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At least 379 records · Page 21

A red algal polysaccharide influences the multicellular development of the choanoflagellate Salpingoeca rosetta

The choanoflagellate Salpingoeca rosetta exemplifies the capacity of marine microeukaryotes to integrate environmental cues into their life histories. Some of the cues that S. rosetta detects are lipids from their bacterial prey that induce the development of multicellular colonies called rosettes. In the aquatic environments where S. rosetta would encounter bacterial cues, microbial communities gain refuge and food from algae. For example, microbes in coastal environments benefit from macroalgae that annually release ∼10 14 g of carbon, primarily in the form of polysaccharides that comprise ∼80% of dissolved organic carbon in surface waters. Despite the ecological impact of algae and historical descriptions of choanoflagellates attached to algae, the influence of algae on choanoflagellate life history transitions has remained unexplored. Here, we report that porphyran, a polysaccharide produced by the red macroalga Porphyra umbilicalis, induces multicellular development in S. rosetta. We first noticed this response when S. rosetta formed multicellular rosette colonies while growing in media prepared from P. umbilicalis, even though rosette-inducing bacteria were absent. By biochemically purifying extracts of P. umbilicalis, we identified porphyran as the rosette-inducing factor. This response provides a biochemical insight that indicates algal polysaccharides may serve as cues to mediate associations between choanoflagellates and algae. Moreover, this observation complements evidence from environmental and geochemical studies that show the impact that algae have exerted on the ecology and evolution of eukaryotes, including a rise in algal productivity during the origin of animals, the closest living relatives of choanoflagellates.

59 BASIC BIOLOGICAL SCIENCES↗

Tapping the treasure trove of atypical phages

With advancements in genomics technologies, a vast diversity of ‘atypical’ phages, that is, with single-stranded DNA or RNA genomes, are being uncovered from different ecosystems. Though these efforts have revealed the existence and prevalence of these nonmodel phages, computational approaches often fail to associate these phages with their specific bacterial host(s), while the lack of methods to isolate these phages has limited our ability to characterize infectivity pathways and new gene function. In this review, we call for the development of generalizable experimental methods to better capture this understudied viral diversity via isolation and study them through gene-level characterization and engineering. Establishing a diverse set of new ‘atypical’ phage model systems has the potential to provide many new biotechnologies, including potential uses of these atypical phages in halting the spread of antibiotic resistance and engineering of microbial communities for beneficial outcomes.

59 BASIC BIOLOGICAL SCIENCES↗

Plant-microbial interplay for organic nitrogen mediated by functional specificity of root compartments

The organic form of nitrogen (N) is a critical intermediate in mutualistic and competitive root-microbial interactions, mediated by extracellular enzymes. Visualization of the hotspots of organic N and proteolytic activity might be valuable for revealing root functional specificity in N acquisition and transformation at the level of individual roots and compartments. For the first time, we used time-lapse amino-mapping and zymography to co-localize and map the spatial distribution of amino-N and leucine aminopeptidase (LAP) activity in the soil and different root parts of maize (Zea mays L.). Amino-N distribution was mainly associated with seminal roots and root tips, where it overlapped with LAP activity hotspots. In the lateral roots and bulk soil, however, LAP activity was decoupled from amino-N. Distinct functional traits revealed themselves as the highest amino-N content and LAP activity in seminal root tips and as the largest relative extent of the rhizosphere in lateral root tips. Co-localized amino-N and LAP activities highlighted different nutrient acquisition strategies mediated by root-microbe interactions, depending on the root compartment. Seminal roots and their tips appeared to adopt mutualistic strategies, potentially attracting root-associated microorganisms through releasing oligo- and polypeptides. In contrast, lateral roots, with amino-N detected only at their tips, demonstrated stronger N competition, relying on the enzyme activity of the rhizosphere microbial community for N acquisition. These insights emphasized the role of root functional specialization in shaping plant-microbe interactions, offering pathways to enhance nutrient use efficiency.

Maize (Zea mays L.)↗

DNA viruses from different stages of a wastewater treatment plant in southwest Ohio

Wastewater treatment plants (WWTPs) house diverse populations of microbial communities that are dynamic across treatment stages. Although viruses, especially non-pathogenic ones are relatively less studied compared to their bacterial counterparts, they are equally important players in WWTP microbiomes. In this study, we sampled influent, activated sludge, and effluent stages of a WWTP in southwest Ohio to sequence their viral population using multiple displacement amplification (MDA) and metagenomic approaches. We recovered 1003 viral genomes, the majority of which were ssDNA (single-stranded DNA) viruses that formed distinct clusters representing novel species and phylogenetic groups not closely related to known human gut or terrestrial viruses. Additionally, tracking viruses across treatment stages showed several persisted across multiple treatment stages. These results suggest that further studies are needed to understand the persistence of viral populations, particularly non-pathogenic ones, across different stages of WWTPs.

