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Abstract: Breeding has enabled the development of wheat genotypes with different quality characteristics and end uses. Despite its numerous positive properties that make it one of the most cultivated crops, wheat is known to induce allergic reactions in predisposed consumers. Genetic modification (GM) technology for crop improvement has recently emerged and its impact on allergenicity must be evaluated, as recommended by the Codex Alimentarius. Our aim was to determine whether the variation in the amount of allergenic polypeptides in five GM wheats, along with their untransformed genotypes, was equivalent to the variation observed among twenty commercial cultivars, either durum or bread wheats. Since the most important factor involved is the amount of allergenic polypeptides, we performed Enzyme-Linked Immunoabsorbent Assays with IgE from twenty-two patients suffering from food or respiratory allergy to wheat on two well characterised soluble protein fractions. Statistical analyses showed a significant effect of the genotypes and sera, both by considering GM lines and untransformed genotypes. This study leads us to conclude that a wide variation exists in the amount of allergenic polypeptides among durum and bread wheat cultivars, and that the differences observed between GM wheats and their parents are within the range of cultivated wheats. [Copyright &y& Elsevier]

A new wheat endosperm protein subunit that was found in accessions belonging to different collections was identified by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Insoluble in 0·5 M NaCl, 70% ethanol, dimethyl sulphoxide (DMSO) and 50% propan-1-ol, it appeared in the pellet corresponding to the polymeric proteins along with high (HMW) and low molecular weight (LMW) glutenin subunits (GS). In the reduced form, it had an electrophoretic mobility between those two types of glutenin subunits. The apparent Mr of this novel protein was estimated by SDS-PAGE to be 71 000. N-terminal sequence and amino acid analyses indicated a composition similar to the ω-gliadins encoded by genes located on chromosome 1B. This protein can be ascribed to the D-subunits of LMW-GS with at least one cysteine residue that allows it to form part of the polymeric structure of glutenin, as shown by reaction with a fluorogenic reagent specific for sulphydryl groups. Fractions collected after size exclusion high-performance liquid chromatography (SE-HPLC) fractionation and further characterised by SDS-PAGE, confirm that the protein participates in the glutenin polymeric structure. An increase in its concentration was observed in fractions collected within the polymeric peak as elution time increased, implying that a larger amount of this protein is present in small size polymers. The role of this protein in the complex relationship between endosperm proteins and quality parameters is discussed in relation to its likely role as a chain terminator. [Copyright &y& Elsevier]

SDS-PAGE analysis of seed proteins of the cultivar 'Red River 68' showed a considerably higher staining intensity of the band corresponding to HMW-GS Bx7 relative to the equivalent band in the cultivars `Chinese Spring' and `Cheyenne'. Southern blots of restriction enzyme fragments from DNA of these three cultivars were analyzed densitometrically to reveal that the band corresponding to the Bx7 gene of `Red River 68' had a double staining intensity compared to the equivalent bands from the other two cultivars, which indicates that in `Red River 68' a duplication of the Bx7 gene has occurred. Although the possibility of the gene copy being a pseudogene was not ruled out, the greater amount of protein corresponding to Bx7 in `Red River 68' most likely is in accord with an increase in active gene number. SDS-PAGE analysis of the proteins showed also that the mobility of Bx7 in 'Cheyenne' was slightly different from the mobilities of the Bx7 subunits of 'Red River 68' and `Chinese Spring'. The same difference was observed at the gene level by PCR amplification of the genes encoding these subunits. [ABSTRACT FROM AUTHOR]

Two patients suffering from eosinophilic gastroenteritis (EG) were treated with sodium cromoglycate (SCG). Before treatment they showed enteric and cutaneous symptoms, such as abdominal pain, nausea, vomiting, diarrhoea and recurrent urticaria and angioedema. The histological findings were a notable amount of eosinophilic infiltration in the lamina propria and gastric glands, a villous shortening and thickening and weak eosinophilic inflammation in the duodenum. The patients were treated with 300 mg SCG, 4 times daily, for ⅘ months. During treatment, the clinical symptoms disappeared and at the end of treatment a reduced inflammation with an almost complete decrease of eosinophilic infiltration was observed. The results provide evidence of SCG efficacy in the treatment of EG and suggest its employment as an alternative to the steroids commonly used in EG. [ABSTRACT FROM AUTHOR]

An abstract of the article "Delayed Fusarium Head Blight Symptoms in Durum Wheat Transgenic Plants Expressing the Xylanase Inhibitor Taxi-III" by S. Mosci and colleagues is presented.

