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At least 343 records · Page 19

Cellulolytic enzyme-aided extraction of hemicellulose from switchgrass and its characteristics

The isolation of biomass components with their intact structure is integral for understanding the characteristics of biomass. Compared to other major biomass components such as cellulose and lignin, hemicellulose remains a challenging component to be isolated from the plant cell wall without significant depolymerization and modification. Here, a novel cellulolytic enzyme-aided hemicellulose (CEH) isolation method was developed to isolate hemicellulose with a near-native branched structure from switchgrass. The structural characteristics of CEH were investigated and compared with hemicelluloses isolated by conventional alkaline extracted hemicellulose (AEH) and DMSO extracted hemicellulose (DMSOH) methods. Gel permeation chromatography (GPC) analysis indicated that CEH had a weight-average molecular weight of 44 kDa, which was comparable to that of AEH (43 kDa) but higher than that of DMSOH (37 kDa). The chemical composition analysis revealed that CEH retained a higher proportion of glucuronic acid compared to AEH and DMSOH. The 2D 13 C– 1 H heteronuclear single quantum coherence (HSQC) NMR spectra containing the β-(1,4)-linked-D-xylan backbone, non-reducing-end peaks and both α- and β-reducing-end peaks in CEH were comparable to the spectra of the commercial beechwood xylan. CEH showed a highly branched hemicellulose structure, which retained methoxyl groups, O -acetyl groups, and 4- O -methyl-glucuronic acid attached to the xylan backbone.

59 BASIC BIOLOGICAL SCIENCES↗

QTL × environment interactions underlie adaptive divergence in switchgrass across a large latitudinal gradient

Local adaptation is the process by which natural selection drives adaptive phenotypic divergence across environmental gradients. Theory suggests that local adaptation results from genetic trade-offs at individual genetic loci, where adaptation to one set of environmental conditions results in a cost to fitness in alternative environments. However, the degree to which there are costs associated with local adaptation is poorly understood because most of these experiments rely on two-site reciprocal transplant experiments. Here, we quantify the benefits and costs of locally adaptive loci across 17° of latitude in a four-grandparent outbred mapping population in outcrossing switchgrass ( Panicum virgatum L.), an emerging biofuel crop and dominant tallgrass species. We conducted quantitative trait locus (QTL) mapping across 10 sites, ranging from Texas to South Dakota. This analysis revealed that beneficial biomass (fitness) QTL generally incur minimal costs when transplanted to other field sites distributed over a large climatic gradient over the 2 y of our study. Therefore, locally advantageous alleles could potentially be combined across multiple loci through breeding to create high-yielding regionally adapted cultivars.

59 BASIC BIOLOGICAL SCIENCES↗

Promises and challenges of eco-physiological genomics in the field: tests of drought responses in switchgrass

Identifying the physiological and genetic basis of stress tolerance in plants has proven to be critical to understanding adaptation in both agricultural and natural systems. However, many discoveries were initially made in the controlled conditions of greenhouses or laboratories, not in the field. To test the comparability of drought responses across field and greenhouse environments, we undertook three independent experiments using the switchgrass reference genotype Alamo AP13. We analyzed physiological and gene expression variation across four locations, two sampling times, and three years. Relatively similar physiological responses and expression coefficients of variation across experiments masked highly dissimilar gene expression responses to drought. Critically, a drought experiment utilizing small pots in the greenhouse elicited nearly identical physiological changes as an experiment conducted in the field, but an order of magnitude more differentially expressed genes. However, we were able to define a suite of several hundred genes that were differentially expressed across all experiments. This list was strongly enriched in photosynthesis, water status, and reactive oxygen species responsive genes. The strong across-experiment correlations between physiological plasticity—but not differential gene expression—highlight the complex and diverse genetic mechanisms that can produce phenotypically similar responses to various soil water deficits.

