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At least 469 records · Page 26
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Teredinibacter haidensis sp. nov., Teredinibacter purpureus sp. nov. and Teredinibacter franksiae sp. nov., marine, cellulolytic endosymbiotic bacteria isolated from the gills of the wood-boring mollusc Bankia setacea (Bivalvia: Teredinidae) and emended description of the genus Teredinibacter

Here, we describe three endosymbiotic bacterial strains isolated from the gills of the shipworm, Bankia setacea (Teredinidae: Bivalvia). These strains, designated as Bs08 T , Bs12 T and Bsc2 T , are Gram-stain-negative, microaerobic, gammaproteobacteria that grow on cellulose and a variety of substrates derived from lignocellulose. Phenotypic characterization, phylogeny based on 16S rRNA gene and whole genome sequence data, amino acid identity and percentage of conserved proteins analyses, show that these strains are novel and may be assigned to the genus Teredinibacter. The three strains may be differentiated and distinguished from other previously described Teredinibacter species based on a combination of four characteristics: colony colour (Bs12 T , purple; others beige to brown), marine salt requirement (Bs12 T , Bsc2 T and Teredinibacter turnerae strains), the capacity for nitrogen fixation (Bs08 T and T. turnerae strains) and the ability to respire nitrate (Bs08 T ). Based on these findings, we propose the names Teredinibacter haidensis sp. nov. (type strain Bs08 T =ATCC TSD-121 T =KCTC 62964 T ), Teredinibacter purpureus sp. nov. (type strain Bs12 T =ATCC TSD-122 T =KCTC 62965 T ) and Teredinibacter franksiae sp. nov. (type strain Bsc2 T =ATCC TSD-123 T =KCTC 62966 T ).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

From viruses to protists: temperature response of the neglected components of microbial controls on peatland nutrient cycling

The response of plant-associated microbial communities to rising temperatures likely plays a key role in global Carbon (C) and Nitrogen (N) biogeochemical cycles. Mosses of the genus Sphagnum, in particular, sequester 25% of all terrestrial C as recalcitrant peat. However, their ability to sequester C is mediated by interactions with microbes that fix N and pass it to the moss to grow in otherwise oligotrophic systems. Two important biotic controls on this processes are likely to respond to rising temperatures –predation by protists and infections by viruses– but whether and how this might occur is not well understood. This proposal addressed these questions using a combination of genomics and mathematical modeling with both field data from whole-ecosystem warming experiments, and laboratory-based microcosm experiments. We have discovered 1) mechanistic links between cell traits and thermal performance in protists, which 2) enabled predictions from populations to ecosystems, 3) we have identified eco-phenotypic feedbacks that determine protist–bacterial predator–prey dynamics, 4) have documented concomitant compositional and trait shifts in protist communities with warming in the field, 5) revealed consistent interactive effects of temperature and nutrients on bacterial and protist communities; 6) characterized the Sphagnum virome, and 7) made modeling predictions on its implications for thermal responses of microbial communities in peatlands globally. This award also substantially strengthened U.S. scientific capacity and workforce development.

Gibert, Jean Philippe [Duke University]↗
DOE OSTI2023

Genetic determinants of switchgrass-root-associated microbiota in field sites spanning its natural range

A fundamental goal in plant microbiome research is to determine the relative impacts of host and environmental effects on root microbiota composition, particularly how host genotype impacts bacterial community composition. Most studies characterizing the effect of plant genotype on root microbiota undersample host genetic diversity and grow plants outside of their native ranges, making the associations between host and microbes difficult to interpret. Here, we characterized the root microbiota of a large diversity panel of switchgrass, a North American native C4 bioenergy crop, in three field locations spanning its native range. Our data, composed of 1,961 samples, suggest that field location is the primary determinant of microbiome composition; however, substantial heritable variation is widespread across bacterial taxa, especially those in the Sphingomonadaceae family. Despite diverse compositions, relatively few highly prevalent taxa make up the majority of the switchgrass root microbiota, a large fraction of which is shared across sites. Local genotypes preferentially recruit/filter for local microbes, supporting the idea of affinity between local plants and their microbiota. Using genome-wide association, we identified loci impacting the abundance of >400 microbial strains and found an enrichment of genes involved in immune responses, signaling pathways, and secondary metabolism. We found loci associated with over half of the core microbiota (i.e., microbes in >80% of samples), regardless of field location. Finally, we show a genetic relationship between a basal plant immunity pathway and relative abundances of root microbiota. Furthermore, this study brings us closer to harnessing and manipulating beneficial microbial associations via host genetics.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2020