Abstract Wheat is one of the most important crops in the world in terms of human nutrition. With regards to health, some individuals exhibit wheat-related disorders such as food allergy to wheat (FAW). In this disorder, gluten is involved, particularly the gliadins which are among the main proteins responsible for FAW. Food processing, as well as digestibility and intestinal transport are key factors to consider since they may affect the allergenic potential of food allergens. Wheat is always consumed after heat processing and this step may impact epitope accessibility by inducing aggregation and may irreversibly destroy conformational epitopes. Our aim was to investigate the effects of heating and digestion on the structure of well-known allergens (total gliadins and α-gliadins) and their capacity to maintain their allergenic potential after crossing an intestinal barrier. The sizes of the processed (heated and heated/digested) proteins were characterized by laser light scattering and chromatographic reverse phase. The IgE-binding capacities of native and processed proteins were checked using a dot blot with sera from wheat allergenic patients. Furthermore, the abilities of these samples to cross the intestinal barrier and to induce mast cell degranulation were investigated by combining two in vitro cellular models, Caco-2 and RBL-SX38. The heat treatment of total gliadins and α-gliadins induced the production of large aggregates that were hardly recognized by IgE of patients in dot-blot. However, after limited pepsin hydrolysis, the epitopes were unmasked, and they were able to bind IgE again. Native proteins (gliadins and α-type) and processed forms were able to cross the Caco-2 cells in small amount. Permeability studies revealed the capacity of α-gliadins to increase paracellular permeability. In the RBL assay, the total native gliadins were able to trigger cell degranulation, but none of their processed forms. However after crossing the CaCo-2 monolayer, processed gliadins recovered their degranulation capacity to a certain extent. Total native gliadins remained the best allergenic form compared to α-type. Graphical abstract Unlabelled Image Highlights • Native gliadins are more allergenic in vitro than heat-treated and digested proteins. • Limited pepsin hydrolysis restores IgE-binding capacity of aggregated gliadins. • Gliadins show better capacity to activate basophils after transport across Caco-2 cells. • Processing by Caco-2 epithelium cells restores triggering capacities of aggregated proteins. • α-Gliadins increase paracellular permeability of Caco-2 cells. [ABSTRACT FROM AUTHOR]

Abstract: Glutenin polymers are formed by high (HMW-GS) and low molecular weight glutenin subunits (LMW-GS). The latter group of subunits has been less characterised compared to the former due to their great number and heterogeneity. In order to gain more information on the LMW-GS, we have used a durum wheat line carrying a 1BL.1RS translocation, in which the short arm of the chromosome 1B is replaced by the short arm of the chromosome 1R of rye. This line was obtained using the durum wheat cultivar Cando, in which the translocation is present, crossed and back-crossed four times with the Italian durum wheat cultivar Svevo. Comparative electrophoretic and mass spectrometric analyses carried out on LMW-GS prepared from the durum wheat cultivar Svevo and the line carrying the 1BL.1RS translocation have provided further information on these complex group of proteins. In particular, all the three types of typical LMW-GS (LMW-s, LMW-m and LMW-i) were identified in Svevo, whereas the latter group is the only one present in the line with the 1BL.1RS translocation, thus confirming previous findings of the association of the LMW-i type subunits with genes present on chromosome 1A. [Copyright &y& Elsevier]

This systematic review examines the available clinical data on CD34+ cell mobilization, collection, and engraftment in multiple myeloma patients treated with the anti-CD38 monoclonal antibodies daratumumab and isatuximab in clinical trials and in real life. Twenty-six clinical reports were published between 2019 and February 2024. Most studies documented lower circulating CD34+ cells after mobilization compared to controls, leading to higher plerixafor requirements. Although collection yields were significantly lower in approximately half of the studies, the collection target was achieved in similar proportions of daratumumab- and isatuximab-treated and nontreated patients, and access to autologous stem cell transplant (ASCT) was comparable. This could be explained by the retained efficacy of plerixafor in anti-CD38 monoclonal antibody-treated patients, while no chemotherapy-based or sparing mobilization protocol proved superior. Half of the studies reported slower hematopoietic reconstitution after ASCT in daratumumab- and isatuximab-treated patients, without an excess of infectious complications. While no direct effect on stem cells was observed in vitro, emerging evidence suggests possible dysregulation of CD34+ cell adhesion after daratumumab treatment. Overall, anti-CD38 monoclonal antibodies appear to interfere with CD34+ cell mobilization, without consistently leading to significant clinical consequences. Further research is needed to elucidate the underlying mechanisms and define optimal mobilization strategies in this patient population.

