The Peach v2.0 release: high-resolution linkage mapping and deep resequencing improve chromosome-scale assembly and contiguity.

BACKGROUND: The availability of the peach genome sequence has fostered relevant research in peach and related Prunus species enabling the identification of genes underlying important horticultural traits as well as the development of advanced tools for genetic and genomic analyses. The first release of the peach genome (Peach v1.0) represented a high-quality WGS (Whole Genome Shotgun) chromosome-scale assembly with high contiguity (contig L50 214.2 kb), large portions of mapped sequences (96%) and high base accuracy (99.96%). The aim of this work was to improve the quality of the first assembly by increasing the portion of mapped and oriented sequences, correcting misassemblies and improving the contiguity and base accuracy using high-throughput linkage mapping and deep resequencing approaches. RESULTS: Four linkage maps with 3,576 molecular markers were used to improve the portion of mapped and oriented sequences (from 96.0% and 85.6% of Peach v1.0 to 99.2% and 98.2% of v2.0, respectively) and enabled a more detailed identification of discernible misassemblies (10.4 Mb in total). The deep resequencing approach fixed 859 homozygous SNPs (Single Nucleotide Polymorphisms) and 1347 homozygous indels. Moreover, the assembled NGS contigs enabled the closing of 212 gaps with an improvement in the contig L50 of 19.2%. CONCLUSIONS: The improved high quality peach genome assembly (Peach v2.0) represents a valuable tool for the analysis of the genetic diversity, domestication, and as a vehicle for genetic improvement of peach and related Prunus species. Moreover, the important phylogenetic position of peach and the absence of recent whole genome duplication (WGD) events make peach a pivotal species for comparative genomics studies aiming at elucidating plant speciation and diversification processes.

The starch-bound alpha-amylase/trypsin-inhibitors in Avena.

Oat kernels exhibit an extra-soft texture, a trait recently demonstrated to be largely modulated by starch-bound tryptophan-rich 2S proteins, the vromindolines. In this study, fractionation by two-dimensional electrophoresis of starch-bound proteins in 25 oat (Avena sativa) cultivars and 11 diploid or tetraploid Avena species revealed novel 2S proteins called Avena α-amylase/trypsin-inhibitors (AATI) because of their sequence similarity with wheat α-amylase/trypsin inhibitors. Thirty-seven AATI polypeptides, about 14 kDa in size, were split into three families named AATI-1, AATI-2, and AATI-3 with different primary structures and isoelectric points. AATI-1 and AATI-2 proteins showed 55.5-60.0 % sequence similarity with wheat α-amylase inhibitors CM1, CM2, and CM16, which have been found to cause innate immunity responses in celiac disease and non-celiac gluten sensitivity. Diploid A-genome and tetraploid AC-genome oat species possess three and five genes encoding for the AATI proteins, respectively, whereas hexaploid A. sativa exhibits 12 genes dispersed over the A-, C-, and D-genomes. Some AATI proteins expressed in hexaploid oats were assigned to the A-genome based on similarity to their counterparts in diploid species, contributing to further clarify the genetic origin of hexaploid oats. Moreover, AATI may interact with starch-bound vromindolines in determining the extra-soft texture of oat kernels and, due to their balanced amino acid compositions, may contribute to the biological value of oat proteins in a positive manner.

Biochemical and molecular characterization of Avena indolines and their role in kernel texture.

Among cereals, Avena sativa is characterized by an extremely soft endosperm texture, which leads to some negative agronomic and technological traits. On the basis of the well-known softening effect of puroindolines in wheat kernel texture, in this study, indolines and their encoding genes are investigated in Avena species at different ploidy levels. Three novel 14 kDa proteins, showing a central hydrophobic domain with four tryptophan residues and here named vromindoline (VIN)-1,2 and 3, were identified. Each VIN protein in diploid oat species was found to be synthesized by a single Vin gene whereas, in hexaploid A. sativa, three Vin-1, three Vin-2 and two Vin-3 genes coding for VIN-1, VIN-2 and VIN-3, respectively, were described and assigned to the A, C or D genomes based on similarity to their counterparts in diploid species. Expression of oat vromindoline transgenes in the extra-hard durum wheat led to accumulation of vromindolines in the endosperm and caused an approximate 50 % reduction of grain hardness, suggesting a central role for vromindolines in causing the extra-soft texture of oat grain. Further, hexaploid oats showed three orthologous genes coding for avenoindolines A and B, with five or three tryptophan residues, respectively, but very low amounts of avenoindolines were found in mature kernels. The present results identify a novel protein family affecting cereal kernel texture and would further elucidate the phylogenetic evolution of Avena genus.

