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DOE OSTI2022

Variation in Root Exudate Composition Influences Soil Microbiome Membership and Function

Root exudation is one of the primary processes that mediate interactions between plant roots, microorganisms, and the soil matrix, yet the mechanisms by which exudation alters microbial metabolism in soils have been challenging to unravel. Here, utilizing distinct sorghum genotypes, we characterized the chemical heterogeneity between root exudates and the effects of that variability on soil microbial membership and metabolism. Distinct exudate chemical profiles were quantified and used to formulate synthetic root exudate treatments: a high-organic-acid treatment (HOT) and a high-sugar treatment (HST). To parse the response of the soil microbiome to different exudate regimens, laboratory soil reactors were amended with these root exudate treatments as well as a nonexudate control. Amplicon sequencing of the 16S rRNA gene illustrated distinct microbial diversity patterns and membership in response to HST, HOT, or control amendments. Exometabolite changes reflected these microbial community changes, and we observed enrichment of organic and amino acids, as well as possible phytohormones in the HST relative to the HOT and control. Linking the metabolic capacity of metagenome-assembled genomes in the HST to the exometabolite patterns, we identified microorganisms that could produce these phytohormones. Our findings emphasize the tractability of high-resolution multiomics tools to investigate soil microbiomes, opening the possibility of manipulating native microbial communities to improve specific soil microbial functions and enhance crop production.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Terpenes modulate bacterial and fungal growth and sorghum rhizobiome communities

ABSTRACT Terpenes are among the oldest and largest class of plant-specialized bioproducts that are known to affect plant development, adaptation, and biological interactions. While their biosynthesis, evolution, and function in aboveground interactions with insects and individual microbial species are well studied, how different terpenes impact plant microbiomes belowground is much less understood. Here we designed an experiment to assess how belowground exogenous applications of monoterpenes (1,8-cineole and linalool) and a sesquiterpene (nerolidol) delivered through an artificial root system impacted its belowground bacterial and fungal microbiome. We found that the terpene applications had significant and variable impacts on bacterial and fungal communities, depending on terpene class and concentration; however, these impacts were localized to the artificial root system and the fungal rhizosphere. We complemented this experiment with pure culture bioassays on responsive bacteria and fungi isolated from the sorghum rhizobiome. Overall, higher concentrations (200 µM) of nerolidol were inhibitory to Ferrovibrium and tested Firmicutes. While fungal isolates of Penicillium and Periconia were also more inhibited by higher concentrations (200 µM) of nerolidol, Clonostachys was enhanced at this higher level and together with Humicola was inhibited by the lower concentration tested (100 µM). On the other hand, 1,8-cineole had an inhibitory effect on Orbilia at both tested concentrations but had a promotive effect at 100 µM on Penicillium and Periconia . Similarly, linalool at 100 µM had significant growth promotion in Mortierella , but an inhibitory effect for Orbilia . Together, these results highlight the variable direct effects of terpenes on single microbial isolates and demonstrate the complexity of microbe-terpene interactions in the rhizobiome. Importance Terpenes represent one of the largest and oldest classes of plant-specialized metabolism, but their role in the belowground microbiome is poorly understood. Here, we used a “rhizobox” mesocosm experimental set-up to supply different concentrations and classes of terpenes into the soil compartment with growing sorghum for 1 month to assess how these terpenes affect sorghum bacterial and fungal rhizobiome communities. Changes in bacterial and fungal communities between treatments belowground were characterized, followed by bioassays screening on bacterial and fungal isolates from the sorghum rhizosphere against terpenes to validate direct microbial responses. We found that microbial growth stimulatory and inhibitory effects were localized, terpene specific, dose dependent, and transient in time. This work paves the way for engineering terpene metabolisms in plant microbiomes for improved sustainable agriculture and bioenergy crop production.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Structural Diversity in Eukaryotic Photosynthetic Light Harvesting

Photosynthesis has been using energy from sunlight to assimilate atmospheric CO 2 for at least 3.5 billion years. Through evolution and natural selection, photosynthetic organisms have flourished in almost all aquatic and terrestrial environments. This is partly due to the diversity of light-harvesting complex (LHC) proteins, which facilitate photosystem assembly, efficient excitation energy transfer, and photoprotection. Structural advances have provided angstrom-level structures of many of these proteins and have expanded our understanding of the pigments, lipids, and residues that drive LHC function. In this review, we compare and contrast recently observed cryo-electron microscopy structures across photosynthetic eukaryotes to identify structural motifs that underlie various light-harvesting strategies. We discuss subtle monomer changes that result in macroscale reorganization of LHC oligomers. Additionally, we find recurring patterns across diverse LHCs that may serve as evolutionary stepping stones for functional diversification. Advancing our understanding of LHC protein–environment interactions will improve our capacity to engineer more productive crops.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2020

