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DOE OSTI2018

High resolution time series reveals cohesive but short-lived communities in coastal plankton

Because microbial plankton in the ocean comprise diverse bacteria, algae, and protists that are subject to environmental forcing on multiple spatial and temporal scales, a fundamental open question is to what extent these organisms form ecologically cohesive communities. Here we show that although all taxa undergo large, near daily fluctuations in abundance, microbial plankton are organized into clearly defined communities whose turnover is rapid and sharp. We analyze a time series of 93 consecutive days of coastal plankton using a technique that allows inference of communities as modular units of interacting taxa by determining positive and negative correlations at different temporal frequencies. This approach shows both coordinated population expansions that demarcate community boundaries and high frequency of positive and negative associations among populations within communities. Our analysis thus highlights that the environmental variability of the coastal ocean is mirrored in sharp transitions of defined but ephemeral communities of organisms.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Genomes to ecosystem function: Targeting critical knowledge gaps in methanogenesis and translation to updated global biogeochemical models

Natural freshwater temperate wetland systems currently represent the largest natural source of atmospheric methane, but are relatively understudied using systems biology tools (e.g. meta-omics) compared to other high producing methane systems (e.g. peat, tropical, or reconstructed wetlands). Using field investigations at the NOAA operated sentinel site on Lake Erie, methane producing activities and responses to geochemical conditions will be determined along seasonal and spatial gradients (Objective 1). Here, a combination of high-throughput activity and gas measurements, combined with high-resolution systems biology and analytical methods, will provide in depth knowledge of the microbiological, chemical, and physical constraints on methane production in wetlands. Using laboratory microcosms, the formation of anoxic microsites and their capacity to facilitate methane production in wetland soils will be simulated (Objective 2). This objective will validate the findings from the field investigations, offering a more controlled environment for teasing out the role of different yet interrelated variables. Lastly, these field and laboratory data will be used for multi-scale, process-level evaluation of an ecosystem biogeochemical model that accommodates these newly identified processes and parameterizes representation of these processes along relevant environmental gradients (Objective 3).This research will identify multiple interacting geochemical, ecological, and metabolic constraints that are poorly understood, oversimplified, or missing in global biogeochemical methane models. This proposal targets the role of oxygen limitation on methane processes in soil domains, to improve reactive transport models of microbial carbon cycling across terrestrial-aquatic soils and generate data on nutrient cycling activities in Great Lake wetlands. This information could provide new insights into the microbial controllers of Lake Erie eutrophication.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2018

Diversity of Active Viral Infections within the Sphagnum Microbiome

Sphagnum -dominated peatlands play an important role in global carbon storage and represent significant sources of economic and ecological value. While recent efforts to describe microbial diversity and metabolic potential of the Sphagnum microbiome have demonstrated the importance of its microbial community, little is known about the viral constituents. We used metatranscriptomics to describe the diversity and activity of viruses infecting microbes within the Sphagnum peat bog. The vegetative portions of six Sphagnum plants were obtained from a peatland in northern Minnesota, and the total RNA was extracted and sequenced. Metatranscriptomes were assembled and contigs were screened for the presence of conserved virus marker genes. Using bacteriophage capsid protein gp23 as a marker for phage diversity, we identified 33 contigs representing undocumented phages that were active in the community at the time of sampling. Similarly, RNA-dependent RNA polymerase and the nucleocytoplasmic large DNA virus (NCLDV) major capsid protein were used as markers for single-stranded RNA (ssRNA) viruses and NCLDV, respectively. In total, 114 contigs were identified as originating from undescribed ssRNA viruses, 22 of which represent nearly complete genomes. An additional 64 contigs were identified as being from NCLDVs. Finally, 7 contigs were identified as putative virophage or polinton-like viruses. We developed co-occurrence networks with these markers in relation to the expression of potential-host housekeeping gene rpb1 to predict virus-host relationships, identifying 13 groups. Together, our approach offers new tools for the identification of virus diversity and interactions in understudied clades and suggests that viruses may play a considerable role in the ecology of the Sphagnum microbiome. Sphagnum -dominated peatlands play an important role in maintaining atmospheric carbon dioxide levels by modifying conditions in the surrounding soil to favor the growth of over that of other plant species. This lowers the rate of decomposition and facilitates the accumulation of fixed carbon in the form of partially decomposed biomass. The unique environment produced by Sphagnum enriches for the growth of a diverse microbial consortia that benefit from and support the moss's growth, while also maintaining the hostile soil conditions. While a growing body of research has begun to characterize the microbial groups that colonize Sphagnum , little is currently known about the ecological factors that constrain community structure and define ecosystem function. Top-down population control by viruses is almost completely undescribed. Furthermore, this study provides insight into the significant viral influence on the Sphagnum microbiome and identifies new potential model systems to study virus-host interactions in the peatland ecosystem.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

