The identification of QTL controlling ergot sclerotia size in hexaploid wheat implicates a role for the Rht dwarfing alleles

The fungal pathogen Claviceps purpurea infects ovaries of a broad range of temperate grasses and cereals, including hexaploid wheat, causing a disease commonly known as ergot. Sclerotia produced in place of seed carry a cocktail of harmful alkaloid compounds that result in a range of symptoms in humans and animals, causing ergotism. Following a field assessment of C. purpurea infection in winter wheat, two varieties ‘Robigus’ and ‘Solstice’ were selected which consistently produced the largest differential effect on ergot sclerotia weights. They were crossed to produce a doubled haploid mapping population, and a marker map, consisting of 714 genetic loci and a total length of 2895 cM was produced. Four ergot reducing QTL were identified using both sclerotia weight and size as phenotypic parameters; QCp.niab.2A and QCp.niab.4B being detected in the wheat variety ‘Robigus’, and QCp.niab.6A and QCp.niab.4D in the variety ‘Solstice’. The ergot resistance QTL QCp.niab.4B and QCp.niab.4D peaks mapped to the same markers as the known reduced height (Rht) loci on chromosomes 4B and 4D, Rht-B1 and Rht-D1, respectively. In both cases, the reduction in sclerotia weight and size was associated with the semi-dwarfing alleles, Rht-B1b from ‘Robigus’ and Rht-D1b from ‘Solstice’. Two-dimensional, two-QTL scans identified significant additive interactions between QTL QCp.niab.4B and QCp.niab.4D, and between QCp.niab.2A and QCp.niab.4B when looking at sclerotia size, but not between QCp.niab.2A and QCp.niab.4D. The two plant height QTL, QPh.niab.4B and QPh.niab.4D, which mapped to the same locations as QCp.niab.4B and QCp.niab.4D, also displayed significant genetic interactions.

Effects of aphid infestation on Cd and Zn concentration in wheat

The effect of infestation by the aphid Metopolophium dirhodum on the concentration and mass partitioning of Cd and Zn was studied in wheat plants. Results show that infestation did not affect the concentration of either metal in the roots or shoots of wheat, but elevated concentrations in the ears. This appeared to result from the concentration of metal in the smaller ear mass of infested plants. Infestation did not significantly affect the mass partitioning of either metal in any part of the plant, some 10% of both metals being allocated to the roots. However, the two metals contrasted in their partitioning in the aerial parts of the plant, with ca. 60% of Cd mass partitioned in the shoots and ca. 67% of Zn mass partitioned in the ears. The possible effects of infestation on the transfer of Cd and Zn from the soil to cereal aphids are discussed.

Extractability and characteristics of proteins deriving from wheat DDGS

Wheat Distillers’ Dried Grains with Solubles (DDGS) and in-process samples were used for protein extraction. Prolamins were the predominant protein components in the samples. The absence of extractable α- and γ-gliadins in DDGS indicated protein aggregation during the drum drying processing stage. Prolamin extraction was performed using 70% (v/v) ethanol or alkaline-ethanol solution in the presence of reducing agent. DDGS extracts had relatively low protein contents (14-44.9%, w/w), regardless of the condition applied. The wet solids were the most suitable raw material for protein extraction, with recovery yields of ~ 55% (w/w) and protein content of ~58% (w/w) in 70% (v/v) ethanol. Protein extracts from wet solids were significantly rich in glutamic acid and proline. Mass balance calculations demonstrated the high carbohydrate content (~ 50%, w/w) of solid residues. Overall, the feasibility of utilising in-process samples of DDGS for protein extraction with commercial potential was demonstrated.

Valorisation of side streams from wheat milling and confectionery industries for consolidated production and extraction of microbial lipids

Crude enzymes produced via solid state fermentation (SSF) using wheat milling by-products have been employed for both fermentation media production using flour-rich waste (FRW) streams and lysis of Rhodosporidium toruloides yeast cells. Filter sterilization of crude hydrolysates was more beneficial than heat sterilization regarding yeast growth and microbial oil production. The initial carbon to free amino nitrogen ratio of crude hydrolysates was optimized (80.2 g/g) in fed-batch cultures of R. toruloides leading to a total dry weight of 61.2 g/L with microbial oil content of 61.8 % (w/w). Employing a feeding strategy where the glucose concentration was maintained in the range of 12.2 – 17.6 g/L led to the highest productivity (0.32 g/L∙h). The crude enzymes produced by SSF were utilised for yeast cell treatment leading to simultaneous release of around 80% of total lipids in the broth and production of a hydrolysate suitable as yeast extract replacement.