Background: In wheat-derived bakery products, the quantity of free asparagine (fAsn) has been identified as a key factor in acrylamide (AA) formation. Based on this assumption, four varieties of common wheat (Triticum aestivum L.), Stromboli, Montecarlo, Sothys and Cosmic, selected for their different fAsn content inside the grain, were studied to evaluate their potential in the production of pizza with reduced AA levels. To this purpose, wholemeal and refined flours were obtained from each variety., Results: The fAsn content ranged from 0.25 to 3.30 mmol kg -1 , with higher values for wholemeal flours which also showed greater amount of ash, fibre and damaged starch than refined wheat flours. All types of flours were separately used to produce wood oven baked pizza base, according to the Traditional Speciality Guaranteed EU Regulation (97/2010). AA reduction in the range 47-68% was found for all the selected wheat cultivars, compared with a commercial flour, with significantly lower values registered when refined flour was used. Moreover, refined leavened dough samples showed decreased levels of fAsn and reducing sugars due to the fermentation activity of yeasts. Furthermore, it was confirmed that pizza made with wholemeal flours exhibited lower rapidly digestible starch (RDS) and rapidly available glucose (RAG) values compared to that prepared with the refined flour., Conclusion: This study clearly shows that a reduced asparagine content in wheat flour is a key factor in the mitigation of AA formation in pizza base. Unfortunately, at the same time, it is highlighted how it is necessary to sacrifice the beneficial effects of fibre intake, such as lowering the glycaemic index, in order to reduce AA. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

The criteria of "Distinctness, Uniformity and Stability" as well as a high "overall quality index" are used to register the Italian modern varieties to the national register. Differently, local conservation varieties can be certified under different EU Directives that facilitate, as an overall objective, the preservation of biodiversity and the containment of genetic erosion. In recent years, products derived from ancient grains are perceived to be healthier and more sustainable by consumers, especially in Italy, with consequent higher market prices. The ancient tetraploid wheat varieties registered in the national register of conservation varieties amount to 28, 24 of which are Sicilian. They are supposed to have wide genetic variability compared to modern ones, making them vulnerable to fraud because they are difficult to trace. It is therefore important to have tools able to discriminate between autochthonous Sicilian varieties. This can be completed by gluten proteins composition, which also provides information on the technological properties of derived products. Fifty-one accessions belonging to twenty-two ancient varieties of Sicilian tetraploid (mostly durum) wheat were analyzed. Although wide intra-accession and intra-varietal variability measurements were assessed, the gliadin pattern of bulks of seeds belonging to each variety was discriminatory. Moreover, differences in technological attitudes were found between landraces. This paves the way to use gluten protein patterns for traceability, allowing local farmers and producers to valorize their products and assure consumers regarding the transparency of the entire supply chain.

Toxoplasma gondii is a protozoan parasite that infects a broad spectrum of hosts and can colonize many organs and cell types. The ability to reside within a wide range of different niches requires substantial adaptability to diverse microenvironments. Very little is known about how this parasite senses various milieus and adapts its metabolism to survive, replicate during the acute stage, and then differentiate to the chronic stage. T. gondii possesses a lysosome-like organelle known as the plant-like vacuolar compartment (PLVAC), which serves various functions, including digestion, ion storage and homeostasis, endocytosis, and autophagy. Lysosomes are critical for maintaining cellular health and function by degrading waste materials and recycling components. To supply the cell with the essential building blocks and energy sources required for the maintenance of its functions and structures, the digested solutes generated within the lysosome are transported into the cytosol by proteins embedded in the lysosomal membrane. Currently, a limited number of PLVAC transporters have been characterized, with TgCRT being the sole potential transporter of amino acids and small peptides identified thus far. To bridge this knowledge gap, we used lysosomal amino acid transporters from other organisms as queries to search the T. gondii proteome. This led to the identification of four potential amino acid transporters, which we have designated as TgAAT1-4. Assessing their expression and sub-cellular localization, we found that one of them, TgAAT1, localized to the PLVAC and is necessary for normal parasite extracellular survival and bradyzoite differentiation. Moreover, we present preliminary data showing the possible involvement of TgAAT1 in the PLVAC transport of arginine.IMPORTANCE Toxoplasma gondii is a highly successful parasite infecting a broad range of warm-blooded organisms, including about one-third of all humans. Although Toxoplasma infections rarely result in symptomatic disease in individuals with a healthy immune system, the incredibly high number of persons infected, along with the risk of severe infection in immunocompromised patients and the potential link of chronic infection to mental disorders, makes this infection a significant public health concern. As a result, there is a pressing need for new treatment approaches that are both effective and well tolerated. The limitations in understanding how Toxoplasma gondii manages its metabolism to adapt to changing environments and triggers its transformation into bradyzoites have hindered the discovery of vulnerabilities in its metabolic pathways or nutrient acquisition mechanisms to identify new therapeutic targets. In this work, we have shown that the lysosome-like organelle plant-like vacuolar compartment (PLVAC), acting through the putative arginine transporter TgAAT1, plays a pivotal role in regulating the parasite's extracellular survival and differentiation into bradyzoites., Competing Interests: The authors declare no conflict of interest.