54 ENVIRONMENTAL SCIENCES↗

Evaluation of eastern gamagrass as dual-purpose complementary bioenergy and forage feedstock to switchgrass

Switchgrass (SG) is considered a model bioenergy crop and a warm-season perennial grass (WSPG) that traditionally served as forage feedstock in the United States. To avoid the sole dependence on SG for bioenergy production, evaluation of other crops to diversify the pool of feedstock is needed. We conducted a 3-year field experiment evaluating eastern gamagrass (GG), another WSPG, as complementary feedstock to SG in one- and two-cut systems, with or without intercropping with crimson clover or hairy vetch, and under different nitrogen (N) application rates. Our results showed that GG generally produced lower biomass (by 29.5%), theoretical ethanol potential (TEP, by 2.8%), and theoretical ethanol yield (TEY, by 32.9%) than corresponding SG under the same conditions. However, forage quality measures, namely acid detergent fiber (ADF), crude protein (CP), and elements P, K, Ca, and Mg were significantly higher in GG than those in SG. Nitrogen fertilizer significantly enhanced biomass (by 1.54 Mg ha -1 ), lignin content (by 2.10 g kg -1 ), and TEY (787.12 L ha -1 ) in the WSPGs compared to unfertilized treatments. Intercropping with crimson clover or hairy vetch did not significantly increase biomass of the WSPGs, or TEP and TEY in unfertilized plots. This study demonstrated that GG can serve as a complementary crop to SG and could be used as a dual-purpose crop for bioenergy and forage feedstock in farmers' rotations.

09 BIOMASS FUELS↗

Uncertainties in greenhouse gas emission factors: A comprehensive analysis of switchgrass‐based biofuel production

Abstract This study investigates uncertainties in greenhouse gas (GHG) emission factors related to switchgrass‐based biofuel production in Michigan. Using three life cycle assessment (LCA) databases—US lifecycle inventory (USLCI) database, GREET, and Ecoinvent—each with multiple versions, we recalculated the global warming intensity (GWI) and GHG mitigation potential in a static calculation. Employing Monte Carlo simulations along with local and global sensitivity analyses, we assess uncertainties and pinpoint key parameters influencing GWI. The convergence of results across our previous study, static calculations, and Monte Carlo simulations enhances the credibility of estimated GWI values. Static calculations, validated by Monte Carlo simulations, offer reasonable central tendencies, providing a robust foundation for policy considerations. However, the wider range observed in Monte Carlo simulations underscores the importance of potential variations and uncertainties in real‐world applications. Sensitivity analyses identify biofuel yield, GHG emissions of electricity, and soil organic carbon (SOC) change as pivotal parameters influencing GWI. Decreasing uncertainties in GWI may be achieved by making greater efforts to acquire more precise data on these parameters. Our study emphasizes the significance of considering diverse GHG factors and databases in GWI assessments and stresses the need for accurate electricity fuel mixes, crucial information for refining GWI assessments and informing strategies for sustainable biofuel production.

Kim, Seungdo↗

Corn and Switchgrass Derived Sustainable Aviation Fuel Carbon Intensity

The production and use of sustainable aviation fuel (SAF) could lower aviation's carbon footprint. Feedstock agricultural management can optimize—or potentially determine—SAF sustainability and profitability, but long-term SAF feedstock comparisons are limited. Life cycle assessments (LCA) are commonly used to estimate the carbon intensity of producing SAF from emerging feedstocks due to the limited availability of long-term, primary field data. We quantified the agricultural phase and well-to-wake (WTWa) carbon intensities of producing SAF from no-till, continuous corn (Zea mays L.) under 120 kg N ha−1 year−1 (120 N corn) and switchgrass (Panicum virgatum L.) under 60 or 120 kg N ha−1 year−1 (60 N and 120 N SWG, respectively) via LCAs based on measured data from a long-term (1998-present) field experiment on marginally productive cropland located in Eastern NE, USA. Measured data included crop productivity, long-term changes in soil organic carbon (SOC) stocks (0–150 cm), annual soil N2O emissions, and field management practices. Both 120 N corn (2078 L ha−1) and 120 N SWG (1752 L ha−1) produced more SAF than 60 N SWG (1229 L ha−1) (p < 0.0001). However, making SAF from corn had greater life cycle GHG emissions (i.e., WTWa carbon intensity) than making SAF from 120 N and 60 N SWG (61.5 > −14.5 and −27.5 g CO2e MJ−1 SAF, respectively; p = 0.0007), primarily due to SOC accrual during crop production. Here, we demonstrate that agricultural management (e.g., crop type, fertilizer use) can determine the sustainably and productivity of producing SAF by using long-term, measured field data in LCAs.

