Effect of Water Deficit on Germination, Growth and Biochemical Responses of Four Potentially Invasive Ornamental Grass Species.

Ornamental plant species introduced into new environments can exhibit an invasive potential and adaptability to abiotic stress factors. In this study, the drought stress responses of four potentially invasive ornamental grass species (Cymbopogon citratus, Cortaderia selloana, Pennisetum alopecuroides and P. setaceum) were analysed. Several seed germination parameters were determined under increasing polyethylene glycol (PEG 6000) concentrations. Additionally, plants in the vegetative stage were subjected to intermediate and severe water stress treatments for four weeks. All species registered high germination rates in control conditions (no stress treatment), even at high PEG concentrations, except C. citratus, which did not germinate at -1 MPa osmotic potential. Upon applying the water stress treatments, P. alopecuroides plants showed the highest tolerance, and C. citratus appeared the most susceptible to drought. Stress-induced changes in several biochemical markers (photosynthetic pigments, osmolytes, antioxidant compounds, root and shoot Na+ and K+ contents), highlighted different responses depending on the species and the stress treatments. Basically, drought tolerance seems to depend to a large extent on the active transport of Na+ and K+ cations to the aerial part of the plants, contributing to osmotic adjustment in all four species and, in the case of the most tolerant P. alopecuroides, on the increasing root K+ concentration under water deficit conditions. The study shows the invasive potential of all species, except C. citratus, in dry areas such as the Mediterranean region, especially in the current climate change scenario. Particular attention should be given to P. alopecuroides, which is widely commercialised in Europe as ornamental.

Syntenic Cell Wall QTLs as Versatile Breeding Tools: Intraspecific Allelic Variability and Predictability of Biomass Quality Loci in Target Plant Species.

Syntenic cell wall QTLs (SQTLs) can identify genetic determinants of biomass traits in understudied species based on results from model crops. However, their effective use in plant breeding requires SQTLs to display intraspecific allelic variability and to predict causative loci in other populations/species than the ones used for SQTLs identification. In this study, genome assemblies from different accessions of Arabidopsis, rapeseed, tomato, rice, Brachypodium and maize were used to evaluate the intraspecific variability of SQTLs. In parallel, a genome-wide association study (GWAS) on cell wall quality traits was performed in miscanthus to verify the colocalization between GWAS loci and miscanthus SQTLs. Finally, an analogous approach was applied on a set of switchgrass cell wall QTLs retrieved from the literature. These analyses revealed large SQTLs intraspecific genetic variability, ranging from presence-absence gene variation to SNPs/INDELs and changes in coded proteins. Cell wall genes displaying gene dosage regulation, such as PAL and CAD, displayed presence-absence variation in Brachypodium and rapeseed, while protein INDELs were detected for the Brachypodium homologs of the rice brittle culm-like 8 locus, which may likely impact cell wall quality. Furthermore, SQTLs significantly colocalized with the miscanthus and switchgrass QTLs, with relevant cell wall genes being retained in colocalizing regions. Overall, SQTLs are useful tools to screen germplasm for relevant genes and alleles to improve biomass quality and can increase the efficiency of plant breeding in understudied biomass crops.

Novel Miscanthus hybrids: Modelling productivity on marginal land in Europe using dynamics of canopy development determined by light interception.

New biomass crop hybrids for bioeconomic expansion require yield projections to determine their potential for strategic land use planning in the face of global challenges. Our biomass growth simulation incorporates radiation interception and conversion efficiency. Models often use leaf area to predict interception which is demanding to determine accurately, so instead we use low-cost rapid light interception measurements using a simple laboratory-made line ceptometer and relate the dynamics of canopy closure to thermal time, and to measurements of biomass. We apply the model to project the European biomass potentials of new market-ready hybrids for 2020-2030. Field measurements are easier to collect, the calibration is seasonally dynamic and reduces influence of weather variation between field sites. The model obtained is conservative, being calibrated by crops of varying establishment and varying maturity on less productive (marginal) land. This results in conservative projections of miscanthus hybrids for 2020-2030 based on 10% land use conversion of the least (productive) grassland and arable for farm diversification, which show a European potential of 80.7-89.7 Mt year-1 biomass, with potential for 1.2-1.3 EJ year-1 energy and 36.3-40.3 Mt year-1 carbon capture, with seeded Miscanthus sacchariflorus × sinensis displaying highest yield potential. Simulated biomass projections must be viewed in light of the field measurements on less productive land with high soil water deficits. We are attempting to model the results from an ambitious and novel project combining new hybrids across Europe with agronomy which has not been perfected on less productive sites. Nevertheless, at the time of energy sourcing issues, seed-propagated miscanthus hybrids for the upscaled provision of bioenergy offer an alternative source of renewable energy. If European countries provide incentives for growers to invest, seeded hybrids can improve product availability and biomass yields over the current commercial miscanthus variety.