Complex subsurface hydrothermal fluid mixing at a submarine arc volcano supports distinct and highly diverse microbial communities

Hydrothermally active submarine volcanoes are mineral-rich biological oases contributing significantly to chemical fluxes in the deep sea, yet little is known about the microbial communities inhabiting these systems. Here we investigate the diversity of microbial life in hydrothermal deposits and their metagenomics-inferred physiology in light of the geological history and resulting hydrothermal fluid paths in the subsurface of Brothers submarine volcano north of New Zealand on the southern Kermadec arc. From metagenome-assembled genomes we identified over 90 putative bacterial and archaeal genomic families and nearly 300 previously unknown genera, many potentially endemic to this submarine volcanic environment. While magmatically influenced hydrothermal systems on the volcanic resurgent cones of Brothers volcano harbor communities of thermoacidophiles and diverse members of the superphylum “DPANN,” two distinct communities are associated with the caldera wall, likely shaped by two different types of hydrothermal circulation. The communities whose phylogenetic diversity primarily aligns with that of the cone sites and magmatically influenced hydrothermal systems elsewhere are characterized predominately by anaerobic metabolisms. These populations are probably maintained by fluids with greater magmatic inputs that have interacted with different (deeper) previously altered mineral assemblages. However, proximal (a few meters distant) communities with gene-inferred aerobic, microaerophilic, and anaerobic metabolisms are likely supported by shallower seawater-dominated circulation. Furthermore, mixing of fluids from these two distinct hydrothermal circulation systems may have an underlying imprint on the high microbial phylogenomic diversity. Collectively our results highlight the importance of considering geologic evolution and history of subsurface processes in studying microbial colonization and community dynamics in volcanic environments.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2015

Integrated ‘omics analysis for studying the microbial community response to a pH perturbation of a cellulose-degrading bioreactor culture

Propionate accumulation is a common indicator of process imbalances in anaerobic bioreactor systems. The accumulation of propionate can occur due to low retention rates, hydrogen accumulation, or mechanical changes affecting the proximity between propionate oxidizers and partner species, thereby preventing necessary electron transfer. Few studies, however, have observed the changes in microbial community structure during propionate accumulation. We used 454 pyrosequencing of 16S rDNA to evaluate the community membership during propionate accumulations in replicate bioreactors with rumen based cultures. Half of the culture volume from a parent reactor was transferred to a sterile “daughter” reactor, and both systems were run identically. Both reactors experienced a propionate accumulation after roughly 10 days, with the propionate accumulation being less pronounced in the parent reactor as compared to the daughter reactor. Non-metric multidimensional scaling (NMDS) was used to determine clustering patterns of the samples, and correlative methods were used to determine which OTUs were significantly associated with the movements of samples along the NMDS axes. The presence of Saccharofermentans characterized the position of early samples, whereas the presence of Ruminococcus and Succiniclasticum were more indicative of the positions of later samples. Hydrogen accumulation and low sequence counts indicated low methanogen activity. Although both reactor systems were closed to microbial inputs due to the sterilization of influent media, we recorded significant increases in reactor diversity over time. This suggests that changes in the abundances of dominant community members may affect the sequencing of rare taxa within samples.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Environmental controls on observed spatial variability of soil pore water geochemistry in small headwater catchments underlain with permafrost