Phage-specific metabolic reprogramming of virocells

Ocean viruses are abundant and infect 20–40% of surface microbes. Infected cells, termed virocells, are thus a predominant microbial state. Yet, virocells and their ecosystem impacts are understudied, thus precluding their incorporation into ecosystem models. Here we investigated how unrelated bacterial viruses (phages) reprogram one host into contrasting virocells with different potential ecosystem footprints. We independently infected the marine Pseudoalteromonas bacterium with siphovirus PSA-HS2 and podovirus PSA-HP1. Time-resolved multi-omics unveiled drastically different metabolic reprogramming and resource requirements by each virocell, which were related to phage–host genomic complementarity and viral fitness. Namely, HS2 was more complementary to the host in nucleotides and amino acids, and fitter during infection than HP1. Functionally, HS2 virocells hardly differed from uninfected cells, with minimal host metabolism impacts. HS2 virocells repressed energy-consuming metabolisms, including motility and translation. Contrastingly, HP1 virocells substantially differed from uninfected cells. They repressed host transcription, responded to infection continuously, and drastically reprogrammed resource acquisition, central carbon and energy metabolisms. Ecologically, this work suggests that one cell, infected versus uninfected, can have immensely different metabolisms that affect the ecosystem differently. Finally, we relate phage–host genome complementarity, virocell metabolic reprogramming, and viral fitness in a conceptual model to guide incorporating viruses into ecosystem models.

Environmental Sciences & Ecology↗
DOE OSTI2024

A novel conjugative transposon carrying an autonomously amplified plasmid

Tetracyclines serve as broad-spectrum antibiotics to treat bacterial infections. The discovery of new tetracycline resistance genes has led to new questions about the underlying mechanisms of resistance, gene transfer, and their relevance to human health. We tracked changes in the abundance of a 55-kbp conjugative transposon (CTn214) carrying tetQ, a tetracycline resistance gene, within a Bacteroides fragilis metagenome-assembled genome derived from shotgun sequencing of microbial DNA extracted from the ileal pouch of a patient with ulcerative colitis. The mapping of metagenomic reads to CTn214 revealed the multi-copy nature of a 17,044-nt region containing tetQ in samples collected during inflammation and uninflamed visits. B. fragilis cultivars isolated from the same patient during periods of inflammation harbored CTn214 integrated into the chromosome or both a circular, multi-copy, extrachromosomal region of the CTn214 containing tetQ and the corresponding integrated form. The tetracycline-dependent mechanism for the transmission of CTn214 is nearly identical to a common conjugative transposon found in the genome of B. fragilis (CTnDOT), but the autonomously amplified nature of a circular 17,044-nt region of CTn214 that codes for tetQ and the integration of the same sequence in the linear chromosome within the same cell is a novel observation. Genome and transcriptome sequencing of B. fragilis cultivars grown under different concentrations of tetracycline and ciprofloxacin indicates that tetQ in strains containing the circular form remains actively expressed regardless of treatment, while the expression of tetQ in strains containing the linear form increases only in the presence of tetracycline.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

PlasmidHostFinder: Prediction of Plasmid Hosts Using Random Forest

Plasmids play a major role facilitating the spread of antimicrobial resistance between bacteria. Understanding the host range and dissemination trajectories of plasmids is critical for surveillance and prevention of antimicrobial resistance. Identification of plasmid host ranges could be improved using automated pattern detection methods compared to homology-based methods due to the diversity and genetic plasticity of plasmids. In this study, we developed a method for predicting the host range of plasmids using machine learning—specifically, random forests. We trained the models with 8,519 plasmids from 359 different bacterial species per taxonomic level; the models achieved Matthews correlation coefficients of 0.662 and 0.867 at the species and order levels, respectively. Our results suggest that despite the diverse nature and genetic plasticity of plasmids, our random forest model can accurately distinguish between plasmid hosts. This tool is available online through the Center for Genomic Epidemiology (https://cge.cbs.dtu.dk/services/PlasmidHostFinder/).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Data for Immediate Impacts of Soybean Cover Crop on Bacterial Community Composition and Diversity in Soil Under Long-Term Saccharum Monoculture

