Synthetic polyploid production of Miscanthus sacchariflorus,Miscanthus sinensis,and Miscanthus x giganteus

Plants from the genus Miscanthus are potential renewable sources of lignocellulosic biomass for energy production. A potential strategy for Miscanthus crop improvement involves interspecific manipulation of ploidy levels to generate superior germplasm and to circumvent reproductive barriers for the introduction of new genetic variation into core germplasm. Synthetic autotetraploid lines of Miscanthus sacchariflorus and Miscanthus sinensis, and autoallohexaploid Miscanthus x giganteus were produced in tissue culture from oryzalin treatments to seed‐ and immature inflorescence‐derived callus lines. This is the first report of the genome doubling of diploid M. sacchariflorus. Genome doubling of diploid M. sinensis, M. sacchariflorus, and triploid M. x giganteus to generate tetraploid and hexaploid lines was confirmed by stomata size, nuclear DNA content, and chromosome counts. A putative pentaploid line was also identified among the M. x giganteus synthetic polyploid lines by nuclear DNA content and chromosome counts. Comparisons of phenotypic performance of synthetic polyploid lines with their diploid and triploid progenitors in the greenhouse found species‐specific differences in plant tiller number, height, and flowering time among the doubled lines. Stem diameter tended to increase after polyploidization but there were no significant improvements in biomass traits. Under field conditions, M. x giganteus synthetic hexaploid lines showed greater phenotypic variation, in terms of plant height, stem diameter, and tiller number, than their progenitor lines. Production of synthetic autopolyploid lines displaying significant phenotypic variation suggests that ploidy manipulation can introduce useful genetic diversity in the limited Miscanthus germplasm currently available in the United States. The role of polyploidization in the evolution and breeding of the genus Miscanthus is discussed.     

Development of SCAR marker for simultaneous identification of Miscanthus sacchariflorus, M. sinensis and M. x giganteus

The sequence-characterized amplified region (SCAR) marker for simultaneous identification of Miscanthus sacchariflorus, Miscanthus sinensis, and Miscanthus x giganteus was developed. In this study, it was attempted for the first time to develop the SCAR marker for detecting the molecular phenotypes among Miscanthus species. Randomly amplified polymorphic DNA technique was applied for this study and one fragment which is unique to M. sacchariflorus was identified and then sequenced. Based on the specific fragment, one SCAR primer pair designated as MS62-5F and MS62-5R was designed to amplify an approximately 1,000 bp DNA fragment within the sequenced region. Diagnostic PCR was performed using the primer pair. Using this SCAR marker, approximately 1,000 bp and 1,200 bp DNA fragments were obtained in M. sacchariflorus and M. sinensis, respectively. Moreover, M. x giganteus was obtained both bands at the same time. The result showed that this SCAR marker can clearly distinguish the M. sacchariflorus, M. sinensis, and M. x giganteus, respectively.     

Radiation capture and conversion efficiencies of Miscanthus sacchariflorus,M. sinensis and their naturally occurring hybrid M. × giganteus

Miscanthus is a rhizomatous C4 grass of great interest as a biofuel crop because it has the potential to produce high yields over a wide geographical area with low agricultural inputs on marginal land less suitable for food production. At the moment, a clonal interspecific hybrid Miscanthus × giganteus is the most widely cultivated and studied in Europe and the United States, but breeding programmes are developing newer more productive varieties. Here, we quantified the physiological processes relating to whole season yield in a replicated plot trial in Wales, UK. Light capture and conversion efficiency were parameterized for four carefully selected genotypes (M. sinensis, M. sacchariflorus and Miscanthus × giganteus). Differences in the canopy architecture in mature stands as measured by the extinction coefficient (k) were small (0.55–0.65). Sensitivity analysis on a mathematical model of Miscanthus was performed to quantify the accumulative intercepted photosynthetically active radiation (iPAR) in the growing season using (i) k, (ii) variation in the thermal responses of leaf expansion rate, (iii) base temperature for degree days and (iv) date start of canopy expansion. A 10% increase in k or leaf area per degree day both had a minimal effect on iPAR (3%). Decreasing base temperature from 10 to 9 °C gave an 8% increase in iPAR. If the starting date for canopy expansion was the same as shoot emergence date, then the iPAR increases by 12.5%. In M. × giganteus, the whole season above ground and total (including below ground) radiation‐use efficiency (RUE) ranged from 45% to 37% higher than the noninterspecific hybrid genotypes. The greater yields in the interspecific hybrid M. × giganteus are explained by the higher RUE and not by differences in iPAR or partitioning effects. Studying the mechanisms underlying this complex trait could have wide benefits for both fuel and food production.     

