-expression. We studied gene transcription in a soil microbial community before, and 8, 24, and 48h after glucose addition (0.7 mg C g-1 dry soil) to understand how microbes react to periods of substrate excess and subsequent starvation. The relative transcript abundance of genes associated with energy metabolism and biosynthesis of amino acids, lipids, nucleotides, and cell wall components increased 8h after glucose addition. By 24 and 48h, the abundances of these transcripts reversed. Transcript abundance for genes associated with degradation of lipids, nucleotides, and (hetero)cyclic hydrocarbons decreased at 8h, but increased 24 and 48h after glucose addition. Simultaneously with a rise in transcripts for energy production and biosynthesis at 8h, transcription of regulatory genes for the exponential growth phase and ribosome assembly and maturation increased. In contrast, at 24 and 48h, transcript abundance for genes associated with ribosomal hibernation, sporulation, and regulation of the stationary phase increased, while transcripts for regulators for the exponential phase, and ribosome activation decreased. Based on changes in transcript abundance of phosphoenolpyruvate carboxylase and pyruvate carboxylase, it appeared that 8h after glucose addition glycolytic activity was high, however, gluconeogenesis returned at 24 and 48h. High levels of transcripts for nrtC-ntrB indicated N limitation 8 and 24h after glucose addition. Transcripts associated with Type VI Secretion Systems increased 24 and 48h after start of the experiment, suggesting a short lag between primary consumers and predatory bacteria. These results illustrate how metatranscriptome analysis can be used to study the ecophysiology of soil microbes providing details on the timing of exponential and stationary phase processes, coordination between anabolism and catabolism, and emerging nutrient limitations in natural soil communities. Research HighlightsO_LIWe studied gene transcription of a soil microbial community after glucose addition C_LIO_LITranscript abundances for biosynthesis and energy production initially increased, while those for degradation decreased C_LIO_LITranscripts of regulators and sporulation genes indicated start of stationary phase at 24h C_LIO_LINitrogen limitation induced transcription of nitrogen stress genes C_LI

Emergence of multi-drug resistant (MDR) pathogens is facilitated by the mobilization of resistance genes from bacteria in animal and environmental habitats, a process often mediated by plasmids. While fertilization of agricultural soils with manure is hypothesized to serve as a pathway for transferring antimicrobial resistance plasmids to soil and crop bacteria, evidence is limited. In this study, we aimed to determine whether MDR-plasmids from manure transfer in soil, leading to the formation of long-term agricultural resistance reservoirs. To this end, we introduced a known MDR plasmid to agricultural soil where barley was subsequently grown and monitored spread of the plasmid over the course of a growing season (up to 190 days). Our experimental design mimicked conventional agricultural practices at a microcosm scale. A digital droplet PCR approach indicated plasmid transfer in the rhizosphere, which was confirmed by a targeted Hi-C method (termed Hi-C+). This demonstrated transfer of the plasmid to soil bacteria 10 days after barley planting but was not observed afterwards. The new plasmid hosts could not be identified, as plasmid-associated host Hi-C reads were absent from existing databases. This implies these hosts were rare and likely unculturable members of the soil microbiome. Our findings demonstrate that plasmid transfer from manure to soil can occur under conditions reflecting those found in agricultural settings. Furthermore, rare and uncharacterized members of the soil microbiomes may participate in acquiring MDR plasmids from manure bacteria, raising important questions about their role in spreading resistance plasmids.

