Exploring the potential of endophyte-plant interactions for improving crop sustainable yields in a changing climate.

Climate change poses a major threat to global food security, significantly reducing crop yields as cause of abiotic stresses, and for boosting the spread of new and old pathogens and pests. Sustainable crop management as a route to mitigation poses the challenge of recruiting an array of solutions and tools for the new aims. Among these, the deployment of positive interactions between the micro-biotic components of agroecosystems and plants can play a highly significant role, as part of the agro-ecological revolution. Endophytic microorganisms have emerged as a promising solution to tackle this challenge. Among these, Arbuscular Mycorrhizal Fungi (AMF) and endophytic bacteria and fungi have demonstrated their potential to alleviate abiotic stresses such as drought and heat stress, as well as the impacts of biotic stresses. They can enhance crop yields in a sustainable way also by other mechanisms, such as improving the nutrient uptake, or by direct effects on plant physiology. In this review we summarize and update on the main types of endophytes, we highlight several studies that demonstrate their efficacy in improving sustainable yields and explore possible avenues for implementing crop-microbiota interactions. The mechanisms underlying these interactions are highly complex and require a comprehensive understanding. For this reason, omic technologies such as genomics, transcriptomics, proteomics, and metabolomics have been employed to unravel, by a higher level of information, the complex network of interactions between plants and microorganisms. Therefore, we also discuss the various omic approaches and techniques that have been used so far to study plant-endophyte interactions.

Seed priming mitigates high salinity impact on germination of bread wheat (Triticum aestivum L.) by improving carbohydrate and protein mobilization.

Salinity is increasingly considered as a major environmental issue, which threatens agricultural production by decreasing yield traits of crops. Seed priming is a useful and cost-effective technique to alleviate the negative effects of salinity and to enable a fast and uniform germination. In this context, we quantified the effects of priming with gibberellic acid (GP), calcium chloride (CP), and mannitol (MP) on seed germination of three bread wheat cultivars and investigated their response when grown at high salinity conditions (200 mM NaCl). Salt exposure strongly repressed seed imbibition and germination potential and extended germination time, whereas priming enhanced uniformity and seed vigor. Seed preconditioning alleviated the germination disruption caused by salt stress to varying degrees. Priming mitigating effect was agent-dependent with regard to water status (CP and MP), ionic imbalance (CP), and seed reserve mobilization (GP). Na+ accumulation in seedling tissues significantly impaired carbohydrate and protein mobilization by inhibiting amylase and proteases activities but had lesser effects on primed seeds. CP attenuated ionic imbalance by limiting sodium accumulation. Gibberellic acid was the most effective priming treatment for promoting the germination of wheat seeds under salt stress. Moreover, genotypic differences in wheat response to salinity stress were observed between varieties used in this study. Ardito, the oldest variety, seems to tolerate better salinity in priming-free conditions; Aubusson resulted the most salt-sensitive cultivar but showed a high germination recovery under priming conditions; Bologna showed an intermediate behavior.

Marker-Assisted Pyramiding of Blast-Resistance Genes in a japonica Elite Rice Cultivar through Forward and Background Selection.

Rice blast, caused by Pyricularia oryzae, is one of the main rice diseases worldwide. The pyramiding of blast-resistance (Pi) genes, coupled to Marker-Assisted BackCrossing (MABC), provides broad-spectrum and potentially durable resistance while limiting the donor genome in the background of an elite cultivar. In this work, MABC coupled to foreground and background selections based on KASP marker assays has been applied to introgress four Pi genes (Piz, Pib, Pita, and Pik) in a renowned japonica Italian rice variety, highly susceptible to blast. Molecular analyses on the backcross (BC) lines highlighted the presence of an additional blast-resistance gene, the Pita-linked Pita2/Ptr gene, therefore increasing the number of blast-resistance introgressed genes to five. The recurrent genome was recovered up to 95.65%. Several lines carrying four (including Pita2) Pi genes with high recovery percentage levels were also obtained. Phenotypic evaluations confirmed the effectiveness of the pyramided lines against multivirulent strains, which also had broad patterns of resistance in comparison to those expected based on the pyramided Pi genes. The developed blast-resistant japonica lines represent useful donors of multiple blast-resistance genes for future rice-breeding programs related to the japonica group.