Saccharification of Miscanthus x giganteus, incorporation of lignocellulosic by-product in cementitious matrix

Given the non competition of miscanthus with food and animal feed, this lignocellulosic species has attracted attention as a possible biofuel resource. However, sustainability of ethanol production from lignocelluloses biomass would imply reduction in the consumption of chemicals and/or energetic means, but also valorization of the lignocellulosic by-product remaining from enzymatic saccharification. Introduction of these by-products into a cementitious matrix could be used in manufacturing a lightweight composite. Miscanthus biomass was submitted to chemical pretreatments followed by saccharification using an enzymatic cocktail. Residues from saccharification were then mixed with a cementitious matrix. Given their mechanical properties and a good adherence between cement and by-product, the hardened materials could be used. However, the delay in the beginning of setting time is too long, which prevents the direct use of by-product into cementitious matrix. Preliminary experiments using a setting accelerator in the cementitious matrix permitted significant reduction in the setting time delay.     

芒和五节芒在中国的潜在分布

利用最大熵模型, 将我国现有芒(Miscanthus sinensis)和五节芒(M. floridulus)的地理分布信息与19个降水及温度等气候因子相拟合, 预测了芒和五节芒在我国的潜在分布区域, 并推测出芒和五节芒的基本生态位。结果显示: 芒的潜在适生区包括四川西部、陕西北部、宁夏、内蒙古中部、黑龙江、吉林西部、辽宁西部、青海东南部等地区, 其基本生态位参数为: 最暖季节降水量为400-1 000 mm, 平均8月降水量为100-350 mm, 7月平均最低气温为15 ℃, 平均7月降水量为100-350 mm, 11月平均最高气温为-10-22 ℃, 最干月平均气温为-15-20 ℃, 平均12月降水量为100 mm以下; 五节芒的潜在适生区为云南、陕西、山西、宁夏、河南、山东、吉林、辽宁以及四川西部、甘肃南部和内蒙古东部等地区, 其基本生态位参数为: 最暖季节降水量至少在400 mm以上, 平均6月降水量为150-550 mm, 7月平均最低气温在15-30 ℃之间, 6月平均最低气温为10 ℃, 平均4月降水量为50-100 mm。结果表明, 在进行遗传改良的前提下, 我国有丰富的适合栽植芒和五节芒的土地资源。     

Rhizobacteria associated with Miscanthus x giganteus improve metal accumulation and plant growth in the flotation tailings

Plant and Soil - Flotation tailings represent an extremely unfriendly substrate for plant colonization due to toxic metal concentrations and marked macronutrient deficiencies. The perennial grass...     

Evolution of soil carbon stocks under Miscanthus × giganteus and Miscanthus sinensis across contrasting environmental conditions

Miscanthus is a C4 bioenergy perennial crop characterized by its high potential yield. Our study aimed to compare the carbon storage capacities of Miscanthus sinensis (M. sinensis) with that of Miscanthus × giganteus (M. × giganteus) in field conditions in different types of soils in France. We set up a multi‐environment experimental network. On each trial, we tested two treatments: M. × giganteus established from rhizomes (Gr) and M. sinensis transplanted seedlings (Sp). We quantified the soil organic carbon (SOC) stock at equivalent soil mass for both genotypes in 2014 and 2019 and for two sampling depths: L1 (ca. 0–5 cm) and L1‐2 (ca. 0–30 cm). We also calculated the total and annual variation of the SOC stock and investigated factors that could explain the variation and the initial state of the SOC stock. ANOVAs were performed to compare the SOC stock, as well as the SOC stock variation rates across treatments and soil layers. Results showed that the soil bulk density did not vary significantly between 2014 and 2019 for both treatments (Gr and Sp). The SOC concentration (i.e. SOC expressed in g/kg) increased significantly between 2014 and 2019 in L1, whereas no significant evolution was found in L2 (ca. 5–30 cm). The SOC stock (i.e. SOC expressed in t/ha) increased significantly in the superficial layer L1 for M. × giganteus and M. sinensis, by 0.48 ± 0.41 and 0.54 ± 0.25 t ha^?1 year^?1 on average, respectively, although no significant change was detected in the layer L1‐2 for both genotypes. Moreover, SOC stocks in 2019 did not differ significantly between M. × giganteus and M. sinensis in the soil layers L1 and L1‐2. Lastly, our results showed that the initial SOC stock was significantly higher when miscanthus was grown after set‐aside than after annual crops.     