carbon intensity↗

SPL 7 and SPL 8 represent a novel flowering regulation mechanism in switchgrass

Summary The aging pathway in flowering regulation is controlled mainly by micro RNA 156 (miR156). Studies in Arabidopsis thaliana reveal that nine miR156‐targeted SQUAMOSA PROMOTER BINDING ‐ LIKE ( SPL ) genes are involved in the control of flowering. However, the roles of SPL s in flowering remain elusive in grasses. Inflorescence development in switchgrass was characterized using scanning electron microscopy ( SEM ). Microarray, quantitative reverse transcription polymerase chain reaction (q RT ‐ PCR ), chromatin immunoprecipitation (Ch IP )‐ PCR and EMSA were used to identify regulators of phase transition and flowering. Gene function was characterized by downregulation and overexpression of the target genes. Overexpression of SPL 7 and SPL 8 promotes flowering, whereas downregulation of individual genes moderately delays flowering. Simultaneous downregulation of SPL 7 / SPL 8 results in extremely delayed or nonflowering plants. Furthermore, downregulation of both genes leads to a vegetative‐to‐reproductive reversion in the inflorescence, a phenomenon that has not been reported in any other grasses. Detailed analyses demonstrate that SPL 7 and SPL 8 induce phase transition and flowering in grasses by directly upregulating SEPALLATA3 ( SEP 3 ) and MADS 32 . Thus, the SPL 7 / 8 pathway represents a novel regulatory mechanism in grasses that is largely different from that in Arabidopsis . Additionally, genetic modification of SPL 7 and SPL8 results in much taller plants with significantly increased biomass yield and sugar release.

Gou, Jiqing↗

Transgenic switchgrass ( Panicum virgatum L.) targeted for reduced recalcitrance to bioconversion: A two-year comparative analysis of field-grown lines modified for target gene or genetic element expression

Five different types of transgenic ( GAUT4 , miRNA, MYB4, COMT and FPGS ) Panicum virgatum L. (switchgrass) were grown in a field in Knoxville, Tenn., USA over two consecutive years between 2011 and 2015 in separate experiments. Clonal replicates were established (year-one) and produced much greater biomass during the second year. After each growing season the above ground biomass was analyzed for cell wall sugars and for recalcitrance to enzymatic digestibility, and biofuel using a separate hydrolysis and fermentation (SHF) screen. Here, each transgenic event and control had more glucan, xylan and less ethanol (g/g basis) from the second year of growth relative to the first year plants. There was no correlation between plant carbohydrate content and biofuel production. In each of cell wall-targeted transgenics, GAUT4 , MYB4, COMT and FPGS , the second year of growth resulted in increased carbohydrate abundance (up to 12%) and reduced recalcitrance through higher ethanol yields (up to 21%) over the non-transgenic control plants.

