Purification and identification of barley (Hordeum vulgare L.) proteins that inhibit the alkaline serine proteinases of Fusarium culmorum

It has been proposed that microbial proteinase inhibitors, which are present in abundance in cereal grains, protect the seed against plant pathogens. So far, however, very little is known about the interactions of those inhibitors with the proteinases of phytopathogenic microbes. The increased alkaline proteinase activities of Fusarium head blight (FHB) diseased wheat and barley grain imply that the Fusarium fungi synthesize those enzymes during the colonization of the kernel. To study which barley proteins can inhibit Fusarium proteinases, and hence, possibly protect the seed from FHB, the proteins of a grain extract have been separated and tested for their abilities to inhibit two alkaline serine proteinases that we previously isolated from F. culmorum. The proteins were separated by size exclusion, ion exchange, and reversed-phase-HPLC chromatographies. The purified inhibitors were identified by their molecular masses and N-terminal amino acid sequences. The proteins that inhibited the subtilisin-like Fusarium proteinase were the chymotrypsin/subtilisin (CI) inhibitors 1A, 1B, and 2A and the barley alpha-amylase/subtilisin inhibitor (BASI). Only one of the purified proteins inhibited the trypsin-like proteinase, the barley Bowman-Birk inhibitor (BBBI). No novel inhibitors were detected.     

The barley (Hordeum vulgare L.) of Sardinia, Italy

Since ancient times, barley has been an important food resource for the people of Sardinia. The oldest traces of its cultivation are from the mid-Neolithic (fourth millennium B.C.). Archaeological, historical and anthropological aspects of barley cultivated in Sardinia are discussed in this paper. We describe the traditional process for making barley bread (orgiathu) in Sardinia, where a special starter called ghimisone was prepared. Today, barley is cultivated only as animal feed, with two uses, grain yield and grazing. Many farmers prefer to grow local populations belonging to landrace locally known as S' orgiu sardu. Local Sardinian populations of barley evolved in diverse environments, being cultivated from sea-level up to 1000 m elevation, on various soil types at different intensities of abiotic stresses, and with climates and environments associated with various agricultural practices, depending both on production strategies and climatic conditions. These barley materials are thought to be valuable genetic and cultural inheritance which must be preserved and used for both productive and research purposes.     

Regrowth in barley (Hordeum vulgare L.) and rye (Secale cereale L.)

Regrowth after cutting at four development stages, from heading to grain maturity, was investigated in a pot experiment containing three rye and four barley varieties. Regrowth in the barley varieties decreased strongly from heading to grain maturity. Rye generally showed stronger regrowth than barley after late cutting, but only the perennial variety ‘Soperta’ regenerated as many tillers at cutting as formed when harvested at the ripe grain stage. In both species, significant differences were found between varieties. The level of soluble carbohydrates reached a maximum between heading and maturity, but differences in regrowth could not be explained by such differences. Total N decreased from heading to maturity, except for perennial rye, where an increase was observed towards ripening. It is, however, uncertain whether this was an effect rather than a cause of the regrowth.     

Interactions of malt and barley (Hordeum vulgare L.) endoproteinases with their endogenous inhibitors.

For producing worts that are optimal for beer production, some, but not all, of the barley proteins must be degraded during malting and mashing. This protein hydrolysis is controlled by endoproteinases, and, in turn, is partially regulated by the presence of low-molecular-weight (LMW) proteinaceous inhibitors. This paper reports studies of the interactions between the proteinases and inhibitors and an "affinity" method for concentrating the inhibitors. The malt inhibitors (I) and proteinases (E) quickly formed strong (E-I) complexes when dissolved together, and all of the I was complexed. Heating at 100 degrees C, but not 70 degrees C, dissociated the complex, even though the enzyme activities were destroyed at 70 degrees C. The released I readily recomplexed with fresh E. Barley, however, contained insufficient E to complex all of its I complement. The E-I complex was treated with salts, detergents, and reducing agents to release active E molecules, but none disrupted the complex. By removing the LMW proteins from a malt E-I extract and dissociating the complex by heating, the concentration of I molecules was greatly increased. This "affinity" method can thus be used to concentrate the I molecules for further purification.     