Heteroalleles in Common Wheat: Multiple Differences between Allelic Variants of the Gli-B1 Locus.

The Gli-B1-encoded γ-gliadins and non-coding γ-gliadin DNA sequences for 15 different alleles of common wheat have been compared using seven tests: electrophoretic mobility (EM) and molecular weight (MW) of the encoded major γ-gliadin, restriction fragment length polymorphism patterns (RFLPs) (three different markers), Gli-B1-γ-gliadin-pseudogene known SNP markers (Single nucleotide polymorphisms) and sequencing the pseudogene GAG56B. It was discovered that encoded γ-gliadins, with contrasting EM, had similar MWs. However, seven allelic variants (designated from I to VII) differed among them in the other six tests: I (alleles Gli-B1i, k, m, o), II (Gli-B1n, q, s), III (Gli-B1b), IV (Gli-B1e, f, g), V (Gli-B1h), VI (Gli-B1d) and VII (Gli-B1a). Allele Gli-B1c (variant VIII) was identical to the alleles from group IV in four of the tests. Some tests might show a fine difference between alleles belonging to the same variant. Our results attest in favor of the independent origin of at least seven variants at the Gli-B1 locus that might originate from deeply diverged genotypes of the donor(s) of the B genome in hexaploid wheat and therefore might be called "heteroallelic". The donor's particularities at the Gli-B1 locus might be conserved since that time and decisively contribute to the current high genetic diversity of common wheat.

A Cross between Bread Wheat and a 2D(2R) Disomic Substitution Triticale Line Leads to the Formation of a Novel Disomic Addition Line and Provides Information of the Role of Rye Secalins on Breadmaking Characteristics.

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 BC4F8 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.

Elevated CO2 Impact on Common Wheat (Triticum aestivum L.) Yield, Wholemeal Quality, and Sanitary Risk.

The rising atmospheric CO2, concentration is expected to exert a strong impact on crop production, enhancing crop growth but threatening food security and safety. An improver wheat, a hybrid, and its parents were grown at elevated CO2, e[CO2] in open field, and their yield and rheological, nutritional, and sanitary quality were assessed. For all cultivars, grain yield increased (+16%) and protein content decreased (-7%), accompanied by a reduction in dough strength. Grain nitrogen yield increased (+24%) only in ordinary bread making cultivars. e[CO2] did not result in significant changes in phenolic acid content and composition, whereas it produced a significant increase in the deoxynivalenol content. Different responses to e[CO2] between cultivars were found for yield parameters, while the effect on qualitative traits was quite similar. In the upcoming wheat cropping systems, agronomic practices and cultivar selection suited to guarantee higher nitrogen responsiveness and minimization of sanitary risk are required.

Co-Occurrence of Moniliformin and Regulated Fusarium Toxins in Maize and Wheat Grown in Italy.

The co-occurrence of moniliformin (MON), fumonisins (FBs), and deoxynivalenol (DON) was evaluated in maize, durum, and common wheat grown in different experimental fields located in several Italian regions. MON was quantified using a LC-MS/MS method adding lanthanum ions in the mobile phase. In maize, MON contamination was widespread and considerable; the toxin was detected in almost all the samples (95.1%) and exceeded 500 and 1000 µg kg-1 in 42.0% and in 18.5% of samples, respectively. Significant positive correlation was found between MON and FB contamination levels. When there were not droughty climate conditions, a positive significant correlation was found between growing degree days (GDD) and MON values. In wheat, MON contamination was not widespread like in maize and it was lower in common wheat than in durum wheat. In durum wheat, MON was detected in 45.0% of the samples with only 6 samples (7.5%) exceeding 500 µg kg-1, while in common wheat the toxin was detected above the LOD in 18.7% of samples exceeding 100 µg kg-1 in only two samples (2.5%). No correlation was found with DON contamination. Climate conditions influenced both MON and DON occurrence.