Plant roots host defined microbial communities that differ from those found in the surrounding soil and these communities shift dynamically in response to plant development and environmental changes. Whilst it is widely accepted that root exudates play a key role in the assembly and dynamics of root-associated microbial communities, the underlying mechanisms are not well understood. This is partly due to a lack of controlled experimental systems that monitor both exudate- and microbiome-dynamics simultaneously. Here, we compared two microcosm systems commonly used in either root microbiome (clay particle-based) or root exudate studies (glass bead-based) for their suitability to simultaneously monitor both aspects. We evaluated these systems based on plant performance, bacterial growth, and time-resolved community and exudate profiling. In both systems, we reveal an exudate effect, characterised by higher bacterial diversity and Pseudomonas abundances in proximity to plant roots. While clay particles impeded exudate recovery, even when plants were removed from microcosms for exudate collection, the glass bead set-up allowed us to uncover dynamic exudate shifts during bacterial community establishment. This highlighted a transient increase of glucosinolates upon root colonisation by initially dominant Pseudomonas species. Overall, the comparison proved only the glass bead-based semi-hydroponic system to be suitable for the paralleled study of exudate and root microbiome dynamics.

The human gut microbiome is a complex community that plays an important role in health, where perturbations can result in dysbiosis and disease. Bacteriophages (phages) can provide treatment for bacterial gastrointestinal disease, and commercial preparations such as the Intesti bacteriophage cocktail can be taken orally to target bacterial pathogens. However, interactions between these phages and the native gut microbiota are understudied. To investigate the impact of phage treatment, we used simulated gut models seeded with healthy donor microbiota from three individuals, sequenced the DNA, and analysed the bacterial and viral portion from samples obtained over time. Each donor had a unique bacterial composition which diverged with time. When comparing phage treated to control samples, we observed that Escherichia coli abundance accounted for the largest portion of bacterial community variance and was more associated with the controls. The lower abundance in phage treated samples may have resulted from the lytic action of phages from the cocktail. Additionally, our analyses of the viral portion revealed a phage bloom exclusive to phage treated samples. A highly abundant phage in this bloom was matched with the Intesti bacteriophage cocktail, showed similarity to Enterobacteria phage phi92, and provided evidence of productive infection within the model. While we did observe fluctuations in relative abundance of additional viral sequences in the presence of the phage cocktail, these changes were often transient. Furthermore, we detected only slight differences to typical members of the virome, and low numbers of active prophages. Our experiments suggest that the phage cocktail had minimal interruption to the native gut microbiota within the model. Impact statementBacteriophages are increasingly investigated and tested for their efficacy in treating infections and are a key component in fight against antimicrobial resistant bacterial infections. Because of their specificity, it has become almost a dogma to state that they do not alter the gut microbiome. We have now tested this in an in vitro study using a commercially available cocktail and real human faecal microbiota. We show minimal effects on the composition of the healthy microbiota with an individual-specific effect on Escherichia coli caused by productive infection of one phage in the cocktail.

Microbial communities can shift under drought in ways that enhance plant performance during drought ("microbe-mediated acclimation"). However, it is also possible for microbial communities to shift in ways that worsen the effects of drought ("mal-acclimation"). It is unclear how and where microbe-mediated acclimation vs. mal-acclimation occurs, or if there are types of soils or microbial communities that are more likely to harbor microbes that enhance plant acclimation and limit mal-acclimation. We tested for microbe-mediated plant acclimation/mal-acclimation to drought in soils from 21 maize farms in the midwestern United States, spanning a range of climate, soil types, and management practices. We first conditioned soil microbial communities to drought vs. well-watered conditions in a greenhouse and then tested for microbe-mediated acclimation by growing maize in soils inoculated with the conditioned microbial communities under drought and well-watered conditions. Drought-conditioned soils did not enhance plant performance under drought. In fact, one third of the farms exhibited mal-acclimation, especially under well-watered conditions where wet-conditioned soils reduced plant performance in well-watered contemporary conditions. Farm management practices, climate, soil texture, and microbial diversity generally did not predict when this microbe-mediated mal-acclimation occurred. Overall, these results suggest that in agricultural soils, microbes may frequently impede-rather than facilitate-plant acclimation to soil moisture levels. Open research statementThe plant and soil data used in this study are available via the Environmental Data Initiative repository at https://doi.org/10.6073/pasta/f4a0db3a076cf6d8cef908947f82736e. The bacterial and fungal amplicon sequence data are available via the European Nucleotide Archive under accessions PRJEB110071 and PRJEB109827, respectively.