Breeding Targets to Improve Biomass Quality in Miscanthus.

Lignocellulosic crops are attractive bioresources for energy and chemicals production within a sustainable, carbon circular society. Miscanthus is one of the perennial grasses that exhibits great potential as a dedicated feedstock for conversion to biobased products in integrated biorefineries. The current biorefinery strategies are primarily focused on polysaccharide valorization and require severe pretreatments to overcome the lignin barrier. The need for such pretreatments represents an economic burden and impacts the overall sustainability of the biorefinery. Hence, increasing its efficiency has been a topic of great interest. Inversely, though pretreatment will remain an essential step, there is room to reduce its severity by optimizing the biomass composition rendering it more exploitable. Extensive studies have examined the miscanthus cell wall structures in great detail, and pinpointed those components that affect biomass digestibility under various pretreatments. Although lignin content has been identified as the most important factor limiting cell wall deconstruction, the effect of polysaccharides and interaction between the different constituents play an important role as well. The natural variation that is available within different miscanthus species and increased understanding of biosynthetic cell wall pathways have specified the potential to create novel accessions with improved digestibility through breeding or genetic modification. This review discusses the contribution of the main cell wall components on biomass degradation in relation to hydrothermal, dilute acid and alkaline pretreatments. Furthermore, traits worth advancing through breeding will be discussed in light of past, present and future breeding efforts.

Qualitative and Quantitative Differences in Osmolytes Accumulation and Antioxidant Activities in Response to Water Deficit in Four Mediterranean Limonium Species.

Limonium is a genus represented in the Iberian Peninsula by numerous halophytic species that are affected in nature by salinity, and often by prolonged drought episodes. Responses to water deficit have been studied in four Mediterranean Limonium species, previously investigated regarding salt tolerance mechanisms. The levels of biochemical markers, associated with specific responses-photosynthetic pigments, mono- and divalent ions, osmolytes, antioxidant compounds and enzymes-were determined in the control and water-stressed plants, and correlated with their relative degree of stress-induced growth inhibition. All the tested Limonium taxa are relatively resistant to drought on the basis of both the constitutive presence of high leaf ion levels that contribute to osmotic adjustment, and the stress-induced accumulation of osmolytes and increased activity of antioxidant enzymes, albeit with different qualitative and quantitative induction patterns. Limonium santapolense activated the strongest responses and clearly differed from Limonium virgatum, Limonium girardianum, and Limonium narbonense, as indicated by cluster and principal component analysis (PCA) analyses in agreement with its drier natural habitat, and compared to that of the other plants. Somewhat surprisingly, however, L. santapolense was the species most affected by water deficit in growth inhibition terms, which suggests the existence of additional mechanisms of defense operating in the field that cannot be mimicked in greenhouses.

Variable Levels of Tolerance to Water Stress (Drought) and Associated Biochemical Markers in Tunisian Barley Landraces.

Due to its high tolerance to abiotic stress, barley (Hordeum vulgare) is cultivated in many arid areas of the world. In the present study, we evaluate the tolerance to water stress (drought) in nine accessions of "Ardhaoui" barley landraces from different regions of Tunisia. The genetic diversity of the accessions is evaluated with six SSR markers. Seedlings from the nine accessions are subjected to water stress by completely stopping irrigation for three weeks. A high genetic diversity is detected among the nine accessions, with no relationships between genetic distance and geographical or ecogeographical zone. The analysis of growth parameters and biochemical markers in the water stress-treated plants in comparison to their respective controls indicated great variability among the studied accessions. Accession 2, from El May Island, displayed high tolerance to drought. Increased amounts of proline in water-stressed plants could not be correlated with a better response to drought, as the most tolerant accessions contained lower levels of this osmolyte. A good correlation was established between the reduction of growth and degradation of chlorophylls and increased levels of malondialdehyde and total phenolics. These biochemical markers may be useful for identifying drought tolerant materials in barley.