Endosperm structure affects the malting quality of barley (Hordeum vulgare L.)

Twenty-seven barley (Hordeum vulgare L.) samples collected from growing sites in Scandinavia in 2001 and 2002 were examined to study the effect of endosperm structure on malting behavior. Samples were micromalted, and several malt characteristics were measured. Samples were classified as having a mealier or steelier endosperm on the basis of light transflectance (LTm). Because endosperm structure is greatly dependent on protein content, three barley sample pairs with similar protein contents were chosen for further analysis. During malting, the steelier barley samples produced less root mass, but showed higher respiration losses and higher activities of starch-hydrolyzing enzymes. Malts made from steelier barley had a less friable structure, with more urea-soluble D hordein and more free amino nitrogen and soluble protein. The reason for these differences may lie in the structure or localization of the hordeins as well as the possible effects of endosperm packing on water uptake and movement of enzymes.     

Metabolomic profiling of five hulless barley (Hordeum vulgare L.) with different

Genetic Resources and Crop Evolution - Colored hulless barley (Hordeum vulgare L.) is a high-quality germplasm resource rich in nutrients, such as protein, β-glucan, flavonoids, amino acids,...     

Identification and selection of low-phosphate-tolerant germplasm in barley (Hordeum vulgare L.)

Phosphorus (P) deficiency is one of the major constraints to crop yield worldwide, and genotypes or cultivars with high phosphate use efficiency (PUE) sustain growth when exposed to phosphate stress. Therefore, it is imperative to develop the genotypes or cultivars with high PUE. A pot experiment was conducted to evaluate the PUE among 150 barley (Hordeum vulgare L.) genotypes. Two high-tolerant and -sensitive accessions were selected. These two candidate materials were used to investigate the differences among the root morphology characteristics, antioxidant enzyme activity, inorganic phosphate (Pi) content and gene expression of HvPT5 under P-deficiency and P-sufficiency conditions. The values of these parameters were higher in the low-P-tolerant genotype than in the sensitive one. In pot experiment 1, all genotypes showed a significant difference in low-P tolerance, with variety GN121 achieving the highest tolerance, and GN42 being most sensitive. The results of this study may provide elite genetic germplasms for future work on isolation of P-related genes, and the improvement of PUE in barley.

Abbreviations: PUE: phosphate use efficiency; CAT: catalase; POD: peroxidase; SOD: superoxide dismutase; DMSO: dimethyl sulphoxide; MDA: malondialdehyde; TOPSIS: technique for order preference by similarity to an ideal solution; MCDM/MADM: multi-criteria (or attribute) decision making     

Potassium placement effect on dynamics of barley (Hordeum vulgare L.) nutrition

The effects of local placement of potassium (K) on mineral nutrition dynamics of barley (Hordeum vulgare L.) in fertile Сhernozem were studied. A pot experiment with local K-placement at 4–5 cm soil depth was carried out and the dynamics of nitrogen, phosphorus, potassium (NPK) concentrations in tillers, organs and parts of spring barley was measured. K-placement increased the productivity index from 0.49 to 0.54, despite optimal and slightly varying NPK concentrations during the second half of the vegetation period (60–100 d). This occurs due to partitioning of assimilates, N, K, and especially P in generative organs of primary and secondary tillers forming quality grains. Nutrient concentrations in certain primary tiller parts of a 60-d plant (senescing leaves and the main spike axis) proved to be more sensitive indicators of the K-placement effect than average whole-plant characteristics. While being beneficial, K-placement had little impact on the overall NPK removal in barley, which shows a significant role of factors related to K uptake kinetics. Thus, the chosen parameters in the soil–plant system (the high content of available K in Chernozem, in the second part of the vegetation period) have for the first time allowed the evaluation of the impact of local K-placement on mineral nutrition dynamics in barley.     