59 BASIC BIOLOGICAL SCIENCES↗

Modeling the Impacts of Hydrogen Extraction on Anaerobic Fermentation Systems Using a Modified ADM1 Model

Here, this paper presents a modified ADM1 (mADM1) model for use in assessing the influence of active dissolved H 2 extraction on biogas production from anaerobic fermentation reactors, specifically in the context of high-strength brewery wastewater as the feed. The modified model adds regulation factors to account for the varying product stoichiometry of glucose degradation reactions as a function of H 2 concentration in anaerobic digestion, and incorporates the use of both lactate and ethanol as intermediates. The changes to the model are expected to better predict the effects of H 2 removal on the production of H 2 in fermentation processes. Both ADM1 and mADM1 were calibrated against experimental data from a fluidized-bed reactor containing encapsulated acidogenic-acetogenic microbial communities treating real brewery wastewater. The calibrated models were validated against another fluidized-bed reactor identical to the calibration reactor, with the addition of a hollow-fiber membrane module that was used to reduce dissolved H 2 concentrations in the validation reactor. It was found that ADM1 over-estimated the H 2 production rates of the validation reactor (117 mL/day simulated vs. 16.48 mL/day experimental), whereas mADM1 was able to successfully predict fermentation, and specifically the H 2 production rate (17 mL/day). The presented model is a first step towards a model that can predict the effects of dissolved H 2 removal on fermentation processes.

Anaerobic Digestion↗

Optimized CRISPR Interference System for Investigating Pseudomonas alloputida Genes Involved in Rhizosphere Microbiome Assembly

Pseudomonas alloputida KT2440 (formerly P. putida) has become both a well-known chassis organism for synthetic biology and a model organism for rhizosphere colonization. Here, we describe a CRISPR interference (CRISPRi) system in KT2440 for exploring microbe–microbe interactions in the rhizosphere and for use in industrial systems. Our CRISPRi system features three different promoter systems (XylS/P m , LacI/P lac , and AraC/P BAD ) and a dCas9 codon-optimized for Pseudomonads, all located on a mini-Tn7-based transposon that inserts into a neutral site in the genome. It also includes a suite of pSEVA-derived sgRNA expression vectors, where the expression is driven by synthetic promoters varying in strength. We compare the three promoter systems in terms of how well they can precisely modulate gene expression, and we discuss the impact of environmental factors, such as media choice, on the success of CRISPRi. We demonstrate that CRISPRi is functional in bacteria colonizing the rhizosphere, with repression of essential genes leading to a 10–100-fold reduction in P. alloputida cells per root. Finally, we show that CRISPRi can be used to modulate microbe–microbe interactions. When the gene pvdH is repressed and P. alloputida is unable to produce pyoverdine, it loses its ability to inhibit other microbes in vitro. Furthermore, our design is amendable for future CRISPRi-seq studies and in multispecies microbial communities, with the different promoter systems providing a means to control the level of gene expression in many different environments.

Bacteria↗

Distinct Fracture Mineralogy That is Out of Equilibrium With Modern Groundwaters Provides Important Context for Subsurface Life

Rock fracture surfaces in the crust are essential habitat for microorganisms. Fracture‐groundwater interfaces provide physical substrates for biofilm growth and are sources of carbon, nutrients, and electron donors and acceptors. To better understand geochemical processes impacting fracture surfaces and the subsurface microbiome, we identified fractures in archived rock cores from the Soudan formation, which is known to host saline groundwaters and isolated microbial communities dependent on rock‐water interactions. Cores with open fractures were thin sectioned and studied via electron microprobe and synchrotron X‐ray fluorescence microprobe. Most fracture surfaces had mineralogy distinct from that of the bulk rock. Chlorite minerals were abundant on fracture surfaces and had elemental compositions suggesting deposition during late‐stage hydrothermal alteration. Fracture‐lining chlorites likely limit access to iron oxide and sulfide minerals that are active in subsurface biogeochemical cycles. Calcium‐rich rinds were also observed along fracture edges. These rinds were too thin and poorly ordered to be identified via light microscopy or X‐ray diffraction; however, Ca K‐edge micro‐X‐ray absorption near‐edge structure spectroscopy identified them as carbonates, minerals not observed in the bulk rock. Thermodynamic modeling shows that carbonate precipitation is largely unfavorable in Soudan groundwaters, indicating that fracture edge conditions differed from those in modern water samples. Because of the low carbon concentrations in Soudan groundwaters, carbonate rinds likely play an important role in subsurface carbon cycling and may mark fracture surfaces that once hosted biofilms. Overall, this study suggests that fracture alteration can both play an active role in and suppress rock‐water interactions essential to subsurface life.