Chlorophyll fluorescence imaging captures photochemical efficiency of grain sorghum (Sorghum bicolor) in a field setting

Abstract Background Photosynthesis is one of the most important biological reactions and forms the basis of crop productivity and yield on which a growing global population relies. However, to develop improved plant cultivars that are capable of increased productivity, methods that can accurately and quickly quantify photosynthetic efficiency in large numbers of genotypes under field conditions are needed. Chlorophyll fluorescence imaging is a rapid, non-destructive measurement that can provide insight into the efficiency of the light-dependent reactions of photosynthesis. Results To test and validate a field-deployed fluorescence imaging system on the TERRA-REF field scanalyzer, leaves of potted sorghum plants were treated with a photosystem II inhibitor, DCMU, to reduce photochemical efficiency (F V /F M ). The ability of the fluorescence imaging system to detect changes in fluorescence was determined by comparing the image-derived values with a handheld fluorometer. This study demonstrated that the imaging system was able to accurately measure photochemical efficiency (F V /F M ) and was highly correlated ( r = 0.92) with the handheld fluorometer values. Additionally, the fluorescence imaging system was able to track the decrease in photochemical efficiency due to treatment of DCMU over a 7 day period. Conclusions The system’s ability to capture the temporal dynamics of the plants’ response to this induced stress, which has comparable dynamics to abiotic and biotic stressors found in field environments, indicates the system is operating correctly. With the validation of the fluorescence imaging system, physiological and genetic studies can be undertaken that leverage the fluorescence imaging capabilities and throughput of the field scanalyzer.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

DOE OSTI2022

Data for Responsiveness of Miscanthus and Switchgrass Yields to Stand Age and Nitrogen Fertilization: A Meta-regression Analysis

The compiled datasets include plot level observations of energy crops (miscanthus and switchgrass) from recent experimental field trials in the US including dry biomass yield, location, state, region, harvest year, growing season degree days (GDD), winter season heating degree days (HDD), growing season cumulative precipitation, annual nitrogen application rate, age of the pant when harvested, National Commodity Crop Productivity Index (NCCPI) values, and cultivar type (switchgrass) from various published and unpublished sources. The stata codes include estimation procedures for four different specifications, i.e., Model A includes deterministic effect without interaction terms; Model B includes deterministic effect with interaction terms (N2, age2, N × age, GDD2, precip2, N × NCCPI); Model C includes deterministic effect with interaction terms, study, and location random effect; Model D includes deterministic effect with interaction terms, harvest year augmented study, and location random effect.

Age↗

DOE OSTI2024

Biochar-compost-based controlled-release nitrogen fertilizer intended for an active microbial community

Nitrogen (N) fertilizers in agriculture suffer losses by volatilization of N to the air, surface runoff and leaching into the soil, resulting in low N use efficiency (NUE) (< 50%) and raising severe environmental pollutions. Controlled- release nitrogen fertilizers (CRNFs) can control the release of N nutrients to NUE in crop production. Different methods were used to develop new CRNFs. However, different CRNF technologies are still underdeveloped due to inadequate controlling on N releasing time and/or unsustainable diffusion. The study on the influences of CRNF processing parameters on microbial conditions are lacking when the CRNFs composed of various bio-ingredients such as biochar, composts, and biowaste. The complexity of processing methods, material biodegradability, and other physical properties make current CRNFs of questionable value in agricultural production. This research aims to develop a novel biochar-compost-based controlled-release urea fertilizer (BCRUF) to preserve microbial properties carried by the compost. The BCRUF was synthesized by pelletizing the 50:50 (dry, wt/wt) mixture of biochar and compost. BCRUF was loaded with urea and then spray-coated with polylactic acid (PLA). The releasing time of two types of BCRUFs, coated and uncoated with PLA, for 80% of N release in water was up to 6 h at three different temperatures (4, 23, and 40 °C), compared to conventional urea fertilizer and commercial environmentally smart N (ESN) fertilizer. The releasing time of coated BCRUF for 80% N release in soil was up to 192 h (8 days). Fourier-transform infrared spectroscopy (FTIR) analysis revealed that no new functional groups were found in the release solution, indicating no new chemical hazards generated. The differential scanning calorimetry (DSC) tests also verified that its thermal stability could be up to 160 °C. The microbe populations in the BCRUF pellets were reduced after the pelleting and drying processes in BCRUF fabrication, but a few bacteria can endure in the air-drying process. BCRUF pellets soaked in water for 4 days retained some bacteria. The BCRUF showed very promising characteristics to improve NUE and sustainability in agricultural production.

58 GEOSCIENCES↗

DOE OSTI2022

AmeriFlux US-Mo1 LTAR CMRB Field 1 (CMRB ASP)

This is the AmeriFlux version of the carbon flux data for the site US-Mo1 LTAR CMRB Field 1 (CMRB ASP). Site Description - Site is in a 36 ha agricultural research field near Centralia, MO on a broad, slightly sloped landscape. Currently farmed in a corn-soybean-wheat-hay rotation. Used for research since 1991; prior to that was managed for commercial crop production.