Nutritional Traits, Pasting Properties and Antioxidant Profile of Selected Genotypes of Sorghum, Oat and Maize Eligible for Gluten-Free Products.

The technological and nutritional traits of food-grade sorghum hybrids, hulled/naked oat varieties and maize genotypes of different colors were studied for novel and healthier gluten-free foods. Oat genotypes showed the highest protein content, followed by maize and sorghum. The total starch and the total dietary fiber content were quite similar among the three species. Great variation was found in the amylose content, and the highest was in sorghum (27.12%), followed by oat 16.71% and maize 10.59%. Regarding the pasting profile, the rank of Peak Viscosity was sorghum (742.8 Brabender Unit, BU), followed by maize (729.3 BU) and oat (685.9 BU). Oat and sorghum genotypes had similar average breakdown (407.7 and 419.9 BU, respectively) and setback (690.7 and 682.1 BU, respectively), whereas maize showed lower values for both parameters (384.1 BU and 616.2 BU, respectively). The total antioxidant capacity, only in maize, significantly correlated with total flavonoid, phenolic and proanthocyanidin contents, indicating that all the measured compounds contributed to antioxidant capacity. The study indicated the importance of sounding out the nutritional and technological characteristics of gluten-free cereals in order to select suitable cultivars to be processed in different gluten-free foods with better and healthier quality.

The Contribution of Minor Cereals to Sustainable Diets and Agro-Food Biodiversity.

Since the second half of the 20th century, the intensification of agriculture by increasing external inputs (fertilizers, pesticides), cropland expansion, and the cultivation of only a few selected cereal species or varieties have caused the loss of biodiversity and ecosystem services on farmland [...].

Valorization of Two African Typical Crops, Sorghum and Cassava, by the Production of Different Dry Pasta Formulations.

Mediterranean diet is changing to keep up with the increasingly multiethnic Italian society. With food being considered as a means of integration, innovative foods capable of mixing different raw materials could be of interest. In this work, some of the most consumed African foods such as sorghum, cassava, and durum wheat were used to produce wholegrain spaghetti to valorize their nutritional and sensorial aspects and to combine Italian and foreign tastes. Different pasta formulations (cassava, semolina, cassava:semolina, cassava:sorghum, cassava:durum wheat whole meal, sorghum:semolina) were developed and compared for their content of proteins, total starch, resistant starch, amylose, fiber, total antioxidant capacity, ash, cooking quality and sensorial characteristics. The enrichment of cassava flour with durum wheat and sorghum wholegrain enhanced the total antioxidant capacity, protein, and fiber content with respect to 100% cassava pasta. The presence of cassava or sorghum resulted in a high diameter variability of pasta samples, lower water absorption, and shorter optimal cooking time with respect to semolina pasta. Sensory evaluation of cooked pasta revealed better scores in blends containing semolina. Although the obtained pasta samples were interesting for their nutritional aspects, further adjustments are required in the pasta-making process to improve pasta quality.

Perenniality, more than genotypes, shapes biological and chemical rhizosphere composition of perennial wheat lines.

Perennial grains provide various ecosystem services compared to the annual counterparts thanks to their extensive root system and permanent soil cover. However, little is known about the evolution and diversification of perennial grains rhizosphere and its ecological functions over time. In this study, a suite of -OMICSs - metagenomics, enzymomics, metabolomics and lipidomics - was used to compare the rhizosphere environment of four perennial wheat lines at the first and fourth year of growth in comparison with an annual durum wheat cultivar and the parental species Thinopyrum intermedium. We hypothesized that wheat perenniality has a greater role in shaping the rhizobiome composition, biomass, diversity, and activity than plant genotypes because perenniality affects the quality and quantity of C input - mainly root exudates - hence modulating the plant-microbes crosstalk. In support of this hypothesis, the continuous supply of sugars in the rhizosphere along the years created a favorable environment for microbial growth which is reflected in a higher microbial biomass and enzymatic activity. Moreover, modification in the rhizosphere metabolome and lipidome over the years led to changes in the microbial community composition favoring the coexistence of more diverse microbial taxa, increasing plant tolerance to biotic and abiotic stresses. Despite the dominance of the perenniality effect, our data underlined that the OK72 line rhizobiome distinguished from the others by the increase in abundance of Pseudomonas spp., most of which are known as potential beneficial microorganisms, identifying this line as a suitable candidate for the study and selection of new perennial wheat lines.