Effects of several establishment modes of Miscanthus × giganteus and Miscanthus sinensis on yields and yield trends

Miscanthus is a C4 perennial grass originating from East Asia, the yields of which progressively increase in the first years of growth. Several species for bioenergy have been studied since the mid‐1980s in Europe, in particular (Miscanthus × giganteus [M. × giganteus]), due to its high yields. M. × giganteus is mainly cultivated in France and established from rhizomes. Our study aimed to assess, in field conditions, alternative establishment methods combined with an alternative species, Miscanthus sinensis (M. sinensis). We set up a multi‐environment experimental network. On each trial, we tested two treatments with M. × giganteus, established from rhizomes (G_r‐sd) and from plantlets obtained from rhizomes (G_p‐sd), and two treatments with M. sinensis seedlings transplanted in single (S_p‐sd) and double density (S_p‐dd). ANOVA was performed to compare establishment and regrowth rates across treatments, as well as yields across treatments and site‐years. A logistic model was used to describe yield trends and to compare the maximum yield reached and the rate of yield increase of both species. Results showed that miscanthus establishment from plantlets resulted in higher establishment (between 87% and 92%) and regrowth (between 91% and 94%) rates compared to establishment from rhizomes. Treatments with M. × giganteus obtained higher average yields across site‐years than those with M. sinensis, but more variable yields across site‐years. We showed a strong species effect on yields, yield components (shoot weight, shoot density and shoot number per plant) and light interception (through leaf area index). Lastly, to use M. sinensis established from transplanted plantlets as an alternative to M. × giganteus, research would be required on the breeding of M. sinensis sterile seeds to avoid risks of invasiveness.     

Leaf photosynthesis in the C4-grass Miscanthus x giganteus, growing in the cool temperate climate of southern England

Previous studies have shown that both photoinhibition and lowtemperature impairment of leaf development occur in C₄ speciesgrowing in temperate climates. These result in reductions inthe maximum quantum efficiency (     

SSR-based genetic maps of Miscanthus sinensis and M. sacchariflorus, and their comparison to sorghum

We present SSR-based genetic maps from a cross between Miscanthus sacchariflorus Robustus and M. sinensis, the progenitors of the promising cellulosic biofuel feedstock Miscanthus × giganteus. cDNA-derived SSR markers were mapped by the two-way pseudo-testcross model due to the high heterozygosity of each parental species. A total of 261 loci were mapped in M. sacchariflorus, spanning 40 linkage groups and 1,998.8 cM, covering an estimated 72.7% of the genome. For M. sinensis, a total of 303 loci were mapped, forming 23 linkage groups and 2,238.3 cM, covering 84.9% of the genome. The use of cDNA-derived SSR loci permitted alignment of the Miscanthus linkage groups to the sorghum chromosomes, revealing a whole genome duplication affecting the Miscanthus lineage after the divergence of subtribes Sorghinae and Saccharinae, as well as traces of the pan-cereal whole genome duplication. While the present maps provide for many early research needs in this emerging crop, additional markers are also needed to improve map density and to further characterize the structural changes of the Miscanthus genome since its divergence from sorghum and Saccharum.     