Schreiner-McGraw, Adam↗

DOE OSTI2022

AmeriFlux US-Mo3 LTAR CMRB Field 3 (CMRB BAU)

This is the AmeriFlux version of the carbon flux data for the site US-Mo3 LTAR CMRB Field 3 (CMRB BAU). Site Description - Site is in a 20 ha agricultural field near Centralia, MO on a broad, slightly sloped landscape. Currently managed in a corn-soybean rotation for commercial crop production

Schreiner-McGraw, Adam↗

DOE OSTI2023

AmeriFlux FLUXNET-1F US-Mo1 LTAR CMRB Field 1 (CMRB ASP)

This is the AmeriFlux Management Project (AMP) created FLUXNET-1F version of the carbon flux data for the site US-Mo1 LTAR CMRB Field 1 (CMRB ASP). This is the FLUXNET version of the carbon flux data for the site US-Mo1 LTAR CMRB Field 1 (CMRB ASP) produced by applying the standard ONEFlux (1F) software. Site Description - Site is in a 36 ha agricultural research field near Centralia, MO on a broad, slightly sloped landscape. Currently farmed in a corn-soybean-wheat-hay rotation. Used for research since 1991; prior to that was managed for commercial crop production.

Schreiner-McGraw, Adam↗

DOE OSTI2023

AmeriFlux FLUXNET-1F US-Mo3 LTAR CMRB Field 3 (CMRB BAU)

This is the AmeriFlux Management Project (AMP) created FLUXNET-1F version of the carbon flux data for the site US-Mo3 LTAR CMRB Field 3 (CMRB BAU). This is the FLUXNET version of the carbon flux data for the site US-Mo3 LTAR CMRB Field 3 (CMRB BAU) produced by applying the standard ONEFlux (1F) software. Site Description - Site is in a 20 ha agricultural field near Centralia, MO on a broad, slightly sloped landscape. Currently managed in a corn-soybean rotation for commercial crop production

Schreiner-McGraw, Adam↗

DOE OSTI2020

The Brachypodium ENCODE Project – from sequence to function: Predicting physiological responses in grasses to facilitate engineering of biofuel crops

The overarching goal of this proposal was to gain deep knowledge of chromatin dynamics and gene networks related to stress responses in grasses. To accomplish this goal our Brachypodium ENCODE project (1) coupled drought-focused physiological analysis with multiple layers of ENCODE ‘omics’ profiling; (2) obtained fundamental information about the Brachypodium transcriptome, epigenome, chromatin dynamics, and gene regulatory networks; and (3) elucidated the regulatory dynamics of gene networks that can be manipulated to increase energy grass crop productivity.

09 BIOMASS FUELS↗

DOE OSTI2021

Plant Synthetic Biology Conference – Travel Awardees and Speakers

The overarching goals of the Plant Synthetic Biology (SynBio) Conference, convened by the American Society of Plant Biologists (ASPB), were to raise community consciousness of what SynBio is and what it can do in key areas, including crop productivity, specialty chemicals, biosensors, and microbe-plant interactions. Within that context, the conference had two broad objectives. First, to feature a set of leading university and industry speakers from the US and Europe whose research on plants and microbes spans the SynBio spectrum with respect both to technology and to applications, as well as short talks (selected from submitted abstracts) on cutting-edge topics. And second, to foster exchanges among public sector scientists and industry scientists, whose participation as poster presenters, exhibitors, and sponsors was actively encouraged because so much innovative SynBio work is being carried out at dynamic young companies.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2022

Step Catalysis to Synthesize Fossil-Free Ammonia at Atmospheric Pressure (Final Technical Report)

Affordable fossil-free ammonia synthesis will enable high-yield sustainable crop production for a growing world population while preserving the planet and making the best use of water and land. Simple technology amenable to intermittent operation using renewable energy will be made available through step catalysis activating dinitrogen at atmospheric pressure by nitride formation. Project objectives: (1) At the single particle level, quantify and analyze N2 activation by doped or alloyed manganese macroscopic- and nano particles at atmospheric pressure, (2) Quantify and analyze ammonia synthesis from doped or alloyed metal nitride particles when contacting them with hydrogen at atmospheric pressure, and (3) Integrate experimental data and atomic-level computational models to predict the behavior of metal alloys in nitrogen activation and subsequent ammonia synthesis.