Special Issue: Innovative Pasta with High Nutritional and Health Potential.

This editorial summarizes some of the key challenges in the production of novel pasta formulations in order to obtain high nutritional and healthy products [...].

Ancient Caucasian Wheats: A Contribution for Sustainable Diets and Food Diversity.

Through the centuries, the domestication and modern breeding of wheat led to a significant loss of genetic variation in the cultivated gene pool with a consequent decrease in food diversity. Current trends towards low-input and sustainable agriculture call for the revitalization and exploitation of ancient wheats, which represent a reservoir of biodiversity useful to ensure sustainable wheat production in the context of climate change and low-input farming systems. Ancient Caucasian wheat species, such as the hulled wheats Triticum timopheevii (tetraploid AuAuGG) and Triticum zhukovskyi (hexaploid AuAuAmAmGG), are still grown to a limited extent in the Caucasus for the production of traditional foods. These Caucasian wheats were grown in Italy and were analyzed for physical, nutritional and technological characteristics and compared to durum wheat. Both Caucasian species revealed a high protein content (on average 18.5%) associated with a low gluten index, mainly in T. zhukovskyi, and test weight values comparable to commercial wheats. The total antioxidant capacity was revealed to be the double of that in durum wheat, suggesting the use of ancient Caucasian wheats for the production of healthy foods. Finally, the technological and rheological results indicated that Caucasian wheats could be potential raw material for the formulation of flat breads, biscuits and pasta.

Cooking Quality and Chemical and Technological Characteristics of Wholegrain Einkorn Pasta Obtained from Micronized Flour.

The increased demand for healthier foods, the recognition of dry pasta as an ideal carrier of functional ingredients, and the current interest for ancient wheats such as einkorn motivated the present research. Two varieties of Triticum monococcum, namely cv Norberto and the free-threshing cv Hammurabi, were milled by ultra-fine milling process (micronization) to produce wholegrain spaghetti. Einkorn pasta was assessed in terms of technological and biochemical properties and cooking and sensorial quality and compared to durum wheat semolina pasta. Wholewheat einkorn pasta showed a threefold increase in total dietary fibre content as well as in total antioxidant capacity in comparison to the control. The level of resistant starch in cv Norberto resulted significantly higher respect to semolina and einkorn cv Hammurabi pasta. Despite the very weak einkorn gluten network, the sensory and instrumental assessment of pasta quality highlighted that einkorn spaghetti presented good sensorial properties related to their technological quality, in particular, for the overall judgment and firmness. Cultivar Hammurabi emerged as the preeminent compromise on the basis of technological performances together with chemical and sensorial aspects.

Immunogenicity of two oat varieties, in relation to their safety for celiac patients.

OBJECTIVE: Most of the recent studies suggest that oats are well tolerated by celiac disease (CD) patients. However, it is still possible that different oat cultivars may display different biological properties relevant for CD pathogenesis. We aimed to investigate biological and immunological properties of two oat varieties, Avena genziana and Avena potenza, in relation to their safety for CD patients. MATERIAL AND METHODS: Phosphorylation of extracellular signal-regulated kinase (ERK) and trans-epithelial electrical resistance (TEER) were evaluated in CaCo-2 cells treated with peptic-tryptic (PT) digests from the two oats and from gliadin (PTG). With the same PT-digests, duodenal biopsies from 22 CD patients were treated in vitro for 24 h and density of CD25+ cells in lamina propria and of intraepithelial CD3+ T cells was measured, as well as crypt cell proliferation and epithelial expression of interleukin 15. Finally, interferon γ (IFN-γ) production was measured as evidence of gliadin-specific T-cell activation by PT-digests. RESULTS: In contrast to PTG, oats PT-digests were not able to induce significant increase in ERK phosphorylation and decrease in TEER in CaCo-2 cells. In the organ culture system, oats PT-digests, unlike PTG, did not induce significant increase in crypt enterocyte proliferation, increase in interleukin 15 expression or in lamina propria CD25+ cells. Nevertheless Avena potenza increased intraepithelial T-cell density, while Avena genziana-induced IFN-γ production in 3/8 CD intestinal T cell lines. CONCLUSIONS: Our data show that Avena genziana and Avena potenza do not display in vitro activities related to CD pathogenesis. Some T-cell reactivity could be below the threshold for clinical relevance.