Saccharum yield decline results from long-term monoculture practices. Changes in cropping management can improve soil health and productivity. Below-ground bacterial community diversity and composition across soybean (Glycine max (L.) Merr) cover crop, Saccharum monoculture (30+ year) and fallowed soil were determined. Near full length (~1,400 base pairs) of 16S rRNA gene sequences were extracted from the rhizospheres of sugarcane and soybean and fallowed soil were compared. Higher soil bacterial diversity was observed in the soybean cover crop than sugarcane monoculture across all measured indices (observed operationational taxonomic units, Chao1, Shannon, reciprocal Simpson and Jackknife). Acidocateria, Proteobacteria, Bacteroidetes and Planctomycetes were the most abundant bacterial phyla across the treatments. Indicator species analysis identified nine indicator phyla. Planctomycetes, Armatimonadetes and candidate phylum FBP were associated with soybean; Proteobacteria and Firmicutes were linked with sugarcane and Gemmatimonadetes, Nitrospirae, Rokubacteria and unclassified bacteria were associated with fallowed soil. Non-metric multidimensional scaling analysis showed distinct groupings of bacterial operational taxonomic units (97% identity) according to management system (soybean, sugarcane or fallow) indicating compositional differences among treatments. This is confirmed by the results of the multi-response permutation procedures (A = 0.541, p = 0.00045716). No correlation between soil parameters and bacterial community structure was observed according to Mantel test (r = 211865, p = 0.14). Use of soybean cover-crop fostered bacterial diversity and altered community structure. This indicates cover crops could have a restorative effect and potentially promote sustainability in long-term Saccharum production systems.

Field Data↗
DOE OSTI2020

WHONDRS Summer 2019 Sampling Campaign: Global River Corridor Sediment FTICR-MS, Dissolved Organic Carbon, Aerobic Respiration, Elemental Composition, Grain Size, Total Nitrogen and Organic Carbon Content, Bacterial Abundance, and Stable Isotopes (v10)

The WHONDRS Summer 2019 Sampling (S19S) Campaign collected samples in 97 globally distributed river corridor systems between July and September 2019. Surficial streambed sediments were collected at three locations within each site (upstream, midstream, and downstream). Surface water was collected at the downstream site. This dataset includes a portion of the data types produced from the sediment samples and does not include any results from the surface water. Surface water data can be found at https://data.ess-dive.lbl.gov/view/doi:10.15485/1603775. The S19S campaign was designed with the science community to ask questions associated with links among core/transient metabolomes, microbial metabolism, biogeochemical function, and physical properties of watershed and river corridor systems. This data package was originally published in December 2020. It was updated in April 2021 (v2; modified files), May 2022 (v3; new and modified files), November 2022 (v4; new and modified files), January 2023 (v5 and v6; new and modified files), February 2023 (v7; new and modified files), August 2023 (v8; new and modified files), July 2025 (v9; new and modified files), and June 2026 (v10; new and modified files). Version 10 includes major restructuring of the files and their formats. See the change history in the readme for details. In addition to a readme, this data package also includes a file-level metadata (FLMD) file that describes each file and a data dictionary (DD) that describes all column/row headers and variable definitions. This dataset contains (1) readme, (2) data dictionary, (3) file level metadata, (4) sampling protocol, (5) a data checks report, (6) field metadata, (7) international generic sample number (IGSN) mapping file, (8) joint genome institute (JGI) mapping file, (9) a folder of site photos, and (10) a folder of sample data. The sample data folder contains (1) dissolved organic carbon (DOC, measured as non-purgeable organic carbon, NPOC) and FTICR methods; (2) respiration rates and normalized respiration rates calculated from laboratory sediment incubations, (3) X-ray fluorescence (XRF) elemental composition, (4) grain size distribution from the < 2 millimeter fraction of sediment, (5) total organic carbon and total nitrogen content, (6) bacterial abundance flow cytometry data, (7) sediment stable isotopes (C and N), (8) laboratory methods, (9) water and sediment mass and volume used in incubations, (10) a folder with code, inputs, and plots for the respiration rates, (11) a folder containing XRF data, (12) a folder containing unaligned, peak-picked high resolution characterization of dissolved organic matter from sediment via 12 Tesla Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) through the Environmental Molecular Sciences Laboratory (EMSL; https://www.pnnl.gov/environmental-molecular-sciences-laboratory), (12) and a folder containing processed FTICR-MS data. All files are .msa, .txt, .spx, .xml, .csv, .pdf, .txt, .jpg, or .jpeg.