Distribution of lipids in the grain of wheat (cv Hereward) determined by lipidomic analysis of milling and pearling fractions

Lipidomic analyses of milling and pearling fractions from wheat grain were carried out to determine differences in composition which could relate to the spatial distribution of lipids in the grain. Free fatty acids and triacylglycerols were major components in all fractions, but the relative contents of polar lipids varied, particularly lysophosphatidyl choline and digalactosyldiglyceride, which were enriched in flour fractions. By contrast, minor phospholipids were enriched in bran and offal fractions. The most abundant fatty acids in the analysed acyl lipids were C16:0 and C18:2 and their combinations, including C36:4 and C34:2. Phospholipids and galactolipids have been reported to have beneficial properties for bread making, while free fatty acids and triacylglycerols are considered detrimental. The subtle differences in the compositions of fractions determined in the present study could therefore underpin the production of flour fractions with optimised compositions for different end uses.

Effect of simulated rainfall during wheat seed development and maturation on subsequent seed longevity is reversible

Poor wheat seed quality in temperate regions is often ascribed to wet production environments. We investigated the possible effect of simulated rain during seed development and maturation on seed longevity in wheat (Triticum aestivum L.) cv. Tybalt grown in the field (2008, 2009) or a polythene tunnel house (2010). To mimic rain, the seed crops were wetted from above with the equivalent of 30mm (2008, 2009) or 25mm rainfall (2010) at different stages of seed development and maturation (17 to 58 DAA, days after 50% anthesis), samples harvested serially, and subsequent air-dry seed longevity estimated. No pre-harvest sprouting occurred. Seed longevity (p50, 50% survival period in experimental hermetic storage at 40°C with c. 15% moisture content) in field-grown controls increased during seed development and maturation attaining maxima at 37 (2008) or 44 DAA (2009); it declined thereafter. Immediate effects of simulated rain at 17-58 DAA in field studies (2008, 2009) on subsequent seed longevity were negative but small, e.g. a 1-4 d delay in seed quality improvement for treatments early in development but with no damage detected at final harvests. In rainfall-protected conditions (2010), simulated rain close to harvest maturity (55-56 DAA) reduced longevity immediately and substantially, with greater damage from two sequential days of wetting than one; again, later harvests provided evidence of recovery in subsequent longevity. In the absence of pre-harvest sprouting, the potentially deleterious effects of rainfall to wheat seed crops on subsequent seed longevity may be reversible in full or in part.

Decimal growth stages for precision wheat production in changing environments?

The utility of the decimal growth stage (DGS) scoring system for cereals is reviewed. The DGS is the most widely used scale in academic and commercial applications because of its comprehensive coverage of cereal developmental stages, the ease of use and definition provided and adoption by official agencies. The DGS has demonstrable and established value in helping to optimise the timing of agronomic inputs, particularly with regard to plant growth regulators, herbicides, fungicides and soluble nitrogen fertilisers. In addition, the DGS is used to help parameterise crop models, and also in understanding the response and adaptation of crops to the environment. The value of the DGS for increasing precision relies on it indicating, to some degree, the various stages in the development of the stem apex and spike. Coincidence of specific growth stage scores with the transition of the apical meristem from a vegetative to a reproductive state, and also with the period of meiosis, is unreliable. Nonetheless, in pot experiments it is shown that the broad period of booting (DGS 41–49) appears adequate for covering the duration when the vulnerability of meiosis to drought and heat stress is exposed. Similarly, the duration of anthesis (61–69) is particularly susceptible to abiotic stresses: initially from a fertility perspective, but increasingly from a mean grain weight perspective as flowering progresses to DGS 69 and then milk development. These associations with DGS can have value at the crop level of organisation: for interpreting environmental effects, and in crop modelling. However, genetic, biochemical and physiological analysis to develop greater understanding of stress acclimation during the vegetative state, and tolerance at meiosis, does require more precision than DGS can provide. Similarly, individual floret analysis is needed to further understand the genetic basis of stress tolerance during anthesis.

Photoperiod sensitivity affects flowering duration in wheat

Flowering and successful pollination in wheat are key determinants of both quantity and quality of grain. Bread wheat line ‘Paragon’, introgressed with single or multiple day length insensitivity alleles was used to dissect the effects on the timing and duration of flowering within a hierarchical plant architecture. Flowering of wheat plants was observed in a series of pot-based and field experiments. Ppd-D1a was the most potent known allele affecting the timing of flowering, requiring the least thermal time to flowering across all experiments. The duration of flowering for individual lines was dominated by the shift in the start of flowering in later tillers and the number of tillers per plant, rather than variation in flowering duration of individual spikes. There was a strong relationship between flowering duration and the start of flowering with the earliest lines flowering for the longest. The greatest flowering overlap between tillers was recorded for the Ppd-1b. Across all lines, a warmer environment significantly reduced the duration of flowering and the influence of Ppd-1a alleles on the start of flowering. These findings provide evidence of pleiotropic effects of the Ppd-1a alleles, and have direct implications for breeding for increased stress resilient wheat varieties.