Innovative Milling Processes to Improve the Technological and Nutritional Quality of Parboiled Brown Rice Pasta from Contrasting Amylose Content Cultivars.

The demand for gluten-free products, including pasta, is increasing and rice pasta accounts for the largest share of this market. Usually, the production of rice pasta requires additives or specific technological processes able to improve its texture, cooking quality, and sensory properties. In this work, two rice cultivars, with different amylose content, were subjected to parboiling, micronization, and flour air fractionation to obtain brown rice pasta, without any supplement but rice itself. In particular, two types of pasta (spaghetti shape) were produced, one from 100% micronized wholemeal, and the other from refined rice flour replaced with 15% of the air-fractionated fine fraction. Regardless of the cultivar, pasta from wholemeal micronized flour showed higher protein and fiber content than refined flour enriched with fine fraction, whereas no differences were revealed in resistant starch and antioxidant capacity. Pasta from the high amylose content genotype showed the highest resistant starch content and the lowest predicted glycemic index along with sensorial characteristics as good as durum semolina pasta in fine fraction enriched pasta. Besides the technological processes, pasta quality was affected the most by the genotype, since pasta obtained from high amylose cv Gladio resulted in the best in terms of technological and sensory quality.

Using Einkorn and Tritordeum Brewers' Spent Grain to Increase the Nutritional Potential of Durum Wheat Pasta.

Brewers' spent grain (BSG), the major by-product of the brewing industry, can be used as a functional ingredient to increase the nutritional value of cereal-based products. In this work, micronized BSG from the einkorn and tritordeum brewing processes were characterized and used to produce four macaroni pasta formulations enriched with BSG at ratios of 5 g and 10 g/100 g of semolina. Einkorn BSG showed the highest values for all the parameters analyzed-proteins, total dietary fiber (TDF) and total antioxidant capacity (TAC)-except for ß-glucan. TDF increased up to 42 and 68% in pasta samples enriched with 10% of BSG from tritordeum and einkorn, respectively. The replacement of 10% of semolina with BSG from both cereals significantly increased the ß-glucan content and TAC values. Finally, the addition of BSG from einkorn and tritordeum affected to a minimal extent the sensory properties of cooked pasta, which showed higher values of optimal cooking time and cooking loss, but lower total organic matter compared to semolina pasta. Results from the sensorial judgment fell in the good quality ranges for durum wheat pasta; the incorporation of 10% of einkorn BSG resulted in the best compromise in terms of technological, nutritional and sensorial aspects of enriched pasta.

Two prolamin peptides from durum wheat preclude celiac disease-specific T cell activation by gluten proteins.

PURPOSE: Celiac disease (CD) is a permanent intolerance to wheat prolamins and related proteins displayed by genetically susceptible individuals. Blocking or modulation of CD-specific T cell response by altered prolamin peptides are currently considered as a potential alternative to the only effective therapy of CD based on a life-long gluten-free diet. Two prolamin peptides, the 9-mer ASRVAPGQQ and the 10-mer GTVGVAPGQQ sequences, were identified by mass spectrometry in the peptic/tryptic digest of prolamins (PTP) from durum wheat (Triticum turgidum ssp. durum) cv. Adamello, and investigated for their ability to preclude the stimulation of CD-specific mucosal T cells by gluten proteins. METHODS: Gluten-specific polyclonal intestinal T cell lines from five CD children (mean age 5 years) were exposed to 50 microg/ml of a deamidated PTP from whole flour of common wheat (T. aestivum) cv. San Pastore, and tested for proliferation and production of interferon-gamma (INF-gamma) and interleukin 10 (IL-10). The same experiment was performed in the presence of 20 microg/ml of the 9-mer or the 10-mer peptide. RESULTS: T cells exposed to PTP showed a threefold increase in proliferation and INF-gamma production, and a significant (P

Multi-block classification of Italian semolina based on Near Infrared Spectroscopy (NIR) analysis and alveographic indices.