Introduction: Wheat is a staple food, with the two most common species being Triticum aestivum and Triticum turgidum ssp . durum . Moreover, the latter, T. turgidum , includes other tetraploid subspecies, among which the sspp. turanicum (Khorasan wheat) and polonicum (Polish wheat), whose importance has increased in the last decades, representing alternative crops for marginal areas, in addition to being a source of genetic diversity., Methods: In this work, different accessions of these three subspecies of T. turgidum have been grown in 2 years in the same environment and have been characterized for technological properties and factors affecting nutritional quality, such as fiber amount and the content of micro- and macro-nutrients in grains, and for root morphological traits., Results: These analyses allowed the identification, in particular, of a Polish wheat accession showing better technological performances, a higher amount of positive micro- and macro-elements, and a lower amount of toxic cadmium. The modern variety Svevo and the Polish Pol2 showed the lowest and the highest shoot:root ratio, respectively. The high shoot:root ratio in Pol2 was mainly attributable to the decrease in root growth. Although Pol2 had a lower root biomass, its particular root morphology made it more efficient for nutrient uptake, as evident from the greater accumulation of micro- and macro-nutrients., Discussion: These results underline that it is not possible to draw general conclusions about the difference between primitive and modern wheats, but rather a case-by-case approach should be chosen., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Palombieri, Bonarrigo, Cammerata, Quagliata, Astolfi, Lafiandra, Sestili and Masci.)

The development of solid-state proton conductors with high proton conductivity at low temperatures is crucial for the implementation of hydrogen-based technologies for portable and automotive applications. Here, we report on the discovery of a new crystalline metal acid triphosphate, ZrH 5 (PO 4 ) 3 (ZP3), which exhibits record-high proton conductivity of 0.5-3.1×10 -2  S cm -1 in the range 25-110 °C in anhydrous conditions. This is the highest anhydrous proton conductivity ever reported in a crystalline solid proton conductor in the range 25-110 °C. Superprotonic conductivity in ZP3 is enabled by extended defective frustrated hydrogen bond chains, where the protons are dynamically disordered over two oxygen centers. The high proton conductivity and stability in anhydrous conditions make ZP3 an excellent candidate for innovative applications in fuel cells without the need for complex water management systems, and in other energy technologies requiring fast proton transfer., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Acrylamide, a suspected human carcinogen, is generated during food processing at high temperatures in the Maillard reaction, which involves reducing sugars and free asparagine. In wheat derivatives, free asparagine represents a key factor in acrylamide formation. Free asparagine levels in the grain of different wheat genotypes has been investigated in recent studies, but little is known about elite varieties that are cultivated in Italy. Here, we analysed the accumulation of free asparagine in a total of 54 bread wheat cultivars that are relevant for the Italian market. Six field trials in three Italian locations over two years were considered. Wholemeal flours obtained from harvested seeds were analysed using an enzymatic method. Free asparagine content ranged from 0.99 to 2.82 mmol/kg dry matter in the first year, and from 0.55 to 2.84 mmol/kg dry matter in the second year. Considering the 18 genotypes that were present in all the field trials, we evaluated possible environment and genetic influences for this trait. Some cultivars seemed to be highly affected by environment, whereas others showed a relative stability in free asparagine content across years and locations. Finally, we identified two varieties showing the highest free asparagine levels in our analysis, representing potential useful materials for genotype x environment interaction studies. Two other varieties, which were characterized by low amounts of free asparagine in the considered samples, may be useful for the food industry and for future breeding programs aimed to reduce acrylamide-forming potential in bread wheat.

Micronutrient malnutrition affects more than half of the world population. Reduced bioavailability of microelements in the raw materials is considered one of the main causes of mineral deficiency in populations whose diet is largely based on the consumption of staple crops. In this context, the production of low phytic acid ( lpa ) cereals is a main goal of the breeding programs, as phytic acid (PA) binds essential mineral cations such as iron (Fe), zinc (Zn), manganese (Mn), potassium (K), calcium (Ca) and magnesium (Mg) precipitating in the form of phytate salts poorly digested by monogastric animals, including humans, due to the lack of phytases in the digestive tract. Since PA limits the bioavailability of microelements, it is widely recognized as an anti-nutritional compound. A Targeting Induced Local Lesions IN Genomes (TILLING) approach has been undertaken to silence the genes encoding the TdABCC13 proteins, known as Multidrug-Resistance associated Proteins 3 (TdMRP3), transporters involved in the accumulation of PA inside the vacuole in durum wheat. The TdMRP3 complete null genotypes showed a significant reduction in the content of PA and were able to accumulate a higher amount of essential micronutrients (Fe, Zn, Mn) compared to the control. The number of spikelets and seeds per spike, traits associated with the agronomic performances, were reduced compared to the control, but the negative effect was in part balanced by the increased grain weight. The TdMRP3 mutant lines showed morphological differences in the root apparatus such as a significant decrease in the number of root tips, root length, volume and surface area and an increase in root average diameter compared to the control plants. These materials represent a promising basis for obtaining new commercial durum wheats with higher nutritional value., Competing Interests: Author MV is employed by Grandi Molini Italiani SpA. FS, AF, EB, MV are coinventors of the Italian patent application no. 102022000008315 “Wheat mutant plant with a reduced content of phytic acid”. The authors declare that this study received funding from Grandi Molini Italiani SpA. The funder was not involved in the study design, collection, analysis, interpretation of data, the writing of this article, or the decision to submit it for publication., (Copyright © 2023 Frittelli, Botticella, Palombieri, Masci, Celletti, Fontanella, Astolfi, De Vita, Volpato and Sestili.)