Investigation and optimization of parameters affecting the multiply charged ion yield in AP-MALDI MS

Liquid matrix-assisted laser desorption/ionization (MALDI) allows the generation of predominantly multiply charged ions in atmospheric pressure (AP) MALDI ion sources for mass spectrometry (MS) analysis. The charge state distribution of the generated ions and the efficiency of the ion source in generating such ions crucially depend on the desolvation regime of the MALDI plume after desorption in the AP-tovacuum inlet. Both high temperature and a flow regime with increased residence time of the desorbed plume in the desolvation region promote the generation of multiply charged ions. Without such measures the application of an electric ion extraction field significantly increases the ion signal intensity of singly charged species while the detection of multiply charged species is less dependent on the extraction field. In general, optimization of high temperature application facilitates the predominant formation and detection of multiply charged compared to singly charged ion species. In this study an experimental setup and optimization strategy is described for liquid AP-MALDI MS which improves the ionization effi- ciency of selected ion species up to 14 times. In combination with ion mobility separation, the method allows the detection of multiply charged peptide and protein ions for analyte solution concentrations as low as 2 fmol/lL (0.5 lL, i.e. 1 fmol, deposited on the target) with very low sample consumption in the low nL-range.

Does fertilizer type and method of application cause significant differences in essential oil yield and composition in rosemary (Rosmarinus officinalis L.)?

Organic fertilizers based on seaweed extract potentially have beneficial effects on many crop plants. Herewe investigate the impact of organic fertilizer on Rosmarinus officinalis measured by both yield and oilquality. Plants grown in a temperature-controlled greenhouse with a natural photoperiod and a controlledirrigation system were treated with seaweed fertilizer and an inorganic fertilizer of matching mineralcomposition but with no organic content. Treatments were either by spraying on to the foliage or wateringdirect to the compost. The essential oil was extracted by hydro-distillation with a Clevenger apparatusand analysed by gas-chromatography mass-spectrometry (GC–MS) and NMR. The chemical composi-tions of the plants were compared, and qualitative differences were found between fertilizer treatmentsand application methods. Thus sprayed seaweed fertilizer showed a significantly higher percentage of�-pinene, �-phellandrene, �-terpinene (monoterpenes) and 3-methylenecycloheptene than other treat-ments. Italicene, �-bisabolol (sesquiterpenes), �-thujene, and E-isocitral (monoterpenes) occurred insignificantly higher percentages for plants watered with the seaweed extract. Each was significantly dif-ferent to the inorganic fertilizer and to controls. The seaweed treatments caused a significant increasein oil amount and leaf area as compared with both inorganic treatments and the control regardless ofapplication method.

High-throughput phenotyping (HTP) identifies seedling root traits linked to variation in seed yield and nutrient capture in field-grown oilseed rape (Brassica napusL.)

Background and Aims Root traits can be selected for crop improvement. Techniques such as soil excavations can be used to screen root traits in the field, but are limited to genotypes that are well-adapted to field conditions. The aim of this study was to compare a low-cost, high-throughput root phenotyping (HTP) technique in a controlled environment with field performance, using oilseed rape (OSR; Brassica napus) varieties. Methods Primary root length (PRL), lateral root length and lateral root density (LRD) were measured on 14-d-old seedlings of elite OSR varieties (n = 32) using a ‘pouch and wick’ HTP system (∼40 replicates). Six field experiments were conducted using the same varieties at two UK sites each year for 3 years. Plants were excavated at the 6- to 8-leaf stage for general vigour assessments of roots and shoots in all six experiments, and final seed yield was determined. Leaves were sampled for mineral composition from one of the field experiments. Key Results Seedling PRL in the HTP system correlated with seed yield in four out of six (r = 0·50, 0·50, 0·33, 0·49; P < 0·05) and with emergence in three out of five (r = 0·59, 0·22, 0·49; P < 0·05) field experiments. Seedling LRD correlated positively with leaf concentrations of some minerals, e.g. calcium (r = 0·46; P < 0·01) and zinc (r = 0·58; P < 0·001), but did not correlate with emergence, general early vigour or yield in the field. Conclusions Associations between PRL and field performance are generally related to early vigour. These root traits might therefore be of limited additional selection value, given that vigour can be measured easily on shoots/canopies. In contrast, LRD cannot be assessed easily in the field and, if LRD can improve nutrient uptake, then it may be possible to use HTP systems to screen this trait in both elite and more genetically diverse, non-field-adapted OSR.