13C↗
DOE OSTI2026

Multi-strain analysis of Pseudomonas putida reveals the metabolic and genetic diversity of the species

Pseudomonas putida is a gram-negative bacterial species increasingly utilized in biotechnology due to its robust growth, ability to degrade aromatic compounds, solvent tolerance, and genetic tractability. In this study, we report a comprehensive multi-strain analysis of 164 P. putida strains based on the reconstruction of a pan-putida metabolic network and the formulation of strain-specific genome-scale metabolic models (GEMs). We performed whole-genome sequencing and hybrid assembly for 40 strains, contributing a ~8% increase to the available genomic data for P. putida . Furthermore, high-throughput phenotypic profiling using the Biolog phenotype microarray system for 24 strains on 190 unique carbon sources, along with 15 aromatic compounds not present on Biolog plates, yielded 4,920 unique strain-phenotype measurements. These data were leveraged to curate GEMs for 24 representative strains, including a refined model for strain KT2440, which comprised 1,480 genes and 2,191 metabolites, achieving a prediction accuracy of 91.2% in carbon utilization. Systematic comparison of genomes and GEMs revealed both conserved core pathways and significant allelic and functional divergence across strains, highlighting strain-specific variation in aromatic degradation. While pathways for protocatechuate and phenylacetate degradation were widely conserved, metabolic capabilities for compounds such as ferulate, phenol, and cresols varied markedly, suggesting adaptation to distinct ecological niches. Alleleome analysis of enzymes, such as PcaI and PcaJ, revealed distinct, functionally similar clades, indicating possible convergent evolution or horizontal gene transfer. These results provide computable resources and informative models for selecting P. putida strains with desired traits for biomanufacturing and bioremediation and offer insights into the evolution and phylogeny of the P. putida species.

aromatics utilization↗
DOE OSTI2026

MassIVE MSV000100641 - Multilayered regulation by RNA thermometers enables precise control of Cas9 expression in E. coli: Cas9-based genome editing technologies can rapidly generate mutations to probe a diverse array of mutant genotypes. However, aberrant Cas9 nuclease translation and activity can occur despite the use of inducible promoters to control expression, leading to extensive cell death. This background killing caused by promoter leakiness severely limits the application of Cas9 for generating mutant libraries because of the potential for population skew. We demonstrate the utility of temperature sensitive RNA elements as a layer of post-transcriptional regulation to reduce the impact of promoter leak. We observe significant temperature-dependent increases in cell survival when certain RNA thermometers (RNATs) are placed upstream of the cas9 coding sequence. We also show that the most highly repressing RNAT, hsp17rep, significantly reduces population skew with a library of characterized guide RNAs in Escherichia coli. This strategy should be applicable to all bacterial Cas9-based methods and technologies.

Explore the source record for details and available documents.

Alexander, William G. [Oak Ridge National Laborato↗
DOE OSTI2023

Expression and characterization of spore coat Cot$\mathrm{H}$ kinases from the cellulosomes of anaerobic fungi ( Neocallimastigomycetes )

Anaerobic fungi (Neocallimastigomycetes) found in the guts of herbivores are biomass deconstruction specialists with a remarkable ability to extract sugars from recalcitrant plant material. Anaerobic fungi, as well as many species of anaerobic bacteria, deploy multi-enzyme complexes called cellulosomes, which modularly tether together hydrolytic enzymes, to accelerate biomass hydrolysis. While the majority of genomically encoded cellulosomal genes in Neocallimastigomycetes are biomass degrading enzymes, the second largest family of cellulosomal genes encode spore coat CotH domains, whose contribution to fungal cellulosome and/or cellular function is unknown. Structural bioinformatics of CotH proteins from the anaerobic fungus Piromyces finnis shows anaerobic fungal CotH domains conserve key ATP and Mg 2+ binding motifs from bacterial Bacillus CotH proteins known to act as protein kinases. Experimental characterization further demonstrates ATP hydrolysis activity in the presence and absence of substrate from two cellulosomal P. finnis CotH proteins when recombinantly produced in E. coli. These results present foundational evidence for CotH activity in anaerobic fungi and provide a path towards elucidating the functional contribution of this protein family to fungal cellulosome assembly and activity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Emergence and disruption of cooperativity in a denitrifying microbial community