Bacteria and fungi on roots of different barley varieties (Hordeum vulgare L.)

Summary We investigated the abundance of bacteria and fungi on roots of different barley varieties grown in soil and in a nutrient solution. Measurements were made on the rhizoplane and, for soil-grown plants, also in the rhizosphere soil. Further, the influence of plant age was investigated. Barley variety, had a significant influence both for plants grown in soil and in the nutrient solution, and the effects were most pronounced on the rhizoplane. There were no significant differences among varieties in fungal hyphal lengths on the roots. Bacterial abundance on the rhizoplane was significantly decreased with increasing plant age. Varietal differences were maintained over different plant ages.     

Biological nitrogen fixation associated with roots of field-grown barley (Hordeum vulgare L.)

Nitrogenase (C₂H₂) activity was measured in microbial media inoculated with barley root segments or corresponding rhizosphere soil. Three different media were used, Döbereiner's malate medium, a modified Ashby medium, and an acid nitrogen-free medium. Only Döbereiner's medium gave consistently positive results, and cultures inoculated with roots showed higher activity than cultures inoculated with corresponding rhizosphere soil. Similar experiments with roots and rhizosphere soil from wheat gave only negligible nitrogenase activity, whereas the tropical grass, Cynodon dactylon, gave higher activity than barley. Measurements on intact soil cores containing barley root systems showed an initial lag phase followed by a rather stable activity level over a period from 12 h to 48 h, and then the activity again decreased. The activity during the stable period corresponded to fixation of about 100 to 200 g N₂ ha^?1 24 h^?1. Measurements on isolated, washed barley roots showed only negligible nitrogenase activity.     

Multiple element concentration in the grain of spring barley (Hordeum vulgare L.) collection

Multiple element analyses were carried out to investigate variation in element concentrations in barley grains of 336 genotypes. Of 13 elements analyzed, Ba ranged from 0.2 to 8.9?mg kg^?1, Ca from 186.4 to 977.5?mg kg^?1, Cu from 1.5 to 9.8?mg kg^?1, K from 353.2 to 7721.5?mg kg^?1, Mg from 1049.8 to 2024.2?mg kg^?1, Mn from 8.1 to 22.9?mg kg^?1, Na from 55.9 to 627.9?mg kg^?1, P from 2272.9 to 5428.8?mg kg^?1, S from 880.7 to 1898.0?mg kg^?1, Si from 19.1 to 663.2?mg kg^?1, and Sr from 0.35 to 2.62?mg kg^?1 in the barley grain. The least square means showed high Zn, Fe, Mg, P, and S concentration in AM-64 and AM-228 genotypes. The principal component analysis of element concentration showed four PCs explained 64.3% total variance. Strong positive correlations (p?<?0.001) of Fe-Mn, Fe-S, S-Mn, Zn-P, Zn-Mg, Mg-P, Mg-Mn, and Ca-Sr were found. The identification barley genotypes that showed high elements concentration furnish valuable genetic resources for biofortification in future.     

Molecular classification of barley (Hordeum vulgare L.) mutants using derivative NIR spectroscopy

Near-infrared reflectance (NIR) spectroscopy was used in the characterization of grain morphology mutants of barley ( Hordeum vulgare L.) in relation to grain nitrogen (N) content and protein composition. Derivative spectroscopy provided spectra with enhanced resolution, allowing wavelengths to be identified with clear differences in contribution from associated chemical bonds. Comparisons of fourth-derivative spectra of wholemeal flour from high-N grains with flour from low-N grains identified wavelengths at which there were statistically significant differences between the groups. Their importance was independently confirmed by step-up regression using these wavelengths to generate an equation predicting N content (R(2) = 0.98). Fourth-derivative spectral comparisons also allowed novel biochemical differences to be predicted. Visual assessment of the spectra of all mutants revealed a variable region (1470-1520 nm, corresponding to N-H stretch vibrations) that allowed two extreme sets to be defined. The protein extracted from these two sets differed markedly in hordein content.     