Using NMR-based metabolomics to evaluate postprandial urinary responses following consumption of minimally processed wheat bran or wheat aleurone by men and women

Wheat bran, and especially wheat aleurone fraction, are concentrated sources of a wide range of components which may contribute to the health benefits associated with higher consumption of whole-grain foods. This study used NMR metabolomics to evaluate urine samples from baseline at one and two hours postprandially, following the consumption of minimally processed bran, aleurone or control by 14 participants (7 Females; 7 Males) in a randomized crossover trial. The methodology discriminated between the urinary responses of control, and bran and aleurone, but not between the two fractions. Compared to control, consumption of aleurone or bran led to significantly and substantially higher urinary concentrations of lactate, alanine, N-acetylaspartate acid and N-acetylaspartylglutamate and significantly and substantially lower urinary betaine concentrations at one and two hours postprandially. There were sex related differences in urinary metabolite profiles with generally higher hippurate and citrate and lower betaine in females compared to males. Overall, this postprandial study suggests that acute consumption of bran or aleurone is associated with a number of physiological effects that may impact on energy metabolism and which are consistent with longer term human and animal metabolomic studies that used whole-grain wheat diets or wheat fractions.

Wheat seed embryo excision enables the creation of axenic seedlings and Koch’s postulates testing of putative bacterial endophytes

Early establishment of endophytes can play a role in pathogen suppression and improve seedling development. One route for establishment of endophytes in seedlings is transmission of bacteria from the parent plant to the seedling via the seed. In wheat seeds, it is not clear whether this transmission route exists, and the identities and location of bacteria within wheat seeds are unknown. We identified bacteria in the wheat (Triticum aestivum) cv. Hereward seed environment using embryo excision to determine the location of the bacterial load. Axenic wheat seedlings obtained with this method were subsequently used to screen a putative endophyte bacterial isolate library for endophytic competency. This absence of bacteria recovered from seeds indicated low bacterial abundance and/or the presence of inhibitors. Diversity of readily culturable bacteria in seeds was low with 8 genera identified, dominated by Erwinia and Paenibacillus. We propose that anatomical restrictions in wheat limit embryo associated vertical transmission, and that bacterial load is carried in the seed coat, crease tissue and endosperm. This finding facilitates the creation of axenic wheat plants to test competency of putative endophytes and also provides a platform for endophyte competition, plant growth, and gene expression studies without an indigenous bacterial background.

Effects of rain shelter or simulated rain during grain filling and maturation on subsequent wheat grain quality in the UK

The effects of simulated additional rain (ear wetting, 25 mm) or of rain shelter imposed at different periods after anthesis on grain quality at maturity and the dynamics of grain filling and desiccation were investigated in UK field-grown crops of wheat (Triticum aestivum L., cvar Tybalt) in 2011 and in 2012 when June–August rainfall was 255.0 and 214.6 mm, respectively, and above the decadal mean (157.4 mm). Grain filling and desiccation were quantified well by broken-stick regressions and Gompertz curves, respectively. Rain shelter for 56 (2011) or 70 d (2012) after anthesis, and to a lesser extent during late maturation only, resulted in more rapid desiccation and hence progress to harvest maturity whereas ear wetting had negligible effects, even when applied four times. Grain-filling duration was also affected as above in 2011, but with no significant effect in 2012. In both years, there were strong positive associations between final grain dry weight and duration of filling. The treatments affected all grain quality traits in 2011: nitrogen (N) and sulphur (S) concentrations, N:S ratio, sodium dodecyl sulphate (SDS) sedimentation volume, Hagberg Falling Number (HFN), and the incidence of blackpoint. Only N concentration and blackpoint were affected significantly by treatments in 2012. Rain shelter throughout grain filling reduced N concentration, whereas rain shelter reduced the incidence of blackpoint and ear wetting increased it. In 2011, rain shelter throughout reduced S concentration, increased N:S ratio and reduced SDS. Treatment effects on HFN were not consistent within or between years. Nevertheless, a comparison between the extreme treatment means in 2012 indicated damage from late rain combined with ear wetting resulted in a reduction of c. 0.7 s in HFN/mm August rainfall, whilst that between samples taken immediately after ear wetting at harvest maturity or 7 d later suggested recovery from damage to HFN upon re-drying in planta. Hence, the incidence of blackpoint was the only grain quality trait affected consistently by the diverse treatments. The remaining aspects of grain quality were comparatively resilient to rain incident upon developing and maturing ears of cvar Tybalt. No consistent temporal patterns of sensitivity to shelter or ear wetting were detected for any aspect of grain quality.