Both Glycaemic index (GI) and Glycaemic Load (GL) were introduced to measure the impact of a carbohydrate-containing food on blood glucose. From this perspective, high-amylose (HA) flours, with a higher percentage of resistant starch (RS), may represent a suitable raw material to improve the glycaemic response. The present work aims to investigate the GI of HA bakery products (biscuits, taralli and bread) compared to products obtained from conventional flour. Ten healthy volunteers were enrolled and their capillary blood glucose was measured every 15 min for 2 h after the consumption of HA and control products containing 50 g of available carbohydrates. On average, in the three bakery products, the amount of total starch replaced by RS was equal to 12%. HA biscuits and HA bread showed significantly lower GI than their control counterparts ( p = 0.0116 and p = 0.011, respectively) and better glycaemic control. From the survey to assess liking and willingness to pay on HA snacks, HA packages received an average premium of €0.66 compared to control products. Although HA flour results in lower GI in both biscuits and bread, further studies are needed to evaluate the correct composition of HA products to have beneficial effects on post-prandial glycaemia.

Amylase/trypsin inhibitors (ATIs) are widely consumed in cereal-based foods and have been implicated in adverse reactions to wheat exposure, such as respiratory and food allergy, and intestinal responses associated with coeliac disease and non-coeliac wheat sensitivity. ATIs occur in multiple isoforms which differ in the amounts present in different types of wheat (including ancient and modern ones). Measuring ATIs and their isoforms is an analytical challenge as is their isolation for use in studies addressing their potential effects on the human body. ATI isoforms differ in their spectrum of bioactive effects in the human gastrointestinal (GI), which may include enzyme inhibition, inflammation and immune responses and of which much is not known. Similarly, although modifications during food processing (exposure to heat, moisture, salt, acid, fermentation) may affect their structure and activity as shown in vitro, it is important to relate these changes to effects that may present in the GI tract. Finally, much of our knowledge of their potential biological effects is based on studies in vitro and in animal models. Validation by human studies using processed foods as commonly consumed is warranted. We conclude that more detailed understanding of these factors may allow the effects of ATIs on human health to be better understood and when possible, to be ameliorated, for example by innovative food processing. We therefore review in short our current knowledge of these proteins, focusing on features which relate to their biological activity and identifying gaps in our knowledge and research priorities., (© 2022. The Author(s).)

Quantitation of wheat proteins is still a challenge, especially regarding amylase/trypsin-inhibitors (ATIs). A selection of ATIs was silenced in the common wheat cultivar Bobwhite and durum wheat cultivar Svevo by RNAi and gene editing, respectively, in order to reduce the amounts of ATIs. The controls and silenced lines were analyzed after digestion to peptides by LC-MS/MS with different approaches to evaluate changes in composition of ATIs. First, a targeted method with stable isotope dilution assay (SIDA) using labeled peptides as internal standards was applied. Additionally, four different approaches for relative quantitation were conducted, in detail, iTRAQ labeled and label free quantitation (LFQ) combined with data dependent acquisition (DDA) and data independent acquisition (DIA). Quantitation was performed manually (Skyline and MASCOT) and with different proteomics software tools (PLGS, MaxQuant, and PEAKS X Pro). To characterize the wheat proteins on protein level, complementary techniques as high-performance liquid chromatography (HPLC) and gel electrophoresis were performed. The targeted approach with SIDA was able to quantitate all ATIs, even at low levels, but an optimized extraction is necessary. The labeled iTRAQ approach revealed an indistinct performance. LFQ with low resolution equipment (IonTrap) showed similar results for major ATIs, but low abundance ATIs as CM1, were not detectable. DDA measurements with an Orbitrap system and evaluation using MaxQuant showed that the relative quantitation was dependent on the wheat species. The combination of manual curation of the MaxQuant search with Skyline revealed a very good performance. The DIA approach with analytical flow found similar results compared to absolute quantitation except for some minor ATIs, which were not detected. Comparison of applied methods revealed that peptide selection is a crucial step for protein quantitation. Wheat proteomics faces challenges due to the high genetic complexity, the close relationship to other cereals and the incomplete, redundant protein database requiring sensitive, precise and accurate LC-MS/MS methods., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Geisslitz, Islam, Buck, Grunwald-Gruber, Sestili, Camerlengo, Masci and D’Amico.)