Abstract. Soil pore water (SPW) chemistry can vary substantially across multiple scales in Arctic permafrost landscapes. The magnitude of these variations and their relationship to scale are critical considerations for understanding current controls on geochemical cycling and for predicting future changes. These aspects are especially important for Arctic change modeling where accurate representation of sub-grid variability may be necessary to predict watershed-scale behaviors. Our research goal is to characterize intra- and inter-watershed soil water geochemical variations at two contrasting locations in the Seward Peninsula of Alaska, USA. We then attempt to identify the key factors controlling concentrations of important pore water solutes in these systems. The SPW geochemistry of 18 locations spanning two small Arctic catchments was examined for spatial variability and its dominant environmental controls. The primary environmental controls considered were vegetation, soil moisture and/or redox condition, water–soil interactions and hydrologic transport, and mineral solubility. The sampling locations varied in terms of vegetation type and canopy height, presence or absence of near-surface permafrost, soil moisture, and hillslope position. Vegetation was found to have a significant impact on SPW NO3- concentrations, associated with the localized presence of nitrogen-fixing alders and mineralization and nitrification of leaf litter from tall willow shrubs. The elevated NO3- concentrations were, however, frequently equipoised by increased microbial denitrification in regions with sufficient moisture to support it. Vegetation also had an observable impact on soil-moisture-sensitive constituents, but the effect was less significant. The redox conditions in both catchments were generally limited by Fe reduction, seemingly well-buffered by a cache of amorphous Fe hydroxides, with the most reducing conditions found at sampling locations with the highest soil moisture content. Non-redox-sensitive cations were affected by a wide variety of water–soil interactions that affect mineral solubility and transport. Identification of the dominant controls on current SPW hydrogeochemistry allows for qualitative prediction of future geochemical trends in small Arctic catchments that are likely to experience warming and permafrost thaw. As source areas for geochemical fluxes to the broader Arctic hydrologic system, geochemical processes occurring in these environments are particularly important to understand and predict with regards to such environmental changes.

Conroy, Nathan Alec (ORCID:0000000305973373)↗

DOE OSTI2019

Artificial reforestation produces less diverse soil nitrogen–cycling genes than natural restoration

Reforestation is effective in restoring ecosystem functions and enhancing ecosystem services of degraded land. The three most commonly employed reforestation methods of natural reforestation, artificial reforestation with native Masson pine, and introduced slash pine plantations were equally successful in biomass yield in southern China. However, it is not known whether soil ecosystem functions, such as nitrogen (N) cycling, are also successfully restored. Here, we employed a functional microarray to illustrate soil N-cycling. The composition of N-cycling genes in soils varied significantly with reforestation method and varied with constructive species identity and plant diversity. Artificial reforestation had less superior organization of N-cycling genes than natural reforestation, as indicated by the less diverse and less stable pathways to perform the biogeochemical N-cycling processes. Besides, artificial reforestation had lower functional potential (abundance of ammonification, denitrification, assimilatory, and dissimilatory nitrate reduction to ammonium genes) in soils than natural method. Evaluations of the abundance and interactions of N-cycling genes in soils showed that plantations, especially artificial reforestation with slash pine plantations, possessed a smaller range of ecosystem functions that provide a less diverse array of N-related substrates and nutrients to microbial communities compared with natural restoration. This might lead to a lower independence of N-cycling, which indicated a higher risk of N release in plantations. The unfavorable N-cycling conditions in plantations were corroborated by the lower contents of available N, ammonium N, and nitrate N. These findings demonstrate that reforestation methods could have broad regional and possibly global implications for N-cycling.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Landscape Topography and Regional Drought Alters Dust Microbiomes in the Sierra Nevada of California