36 MATERIALS SCIENCE↗

Origin of biogeographically distinct ecotypes during laboratory evolution

Resource partitioning is central to the incredible productivity of microbial communities, including gigatons in annual methane emissions through syntrophic interactions. Previous work revealed how a sulfate reducer (Desulfovibrio vulgaris, Dv) and a methanogen (Methanococcus maripaludis, Mm) underwent evolutionary diversification in a planktonic context, improving stability, cooperativity, and productivity within 300-1000 generations. Here, we show that mutations in just 15 Dv and 7 Mm genes within a minimal assemblage of this evolved community gave rise to co-existing ecotypes that were spatially enriched within a few days of culturing in a fluidized bed reactor. The spatially segregated communities partitioned resources in the simulated subsurface environment, with greater lactate utilization by attached Dv but partial utilization of resulting H 2 by low affinity hydrogenases of Mm in the same phase. The unutilized H 2 was scavenged by high affinity hydrogenases of planktonic Mm, producing copious amounts of methane. Our findings show how a few mutations can drive resource partitioning amongst niche-differentiated ecotypes, whose interplay synergistically improves productivity of the entire mutualistic community.

59 BASIC BIOLOGICAL SCIENCES↗

CRAGE-RB-PI-seq reveals transcriptional dynamics of plant-associated bacteria during root colonization

Plant roots release a wide array of metabolites into the rhizosphere, shaping microbial communities and their functions. While metagenomics has expanded our understanding of these communities, little is known about the physiology of their members in host environments. Transcriptome analysis via RNA sequencing is a common approach to learning more, but its use has been challenging because of low bacterial biomass and interference from plant RNA. To overcome this, we developed a randomly-barcoded promoter-library insertion sequencing (RB-PI-seq) combined with chassis-independent recombinase-assisted genome engineering (CRAGE). Using Pseudomonas simiae WCS417 as a model rhizobacterium, this method enabled targeted amplification of barcoded transcripts, bypassing plant RNA interference and allowing measurement of thousands of promoter activities during Arabidopsis root colonization. Our analysis revealed temporally resolved transcriptional regulation, including those associated with cell growth, chemotaxis, plant immune suppression, biofilm formation, and stress responses, reflecting the coordinated physiological adaptation to the root environment. Additionally, we discovered that transcriptional activation of xanthine dehydrogenase and a lysozyme inhibitor is crucial for evading plant immune systems. This framework is scalable to other bacterial species and provides new opportunities for understanding rhizobacterial gene regulation in native environments.

59 BASIC BIOLOGICAL SCIENCES↗

Quantitative stable isotope probing (qSIP)-informed metagenomics identifies viruses infecting chemoautotrophs

Aquatic environments absorb ~2.5 gigatonnes of atmospheric carbon each year1, more than the carbon stored in the atmosphere, soils, and all biomass combined. Primary producers transform this dissolved inorganic carbon into biomass that can subsequently flow into other trophic levels, or be released back into the environment through viral lysis. While there is substantial knowledge about the diversity and activity of viruses infecting photoautotrophic primary producers and the ecosystem impact, little is known about viruses infecting chemoautotrophs, representing a gap in our understanding of key processes driving microbial carbon cycling. Here, we combine metagenomics with quantitative 12/13C stable isotopic probing (qSIP) mesocosm experiments in a marine-derived meromictic pond to quantify population-specific isotopic enrichment, identify key chemoautotrophic primary producers, and virus-host dynamics. Isotopically enriched carbon is tracked from the genomes of chemoautotrophs to putative viruses, showing that active populations of hydrogen/sulfur-oxidizing chemoautotrophs (Thiomicrorhabdus, Hydrogenovibrio, Sulfurimonas, Sulfurovum) are targeted by viruses. This work provides the foundation for revealing the diversity and activity of viruses infecting globally-widespread chemoautotrophs. Our study sheds light on trophic interactions that impact microbial carbon cycling in aphotic environments and builds toward biogeochemical models that incorporate viral impacts on chemoautotrophic microbial communities.