Microbial mutualists partially determine many host traits, including traits related to infection by parasites. However, while microbial effects on host trait plasticity is fairly well established, whether microbial mutualists contribute to genetic variation in infection-related remains an open question. Here we paired 10 mutualistic Sinorhizobium meliloti rhizobacteria strains with 20 Medicago truncatula plant genotypes in an incomplete factorial design, and experimentally infected the plants with parasitic root-knot nematodes. We used this design to estimate rhizobia contributions to genetic variation in four infection-related traits: host resistance, parasite virulence, host tolerance, and mutualism robustness. We find that rhizobia contribute to genetic variation in host resistance and mutualism robustness, and to genetic variation in parasite virulence via genotype-by-genotype interactions with the host. Rhizobia did not contribute to variation in host tolerance. The influence of rhizobia strains on parasite resistance was partially explained by their effect on host root growth. These results underscores the influence that microbial mutualists have on their hosts response to parasite infection, and suggests that resource mutualists may impact host-parasite evolution. Teaser TextMicrobial mutualists like nitrogen-fixing rhizobacteria influence their host traits. Past work indicates that different strains of rhizobia may influence their host plants interactions with nematode parasites. But how does this influence compare to the genetic variation present in hosts? We explore the contribution of genetic variation across rhizobia strains to infection-related traits in their host and find that rhizobia contribute to genetic variation for parasite resistance and virulence in their host.

1Microbial symbionts have the potential to contribute to host-plants ecological and evolutionary success, especially in plants adaptions to harsh environments, however their role has often been overlooked. Conversely, how host local adaptation (e.g., trait divergence across elevation) shapes the composition of associated microbial symbiont communities remains poorly understood. We explored how foliar endophytic fungi (FEF) abundance, richness, and community composition in three sympatric Monkeyflowers, an ecologically diverse group of flowering plants, change across elevation in the Sierra Nevada, CA, USA. We asked: Q1) Are there differences in leaf functional traits and FEF communities among sympatric Mimulus species populations at similar elevations? Q2) How do traits and FEF communities change across elevation within species? Q3) Are FEF richness, diversity and community composition correlated with leaf functional traits and/or elevation? Q4) How does FEF community composition differ with geographic distance within each species? We collected M. guttatus, M. nasutus, and M.laciniatus individuals from natural populations across the Sierra Nevada, measured leaf functional traits and used ITS sequencing to describe the leaf endophyte community. We found significant associations of FEF community composition with host species, and elevation, suggesting that these factors influence fungal community composition. Furthermore, FEF community dissimilarity was correlated with geographic distance indicating isolation by distance and limited dispersal of fungal endophytes. We detected the prevalence of Vishniacozyma victoriae, an endophyte found most commonly in Antarctica, across all Sierran Mimulus populations. The presence of V. victoriae could play a role in the adaptation of Mimulus to cold, high elevation environments. Our findings offer novel insights into the intricate interactions between fungal endophyte communities, plant traits, and elevation.

Microorganisms are increasingly recognized as key facilitators of invasion success for introduced species into new environments. The globally invasive seagrass Halophila stipulacea flourishes in mixed environments with native seagrasses, where it exhibits enhanced growth, while, in contrast, native seagrasses in mixed environments experience reduced growth. Here, we hypothesize that microbes may support the success of invasive seagrass in mixed Mediterranean environments. We analyzed 16S rRNA genes to characterize the microbial diversity on the phyllosphere alongside biochemical, morphological, and sediment nutrient measurements of the Mediterranean-native seagrass Cymodocea nodosa and the invasive H. stipulacea from a controlled mesocosm experiment. Overall, C. nodosa in monoculture harbored a microbiome exhibiting higher ASV richness and a distinct community composition than H. stipulacea. Variation in bacterial diversity associated with hydrogen peroxide (H2O2) and internode length suggests that microbial communities of the native seagrass might be shaped by its stress. Conversely, H. stipulaceas microbiome was most abundant in mixed environments, with bacteria significantly reduced in monoculture, and bacterial diversity loosely associated with growth, suggesting that microbes are critical to assisting and possibly facilitating H. stipulacea in mixed environments. Overall, our findings suggest that invasive H. stipulacea in the Mediterranean Sea are capable of recruiting beneficial bacteria, creating microbial interactions that support its success, and undermining the resilience of native seagrasses in mixed beds. Future work should center on the mechanisms driving H. stipulacea bacterial communities and investigating whether H. stipulacea actively determines its own microbiome, or whether its microbiome is passively determined by environmental variables.