Anthropogenic perturbations to the nitrogen cycle, primarily through use of synthetic fertilizers, is driving an unprecedented increase in the emission of nitrous oxide (N 2 O), a potent greenhouse gas and an ozone depleting substance, causing urgency in identifying the sources and sinks of N 2 O. Microbial denitrification is a primary contributor to biotic production of N 2 O in anoxic regions of soil, marine systems, and wastewater treatment facilities. Here, through comprehensive genome analysis, we show that pathway partitioning is a ubiquitous mechanism of complete denitrification within microbial communities. We have investigated mechanisms and consequences of process partitioning of denitrification through detailed physiological characterization and kinetic modeling of a synthetic community of Rhodanobacter thiooxydans FW510-R12 and Acidovorax sp. GW101-3H11. We have discovered that these two bacterial isolates, from a heavily nitrate (NO 3 − ) contaminated superfund site, complete denitrification through the exchange of nitrite (NO 2 − ) and nitric oxide (NO). The process partitioning of denitrification and other processes, including amino acid metabolism, contribute to increased cooperativity within this denitrifying community. We demonstrate that certain contexts, such as high NO 3 − , cause unbalanced growth of community members, due to differences in their substrate utilization kinetics. The altered growth characteristics of community members drives accumulation of toxic NO 2 − , which disrupts denitrification causing N 2 O off gassing.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Signal sequences target enzymes and structural proteins to bacterial microcompartments and are critical for microcompartment formation

ABSTRACT Spatial organization of pathway enzymes has emerged as a promising tool to address several challenges in metabolic engineering, such as flux imbalances and off-target product formation. Bacterial microcompartments (MCPs) are a spatial organization strategy used natively by many bacteria to encapsulate metabolic pathways that produce toxic, volatile intermediates. Several recent studies have focused on engineering MCPs to encapsulate heterologous pathways of interest, but how this engineering affects MCP assembly and function is poorly understood. In this study, we investigated the role of signal sequences, short domains that target proteins to the MCP core, in the assembly of 1,2-propanediol utilization (Pdu) MCPs. We characterized two novel Pdu signal sequences on the structural proteins PduM and PduB, which constitute the first report of metabolosome signal sequences on structural proteins rather than enzymes. We then explored the role of enzymatic and structural Pdu signal sequences on MCP assembly by deleting their encoding sequences from the genome alone and in combination. Deleting enzymatic signal sequences decreased the MCP formation, but this defect could be recovered in some cases by overexpressing genes encoding the knocked-out signal sequence fused to a heterologous protein. By contrast, deleting structural signal sequences caused similar defects to knocking out the genes encoding the full-length PduM and PduB proteins. Our results contribute to a growing understanding of how MCPs form and function in bacteria and provide strategies to mitigate assembly disruption when encapsulating heterologous pathways in MCPs. IMPORTANCE Spatially organizing biosynthetic pathway enzymes is a promising strategy to increase pathway throughput and yield. Bacterial microcompartments (MCPs) are proteinaceous organelles that many bacteria natively use as a spatial organization strategy to encapsulate niche metabolic pathways, providing significant metabolic benefits. Encapsulating heterologous pathways of interest in MCPs could confer these benefits to industrially relevant pathways. Here, we investigate the role of signal sequences, short domains that target proteins for encapsulation in MCPs, in the assembly of 1,2-propanediol utilization (Pdu) MCPs. We characterize two novel signal sequences on structural proteins, constituting the first Pdu signal sequences found on structural proteins rather than enzymes, and perform knockout studies to compare the impacts of enzymatic and structural signal sequences on MCP assembly. Our results demonstrate that enzymatic and structural signal sequences play critical but distinct roles in Pdu MCP assembly and provide design rules for engineering MCPs while minimizing disruption to MCP assembly.