Effects of salinity and drought on growth, ionic relations, compatible solutes and activation of antioxidant systems in oleander (Nerium oleander L.).

Nerium oleander is an ornamental species of high aesthetic value, grown in arid and semi-arid regions because of its drought tolerance, which is also considered as relatively resistant to salt; yet the biochemical and molecular mechanisms underlying oleander's stress tolerance remain largely unknown. To investigate these mechanisms, one-year-old oleander seedlings were exposed to 15 and 30 days of treatment with increasing salt concentrations, up to 800 mM NaCl, and to complete withholding of irrigation; growth parameters and biochemical markers characteristic of conserved stress-response pathways were then determined in stressed and control plants. Strong water deficit and salt stress both caused inhibition of growth, degradation of photosynthetic pigments, a slight (but statistically significant) increase in the leaf levels of specific osmolytes, and induction of oxidative stress-as indicated by the accumulation of malondialdehyde (MDA), a reliable oxidative stress marker-accompanied by increases in the levels of total phenolic compounds and antioxidant flavonoids and in the specific activities of ascorbate peroxidase (APX) and glutathione reductase (GR). High salinity, in addition, induced accumulation of Na+ and Cl- in roots and leaves and the activation of superoxide dismutase (SOD) and catalase (CAT) activities. Apart from anatomical adaptations that protect oleander from leaf dehydration at moderate levels of stress, our results indicate that tolerance of this species to salinity and water deficit is based on the constitutive accumulation in leaves of high concentrations of soluble carbohydrates and, to a lesser extent, of glycine betaine, and in the activation of the aforementioned antioxidant systems. Moreover, regarding specifically salt stress, mechanisms efficiently blocking transport of toxic ions from the roots to the aerial parts of the plant appear to contribute to a large extent to tolerance in Nerium oleander.

Unraveling Salt Tolerance Mechanisms in Halophytes: A Comparative Study on Four Mediterranean Limonium Species with Different Geographic Distribution Patterns.

We have performed an extensive study on the responses to salt stress in four related Limonium halophytes with different geographic distribution patterns, during seed germination and early vegetative growth. The aims of the work were twofold: to establish the basis for the different chorology of these species, and to identify relevant mechanisms of salt tolerance dependent on the control of ion transport and osmolyte accumulation. Seeds were germinated in vitro, in the presence of increasing NaCl concentrations, and subjected to "recovery of germination" tests; germination percentages and velocity were determined to establish the relative tolerance and competitiveness of the four Limonium taxa. Salt treatments were also applied to young plants, by 1-month irrigation with NaCl up to 800 mM; then, growth parameters, levels of monovalent and divalent ions (in roots and leaves), and leaf contents of photosynthetic pigments and common osmolytes were determined in control and stressed plants of the four species. Seed germination is the most salt-sensitive developmental phase in Limonium. The different germination behavior of the investigated species appears to be responsible for their geographical range size: L. narbonense and L. virgatum, widespread throughout the Mediterranean, are the most tolerant and the most competitive at higher soil salinities; the endemic L. santapolense and L. girardianum are the most sensitive and more competitive only at lower salinities. During early vegetative growth, all taxa showed a strong tolerance to salt stress, although slightly higher in L. virgatum and L. santapolense. Salt tolerance is based on the efficient transport of Na+ and Cl- to the leaves and on the accumulation of fructose and proline for osmotic adjustment. Despite some species-specific quantitative differences, the accumulation patterns of the different ions were similar in all species, not explaining differences in tolerance, except for the apparent activation of K+ transport to the leaves at high external salinity, observed only in the most tolerant L. narbonense and L. virgatum. This specific response may be therefore relevant for salt tolerance in Limonium. The ecological implications of these results, which can contribute to a more efficient management of salt marshes conservation/regeneration programs, are also discussed.