Sourdough starters contain simple microbial communities typically consisting of a few bacterial species and one or two yeast species. The yeast Maudiozyma humilis and the lactic acid bacterium Fructilactobacillus sanfranciscensis often co-occur in sourdough starters, and have been presumed to exist in a trophic relationship supported by glucose cross-feeding. However, previous research has highlighted a lack of evidence showing that yeast strains consume the glucose that F. sanfranciscensis produces. We have investigated the interaction between sourdough isolates of M. humilis and F. sanfranciscensis in a synthetic wheat sourdough medium, allowing us to control substrate composition and use flow cytometry to enumerate living and dead cells. M. humilis fitness was found to be lower in co-culture with F. sanfranciscensis than when grown alone. Analysis of spent medium composition highlighted the reliance of M. humilis on glucose rather than maltose for growth. Comparisons of predicted and measured co-culture metabolite content also revealed that F. sanfranciscensis consumed less maltose in co-culture than when grown alone. For the first time, we examined potential amino acid cross-feeding between M. humilis and F. sanfranciscensis, and found that within the pairing, F. sanfranciscensis was the main producer of amino acids. Our findings suggest that the M. humilis-F. sanfranciscensis interaction is likely to be neutral, or even competitive, with the strain identity of F. sanfranciscensis playing a defining role in the observed dominance of the bacteria and spent medium metabolite composition. ImportanceThe association of the yeast Maudiozyma humilis and the bacterium Fructilactobacillus sanfranciscensis in sourdough starters is well-documented, and together this pairing makes key functional and organoleptic contributions to the final bread product. Their relationship has historically been thought to be stabilised by cross-feeding of glucose to M. humilis. However, this theory has been drawn into question by recent research which found no evidence that M. humilis consumes the glucose produced by F. sanfranciscensis. Our understanding of cooperation, coexistence, and competition in microbial consortia affects approaches to ecosystem management in a broad variety of applied fields. The significance of our research is in demonstrating that this pairing does not interact mutualistically within a specified setting, providing support for neutral or competitive interactions as drivers of ecological stability. Research areas:

BackgroundPelagic Sargassum has undergone significant range expansion and dramatic blooms in the Atlantic over the past 15 years. This algaes microbiome provides symbiotic functions that are believed to contribute to its ecological success. Recent research shows that Sargassum-associated bacteria are enriched in integrated prophages compared to the surrounding seawater and that these prophages are inducible by chemical and ultraviolet treatment. ResultsHere, we investigated a Sargassum-derived in vitro multispecies biofilm encompassing the dominant heterotrophic microbial members associated with Sargassum to probe the impacts of prophage induction on the composition of Sargassum microbiomes. Induction was quantified by coverage-based virus-to-host ratios in chemically induced treatments with Mitomycin C and non-induced controls, and the community composition and metabolic profiles were analyzed after a period of recovery post-induction. Chemical induction led to a significant increase in abundance and virus-to-host ratio of viral genomes linked to Vibrio metagenome-assembled genomes. This was accompanied by altered biofilm community composition, with a reduction in Vibrio bacterial abundance that opened niche space for other biofilm members in the genera Pseudoalteromonas, Alteromonas, and Cobetia. The induced Vibrio-associated phages encoded genes involved in quorum sensing, biofilm formation, virulence, and host metabolism. Induction led to a relative loss of 17 metabolic modules, including functions related to energy metabolism and nitrogen utilization. ConclusionDue to the high frequency of lysogeny in the Sargassum microbiome and the susceptibility of prophages to chemical and ultraviolet light induction, these results suggest that prophage integration and induction are mechanisms that significantly contribute to structuring the Sargassum microbiome and its functional profiles, potentially aiding in microbiome flexibility in changing environmental contexts.