Durum wheat (Triticum turgidum ssp. durum) is widely grown in the Mediterranean area. The semolina obtained by this grain is used to prepare pasta, couscous, and baked products all over the world. The growing area affects the characteristics of Durum wheat; consequently, it is relevant to trace this product. The present study aims at developing an analytical methodology which would allow tracing durum semolina harvested in 7 different Italian macro-areas. In order to achieve this goal, 597 samples of semolina have been analysed by Near Infrared Spectroscopy, and by measuring alveographic parameters. Eventually, the information collected have been handled by a multi-block classifier (SO-PLS-LDA) in order to predict the origin of samples. The proposed approach provided extremely satisfactory results (in external validation, on a test set of 140 objects), correctly classifying all samples according to their growing area, confirming it represents a suitable solution for tracing durum wheat semolina.

Comparative Analysis of in vitro Digestibility and Immunogenicity of Gliadin Proteins From Durum and Einkorn Wheat.

Recent studies suggested that gliadin proteins from the ancient diploid einkorn wheat Triticum monococcum retained a reduced number of immunogenic peptides for celiac disease patients because of a high in vitro digestibility with respect to hexaploid common wheat. In this study, we compared the immunological properties of gliadins from two Triticum monococcum cultivars (Hammurabi and Norberto-ID331) with those of a Triticum durum cultivar (Adamello). Gliadins were digested by mimicking the in vitro gastrointestinal digestion process that includes the brush border membrane peptidases. Competitive ELISA, based on R5 monoclonal antibody, showed that gastrointestinal digestion reduced the immunogenicity of Triticum monococcum gliadins; conversely, the immunogenic potential of Triticum durum gliadins remained almost unchanged by the in vitro digestion. The immune stimulatory activity was also evaluated by detecting the IFN-γ production in gliadin-reactive T-cell lines obtained from the small intestinal mucosa of HLA-DQ2+ celiac disease patients. Interestingly, gastrointestinal digestion markedly reduced the capability of Triticum monococcum gliadins (p <0.05) of both cultivars to activate T cells, while it slightly affected the activity of Triticum durum. In conclusion, our results showed that Triticum durum was almost unaffected by the in vitro gastrointestinal digestion, while Triticum monococcum had a marked sensibility to digestion, thus determining a lower toxicity for celiac disease patients.

In Celiac Disease Patients the In Vivo Challenge with the Diploid Triticum monococcum Elicits a Reduced Immune Response Compared to Hexaploid Wheat.

SCOPE: Gluten from the diploid wheat Triticum monococcum (TM) has low content of immunostimulatory sequences and a high gastro-intestinal digestibility. Gluten-reactive T cells elicited by diploid and hexaploid (Triticum aestivum-TA) wheat in celiac disease (CD) patients upon a brief oral challenge are analyzed. METHODS AND RESULTS: Seventeen patients with CD (median age 13 years) consumed for 3 days sandwiches made with TM (cultivar Norberto-ID331, N=11), or TA (cultivar Sagittario, N=11) flours, corresponding to 12 gr of gluten/die. Immunostimulatory properties are assessed in blood by measuring the IFN-γ-secreting T cells by EliSpot and the expression of inflammatory cytokines/receptors (IL-12A, IL-15, IL-18RAP, IFN-γ) by qPCR. TA mobilizes a remarkable number of gliadin-specific, IFN-γ-secreting T cells (p<0.05), while no significant cell mobilization is induced by TM (p=ns). Similar results are obtained in response to five immunogenic peptides from α-, ω-, and γ-gliadins, although with a large individual variability. An increased mRNA expression for IL-12A and IFN-γ is detected in the group eating TA compared to those consuming TM (p<0.05). CONCLUSIONS: Although T. monococcum is a cereal not suitable for the diet of celiacs, this diploid wheat elicits a reduced in vivo T-cell response compared to T. aestivum in celiac patients.