Antioxidant responses under salinity and drought in three closely related wild monocots with different ecological optima.

Some deleterious effects of drought, soil salinity and other abiotic stresses are mediated by the generation of oxidative stress through an increase in reactive oxygen species (ROS) that damage cellular membranes, proteins and DNA. In response to increased ROS, plants activate an array of enzymatic and non-enzymatic antioxidant defences. We have correlated the activation of these responses with the contrasting tolerance to salinity and drought of three species of the genus Juncus, viz. J. maritimus, J. acutus (both halophytes) and J. articulatus (salt-sensitive). Both stresses were given for 8 weeks to 6-week-old seedlings in a controlled environment chamber. Each stress inhibited growth and degraded photosynthetic pigments in the three species with the most pronounced effects being in J. articulatus. Salt and water stress also generated oxidative stress in all three taxa with J. articulatus being the most affected in terms of accumulation of malondialdehyde (a reliable oxidative stress marker). The apparent lower oxidative stress in halophytic J. maritimus and J. acutus compared with salt-sensitive J. articulatus is explained by a more efficient activation of antioxidant systems since salt or water deficiency induced a stronger accumulation of antioxidant phenolic compounds and flavonoids in J. maritimus and J. acutus than in J. articulatus. Qualitative and quantitative differences in antioxidant enzymes were also detected when comparing the three species and the two stress treatments. Accordingly, glutathione reductase and superoxide dismutase activities increased in the two halophytes under both stresses, but only in response to drought in J. articulatus. In contrast, ascorbate peroxidase activity varied between and within species according to treatment. These results show the relative importance of different antioxidant responses for stress tolerance in species with distinct ecological requirements. The salt-sensitive J. articulatus, contrary to the tolerant taxa, did not activate enzymatic antioxidant responses to salinity-induced oxidative stress.

Salinity-Induced Variation in Biochemical Markers Provides Insight into the Mechanisms of Salt Tolerance in Common (Phaseolus vulgaris) and Runner (P. coccineus) Beans.

The evaluation of biochemical markers is important for the understanding of the mechanisms of tolerance to salinity of Phaseolus beans. We have evaluated several growth parameters in young plants of three Phaseolus vulgaris cultivars subjected to four salinity levels (0, 50, 100, and 150 mM NaCl); one cultivar of P. coccineus, a closely related species reported as more salt tolerant than common bean, was included as external reference. Biochemical parameters evaluated in leaves of young plants included the concentrations of ions (Na⁺, K⁺, and Cl-), osmolytes (proline, glycine betaine, and total soluble sugars), and individual soluble carbohydrates. Considerable differences were found among cultivars, salinity levels, and in their interaction for most traits. In general, the linear component of the salinity factor for the growth parameters and biochemical markers was the most important. Large differences in the salinity response were found, with P. vulgaris cultivars "The Prince" and "Maxidor" being, respectively, the most susceptible and tolerant ones. Our results support that salt stress tolerance in beans is mostly based on restriction of Na⁺ (and, to a lesser extent, also of Cl-) transport to shoots, and on the accumulation of myo-inositol for osmotic adjustment. These responses to stress during vegetative growth appear to be more efficient in the tolerant P. vulgaris cultivar "Maxidor". Proline accumulation is a reliable marker of the level of salt stress affecting Phaseolus plants, but does not seem to be directly related to stress tolerance mechanisms. These results provide useful information on the responses to salinity of Phaseolus.

Effects of Salt Stress on Three Ecologically Distinct Plantago Species.