Obligate intracellular parasites have evolved a remarkable assortment of strategies to scavenge nutrients from the host cells they parasitize. Most apicomplexans form a parasitophorous vacuole (PV) within the invaded cell, a replicative niche within which they survive and multiply. As well as providing a physical barrier against host cell defense mechanisms, the PV membrane (PVM) is also an important site of nutrient uptake that is essential for the parasites to sustain their metabolism. This means nutrients in the extracellular milieu are separated from parasite metabolic machinery by three different membranes, the host plasma membrane, the PVM, and the parasite plasma membrane (PPM). In order to facilitate nutrient transport from the extracellular environment into the parasite itself, transporters on the host cell membrane of invaded cells can be modified by secreted and exported parasite proteins to maximize uptake of key substrates to meet their metabolic demand. To overcome the second barrier, the PVM, apicomplexan parasites secrete proteins contained in the dense granules that remodel the vacuole and make the membrane permissive to important nutrients. This bulk flow of host nutrients is followed by a more selective uptake of substrates at the PPM that is operated by specific transporters of this third barrier. In this review, we recapitulate and compare the strategies developed by Apicomplexa to scavenge nutrients from their hosts, with particular emphasis on transporters at the parasite plasma membrane and vacuolar solute transporters on the parasite intracellular digestive organelle.

Key Message: The suppression of the HYD-1 gene by a TILLING approach increases the amount of β-carotene in durum wheat kernel. Vitamin A deficiency is a major public health problem that affects numerous countries in the world. As humans are not able to synthesize vitamin A, it must be daily assimilated along with other micro- and macronutrients through the diet. Durum wheat is an important crop for Mediterranean countries and provides a discrete amount of nutrients, such as carbohydrates and proteins, but it is deficient in some essential micronutrients, including provitamin A. In the present work, a targeting induced local lesions in genomes strategy has been undertaken to obtain durum wheat genotypes biofortified in provitamin A. In detail, we focused on the suppression of the β-carotene hydroxylase 1 (HYD1) genes, encoding enzymes involved in the redirection of β-carotene toward the synthesis of the downstream xanthophylls (neoxanthin, violaxanthin and zeaxanthin). Expression analysis of genes involved in carotenoid biosynthesis revealed a reduction of the abundance of HYD1 transcripts greater than 50% in mutant grain compared to the control. The biochemical profiling of carotenoid in the wheat mutant genotypes highlighted a significant increase of more than 70% of β-carotene compared to the wild-type sibling lines, with no change in lutein, α-carotene and zeaxanthin content. This study sheds new light on the molecular mechanism governing carotenoid biosynthesis in durum wheat and provides new genotypes that represent a good genetic resource for future breeding programs focused on the provitamin A biofortification through non-transgenic approaches., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Durum wheat milling is a key process step to improve the quality and safety of final products. The aim of this study was to characterize three bran-enriched milling fractions (i.e., F250, G230 and G250), obtained from three durum wheat grain samples, by using an innovative micronization and air-classification technology. Milling fractions were characterized for main standard quality parameters and for alveographic properties, starch composition and content, phenolic acids, antioxidant activity and ATIs. Results showed that yield recovery, ash content and particle size distributions were influenced either by the operating conditions (230 or 250) or by the grain samples. While total starch content was lower in the micronized sample and air-classified fractions, the P/L ratio increased in air-classified fractions as compared to semolina. Six main individual phenolic acids were identified through HPLC-DAD analysis (i.e., ferulic acid, vanillic acid, p -coumaric acid, sinapic acid, syringic and p -hydroxybenzoic acids). Compared to semolina, higher contents of all individual phenolic components were found in all bran-enriched fractions. The highest rise of TPAs occurred in the F250 fraction, which was maintained in the derived pasta. Moreover, bran-enriched fractions showed significant reductions of ATIs content versus semolina. Overall, our data suggest the potential health benefits of F250, G230 and G250 and support their use to make durum-based foods.

Amylase/trypsin-inhibitors (ATIs) comprise about 2-4% of the total wheat grain proteins and may contribute to natural defense against pests and pathogens. However, they are currently among the most widely studied wheat components because of their proposed role in adverse reactions to wheat consumption in humans. ATIs have long been known to contribute to IgE-mediated allergy (notably Bakers' asthma), but interest has increased since 2012 when they were shown to be able to trigger the innate immune system, with attention focused on their role in coeliac disease which affects about 1% of the population and, more recently, in non-coeliac wheat sensitivity which may affect up to 10% of the population. This has led to studies of their structure, inhibitory properties, genetics, control of expression, behavior during processing, effects on human adverse reactions to wheat and, most recently, strategies to modify their expression in the plant using gene editing. We therefore present an integrated account of this range of research, identifying inconsistencies, and gaps in our knowledge and identifying future research needs. Note   This paper is the outcome of an invited international ATI expert meeting held in Amsterdam, February 3-5 2020., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Geisslitz, Shewry, Brouns, America, Caio, Daly, D'Amico, De Giorgio, Gilissen, Grausgruber, Huang, Jonkers, Keszthelyi, Larré, Masci, Mills, Møller, Sorrells, Svensson, Zevallos and Weegels.)