59 BASIC BIOLOGICAL SCIENCES↗

Population-level control of two manganese oxidases expands the niche for bacterial manganese biomineralization

Abstract The enzymatic oxidation of aqueous divalent manganese (Mn) is a widespread microbial trait that produces reactive Mn(III, IV) oxide minerals. These biominerals drive carbon, nutrient, and trace metal cycles, thus playing important environmental and ecological roles. However, the regulatory mechanisms and physiological functions of Mn biomineralization are unknown. This challenge arises from the common occurrence of multiple Mn oxidases within the same organism and the use of Mn oxides as indicators of combined gene activity. Through the detection of gene activation in individual cells, we discover that expression ofmnxGandmcoA, two Mn oxidase-encoding genes inPseudomonas putidaGB-1, is confined to subsets of cells within the population, with each gene showing distinct spatiotemporal patterns that reflect local microenvironments. These coordinated intra-population dynamics control Mn biomineralization and illuminate the strategies used by microbial communities to dictate the extent, location, and timing of biogeochemical transformations.

Biotechnology & Applied Microbiology↗

Analysis of biofilm assembly by large area automated AFM

Biofilms are complex microbial communities critical in medical, industrial, and environmental contexts. Understanding their assembly, structure, genetic regulation, interspecies interactions, and environmental responses is key to developing effective control and mitigation strategies. While atomic force microscopy (AFM) offers critically important high-resolution insights on structural and functional properties at the cellular and even sub-cellular level, its limited scan range and labor-intensive nature restricts the ability to link these smaller scale features to the functional macroscale organization of the films. We begin to address this limitation by introducing an automated large area AFM approach capable of capturing high-resolution images over millimeter-scale areas, aided by machine learning for seamless image stitching, cell detection, and classification. Large area AFM is shown to provide a very detailed view of spatial heterogeneity and cellular morphology during the early stages of biofilm formation which were previously obscured. Using this approach, we examined the organization of Pantoea sp. YR343 on PFOTS-treated glass surfaces. Our findings reveal a preferred cellular orientation among surface-attached cells, forming a distinctive honeycomb pattern. Detailed mapping of flagella interactions suggests that flagellar coordination plays a role in biofilm assembly beyond initial attachment. Additionally, we use large-area AFM to characterize surface modifications on silicon substrates, observing a significant reduction in bacterial density. This highlights the potential of this method for studying surface modifications to better understand and control bacterial adhesion and biofilm formation.

59 BASIC BIOLOGICAL SCIENCES↗

Two decades of bacterial ecology and evolution in a freshwater lake

Ecology and evolution are considered distinct processes that interact on contemporary time scales in microbiomes. Here, to observe these processes in a natural system, we collected a two-decade, 471-metagenome time series from Lake Mendota (Wisconsin, USA). We assembled 2,855 species-representative genomes and found that genomic change was common and frequent. By tracking strain composition via single nucleotide variants, we identified cyclical seasonal patterns in 80% and decadal shifts in 20% of species. In the dominant freshwater family Nanopelagicaceae, environmental extremes coincided with shifts in strain composition and positive selection of amino acid and nucleic acid metabolism genes. Further, these genes identify organic nitrogen compounds as potential drivers of freshwater responses to global change. Seasonal and long-term strain dynamics could be regarded as ecological processes or, equivalently, as evolutionary change. Rather than as distinct interacting processes, we propose a conceptualization of ecology and evolution as a continuum to better describe change in microbial communities.

59 BASIC BIOLOGICAL SCIENCES↗

An integrated metagenomic, metabolomic and transcriptomic survey of Populus across genotypes and environments

Abstract Bridging molecular information to ecosystem-level processes would provide the capacity to understand system vulnerability and, potentially, a means for assessing ecosystem health. Here, we present an integrated dataset containing environmental and metagenomic information from plant-associated microbial communities, plant transcriptomics, plant and soil metabolomics, and soil chemistry and activity characterization measurements derived from the model tree species Populus trichocarpa . Soil, rhizosphere, root endosphere, and leaf samples were collected from 27 different P. trichocarpa genotypes grown in two different environments leading to an integrated dataset of 318 metagenomes, 98 plant transcriptomes, and 314 metabolomic profiles that are supported by diverse soil measurements. This expansive dataset will provide insights into causal linkages that relate genomic features and molecular level events to system-level properties and their environmental influences.

59 BASIC BIOLOGICAL SCIENCES↗

Microbial Metagenomes Across a Complete Phytoplankton Bloom Cycle: High-Resolution Sampling Every 4 Hours Over 22 Days

In May and June of 2021, marine microbial samples were collected for DNA sequencing in East Sound, WA, USA every 4 hours for 22 days. This high temporal resolution sampling effort captured the last 3 days of a Rhizosolenia sp. bloom, the initiation and complete bloom cycle of Chaetoceros socialis (8 days), and the following bacterial bloom (2 days). Metagenomes were completed on the time series, and the dataset includes 128 size-fractionated microbial samples (0.22–1.2 µm), providing gene abundances for the dominant members of bacteria, archaea, and viruses. This dataset also has time-matched nutrient analyses, flow cytometry data, and physical parameters of the environment at a single point of sampling within a coastal ecosystem that experiences regular bloom events, facilitating a range of modeling efforts that can be leveraged to understand microbial community structure and their influences on the growth, maintenance, and senescence of phytoplankton blooms.