Comparative studies on the responses to salt stress of taxonomically related taxa should help to elucidate relevant mechanisms of stress tolerance in plants. We have applied this strategy to three Plantago species adapted to different natural habitats, P. crassifolia and P. coronopus-both halophytes-and P. major, considered as salt-sensitive since it is never found in natural saline habitats. Growth inhibition measurements in controlled salt treatments indicated, however, that P. major is quite resistant to salt stress, although less than its halophytic congeners. The contents of monovalent ions and specific osmolytes were determined in plant leaves after four-week salt treatments. Salt-treated plants of the three taxa accumulated Na+ and Cl- in response to increasing external NaCl concentrations, to a lesser extent in P. major than in the halophytes; the latter species also showed higher ion contents in the non-stressed plants. In the halophytes, K+ concentration decreased at moderate salinity levels, to increase again under high salt conditions, whereas in P. major K+ contents were reduced only above 400 mM NaCl. Sorbitol contents augmented in all plants, roughly in parallel with increasing salinity, but the relative increments and the absolute values reached did not differ much in the three taxa. On the contrary, a strong (relative) accumulation of proline in response to high salt concentrations (600-800 mM NaCl) was observed in the halophytes, but not in P. major. These results indicate that the responses to salt stress triggered specifically in the halophytes, and therefore the most relevant for tolerance in the genus Plantago are: a higher efficiency in the transport of toxic ions to the leaves, the capacity to use inorganic ions as osmotica, even under low salinity conditions, and the activation, in response to very high salt concentrations, of proline accumulation and K+ transport to the leaves of the plants.

Screening for drought tolerance in cultivars of the ornamental genus Tagetes (Asteraceae).

Drought tolerance was evaluated in twelve cultivars of three ornamental Tagetes species (T. patula, T. tenuifolia and T. erecta). A stress treatment was performed by completely stopping watering of plants maintained in controlled greenhouse conditions. After three weeks, several plant growth parameters (stem length (SL), fresh weight (FW) and water content (WC)), photosynthetic pigments (chlorophylls and carotenoids (Car)), osmolytes (proline (Pro), glycine betaine (GB) and total soluble sugars (TSS)), an oxidative stress maker (malondialdehyde (MDA)) and antioxidants (total phenolic compounds (TPC) and total flavonoids (TF)) were measured. Considerable differences in the evaluated traits were found among the control and drought-stressed plants. Drought stress generally caused a marked reduction in plant growth and carotenoid pigments, and an increase in soluble solutes and oxidative stress. For most cultivars, proline levels in stressed plants increased between 30 and 70-fold compared to the corresponding controls. According to the different measured parameters, on average T. erecta proved to be more tolerant to drought than T. patula and T. tenuifolia. However, a considerable variation in the tolerance to drought was found within each species. The traits with greater association to drought tolerance as well as the most tolerant cultivars could be clearly identified in a principal components analysis (PCA). Overall, our results indicate that drought tolerant cultivars of Tagetes can be identified at early stages using a combination of plant growth and biochemical markers.

Native-Invasive Plants vs. Halophytes in Mediterranean Salt Marshes: Stress Tolerance Mechanisms in Two Related Species.

Dittrichia viscosa is a Mediterranean ruderal species that over the last decades has expanded into new habitats, including coastal salt marshes, ecosystems that are per se fragile and threatened by human activities. To assess the potential risk that this native-invasive species represents for the genuine salt marsh vegetation, we compared its distribution with that of Inula crithmoides, a taxonomically related halophyte, in three salt marshes located in "La Albufera" Natural Park, near the city of Valencia (East Spain). The presence of D. viscosa was restricted to areas of low and moderate salinity, while I. crithmoides was also present in the most saline zones of the salt marshes. Analyses of the responses of the two species to salt and water stress treatments in controlled experiments revealed that both activate the same physiological stress tolerance mechanisms, based essentially on the transport of toxic ions to the leaves-where they are presumably compartmentalized in vacuoles-and the accumulation of specific osmolytes for osmotic adjustment. The two species differ in the efficiency of those mechanisms: salt-induced increases in Na(+) and Cl(-) contents were higher in I. crithmoides than in D. viscosa, and the osmolytes (especially glycine betaine, but also arabinose, fructose and glucose) accumulated at higher levels in the former species. This explains the (slightly) higher stress tolerance of I. crithmoides, as compared to D. viscosa, established from growth inhibition measurements and their distribution in nature. The possible activation of K(+) transport to the leaves under high salinity conditions may also contribute to salt tolerance in I. crithmoides. Oxidative stress level-estimated from malondialdehyde accumulation-was higher in the less tolerant D. viscosa, which consequently activated antioxidant responses as a defense mechanism against stress; these responses were weaker or absent in the more tolerant I. crithmoides. Based on these results, we concluded that although D. viscosa cannot directly compete with true halophytes in highly saline environments, it is nevertheless quite stress tolerant and therefore represents a threat for the vegetation located on the salt marshes borders, where several endemic and threatened species are found in the area of study.