Crop stand density enhances competitive ability of spring barley (Hordeum vulgare L.)

Abstract

The aim of the research was to establish weediness, competitive ability and productivity of the crop. The experimental object was agrophytocenoses of spring barley – Hordeum vulgare L. – crop of spring barley ‘Aura’ and unsown soil, and weeds growing in them. The crop was formed sowing 0, 120, 200 and 280 kg ha^?1 (0, 2.7, 4.5 and 6.2 million seeds per ha^?1 respectively) seeds of spring barley ‘Aura’. Spring barley crop was not harrowed and herbicides were not applied. In the field experiment estimates were made of changes of weeds and spring barley inter- and within- species competition optimizing crop density. During three years of field experiment in the crop of spring barley annual weeds prevailed at 88–99%, such as Chenopodium album, Stellaria media, Erysimum cheiranthoides. Perennial weeds formed 1–12% of the crop weeds, such as Sonchus arvensis, Cirsium arvense, Equisetum arvense. General number of weed species in spring barley crops varied from 13 to 21. Weed abundance proportionally declined in the crops of higher density, hence, higher seed rate should be recommended for organic agriculture where weeds are controlled in non-chemical ways. Consistently increasing barley stand density, the competition between species (spring barley with weeds) gradually turned into competition within species (between barley plants) when a higher number of weaker and non-productive stems started forming. Spring barley yield did not significantly depend on the stand density. Increasing stand density enhanced cultivated crop yield to a certain level (200 kg ha^?1), since an increase in spring barley plant number resulted in the reduction in weight per plant and 1000 grain weight, which was compensated by an increase in the number of spring barley plants. Different spring barley density had an essential influence on the chemical composition of weeds which was similar to that of spring barley. Weeds accumulated the greatest amount of crude proteins, crude fat and crude fibre growing without spring barley.     

Distribution of microbial biomass across the rhizosphere of barley (Hordeum vulgare L.) in soils

Summary The microbial activity at the soil-root interface (rhizosphere) of barley was examined using a rhizobox system. In this system, the soil was placed in several compartments separated from each other by a 500-mesh nylon cloth. Plants were grown in the central compartment and after a 2-month growing period the roots were still confined to this compartment. The soil from each compartment was then analyzed for ATP, NO₃ ^/–, total N, total C and CO₂ production. The increase in ATP concentration was found in a range of 4 mm around the roots. The ATP content and CO₂ production across the rhizosphere were correlated in all the soils used, but changes in NO₃ ^– were not correlated with ATP changes. The range of NO₃ ^– change was wider than that of the ATP change, indicating that NO₃ ^– production is not influenced by the biological activity in the rhizosphere.     

In vivo modeling of beta-glucan degradation in contrasting barley (Hordeum vulgare L.) genotypes

An important determinative of malt quality is the malt beta-glucan content, which in turn depends on the initial barley beta-glucan content as well as the beta-glucan depolymerization by beta-glucanase (EC 3.2.1.73) during malting. Another enzyme, named beta-glucan solubilase, has been suggested to act prior to beta-glucanase; its existence, however, has not been unequivocally proven. We monitored changes in beta-glucan levels and in the development of beta-glucan-degrading enzymes during malting of five lots of contrasting barley genotypes. Two models of in vivo kinetics for beta-glucan degradation were then compared as follows: (i) a biphasic model based on the sequential action of beta-glucan solubilase and beta-glucanase and (ii) a monophasic model assuming that all beta-glucans are depolymerized by beta-glucanase without the previous intervention of another enzyme. Confirmatory regression analysis was used to test the fit of the models to the observed data. Our results show that beta-glucan degradation is mostly monophasic, although some enzyme other than beta-glucanase seems to be required for the early solubilization of a small fraction of insoluble beta-glucans (on average, 7% of total beta-glucans). Furthermore, the genotype-dependent kinetic rate constant (indicating beta-glucan degradability), in addition to beta-glucanase activity, is suggested to play a major role in malting quality.     