10 SYNTHETIC FUELS↗

DOE OSTI2023

BTRIC Technical Support for Appalachia: FY 2023 Summary Report on Net Negative Carbon Building Demonstration with AgPod

This study investigates the effects of various concentrations of carbon dioxide (CO 2 ) on the photosynthesis and biomass accumulation of crops growing in an Agricultural Pod (AgPod). The initial test crop was kale. It has been found that increasing the CO 2 concentration in the AgPod promotes higher photosynthetic rates in kale plants up to 1300 ppm (highest CO 2 concentration). An interesting finding was that net photosynthetic rates of kale leaves remain unsaturated until unusually high levels of photosynthetically active radiation (PAR, > 1200 µmolm-2s-1) and CO 2 concentration (> 1100 ppm) are reached. Such high levels of saturating PAR and CO 2 concentration values rarely occur under natural conditions, suggesting a promising potential for carbon capture via enriched greenhouse crop production. This task is one part of an overall plan to incorporate an AgPod to utilize carbon captured from the atmosphere to achieve net negative carbon emissions in a small neighborhood or community.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2024

Conserved genetic mechanisms for biotic stress in sorghum (Final report)

Developing durable disease resistance for biofuel crops is crucial, particularly as the range of biofuel crop production expands and pathogens of other plant species evolve to cause diseases of bioenergy feedstocks. Setosphaeria species are significant pathogens of the Andropogonae, and S. turcica can infect both maize and sorghum. Sorghum leaf blight (SLB), caused by S. turcica, is widespread and can decrease grain yields up to 50%, reduce forage quantity and quality, and predispose plants to other diseases, such as anthracnose. Our overall objective was to gain a systems-level understanding of the pathosystem by leveraging natural genetic variation, host specificity of the pathogen, and transcriptome analysis to improve biotic stress resistance in sorghum. We proposed three specific goals: 1) Identify sorghum alleles conferring resistance to S. turcica using natural genetic variation. 2) Characterize genes associated with biotic stress response in sorghum. 3) Identify fungal genes responsible for host specificity.

09 BIOMASS FUELS↗

DOE OSTI2024

Conserved genetic mechanisms for biotic stress in sorghum (Final Report)

09 BIOMASS FUELS↗

DOE OSTI2025

Zero-Power Wireless Infrared Digitizing Sensors for Large Scale Energy-Smart Farm

This project, funded by ARPA-E and led by Northeastern University, developed zeropower infrared digitizing sensors to optimize irrigation and enhance crop yields. Traditional water stress detection methods are costly and require frequent maintenance, limiting their effectiveness. Our research identified shortwave infrared (SWIR) transmittance as the most reliable indicator of plant water stress and developed plasmonically enhanced micromechanical photoswitches (PMPs) that operate with minimal power. The sensors offer low-cost, large-scale deployment, auto-calibration across different crops, and a 10-year battery life, significantly reducing maintenance costs. The system achieved 4x greater accuracy than conventional soil moisture sensors while ensuring economic feasibility. By enabling precision irrigation, this technology conserves water, enhances crop productivity, and lowers operational costs, making it a scalable solution for sustainable agriculture and global food security.

32 ENERGY CONSERVATION, CONSUMPTION, AND UTILIZATI↗

DOE OSTI2023

Thioredoxin VdTrx1, an unconventional secreted protein, is a virulence factor in Verticillium dahliae

Understanding how plant pathogenic fungi adapt to their hosts is of critical importance to securing optimal crop productivity. In response to pathogenic attack, plants produce reactive oxygen species (ROS) as part of a multipronged defense response. Pathogens, in turn, have evolved ROS scavenging mechanisms to undermine host defense. Thioredoxins (Trx) are highly conserved oxidoreductase enzymes with a dithiol-disulfide active site, and function as antioxidants to protect cells against free radicals, such as ROS. However, the roles of thioredoxins in Verticillium dahliae , an important vascular pathogen, are not clear. Through proteomics analyses, we identified a putative thioredoxin (VdTrx1) lacking a signal peptide. VdTrx1 was present in the exoproteome of V. dahliae cultured in the presence of host tissues, a finding that suggested that it plays a role in host-pathogen interactions. We constructed a VdTrx1 deletion mutant Δ VdTrx1 that exhibited significantly higher sensitivity to ROS stress, H 2 O 2 , and tert -butyl hydroperoxide ( t -BOOH). In vivo assays by live-cell imaging and in vitro assays by western blotting revealed that while VdTrx1 lacking the signal peptide can be localized within V. dahliae cells, VdTrx1 can also be secreted unconventionally depending on VdVps36, a member of the ESCRT-II protein complex. The Δ VdTrx1 strain was unable to scavenge host-generated extracellular ROS fully during host invasion. Deletion of VdTrx1 resulted in higher intracellular ROS levels of V. dahliae mycelium, displayed impaired conidial production, and showed significantly reduced virulence on Gossypium hirsutum , and model plants, Arabidopsis thaliana and Nicotiana benthamiana . Thus, we conclude that VdTrx1 acts as a virulence factor in V. dahliae .

Tian, Li↗