Comparative Cellular Analysis of Pathogenic Fungi with a Disease Incidence in Brachypodium distachyon and Miscanthus x giganteus

Based on a growing demand on renewable energy, fast growing perennial grasses have been identified as energy crops with a high capacity in sustainable biomass production. Among these grasses, the giant reed Miscanthus x giganteus delivers one of the highest biomass yields. Despite its potential for an extended cultivation, only little is known about putative fungal pathogens that might cause biomass losses. Molecular targets that are related to fungal resistance have not been identified because cellular and molecular tools have not been established in this energy crop. Therefore, our study was aimed to evaluate a method to compare the penetration process of fungal plant pathogens in the model grass Brachypodium distachyon and M. giganteus. In a screening with 13 different fungal species on detached leaves, we identified four filamentous fungi that infected both B. distachyon and M. giganteus and have not been previously described as M. giganteus pathogens. Spray inoculations with these four fungi on intact M. giganteus leaves of whole plants confirmed their pathogenicity. Microscopic analysis of the fungal infections and the hyphal propagation within the leaf tissue revealed that the four newly identified fungi used very similar strategies for penetration and colonization in B. distachyon and M. giganteus. This suggests that B.?distachyon could be suitable to establish model pathosystems for these fungal pathogens that colonize M. giganteus. The already existing genetic tools for B. distachyon might improve the identification of defense-related targets and mechanisms supporting fungal resistance in M. giganteus.     

The efficiency of C4 photosynthesis under low light conditions in Zea mays,Miscanthus x giganteus and Flaveria bidentis

The efficiency of C₄ photosynthesis in Zea mays, Miscanthus x giganteus and Flaveria bidentis in response to light was determined using measurements of gas exchange, ¹³CO₂ photosynthetic discrimination, metabolite pools and spectroscopic assays, with models of C₄ photosynthesis and leaf ¹³CO₂ discrimination. Spectroscopic and metabolite assays suggested constant energy partitioning between the C₄ and C₃ cycles across photosynthetically active radiation (PAR). Leakiness (φ), modelled using C₄ light‐limited photosynthesis equations (φ_mod), matched values from the isotope method without simplifications (φ_is) and increased slightly from high to low PAR in all species. However, simplifications of bundle‐sheath [CO₂] and respiratory fractionation lead to large overestimations of φ at low PAR with the isotope method. These species used different strategies to maintain similar φ. For example, Z. mays had large rates of the C₄ cycle and low bundle‐sheath cells CO₂ conductance (g_bs). While F. bidentis had larger g_bs but lower respiration rates and M. giganteus had less C₄ cycle capacity but low g_bs, which resulted in similar φ. This demonstrates that low g_bs is important for efficient C₄ photosynthesis but it is not the only factor determining φ. Additionally, these C₄ species are able to optimize photosynthesis and minimize φ over a range of PARs, including low light.     

Nitrogen‐dependent bacterial community shifts in root,rhizome and rhizosphere of nutrient‐efficient Miscanthus x giganteus from long‐term field trials

The perennial energy crop Miscanthus × giganteus is recognized for its extraordinary nitrogen‐use efficiency. While the remobilization of nitrogen (N) to the rhizome after the growth phase contributes to this efficiency, the plant‐associated microbiome might also contribute, as N‐fixing bacterial species had been isolated from this grass. Here, we studied established Miscanthus × giganteus plots in southern Germany that either received 80 kg N ha^?1 a^?1 or that were not N‐fertilized for 14 years. The bacterial communities of the bulk soil, rhizosphere, roots and rhizomes were analysed. Major differences were encountered between plant‐associated fractions. Nitrogen had little effect on soil communities. The roots and rhizomes showed less microbial diversity than soil fractions. In these compartments, Actinobacteria and N‐fixing symbiosis‐associated Proteobacteria depended on N. Intriguingly, N₂‐fixing‐related bacterial families were enriched in the rhizomes in long‐term zero N plots, while denitrifier‐related families were depleted. These findings point to the rhizome as a potentially interesting plant organ for N fixation and demonstrate long‐term differences in the organ‐specific bacterial communities associated with different N supply, which are mainly shaped by the plant.     

Endogenous Gibberellins in Aeginetia indica,a Parasitic Plant,and Its Host,Miscanthus sinensis

Endogenous gibberellins in a parasitic plant, Aeginetia indica L., and its host, Miscanthus sinensis Andress (eulalia) were analyzed. Gibberellins of the early-non-hydroxylation pathway and their putative metabolites were identified as the major endogenous gibberellins from both types of A. indica parasitizing M. sinensis and parasitizing Oryza sativa L. (rice). Members of both the early-non- and early-13-hydroxylation pathways were detected in the host M. sinensis. Since the early-13-hydroxylation pathway has been reported to be the major pathway operating in vegetative tissues of O. sativa, these results suggest that A. indica can biosynthesize gibberellins independent of its hosts.     