Dust provides an ecologically significant input of nutrients, especially in slowly eroding ecosystems where chemical weathering intensity limits nutrient inputs from underlying bedrock. In addition to nutrient inputs, incoming dust is a vector for dispersing dust-associated microorganisms. While little is known about dust-microbial dispersal, dust deposits may have transformative effects on ecosystems far from where the dust was emitted. Using molecular analyses, we examined spatiotemporal variation in incoming dust microbiomes along an elevational gradient within the Sierra Nevada of California. We sampled throughout two dry seasons and found that dust microbiomes differed by elevation across two summer dry seasons (2014 and 2015), which corresponded to competing droughts in dust source areas. Dust microbial taxa richness decreased with elevation and was inversely proportional to dust heterogeneity. Likewise, dust phosphorus content increased with elevation. At lower elevations, early season dust microbiomes were more diverse than those found later in the year. The relative abundances of microbial groups shifted during the summer dry season. Furthermore, mutualistic fungal diversity increased with elevation, which may have corresponded with the biogeography of their plant hosts. Although dust fungal pathogen diversity was equivalent across elevations, elevation and sampling month interactions for the relative abundance, diversity, and richness of fungal pathogens suggest that these pathogens differed temporally across elevations, with potential implications for humans and wildlife. This study shows that landscape topography and droughts in source locations may alter the composition and diversity of ecologically relevant dust-associated microorganisms.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRSDate not supplied

Early Results from a Broad Compatibility Study of Various Materials with Ionic Silver Biocide

Ionic silver is baselined for microbial control in spacecraft potable water systems forfuture exploration missions, but materials compatibility analysis is required to evaluate thepassive depletion of ionic silver concentration onto wetted material surfaces over time.Various articles concerning such testing have been published that examine interactions withwater containing ionic silver biocide, but most tests have focused on only a couple ofmaterials each and comparing results of different evaluations to one another has provedchallenging.This paper reports the first results from static exposure testing of a large arrayof material coupons to a 400 parts per billion (ppb) aqueous silver fluoride (AgF) solution,using a surface to volume ratio of approximately 2 cm-1. The test is designed in two mainstages. Stage 1 is a one-week screening to evaluate silver uptake. Materials that performmodestly to well after that week are promoted to Stage 2, which is a longer test with periodicsampling to examine the silver uptake rates over time; these samples are evaluated for otherwater quality parameters in addition to the remaining silver concentration. In a tangentialinvestigation, select materials that take up some silver in Stage 1 may be “aged” by repeatingthe Stage 1 test to determine whether repeated exposure reduces silver uptake rate, andsuccessfully aged materials may then continue to Stage 2 testing. The materials under testinclude metallic and polymeric materials with various surface finishes, treatments, andcoatings, as well as select other materials historically used in spacecraft water systems. Thistest began in August 2019, and thus only includes early results; future follow-on papers willinclude additional results as the test progresses. The ultimate goal builds a broad, easilycomparable data set that can be used to guide material selections for silver biocide-compatible spacecraft water system design.

Silver Biocide↗

DOE OSTI2018

Impacts of Atmospheric Microbiomes Field Campaign Report

Aerosols of microbial origin (e.g., bacteria, fungi, etc.; also known as “atmospheric microbiome”) are ubiquitous in the atmosphere where they can remain suspended for several days to weeks and be transported over long ranges across continents. There is growing recognition that a significant fraction of total particulates suspended in the atmosphere may host microorganisms. Studies indicate that atmospheric microbial transport can play a significant role in processes associated with climate change and atmospheric chemistry. Furthermore, microbe-bearing particles are continually exchanged between ecosystems through global atmospheric transport pathways with the potential to influence the dispersion, composition, and dynamics of microbial communities across diverse ecosystems, including land managed, modified, and used by humans. The impacts of these atmospheric microbiomes on climate are poorly understood. The goal of this work is to establish fundamental new capabilities that enable prediction of atmospheric microbiome dynamics and interactions from source to final deposition. To achieve these objectives, surface soils (0-5 cm depth) were taken from within a 2.5-m radial area near the likely origin of the airborne particulates. In addition, we collected bulk and size-resolved airborne samples from a sampling line located ~4 m above the central U.S. Department of Energy (DOE) Atmospheric Radiation Measurement (ARM) Southern Great Plains (SGP) site from 25/10 to 31/10 in 2016. The SGP site is located in north-central Oklahoma and experiences a broad range of air mass, cloud, and atmospheric state disturbances. The site is surrounded by various agricultural crops and pasture fields. In total, 20 samples were collected over one week in October 2016, and each sample collection period was again approximately 12 hours. At both locations, the samples were collected on PTFE laminated membrane 0.45-μm filters (Sterlitech Corp.) mounted inside a 90-mm single-stage filter assembly (Savillex Inc.). The air was sampled from the sampling inlet located ~4 m height from the ground at total ~0.23 m3/min, which allowed us to sample total ~166 m3 of air per filter. The filter holder assembly and membrane filter holder tips were cleaned with isopropyl alcohol before a new filter was loaded. Blank filters were also collected to understand the background bacterial loading and composition but without operating the filter assembly. Both ambient air collected and blank filters were stored and shipped frozen at ~-20 °C to avoid the contamination and growth of unwanted bacterial communities. In parallel, we also collected the samples using a Micro-Orifice Uniform Deposit Impactor (MOUDI) for microscopy analysis.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2025