Starch-bound 2S proteins and kernel texture in einkorn, Triticum monococcum ssp monococcum.

The starch granule proteins from 113 einkorn wheat (Triticum monococcum ssp monococcum) accessions were analyzed by acidic, polyacrylamide gel electrophoresis (A-PAGE), and two-dimensional A-PAGE x SDS-PAGE. All accessions were confirmed to contain equal amounts of two polypeptide chains corresponding to puroindoline B (Pin-B), as well as a prominent component plus a faint band corresponding to puroindoline A (Pin-A). When compared with soft-textured common wheat, "monococcum" accessions showed an increase of 3.2- and 2.7-fold in Pin-A and Pin-B levels on the starch granules, respectively. In addition, all accessions contained a novel component of the 2S super-family of seed proteins named Einkorn Trypsin Inhibitor (ETI), which was found to be encoded as a pre-protein 148 residues long. Wild-type ETI encoded by allele Eti-A(m) 1a and "valine-type" ETI encoded by allele Eti-A(m) 1b, which occurred in 107 and six einkorn accessions, respectively, were found to accumulate on starch granules as a mature protein of 121 amino acids with a hydrophobic central domain. The einkorn accessions exhibited an average SKCS index as low as -2.05 +/- 11.4, which is typical of extra-soft kernels. The total surface area of starch granules in "monococcum" wheat, as determined by visual assessments in counting chambers, was estimated at 764 mm(2)/mg of starch, and was about 1.5 times higher than that for common wheat. The results are discussed in relation to the identification of factors that cause the extra-soft texture of einkorn kernels.

GWAS for Starch-Related Parameters in Japonica Rice (Oryza sativa L.).

Rice quality is mainly related to the following two starch components, apparent amylose content (AAC) and resistant starch (RS). The former affects grain cooking properties, while RS acts as a prebiotic. In the present study, a Genome Wide Association Scan (GWAS) was performed using 115 rice japonica accessions, including tropical and temperate genotypes, with the purpose of expanding the knowledge of the genetic bases affecting RS and AAC. High phenotypic variation was recorded for the two traits, which positively correlated. Moreover, both the parameters correlated with seed length (positive correlation) and seed width (negative correlation). A correlational selection according to human preferences has been hypothesized for the two starch traits and grain size. In addition, human selection has been proposed as the causal agent even for the different phenotypes related to starch and grain size showed by the tropical and temperate japonica accessions utilized in this study. The present GWAS led to the identification of 11 associations for RS on seven chromosomes and five associations for AAC on chromosome 6. Candidate genes and co-positional relationships with quantitative trait loci (QTLs) previously identified as affecting RS and AAC were identified for 6 associations. The candidate genes and the new RS- and/or AAC-associated regions detected provide valuable sources for future functional characterizations and for breeding programs aimed at improving rice grain quality.

Bioactive composition and sensory evaluation of innovative spaghetti supplemented with free or α-cyclodextrin chlatrated pumpkin oil extracted by supercritical CO2.

The feasibility of producing durum wheat pasta enriched with a lipophilic phytocomplex, extracted using supercritical carbon dioxide (SC-CO2), from ripe pumpkin, as free oil or as ready-to-mix oil/α-cyclodextrins (α-CDs) powder, was explored. Four types of pasta were prepared: (i) control spaghetti (S-CTRL); (ii) spaghetti supplemented with α-CDs (S-α-CD); (iii) spaghetti supplemented with pumpkin oil (S-Oil) and (iv) spaghetti supplemented with the pumpkin oil/α-CD powder (S-Oil/α-CD). The chemical, antioxidant, textural and sensory attributes of the different pasta were evaluated and compared. S-Oil and S-Oil/α-CD spaghetti were significantly enriched with phytosterols, squalene, carotenoids, tocochromanols and unsaturated fatty acids. Spaghetti containing α-CDs were slightly improved in terms of fiber content. Oil chlatration increased the stability of some bioactives during pasta production and ameliorated poor textural and sensory characteristics of the cooked spaghetti compared with S-Oil sample. S-Oil/α-CD spaghetti might be accepted by customers, if the potential health benefits were also explained.