Johnson, Elizabeth R. (ORCID:0000000179236881)↗
DOE OSTI2022

BMC Caller: a webtool to identify and analyze bacterial microcompartment types in sequence data

Bacterial microcompartments (BMCs) are protein-based organelles found across the bacterial tree of life. They consist of a shell, made of proteins that oligomerize into hexagonally and pentagonally shaped building blocks, that surrounds enzymes constituting a segment of a metabolic pathway. The proteins of the shell are unique to BMCs. They also provide selective permeability; this selectivity is dictated by the requirements of their cargo enzymes. We have recently surveyed the wealth of different BMC types and their occurrence in all available genome sequence data by analyzing and categorizing their components found in chromosomal loci using HMM (Hidden Markov Model) protein profiles. To make this a “do-it yourself” analysis for the public we have devised a webserver, BMC Caller (https://bmc-caller.prl.msu.edu), that compares user input sequences to our HMM profiles, creates a BMC locus visualization, and defines the functional type of BMC, if known. Shell proteins in the input sequence data are also classified according to our function-agnostic naming system and there are links to similar proteins in our database as well as an external link to a structure prediction website to easily generate structural models of the shell proteins, which facilitates understanding permeability properties of the shell. Additionally, the BMC Caller website contains a wealth of information on previously analyzed BMC loci with links to detailed data for each BMC protein and phylogenetic information on the BMC shell proteins. Our tools greatly facilitate BMC type identification to provide the user information about the associated organism’s metabolism and enable discovery of new BMC types by providing a reference database of all currently known examples.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Unravelling the diversity of magnetotactic bacteria through analysis of open genomic databases

Magnetotactic bacteria (MTB) are prokaryotes that possess genes for the synthesis of membrane-bounded crystals of magnetite or greigite, called magnetosomes. Despite over half a century of studying MTB, only about 60 genomes have been sequenced. Most belong to Proteobacteria, with a minority affiliated with the Nitrospirae, Omnitrophica, Planctomycetes, and Latescibacteria. Due to the scanty information available regarding MTB phylogenetic diversity, little is known about their ecology, evolution and about the magnetosome biomineralization process. This study presents a large-scale search of magnetosome biomineralization genes and reveals 38 new MTB genomes. Several of these genomes were detected in the phyla Elusimicrobia, Candidatus Hydrogenedentes, and Nitrospinae, where magnetotactic representatives have not previously been reported. Analysis of the obtained putative magnetosome biomineralization genes revealed a monophyletic origin capable of putative greigite magnetosome synthesis. The ecological distributions of the reconstructed MTB genomes were also analyzed and several patterns were identified. These data suggest that open databases are an excellent source for obtaining new information of interest.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Depth-specific distribution of bacterial MAGs in permafrost active layer in Ny Ålesund, Svalbard (79°N)

Arctic soil microbial communities may shift with increasing temperatures and water availability from climate change. We examined temperature and volumetric liquid water content (VWC) in the upper 80 cm of permafrost-affected soil over 2 years (2018–2019) at the Bayelva monitoring station, Ny Ålesund, Svalbard. We show VWC increases with depth, whereas in situ temperature is more stable vertically, ranging from -5°C to 5°C seasonally. Prokaryotic metagenome-assembled genomes (MAGs) were obtained at 2–4 cm vertical resolution collected while frozen in April 2018 and at 10 cm vertical resolution collected while thawed in September 2019. The most abundant MAGs were Acidobacteriota, Actinomycetota, and Chloroflexota. Actinomycetota and Chloroflexota increase with depth, while Acidobacteriota classes Thermoanaerobaculia Gp7-AA8, Blastocatellia UBA7656, and Vicinamibacteria Vicinamibacterales are found above 6 cm, below 6 cm, and below 20cm, respectively. All MAGs have diverse carbon-degrading genes, and Actinomycetota and Chloroflexota have autotrophic genes. Further, genes encoding β -glucosidase, N-acetyl-β-D-glucosaminidase, and xylosidase increase with depth, indicating a greater potential for organic matter degradation with higher VWC. Acidobacteriota dominate the top 6 cm with their classes segregating by depth, whereas Actinomycetota and Chloroflexota dominate below ~6 cm. This suggests that Acidobacteriota classes adapt to lower VWC at the surface, while Actinomycetota and Chloroflexota persist below 6cm with higher VWC. This indicates that VWC may be as important as temperature in microbial climate change responses in Arctic mineral soils. Here we describe MAG-based Seqcode type species in the Acidobacteriota, Onstottus arcticum, Onstottus frigus, and Gilichinskyi gelida and in the Actinobacteriota, Mayfieldus profundus.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2026