Predicting receptor-ligand pairing preferences in plant-microbe interfaces via molecular dynamics and machine learning

Microbiome assembly, structure, and dynamics significantly influence plant health. Secreted microbial signaling molecules initiate and mediate symbiosis by binding to structurally compatible plant receptors. For example, lipo-chitooligosaccharides (LCOs), produced by nitrogen-fixing rhizobial bacteria and various fungi, are recognized by plant lysin motif receptor-like kinases (LysM-RLKs), which activate the common symbiotic pathway. Accurately predicting these molecular interactions could reveal complementary signatures underlying the initial stages of endosymbiosis. Despite the breakthrough in protein-ligand structure prediction with deep learning-based tools, such as AlphaFold3, the large size and highly flexible nature of signaling compounds like LCOs present major challenges for detailed structural characterization and binding-affinity prediction. Typical structure-/physics-based methods of ligand virtual screening are designed for small, drug-like molecules, often rely on high-resolution, experimentally determined structures of the protein receptors, and rarely achieve sufficient sampling to obtain converged thermodynamic quantities with large ligands. In this study, we developed a hybrid molecular dynamics/machine learning (MD/ML) approach capable of predicting binding affinity rankings with high accuracy in systems involving large, flexible ligands, despite limited experimental structural information. Using coarse initial structural models, the predictions using the MD/ML workflow achieved strong alignment with experimental trends, particularly in the top-affinity tier for four legume LysM-RLKs (LYR3) binding to LCOs and a chitooligosaccharide. Furthermore, the MD-based conformation selection protocol provided critical structural insights into substrate specificity and binding mechanisms. This study demonstrates a powerful method to screen for challenging cognate ligand-receptors and advance our understanding of the molecular basis of microbial colonization in plants.

Lipo-chitooligosaccharides↗

DOE OSTI2022

Microbial functional diversity across biogeochemical provinces in the central Pacific Ocean

Enzymes catalyze key reactions within Earth’s life-sustaining biogeochemical cycles. Here, we use metaproteomics to examine the enzymatic capabilities of the microbial community (0.2 to 3 µm) along a 5,000-km-long, 1-km-deep transect in the central Pacific Ocean. Eighty-five percent of total protein abundance was of bacterial origin, with Archaea contributing 1.6%. Over 2,000 functional KEGG Ontology (KO) groups were identified, yet only 25 KO groups contributed over half of the protein abundance, simultaneously indicating abundant key functions and a long tail of diverse functions. Vertical attenuation of individual proteins displayed stratification of nutrient transport, carbon utilization, and environmental stress. The microbial community also varied along horizontal scales, shaped by environmental features specific to the oligotrophic North Pacific Subtropical Gyre, the oxygen-depleted Eastern Tropical North Pacific, and nutrient-rich equatorial upwelling. Some of the most abundant proteins were associated with nitrification and C1 metabolisms, with observed interactions between these pathways. The oxidoreductases nitrite oxidoreductase (NxrAB), nitrite reductase (NirK), ammonia monooxygenase (AmoABC), manganese oxidase (MnxG), formate dehydrogenase (FdoGH and FDH), and carbon monoxide dehydrogenase (CoxLM) displayed distributions indicative of biogeochemical status such as oxidative or nutritional stress, with the potential to be more sensitive than chemical sensors. Enzymes that mediate transformations of atmospheric gases like CO, CO 2 , NO, methanethiol, and methylamines were most abundant in the upwelling region. Here, we identified hot spots of biochemical transformation in the central Pacific Ocean, highlighted previously understudied metabolic pathways in the environment, and provided rich empirical data for biogeochemical models critical for forecasting ecosystem response to climate change.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2025