Microbial diversity remains largely unexplored across environments and scales, notably because at local scales many microbial taxa exist under a dormant state. Microbial biogeography is shaped by edaphic and ecological drivers, and shifts in microbial community composition are frequently associated with host community structure and health. Nontuberculous mycobacteria represent a striking example of environmental microorganisms with opportunistic pathogenic potential. Unfortunately, data on their diversity, distribution, and ecological interactions in aquatic environments remain limited. However, understanding competition for niche space and the role of abiotic and biotic factors shaping their biogeography is crucial for predicting disease emergence and transmission. Here we aimed at i) identifying microhabitat abiotic and biotic drivers influencing their distribution, ii) assessing the predictability of their diversity and distribution across continents, and iii) examining potential exclusion or associations between pathogenic and nonpathogenic mycobacterial species. By deploying an eDNA-based metabarcoding approach from freshwater samples collected in urban and rural sites in French Guiana and Cote dIvoire, we have boosted our understanding of environmental mycobacteria ecology by highlighting the influence of habitat type, abiotic factors, and microbial interactions on mycobacterial distribution. In addition, the detection of pathogenic species further highlighted the importance of environmental reservoirs in mycobacterial disease transmission.

The study of microbial volatile organic compounds (mVOCs) is a growing area of research, with applications ranging from agriculture to human health. The majority of the mVOC data are from in vitro liquid cultures, while few analyses of bacterial and fungal volatilomes on solid media cultures exist. Studies comparing liquid versus solid cultures of bacteria and fungi show significant changes to the soluble metabolites that are produced, suggesting that large differences would be observed for mVOCs based on the culture conditions. To test this idea, we characterized the volatilomes of Chromobacterium violaceum (strain ATCC(R) 12472) and C. vaccinii (strain MWU328), and those of their isogenic cviR- quorum sensing mutants cultured on solid versus liquid Kings Medium B media. VOCs were sampled using thin-film solid-phase microextraction (TF-SPME) and analyzed by two-dimensional gas chromatography-time-of-flight mass spectrometry (GCxGC-TOFMS). Of the three variables examined - Chromobacterium species, media type, and quorum sensing ability - growth on liquid versus solid media caused the most significant differences in the volatilomes. Bacterial species and quorum sensing ability were also influential, but to a lesser degree. Our findings indicate the importance of growth conditions in microbial volatilomics, and therefore, more consideration should be given to how microorganisms are cultured for volatilome analyses. ImportanceThe purpose of this work is to elucidate the differences in the volatile metabolic profiles of Chromobacterium spp. by exploring them through the lens of three variables: growth conditions, species, and the ability to quorum sense. Work on organismal metabolic differences stemming from factors such as liquid versus solid media types remains broadly overlooked. Understanding these effects will allow future researchers to design more robust experiments that better translate to native microbial ecosystems such as rhizosphere and phyllosphere, where volatile compounds may influence plant-pathogen or plant-saprobe interactions.