Physiological response of culture media-grown barley (Hordeum vulgare L.) to titanium oxide nanoparticles

Since the fate of nanoparticles after their release in the environment and their possible transfer in plants and subsequent impacts is still largely unknown, this paper evaluates the potential phytotoxic effects of up to 20% w/w TiO₂ nanoparticles (nTiO₂) on barley cultivated in hydroponics and agar media. The X-ray diffraction analysis confirmed that nTiO₂ powder corresponds to anatase phase. On agar medium only high concentrations of nTiO₂ (10% and 20% w/w) induced significant inhibition of shoot growth. However, hydroponics treatment with nTiO₂ up to 1000?mg?L^?1 did not show any adverse effect on the shoot growth. In both experiments, (i) root growth inhibition effects became visible with increasing concentration of nTiO₂, (ii) plants treated with nTiO₂ showed no change in chlorophyll a and b content, even though the plants absorbed nTiO₂, (iii) weight of biomass treated with nTiO₂ was not significantly different compared to control. Therefore, we assume that transport of nTiO₂ into the aerial parts is limited due to the presence of effective mechanical or physiological barriers in roots. Overall, it appears that early root growth is a relevant indicator of potential effects of nTiO₂ exposure. Our results also indicate that synthesized nTiO₂ are not significantly toxic to the barley when applied at the concentrations used in this work, even though plants absorb titanium.     

Nitrogenase activity (Acetylene reduction) during the growth cycle of spring barley (Hordeum vulgare L.)

The nitrogenase activity (C₂H₂-reduction) was measured during the growth cycle of field grown spring barley in soil cores both with and without barley plants, and at two levels of nitrogen application, 30 and 120 kg N ha^?1 year^?1 respectively. The main purpose of the investigation was to study the effects of the growing barley plants on nitrogenase activity in the soil, and temperature and moisture contents were kept constant in all experiments. Therefore, the results cannot be used to calculate actual amounts of fixed nitrogen in the field, but should be considered rather as potential values. The nitrogenase activity was found to vary during the growth cycle, and seemed to be correlated to the photosynthetic activity of the plants. Relatively low nitrogenase activity was found in the early growth stages, and the activity increased up to a maximum in the late reproductive stage, followed by a rapid decrease during the grain filling stage. The mean values of nitrogenase activity in samples without barley plants and with barley plants were 40 and 78 nmoles C₂H₄ g soil dwt^?1 24 h^?1 respectively. The positive effect of barley plants on nitrogenase activity was stronger at 120 kg N than at 30 kg N fertilization. As a mean of the whole growth cycle the ratio between samples with and without barley plants was 1.7 with 30 kgN and 2.3 with 120 kg N fertilization. The inhibitory effect of nitrogen application on nitrogenase activity was measurable until 6–7 weeks after application, and it was strongest in cores without plants.     

Uptake of Heavy Metals by Mycorrhizal Barley (Hordeum vulgare L.)

It has been previously indicated that arbuscular mycorrhizal (AM) fungi can enhance the bioremediation abilities of their host plant. Barley (Hordeum vulgare L.) is a crop plant with some unique physiological properties, such as tolerance to salinity. However, its tolerance to other stresses such as heavy metals must be tested. Accordingly, it was hypothesized that barley can be efficiently used to treat heavy metals in symbiotic and non-symbiotic association with AM fungi. In a greenhouse experiment barley plants were inoculated with the AM species Glomus mosseae and grown in a soil polluted with cadmium (Cd), cobalt (Co), and lead (Pb). Relative to Cd and Co, mycorrhizal barley absorbed significantly higher amounts of Pb. AM species also significantly decreased Cd and Co uptake by barley indicating the alleviating effects of G. mosseae on the stress of such heavy metals.     

Trait phenotyping and molecular marker characterization of barley (Hordeum vulgare L.) germplasm from Western Himalayas

Genetic Resources and Crop Evolution - Barley (Hordeum vulgare L.) is one of the principal cereal crops grown in Western-Himalayas of India. A set of 105 barley&nbsp;genotypes including 38...