Two expressed alleles of the 1Ay high-molecular-weight glutenin subunit (HMW-GS), 1Ay21* and 1AyT1 , previously introduced in durum and bread wheat, were separately introgressed into the Australian bread wheat ( Triticum aestivum L.) cv. Livingston. The developed lines had different allelic compositions compared to that of the parental cultivar ( 1Ax1 ), having either 1Ax21 + 1Ay21* or 1Ax1 + 1AyT1 at the Glu-A1 locus. Since 1Ax21 and 1Ax1 are known to have the same effects on quality, differences observed between the two sets of the developed lines are attributed to the two introgressed Ay genes. Yield and agronomic performance of the lines were evaluated in the field, and the protein, dough, and baking quality attributes were evaluated by large-scale quality testing. Results demonstrated that the subunit 1Ay21* increased unextractable polymeric protein by up to 14.3% and improved bread loaf volume by up to 9.2%. On the other hand, subunit 1AyT1 increased total grain protein by up to 9% along with dough elasticity. Furthermore, milling extraction was higher, and flour ash was lower in the 1Ay21* lines compared to the lines integrating 1AyT1 . Both sets of the 1Ay introgression lines reduced dough-mixing time compared to the recurrent parent Livingston. The results also showed that 1Ay21* had a higher potential to improve the baking quality than 1AyT1 under the Livingston genetic background. Both alleles showed the potential to be utilized in breeding programs to improve the breadmaking quality.

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

A bread wheat line (N11) and a disomic 2D(2R) substitution triticale line were crossed and backrossed four times. At each step electrophoretic selection for the seeds that possessed, simultaneously, the complete set of high molecular weight glutenin subunits of N11 and the two high molecular weight secalins of rye, present in the 2D(2R) line, was carried out. Molecular cytogenetic analyses of the BC 4 F 8 generation revealed that the selection carried out produced a disomic addition line (2n = 44). The pair of additional chromosomes consisted of the long arm of chromosome 1R (1RL) from rye fused with the satellite body of the wheat chromosome 6B. Rheological analyses revealed that the dough obtained by the new addition line had higher quality characteristics when compared with the two parents. The role of the two additional high molecular weight secalins, present in the disomic addition line, in influencing improved dough characteristics is discussed.

Although wheat is used worldwide as a staple food, it can give rise to adverse reactions, for which the triggering factors have not been identified yet. These reactions can be caused mainly by kernel proteins, both gluten and non-gluten proteins. Among these latter proteins, α-amylase/trypsin inhibitors (ATI) are involved in baker's asthma and realistically in Non Celiac Wheat Sensitivity (NCWS). In this paper, we report characterization of three transgenic lines obtained from the bread wheat cultivar Bobwhite silenced by RNAi in the three ATI genes CM3 , CM16 and 0.28 . We have obtained transgenic lines showing an effective decrease in the activity of target genes that, although showing a higher trypsin inhibition as a pleiotropic effect, generate a lower reaction when tested with sera of patients allergic to wheat, accounting for the important role of the three target proteins in wheat allergies. Finally, these lines show unintended differences in high molecular weight glutenin subunits (HMW-GS) accumulation, involved in technological performances, but do not show differences in terms of yield. The development of new genotypes accumulating a lower amount of proteins potentially or effectively involved in allergies to wheat and NCWS, not only offers the possibility to use them as a basis for the production of varieties with a lower impact on adverse reaction, but also to test if these proteins are actually implicated in those pathologies for which the triggering factor has not been established yet.

Aims: Self-care, an essential component of heart failure (HF) treatment, is inadequate in most patients. We evaluated if motivational interviewing (MI) (i) improves patient self-care maintenance (primary endpoint; e.g. taking medications), self-care management (e.g. responding to symptoms) and self-care confidence (or self-efficacy) 3 months after enrolment; (ii) changes self-care over 1 year, and (iii) augments patient self-care if informal caregivers are involved., Methods and Results: Parallel randomized controlled trial (1:1:1). A sample of 510 patients (median 74 years, 58% male) and caregivers (median 55 years, 75% female) was randomized to Arm 1 (MI only for patients), Arm 2 (MI for patients and caregivers), or Arm 3 (usual care). The intervention in Arms 1 and 2 consisted of one face-to-face MI session with three telephone contacts. Self-care was evaluated with the Self-Care of HF Index measuring self-care maintenance, management, and confidence. Scores on each scale range from 0 to 100 with higher scores indicating better self-care; ≥70 is considered adequate. At 3 months, self-care maintenance improved 6.99, 7.42 and 2.58 points in Arms 1, 2, and 3, respectively (P = 0.028). Self-care maintenance was adequate in 18.4%, 19.4%, and 9.2% of patients in Arms 1, 2 and 3, respectively (P = 0.016). Over 1 year, self-care maintenance, management, and confidence scores in Arms 1 and 2 were significantly higher than in Arm 3 in several follow-ups. Over 1 year, Arm 2 had the best scores in self-care management., Conclusions: MI significantly improved self-care in HF patients. Including caregivers may potentiate the effect, especially in self-care management. ClinicalTrial.gov, identifier: NCT02894502., (© 2020 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of the European Society of Cardiology.)