Stress tolerance mechanisms in Juncus: responses to salinity and drought in three Juncus species adapted to different natural environments.

Comparative studies on the responses to salinity and drought were carried out in three Juncus species, two halophytes (Juncus maritimus Lam. and Juncus acutus L.) and one more salt-sensitive (Juncus articulatus L.). Salt tolerance in Juncus depends on the inhibition of transport of toxic ions to the aerial part. In the three taxa studied Na+ and Cl- accumulated to the same extent in the roots of salt treated plants; however, ion contents were lower in the shoots and correlated with the relative salt sensitivity of the species, with the lowest levels measured in the halophytes. Activation of K+ transport at high salt concentration could also contribute to salt tolerance in the halophytes. Maintenance of cellular osmotic balance is mostly based on the accumulation of sucrose in the three species. Yet, neither the relative salt-induced increase in sugar content nor the absolute concentrations reached can explain the observed differences in salt tolerance. In contrast, proline increased significantly in the presence of salt only in the salt-tolerant J. maritimus and J. acutus, but not in J. articulatus. Similar patterns of osmolyte accumulation were observed in response to water stress, supporting a functional role of proline in stress tolerance mechanisms in Juncus.

Transcriptome analysis of Phoenix canariensis Chabaud in response to Rhynchophorus ferrugineus Olivier attacks.

UNLABELLED: Red Palm Weevil (RPW, Rhynchophorus ferrugineus Olivier) threatens most palm species worldwide. Until now, no studies have analyzed the gene regulatory networks of Phoenix canariensis (Chabaud) in response to RPW attacks. The aim of this study was to fill this knowledge gap. Providing this basic knowledge is very important to improve its management. RESULTS: A deep transcriptome analysis was performed on fully expanded leaves of healthy non-infested trees and attacked trees at two symptom stages (middle and late infestation). A total of 54 genes were significantly regulated during middle stage. Pathway enrichment analysis showed that phenylpropanoid-related pathways were induced at this stage. More than 3300 genes were affected during late stage of attacks. Higher transcript abundances were observed for lipid fatty acid metabolism (fatty acid and glycerolipids), tryptophan metabolism, phenylpropanoid metabolism. Key RPW-modulated genes involved in innate response mediated by hormone crosstalk were observed belonging to auxin, jasmonate and salicylic acid (SA) pathways. Among transcription factors, some WRKYs were clearly induced. qRT-PCR validation confirmed the upregulation of key genes chosen as validation of transcriptomic analysis. CONCLUSION: A subset of these genes may be further analyzed in future studies to confirm their specificity to be induced by RPW infestations.

Identification of Salt Stress Biomarkers in Romanian Carpathian Populations of Picea abies (L.) Karst.

The Norway spruce (Picea abies), the most important tree species in European forests, is relatively sensitive to salt and does not grow in natural saline environments. Yet many trees are actually exposed to salt stress due to the common practice of de-icing of mountain roads in winter, using large amounts of NaCl. To help develop strategies for an appropriate use of reproductive seed material on reforestation sites, ensuring better chances of seedling survival in salt-affected areas, we have studied the responses of young spruce seedlings to salt treatments. The specific aim of the work was to identify the optimal salt stress biomarkers in Picea abies, using as experimental material seedlings obtained by germination of seeds with origin in seven populations from the Romanian Carpathian Mountains. These responses included general, conserved reactions such as the accumulation of ions and different osmolytes in the seedlings needles, reduction in photosynthetic pigments levels, or activation of antioxidant systems. Although changes in the contents of different compounds involved in these reactions can be associated to the degree of stress affecting the plants, we propose that the (decreasing) levels of total phenolics or total carotenoids and the (increasing) levels of Na+ or K+ ions in Picea abies needles, should be considered as the most reliable and useful biomarkers for salt stress in this species. They all show very high correlation with the intensity of salt stress, independently of the genetic background of the seeds parental population, and relatively easy, quantitative assays are available to determine their concentrations, requiring simple equipment and little amount of plant material.