59 BASIC BIOLOGICAL SCIENCES↗

Characterization of switchgrass ( Panicum virgatum L.) PvKSL1 as a levopimaradiene/abietadiene‐type diterpene synthase

Abstract The diverse class of plant diterpenoid metabolites serves important functions in mediating growth, chemical defence, and ecological adaptation. In major monocot crops, such as maize (Zea mays), rice (Oryza sativa), and barley (Hordeum vulgare), diterpenoids function as core components of biotic and abiotic stress resilience. Switchgrass (Panicum virgatum) is a perennial grass valued as a stress‐resilient biofuel model crop. Previously we identified an unusually large diterpene synthase family that produces both common and species‐specific diterpenoids, several of which accumulate in response to abiotic stress. Here, we report discovery and functional characterization of a previously unrecognized monofunctional class I diterpene synthase (PvKSL1) viain vivoco‐expression assays with different copalyl pyrophosphate (CPP) isomers, structural and mutagenesis studies, as well as genomic and transcriptomic analyses. In particular, PvKSL1 convertsent‐CPP intoent‐abietadiene,ent‐palustradiene,ent‐levopimaradiene, andent‐neoabietadiene via a 13‐hydroxy‐8(14)‐ent‐abietene intermediate. Notably, although featuring a distinctent‐stereochemistry, this product profile is near‐identical to bifunctional (+)‐levopimaradiene/abietadiene synthases occurring in conifer trees. PvKSL1 has three of four active site residues previously shown to control (+)‐levopimaradiene/abietadiene synthase catalytic specificity. However, mutagenesis studies suggest a distinct catalytic mechanism in PvKSL1. Genome localization ofPvKSL1distant from other diterpene synthases, and its phylogenetic distinctiveness from known abietane‐forming diterpene synthases, support an independent evolution of PvKSL1 activity. Albeit at low levels,PvKSL1gene expression predominantly in roots suggests a role of diterpenoid formation in belowground tissue. Together, these findings expand the known chemical and functional space of diterpenoid metabolism in monocot crops.

Plant Sciences↗

Cold acclimation threshold induction temperatures of switchgrass ecotypes grown under a long and short photoperiod

Abstract Plants can cold acclimate to enhance their freezing tolerance by sensing declining temperature and photoperiod cues. However, the factors influencing genotypic variation in the induction of cold acclimation are poorly understood among perennial grasses. We hypothesized that the more northern upland switchgrass ( Panicum virgatum L.) ecotype develops a higher degree of freezing tolerance by initiating cold acclimation at higher temperatures as compared with the coastal and southern lowland ecotypes. First, we determined the optimal method for assessing freezing tolerance and the length of exposure to 8/4°C required to induce the maximum level of freezing tolerance in the most northern upland and most southern lowland genotypes. We characterized the maximum freezing tolerance of eight uplands, three coastal and five lowland genotypes grown for 21 days at 8/4°C and a 10 or 16 h photoperiod. Next, we identified the temperature required to induce cold acclimation by exposing the 16 genotypes for 7 days at 20–6°C constant temperatures under a 10 or 16 h photoperiod. Cold acclimation initiated at temperatures 5 and 7°C higher in upland than in coastal and lowland genotypes. Among upland genotypes the shorter photoperiod induced cold acclimation at a 1°C higher temperature. Genotypes originating from a more northern latitude initiate cold acclimation at higher temperatures and develop higher maximum freezing tolerances. An earlier response to declining temperatures may provide the upland ecotype with additional time to prepare for winter and provide an advantage when plants are subjected to the rapid changes in fall temperature associated with injurious frosts.

60 APPLIED LIFE SCIENCES↗

Draft Genome Sequences of Switchgrass Diazotrophs

We report the draft genome sequences of five native nitrogen-fixing bacteria associated with roots of switchgrass isolated from the Tallgrass Prairies of Oklahoma. Nitrogen-fixing genes, including the nif cluster, are conserved across the Klebsiella and Kosakonia strains.