Advancing Protein Display on Bacterial Spores through an Extensive Survey of Coat Components

The profound stability of bacterial spores makes them a promising platform for biotechnological applications like biocatalysis, bioremediation, drug delivery, etc. However, though the Bacillus subtilis spore is composed of >40 types of proteins, only ∼12 have been explored as fusion carriers for protein display. Here, we assessed the suitability of 33 spore proteins (SPs) as enzyme display carriers by direct allele tagging at native genomic loci. Of the 33 SPs investigated, 26 formed functional fusions with β-glucuronidase (GUS)─a ∼272 kDa homotetramer. This almost triples the number of SPs assessed for enzyme display and doubles the number of functional fusions documented in the literature. We quantitatively assessed 1) SP promoter activation dynamics, 2) GUS activity on spores, 3) surface availability, and 4) protection from thermal and proteolytic degradation. Multicopy expression and pairwise coexpression of the most promising SP-GUS fusions highlighted the complexity of spore structure/assembly and the difficulty in predicting compatibility between different SP fusions. We also assessed the suitability of engineered spores to degrade PET (polyethylene terephthalate) films and found that surface-exposed SPs were most effective. Beyond the broad survey, a key outcome of our work was the identification of SscA (small spore coat assembly protein A) as an effective spore display carrier. SscA supported enzyme activity at least 4-fold higher than any other SP, including the well-established anchor, CotY. We attribute this to its promoter, which demonstrated early and sustained activation relative to other SPs and its small size (∼3 kDa), which likely minimally interferes with enzyme folding, oligomerization, and activity. Labeling and genetic studies, its hydrophobic nature, and low surface availability suggest that SscA assembles within the inner spore coat, which makes it stabilizing and suitable for many biocatalytic applications. Overall, this work serves as a knowledge base to advance the biotechnological utility of B. subtilis spores.

Bacillus subtilis↗
DOE OSTI2021

Genetic and behavioral adaptation of Candida parapsilosis to the microbiome of hospitalized infants revealed by in situ genomics, transcriptomics, and proteomics

Background Candida parapsilosis is a common cause of invasive candidiasis, especially in newborn infants, and infections have been increasing over the past two decades. C. parapsilosis has been primarily studied in pure culture, leaving gaps in understanding of its function in a microbiome context. Results. Here, we compare five unique C. parapsilosis genomes assembled from premature infant fecal samples, three of which are newly reconstructed, and analyze their genome structure, population diversity, and in situ activity relative to reference strains in pure culture. All five genomes contain hotspots of single nucleotide variants, some of which are shared by strains from multiple hospitals. A subset of environmental and hospital-derived genomes share variants within these hotspots suggesting derivation of that region from a common ancestor. Four of the newly reconstructed C. parapsilosis genomes have 4 to 16 copies of the gene RTA3, which encodes a lipid translocase and is implicated in antifungal resistance, potentially indicating adaptation to hospital antifungal use. Time course metatranscriptomics and metaproteomics on fecal samples from a premature infant with a C. parapsilosis blood infection revealed highly variable in situ expression patterns that are distinct from those of similar strains in pure cultures. For example, biofilm formation genes were relatively less expressed in situ, whereas genes linked to oxygen utilization were more highly expressed, indicative of growth in a relatively aerobic environment. In gut microbiome samples, C. parapsilosis co-existed with Enterococcus faecalis that shifted in relative abundance over time, accompanied by changes in bacterial and fungal gene expression and proteome composition. Conclusions The results reveal potentially medically relevant differences in Candida function in gut vs. laboratory environments, and constrain evolutionary processes that could contribute to hospital strain persistence and transfer into premature infant microbiomes.

59 BASIC BIOLOGICAL SCIENCES↗