Quality and decomposition in soil of rhizome, root and senescent leaf from Miscanthus x giganteus, as affected by harvest date and N fertilization

To predict the environmental benefits of energy crop production and use, the nature and fate of biomass residues in the soil need to be quantified. Our objective was to quantify Miscanthus x giganteus biomass recycling to soil and to assess how harvesting time and N fertilization affect their characteristics and subsequent biodegradability. The quantification of aerial and belowground biomass and their sampling were performed on 2- and 3-year-old Miscanthus stands, either fertilized with 120 kg N ha^?1 year^?1 or not fertilized, in autumn (maximal biomass production) and winter (maturity). Plant biomass was chemically characterized (total sugars, Klason lignin, C/N) and incubated in optimum decomposition conditions (15°C, ?80 kPa) for 263 days, for C and N mineralization. Accumulation of carbon in rhizomes and roots was 7.5 to 10 t C ha^?1 and represented about 50% of total plant biomass C. Senescent leaves represented about 1.5 t C ha^?1 year^?1. All residues, especially the roots, had high lignin contents, while the rhizomes also had a high soluble content due to their nutrient storage function. The C mineralization rates were closely related to the chemical characteristics of the residue, higher sugar and lower lignin contents leading to faster decomposition, as observed for rhizomes.     

Chemical Composition of Miscanthus sinensis var. purpurascens Flower Absolute and Its Beneficial Effects on Skin Wound Healing and Melanogenesis-Related Cell Activities

Miscanthus sinensis var. purpurascens (MSP, flame grass) is found in Korea, Japan, and China, and its biological activities include anti-cancer, detoxifying, vasodilatory, antipyretic, and diuretic effects. However, no study has investigated the effects of MSP on skin-related biological activities. In this study, we explored the effects of the absolute extracted from the MSP flowers (MSPFAb) on skin wound healing- and whitening-related responses in keratinocytes or melanocytes. MSPFAb contained 6 components and induced the proliferation, migration, and syntheses of type I and IV collagens in keratinocytes. MSPFAb also increased the phosphorylations of serine/threonine-specific protein kinase, p38 mitogen-activated protein kinase, and extracellular signal-regulated kinase1/2 in keratinocytes. In addition, treatment with MSPFAb decreased serum-induced melanoma cell proliferation and inhibited tyrosinase activity and melanin contents in α-MSH-stimulated melanoma cells. Taken together, this study indicates MSPFAb may promote wound healing- and whitening-associated activities in dermal cells, and suggests that it has potential use as a wound healing and skin whitening agent.     

Evaluation of Agricultural Production Systems Simulator as yield predictor of Panicum virgatum and Miscanthus x giganteus in several US environments

Simulation models for perennial energy crops such as switchgrass (Panicum virgatum L.) and Miscanthus (Miscanthus x giganteus) can be useful tools to design management strategies for biomass productivity improvement in US environments. The Agricultural Production Systems Simulator (APSIM) is a biophysical model with the potential to simulate the growth of perennial crops. APSIM crop modules do not exist for switchgrass and Miscanthus, however, re‐parameterization of existing APSIM modules could be used to simulate the growth of these perennials. Our aim was to evaluate the ability of APSIM to predict the dry matter (DM) yield of switchgrass and Miscanthus at several US locations. The Lucerne (for switchgrass) and Sugarcane (for Miscanthus) APSIM modules were calibrated using data from four locations in Indiana. A sensitivity analysis informed the relative impact of changes in plant and soil parameters of APSIM Lucerne and APSIM Sugarcane modules. An independent dataset of switchgrass and Miscanthus DM yields from several US environments was used to validate these re‐parameterized APSIM modules. The re‐parameterized modules simulated DM yields of switchgrass [0.95 for CCC (concordance correlation coefficient) and 0 for SB (bias of the simulation from the measurement)] and Miscanthus (0.65 and 0% for CCC and SB, respectively) accurately at most locations with the exception of switchgrass at southern US sites (0.01 for CCC and 2% for SB). Therefore, the APSIM model is a promising tool for simulating DM yields for switchgrass and Miscanthus while accounting for environmental variability. Given our study was strictly based on APSIM calibrations at Indiana locations, additional research using more extensive calibration data may enhance APSIM robustness.