Resistant starch (RS) in foods has positive benefits for potentially alleviating lifestyle diseases. RS is correlated positively with starch amylose content. This study aimed to see what level of amylose in durum wheat is needed to lower pasta GI. The silencing of starch synthases IIa (SSIIa) and starch branching enzymes IIa (SBEIIa), key genes involved in starch biosynthesis, in durum wheat cultivar Svevo was performed and spaghetti was prepared and evaluated. The SSIIa and SBEIIa mutants have a 28% and 74% increase in amylose and a 2.8- and 35-fold increase in RS, respectively. Cooked pasta was softer, with higher cooking loss but lower stickiness compared to Svevo spaghetti, and with acceptable appearance and colour. In vitro starch digestion extent (area under the digestion curve) was decreased in both mutants, but much more in SBEIIa, while in vivo GI was only significantly reduced from 50 to 38 in SBEIIa. This is the first study of the glycaemic response of spaghetti prepared from SBEIIa and SSIIa durum wheat mutants. Overall pasta quality was acceptable in both mutants but the SBEIIa mutation provides a clear glycaemic benefit and would be much more appealing than wholemeal spaghetti. We suggest a minimum RS content in spaghetti of ~7% is needed to lower GI which corresponded to an amylose content of ~58%.

Drought stress is becoming more prevalent with global warming, and has been shown tohave large effects on gluten proteins linked to wheat bread making quality. Likewise, lowtemperature stress can detrimentally affect proteins in wheat. This study was done to determine thedifferential abundance of high molecular weight (HMW) glutenin proteins in a drought and lowtemperature stressed high quality hard red spring wheat cultivar (PAN3478), against a control. Thetreatments were applied in the greenhouse at the soft dough stage. HMW glutenin proteins wereextracted from the flour, and were separated by using two-dimensional gel electrophoresis. Proteinspots that had p values lower than 0.05 and fold values equal to or greater than 1.2 were consideredto be significantly differentially abundant. These proteins were further analyzed by using tandemmass spectrometry. There was a 1.3 to 1.8 fold change in 17 protein spots due to the cold treatment.The drought treatment caused a 1.3 to 3.8 fold change in 19 protein spots. These spots matchedeither HMW or low molecular weight (LMW) glutenin subunits. In the latter case, the C subunits ofLMW glutenins were notably found to be up-regulated under both stress conditions. All the proteinsthat have been identified can directly influence dough characteristics. Data are available viaProteomeXchange with the identifier PXD017578.

Macro- and micronutrients, essential for the maintenance of human metabolism, are assimilated daily through the diet. Wheat and other major cereals are a good source of nutrients, such as carbohydrates and proteins, but cannot supply a sufficient amount of essential micronutrients, including provitamin A. As vitamin A deficiency (VAD) leads to several serious diseases throughout the world, the biofortification of a major staple crop, such as wheat, represents an effective way to preserve human health in developing countries. In the present work, a key enzyme involved in the branch of carotenoids pathway producing β-carotene, lycopene epsilon cyclase, has been targeted by a Targeting Induced Local Lesions in Genomes (TILLING) approach in a "block strategy" perspective. The null mutant genotype showed a strong reduction in the expression of the lcyE gene and also interesting pleiotropic effects on an enzyme (β-ring hydroxylase) acting downstream in the pathway. Biochemical profiling of carotenoids in the wheat mutant lines showed an increase of roughly 75% in β-carotene in the grains of the complete mutant line compared with the control. In conclusion, we describe here the production and characterization of a new wheat line biofortified with provitamin A obtained through a nontransgenic approach, which also sheds new light on the molecular mechanism governing carotenoid biosynthesis in durum wheat.

Key Message: Knocking down GW2 enhances grain size by regulating genes encoding the synthesis of cytokinin, gibberellin, starch and cell wall. Raising crop yield is a priority task in the light of the continuing growth of the world's population and the inexorable loss of arable land to urbanization. Here, the RNAi approach was taken to reduce the abundance of Grain Weight 2 (GW2) transcript in the durum wheat cultivar Svevo. The effect of the knockdown was to increase the grains' starch content by 10-40%, their width by 4-13% and their surface area by 3-5%. Transcriptomic profiling, based on a quantitative real-time PCR platform, revealed that the transcript abundance of genes encoding both cytokinin dehydrogenase 1 and the large subunit of ADP-glucose pyrophosphorylase was markedly increased in the transgenic lines, whereas that of the genes encoding cytokinin dehydrogenase 2 and gibberellin 3-oxidase was reduced. A proteomic analysis of the non-storage fraction extracted from mature grains detected that eleven proteins were differentially represented in the transgenic compared to wild-type grain: some of these were involved, or at least potentially involved, in cell wall development, suggesting a role of GW2 in the regulation of cell division in the wheat grain.