Climate warming enhances biodiversity and stability of grassland soil phosphorus-cycling microbial communities

Abstract Climate warming poses significant challenges to global phosphorus sustainability, an essential component of Earth biogeochemistry cycling and water-food-energy nexus. Despite the crucial role of polyphosphate-accumulating organism as key functional microbial agents in phosphorus cycling, the impacts of global climate warming on polyphosphate accumulating organism communities remain largely enigmatic. This study investigates the effects of climate warming on the taxonomic, network, and functional profiles of soil bacterial polyphosphate-accumulating organisms, leveraging fluorescence-activated cell sorting and single-cell Raman spectroscopy. Climate warming enhances both taxonomic and functional biodiversity of polyphosphate-accumulating organisms via biotic interactions and environmental filtering, with observed functionality-biodiversity relationships supporting the functional redundancy theory. Furthermore, polyphosphate-accumulating organism network complexity and stability rise under warming with strengthened positive relationships, supporting stress gradient hypothesis and the belief that complexity begets stability. Finally, polyphosphate-accumulating organisms are significantly correlated to key ecosystem functioning in carbon and phosphorus cycling under warming. Our study suggests that preserving polyphosphate-accumulating organism communities is crucial for maintaining soil ecosystem functioning and sustainable phosphorus management in a warming world and opens avenues for predicting the responses of other functional microbial groups to climate change, beneficially or maliciously.

Environmental Sciences & Ecology↗

DOE OSTI2022

Exploring the roles of microbes in facilitating plant adaptation to climate change

Plants benefit from their close association with soil microbes which assist in their response to abiotic and biotic stressors. Yet much of what we know about plant stress responses is based on studies where the microbial partners were uncontrolled and unknown. Under climate change, the soil microbial community will also be sensitive to and respond to abiotic and biotic stressors. Thus, facilitating plant adaptation to climate change will require a systems-based approach that accounts for the multi-dimensional nature of plant–microbe–environment interactions. In this perspective, we highlight some of the key factors influencing plant–microbe interactions under stress as well as new tools to facilitate the controlled study of their molecular complexity, such as fabricated ecosystems and synthetic communities. When paired with genomic and biochemical methods, these tools provide researchers with more precision, reproducibility, and manipulability for exploring plant–microbe–environment interactions under a changing climate.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Epistasis and evolution: recent advances and an outlook for prediction

As organisms evolve, the effects of mutations change as a result of epistatic interactions with other mutations accumulated along the line of descent. This can lead to shifts in adaptability or robustness that ultimately shape subsequent evolution. Here, we review recent advances in measuring, modeling, and predicting epistasis along evolutionary trajectories, both in microbial cells and single proteins. We focus on simple patterns of global epistasis that emerge in this data, in which the effects of mutations can be predicted by a small number of variables. The emergence of these patterns offers promise for efforts to model epistasis and predict evolution.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Engineering Terpene Production Pathways in Methylobacterium extorquens AM1