59 BASIC BIOLOGICAL SCIENCES↗

Metagenome-assembled-genomes recovered from the Arctic drift expedition MOSAiC

The Multidisciplinary Observatory for Study of the Arctic Climate (MOSAiC) expedition consisted of a year-long drifting survey of the Central Arctic Ocean. The ecosystems component of MOSAiC included the sampling of molecular data, with metagenomes collected from a diverse range of environments. The generation of metagenome-assembled-genomes (MAGs) from metagenomes are a starting point for genome-resolved analyses. This dataset presents a catalogue of MAGs recovered from a set of 73 samples from MOSAiC, including 2407 prokaryotic and 56 eukaryotic MAGs, as well as annotations of a near complete eukaryotic MAG using the Joint Genome Institute (JGI) annotation pipeline. The metagenomic samples are from the surface ocean, chlorophyll maximum, mesopelagic and bathypelagic, within leads and under-ice ocean, as well as melt ponds, ice ridges, and first- and second-year sea ice. This set of MAGs can be used to benchmark microbial biodiversity in the Central Arctic Ocean, compare individual strains across space and time, and to study changes in Arctic microbial communities from the winter to summer, at a genomic level.

59 BASIC BIOLOGICAL SCIENCES↗

Dynamic soil columns simulate Arctic redox biogeochemistry and carbon release during changes in water saturation

Thawing Arctic permafrost can induce hydrologic change and alter redox conditions, shifting the balance of soil organic matter (SOM) decomposition. There remains uncertainty about how soil saturation and redox transitions impact dissolved and gas phase carbon fluxes, and efforts to link hydrobiogeochemical processes to ecosystem-scale models are limited. This study evaluates SOM decomposition of Arctic tundra soils using column experiments, water chemistry measurements, microbial community analysis, and a PFLOTRAN reactive transport model. Soil columns from a thermokarst channel (TC) and an upland tundra (UC) were exposed to cycles of saturation and drainage, which controlled carbon emissions. During saturation, an outflow of dissolved organic carbon from the UC soil correlated with elevated reduced iron and decreased pH; during drainage, UC carbon dioxide fluxes were 70% higher than TC fluxes. Intermittent methane release was observed for TC, consistent with higher methanogen abundance. Slower drainage in the TC soil correlated with more subtle biogeochemical changes. PFLOTRAN simulations captured experimental trends in soil carbon fluxes, oxygen concentrations, and water contents. The model was then used to evaluate additional soil water drainage rates. This study emphasizes the importance of considering hydrologic change when evaluating and simulating SOM decomposition in dynamic Arctic tundra environments.

54 ENVIRONMENTAL SCIENCES↗

The oleaginous yeast Cutaneotrichosporon oleaginosum modifies corn stover alkali lignin

The current paradigm in synthetic biology for lignin bioconversion platforms includes primarily bacteria and filamentous fungi. Yeast are notoriously understudied for their role in lignin degradation and utilization, despite their ubiquity in saprophytic microbial communities. A few publications report lignin-modifying yeasts, but investigations to date have relied on model aromatic compounds or lignin-containing substrates replete with other carbon sources. In this work, we use a suite of analytical tools to evaluate interactions between corn stover-extracted lignin and the oleaginous yeast Cutaneotrichosporon oleaginosum. Notably, 2D-NMR analysis showed a significant decrease in the H-lignin component as well as resinol (β-β) and phenylcoumaran (β-5) linkages. Using super-resolution fluorescence microscopy, we demonstrated that this yeast may uptake polymeric lignin and/or undertakes interactions at the cellular envelope. To explore mechanisms of lignin modification, transport, and aromatics catabolism, extensive secretomics and proteomics analyses were conducted. Compared to carbon-limited glucose and “No Carbon” controls, several putative laccases, quinone reductases, superoxide dismutases, and glyoxal/oxalate oxidases were upregulated in the lignin condition. Excitingly, two ferric reductases and an oxalate exchanger were only observed in the lignin condition. These results indicate that C. oleaginosum may perform extracellular quinone redox cycling to generate lignin-modifying reactive oxygen species. These findings enhance our understanding of yeast-lignin interactions and provide valuable insights for validation studies and metabolic engineering.

09 BIOMASS FUELS↗