Jones, Lauren B.↗

Transcriptomic Data Sets for Zymomonas mobilis 2032 during Fermentation of Ammonia Fiber Expansion (AFEX)-Pretreated Corn Stover and Switchgrass Hydrolysates

The transcriptomes of Zymomonas mobilis 2032 were captured during the fermentation of ammonia fiber expansion (AFEX)-pretreated corn stover and switchgrass hydrolysates containing different concentrations of glucose and xylose. RNA samples were collected when Z. mobilis was fermenting glucose or xylose. Here, we present transcriptome sequencing (RNA-Seq) data obtained during separate phases of glucose or xylose consumption.

59 BASIC BIOLOGICAL SCIENCES↗

Bacterial isolate collection from switchgrass rhizosphere

ABSTRACT We provide a collection of 78 bacterial isolates from the rhizosphere of switchgrass ( Panicum virgatum L .) at the Lux Arbor Reserve in Delton, MI, a site of the Great Lakes Bioenergy Research Center (GLBRC), Michigan State University, MI, USA. We include information on isolation conditions and full-length 16S rRNA sequences.

Grady, Keara L.↗

Low-Moisture Switchgrass Bulk-Format Logistics Costs Based on Engineering Data

Abstract. Logistics costs that spanned harvest, transport, storage, preprocessing, and year-round delivery of 371,870 dMg year -1 of milled, low-moisture switchgrass (SG) to a local biorefinery were determined for bulk-format versus round-bale systems. Separate spreadsheet analyses were conducted for three diesel fuel prices ($0.53 L -1 , $0.92 L -1 , and $1.32 L -1 ) because of the wide range of fuel prices during the project. A detailed analysis scenario identified a common harvest period, equipment annual use, other equipment uses, and protected SG storage on gravel pad, along with equipment performances based on previously published tests primarily for commercially available equipment. The bulk-format system used a depot equipped with stacker/reclaimer technology for storage, whereas the round-bale system used field-side bale stacks for storage, with both systems assuming 6% losses, primarily attributed to storage. Results indicated total costs for bulk-format versus round-bale systems of $54.63 per dry Mg (dMg -1 ), $60.41 dMg -1 , and $66.21 dMg -1 versus $54.59 dMg -1 , $62.10 dMg -1 , and $67.93 dMg -1 corresponding with diesel fuel costs of $0.53 L -1 , $0.92 L -1 , and $1.32 L -1 , respectively. The most significant bulk-format system total cost of $22.79 dMg -1 was associated with a stacker/reclaimer, conveyance, and dust collection at the depot, which was 42% of total costs at a diesel fuel cost of $0.53 L -1 . For the round-bale system, bale chopping total costs was highest and represented an average of 27% of total costs, with costs ranging from $14.63 dMg -1 to $19.08 dMg -1 for diesel fuel input costs ranging from $0.53 L -1 to $1.32 L -1 , respectively. For comparison, the forage harvester provided harvesting and chopping in the bulk-format system at total costs ranging from $6.47 dMg -1 to $8.80 dMg -1 across the same range of diesel fuel input cost. The forage harvester provided an economic advantage in size reduction compared to tub grinding. Truck hauling for both the bulk-format and round-bale systems represented significant expenses at all selected fuel costs. In addition, the multiple times that round bales were handled represented accumulating costs that were sensitive to diesel fuel costs. Keywords: Bales, Bioenergy, Bulk format, Densification, Field chop, Logistics, Transportation.

Agriculture↗

Techno-Economic Impact of Diesel Fuel, Labor, and Interest Rates on Bulk-Format and Round Bale Logistics Expenses for Switchgrass