Terpenes are diverse specialized metabolites naturally found within plants and have important roles in inter-species communication, adaptation and interaction with the environment. Their industrial applications span a broad range, including fragrances, flavors, cosmetics, natural colorants to agrochemicals and therapeutics, yet formal chemical synthesis is economically challenging due to structural complexities. Engineering terpene biosynthesis could represent an alternative in microbial biotechnological workhorses, such as Saccharomyces cerevisiae or Escherichi coli, utilizing sugars or complex media as feedstocks. Host species that metabolize renewable and affordable carbon sources may offer unique sustainable biotechnological alternatives. Methylotrophs are bacteria with the capacity to utilize one-carbon feedstocks, such as methanol or formate. They colonize the phyllosphere (above-ground area) of plants, and many accumulate abundant carotenoid pigments. Methylotrophs have the capacity to take up and use a subset of the rare earth elements known as lanthanides. These metals can enhance one-carbon (methylotrophic) metabolism. Here, we investigated whether manipulating the metabolism enables and enhances terpene production. A carotenoid-deficient mutant potentially liberates carbon, which may contribute to bioproduct accumulation. To test this hypothesis, terpene-producing bacterial strains regulated by two distinct promoters were generated. Wildtype Methylobacterium extorquens, ∆Meta1_3665, a methylotrophic mutant lacking the carotenoid pathway, and an E. coli strain were transformed with an exogenous terpene pathway and grown both in the presence and absence of lanthanides. The extraction, and the comparison of analytical profiles, provided evidence that engineered cultured M. extorquens under control of a native, inducible methylotrophic promoter can yield the sesquiterpene patchoulol when supplemented with lanthanide. In contrast, using a moderate-strength constitutive promoter failed to give production. We demonstrated colonization of the phyllosphere with the engineered strains, supporting the future engineering of selected species of the plant microbiome and with promising implications for the synthetic biology of small molecules.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS2001

The Role of Cyanobacteria in Stromatolite Morphogenesis, Highborn Cay Bahamas: An Integrated Field and Laboratory Simulation Study

Geomicrobiological phenomena are among the most fundamental of interactions between Earth and its biosphere. Actively growing and lithifying stromatolites at Highborne Cay Bahamas, have recently been documented and allow for detailed examination of the roles microbes play in the mineralization process. These stromatolites contain a variety of complex microbial communities with distinct distribution patterns for different microbial groups. Cyanobacteria are the primary producers in this system providing energy, directly or indirectly, for the entire stromatolite microbial community. They also play key roles in the trapping and binding of sediments. Most of these species are highly motile and can adjust their position and orientation within the sediment matrix in order to optimize their access to irradiance and nutrients. As individual species have different physical and metabolic properties, this motility generally results in segregated distributions of species, which in turn contributes to the laminated textures observed in these actively forming stromatolites. Increasingly our studies suggest that the activities and locations of various cyanobacterial species also contribute greatly to the localization of new mineral precipitation through a variety of processes. We are investigating these contributions using an integrated approach combining detailed observations of field samples with manipulative experiments using both field samples and cultures of specific organisms isolated from these stromatolites. Experiments are conducted both in standard laboratory conditions and in outdoor running seawater flumes. A variety of standard techniques; SEM (scanning electron microscopy), petrographic analyses, TEM (transmission electron microscopy), are used to compare mineralization processes in field samples with those generated in laboratory-flume simulations. Using this approach we are able to more thoroughly investigate the effects of irradiance, CaCO3 saturation, and hydrodynamic regime on cyanobacterial distribution, trapping and binding and mineral precipitation. Simulation results will be presented and compared with community and mineralization distribution patterns seen in the field samples from which these communities were isolated.

Prufert-Bebout, Leslie↗

DOE OSTI2015

Site-specific programming of the host epithelial transcriptome by the gut microbiota

The intestinal epithelium separates us from the microbiota but also interacts with it and thus affects host immune status and physiology. Previous studies investigated microbiota-induced responses in the gut using intact tissues or unfractionated epithelial cells, thereby limiting conclusions about regional differences in the epithelium. Here, we sought to investigate microbiota-induced transcriptional responses in specific fractions of intestinal epithelial cells. To this end, we used microarray analysis of laser capture microdissection (LCM)-harvested ileal and colonic tip and crypt epithelial fractions from germ-free and conventionally raised mice and from mice during the time course of colonization. We found that about 10% of the host’s transcriptome was microbially regulated, mainly including genes annotated with functions in immunity, cell proliferation, and metabolism. The microbial impact on host gene expression was highly site specific, as epithelial responses to the microbiota differed between cell fractions. Specific transcriptional regulators were enriched in each fraction. In general, the gut microbiota induced a more rapid response in the colon than in the ileum. Our study indicates that the microbiota engage different regulatory networks to alter host gene expression in a particular niche. Understanding host-microbiota interactions on a cellular level may facilitate signaling pathways that contribute to health and disease and thus provide new therapeutic strategies.

59 BASIC BIOLOGICAL SCIENCES↗