IntroductionAntimicrobial resistance has existed in the environment long before its rapid emergence and detection in clinically relevant pathogens. Studying the resistance of environmental bacterial strains may allow novel resistance mechanisms to be identified before they appear in pathogenic strains. Gap StatementSearching for antimicrobial resistance genes in environmental bacteria represents an understudied research area compared to resistance within clinically relevant pathogens. AimTo evaluate resistance genes present within environmental non-aeruginosa Pseudomonas spp. isolates. MethodologyWe screened a set of bacterial isolates from untreated wastewater from Liverpool, UK, for the presence of extended spectrum {beta}-lactamases and carbapenemases. A sub-set of three resistant Pseudomonas spp. isolates were selected for whole-genome sequencing. We performed minimum inhibitory concentration assays against several {beta}-lactams, and ectopic expression of four novel resistance genes within Escherichia coli. ResultsHere, we report the discovery of novel class C {beta}-lactamase genes blaPFL7, blaPFL8 and blaPFL9, as well as a novel subclass B2 metallo-{beta}-lactamase blaPFM5 present within these strains. The class C genes encoded proteins with between 61-71% amino acid identity to the closest known match, blaPFL-1. These novel {beta}-lactamases degraded the cephalosporin nitrocefin and confer piperacillin and ceftazidime resistance to susceptible Escherichia coli when ectopically expressed. The {beta}-lactamase inhibitor tazobactam was effective at inhibiting these enzymes. The sub-class B2 metallo-{beta}-lactamase had 88% amino acid identity to its closet match blaPFM-1 and conferred carbapenem resistance to susceptible E. coli. The {beta}-lactamase inhibitors relebactam, vaborbactam, xeruborbactam and captopril had no impact on the carbapenem resistance phenotype. Analogues of all these novel genes (>95% nucleotide sequence identity) were identified within publicly available whole-genome sequencing data, suggesting they are found sporadically. ConclusionOur analysis adds to the growing number of {beta}-lactamase genes found from environmental Pseudomonas spp. and suggests that continued surveillance of this environmental reservoir for novel, clinically relevant, {beta}-lactamase genes is warranted.

BackgroundGlobally, seagrass ecosystems are threatened by anthropogenic activities that are leading to increased levels of eutrophication, coastal pollution and thermal conditions. Consequently, there is a growing need to develop new approaches that work to mitigate these stressors and enhance restoration efforts in seagrass meadows. One promising strategy is to identify, isolate and characterize microbial consortia that are likely to support seagrass productivity. However, our current understanding of key microbial functions that support plant growth in marine systems is limited. Based on evidence from terrestrial plant-microbe systems, seagrass-associated bacteria are expected to provide the plant with nitrogen and phosphorus resources while detoxifying sulfur and producing phytohormones. Here, we sequenced 61 bacterial cultures isolated from the rhizosphere, rhizoplane, and endosphere of the seagrass, Zostera marina to identify a consortium of six putative plant growth promoting (PGP) candidates. ResultsOur cultivation approach using plant-based media allowed us to isolate 201 bacteria from Z. marina, which reflected 18% of the total microbial diversity of the starting inoculum. Genomic and phenotypic analyses of the 61-sequenced pure-cultures revealed that most of the sequenced taxa were able to mobilize nitrogen primarily through catabolic pathways, including denitrification (51%), dissimilatory nitrate reduction to ammonia (71%), and C-N bond cleavage (83%). Six of the isolates, which represent new lineages of Agarivorans, coded for the nitrogenase gene cassette. Additionally, 52% of the genomes had genes for sulfur and/or thiosulfate oxidation, 88.5% for phosphorus solubilization, and 60.5% for IAA production. Genomic analysis also revealed that some pathways, including denitrification and dissimilatory nitrite to ammonia DNRA, required cross-species cooperation as no one taxa contained all the genes needed to complete these metabolic pathways. Based on draft genome models and results from phenotypic assays, isolates Streptomyces sp. (Iso23 and Iso384), Mesobacillus sp (Iso127), Roseibuim sp. (Iso195), Peribacillus sp. (Iso49), and Agarivorans sp. (Iso311) represent a minimal microbial community that is likely to promote seagrass growth and enhance restoration efforts. ConclusionOur work provides a detailed genomic and phenotypic analysis of bacteria isolated from Z. marina and identifies a minimal microbial community with complementary PGP traits. Isolating, identifying and characterizing bacteria that promote seagrass growth is critical towards enhancing restoration efforts of seagrass meadows.