Highlights System-wide sensitivity of bulk-format and round bale logistics responded to different degrees of expense inputs. Biomass size-reduction expenses for bulk-format were consistently less than round-bale chopping. Stack/reclaim operations with high-density-capacity equipment for bulk-format were the highest expenses. Clean-sheet design of stacker/reclaimer and self-compacting bulk trailer may significantly reduce expenses. Abstract . Deterministic cost analyses were conducted for a harvest, storage, preprocessing, and supply system providing 371,870 dry Mg (dMg) year-1 of milled switchgrass (SG) to a biorefinery from 21,929 ha of production fields within 72 km of the biorefinery. Varied input costs of diesel fuel, labor, and interest rate were input into a highly detailed cost analysis for bulk-format and baseline round bale logistics. These operational input factors, potentially reflecting an evolving economic climate or paradigm shift, were examined to provide insight into the cost to deliver preprocessed biomass feedstock for centralized commercial-scale conversions. Total annual costs per dry unit of SG were determined for all combinations of diesel fuel prices of $0.53 L-1, $0.92 L-1, and $1.32 L-1; total labor costs of $20.00 h-1, $28.85 h-1, and $40.00 h-1 singly and uniformly applied across all labor inputs; and three representative interest rates of 6%, 8%, and 12%. Increases in total cost most depended on increases in diesel fuel cost, followed by labor rate, and finally interest rate. Increases in fuel costs from $0.53 L-1 to $1.32 L-1 yielded a total cost increase of about $11.56 dMg-1 and $13.36 dMg-1 for bulk-format and round bale logistics, respectively, for all combinations of labor and interest rates. The overall increase in labor rate ($20.00 h-1 to $40.85 h-1) resulted in increased total costs of about $10.32 dMg-1 for bulk-format logistics and $9.94 dMg-1 for round bale logistics for all fuel and interest rates. The overall increase in interest rate (6% to 12%) resulted in increased total costs of about $6.72 dMg-1 for bulk-format logistics and $4.21 dMg-1 for round bale logistics for all fuel and labor rates. Increases in total costs for combined increases in fuel cost, labor rate, and interest rate were $28.60 dMg-1 and $27.48 dMg-1, yielding maximum total costs of $87.87 dMg-1 and $86.53 dMg-1 for bulk-format and round bale logistics, respectively. Costs were determined for existing equipment systems, including novel use of waste compaction and ejector semi-trailers. The highest potential for cost reductions may require clean-sheet equipment designs specifically tailored for SG, such as forage harvesters that direct-cut standing SG, mobilized stacker-reclaimer technology to not inhibit stack-area footprint, self-compacting ejector trailer technology, and bale grinding technology incorporating controlled mass feeding and shear failure elements. Keywords: Bales, Bioenergy, Bulk-format, Densification, Field chop, Logistics, Transportation

Agriculture↗

Proteogenomic Analysis of a Thermophilic Bacterial Consortium Adapted to Deconstruct Switchgrass

Thermophilic bacteria are a potential source of enzymes for the deconstruction of lignocellulosic biomass. However, the complement of proteins used to deconstruct biomass and the specific roles of different microbial groups in thermophilic biomass deconstruction are not well-explored. Here we report on the metagenomic and proteogenomic analyses of a compost-derived bacterial consortium adapted to switchgrass at elevated temperature with high levels of glycoside hydrolase activities. Near-complete genomes were reconstructed for the most abundant populations, which included composite genomes for populations closely related to sequenced strains of Thermus thermophilus and Rhodothermus marinus, and for novel populations that are related to thermophilic Paenibacilli and an uncultivated subdivision of the littlestudied Gemmatimonadetes phylum. Partial genomes were also reconstructed for a number of lower abundance thermophilic Chloroflexi populations. Identification of genes for lignocellulose processing and metabolic reconstructions suggested Rhodothermus, Paenibacillus and Gemmatimonadetes as key groups for deconstructing biomass, and Thermus as a group that may primarily metabolize low molecular weight compounds. Mass spectrometry-based proteomic analysis of the consortium was used to identify .3000 proteins in fractionated samples from the cultures, and confirmed the importance of Paenibacillus and Gemmatimonadetes to biomass deconstruction. These studies also indicate that there are unexplored proteins with important roles in bacterial lignocellulose deconstruction.

Environmental Molecular Sciences Laboratory↗