Genome-wide association study of agronomic and quality traits in a world collection of the wild wheat relative Triticum urartu.

Triticum urartu (2n = 2x = 14, subgenome Au Au ), a wild diploid wheat progenitor, features broad allelic diversity for a number of traits of agronomic relevance. A thorough characterization of the diversity of T. urartu natural accessions may provide wheat breeders with new alleles potentially contributing to wheat improvement. In this study, we performed an extensive genotypic and phenotypic characterization of a world collection of 299 T. urartu ex situ accessions, developing 441 327 single nucleotide polymorphisms and recording trait values for agronomic and quality traits. The collection was highly diverse, with broad variation in phenology and plant architecture traits. Seed features were also varied, and analyses of flour quality reported 18 distinct patterns of glutenins, and carotenoid concentrations and sedimentation volumes in some cases surpassing those of cultivated materials. The genome-wide molecular markers developed on the collection were used to conduct a genome-wide association study reporting 25 highly significant quantitative trait nucleotides for the traits under examination, only partially overlapping loci already reported in wheat. Our data show that T. urartu may be considered a valuable allele pool to support the improvement of wheat agronomy and quality.

Redox poise and metabolite changes in bread wheat seeds are advanced by priming with hot steam.

Fast and uniform germination is key to agricultural production and can be achieved by seed 'priming' techniques. Here, we characterised the responses of bread wheat (Triticum aestivum L.) seeds to a hot steam treatment ('BioFlash'), which accelerated water uptake, resulting in faster germination and seedling growth, typical traits of primed seed. Before the completion of germination, metabolite profiling of seeds revealed advanced accumulation of several amino acids (especially cysteine and serine), sugars (ribose, glucose), and organic acids (glycerate, succinate) in hot steam-treated seeds, whereas sugar alcohols (e.g. arabitol, mannitol) and trehalose decreased in all seeds. Tocochromanols (the 'vitamin E family') rose independently of the hot steam treatment. We further assessed shifts in the half-cell reduction potentials of low-molecular-weight (LMW) thiol-disulfide redox couples [i.e. glutathione disulfide (GSSG)/glutathione (GSH) and cystine/cysteine], alongside the activities of the reactive oxygen species (ROS)-processing enzyme superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase. Upon the first 4 h of imbibition, a rapid conversion of LMW disulfides to thiols occurred. Completion of germination was associated with a re-oxidation of the LMW thiol-disulfide cellular redox environment, before more reducing conditions were re-established during seedling growth, accompanied by an increase in all ROS-processing enzyme activities. Furthermore, changes in the thiol-disulfide cellular redox state were associated to specific stages of wheat seed germination. In conclusion, the priming effect of the hot steam treatment advanced the onset of seed metabolism, including redox shifts associated with germination and seedling growth.

Changes in low-molecular-weight thiol-disulphide redox couples are part of bread wheat seed germination and early seedling growth.

The tripeptide antioxidant glutathione (γ-l-glutamyl-l-cysteinyl-glycine; GSH) essentially contributes to thiol-disulphide conversions, which are involved in the control of seed development, germination, and seedling establishment. However, the relative contribution of GSH metabolism in different seed structures is not fully understood. We studied the GSH/glutathione disulphide (GSSG) redox couple and associated low-molecular-weight (LMW) thiols and disulphides related to GSH metabolism in bread wheat (Triticum aestivum L.) seeds, focussing on redox changes in the embryo and endosperm during germination. In dry seeds, GSH was the predominant LMW thiol and, 15 h after the onset of imbibition, embryos of non-germinated seeds contained 12 times more LMW thiols than the endosperm. In germinated seeds, the embryo contained 17 and 11 times more LMW thiols than the endosperm after 15 and 48 h, respectively. This resulted in the embryo having significantly more reducing half-cell reduction potentials of GSH/GSSG and cysteine (Cys)/cystine (CySS) redox couples (EGSSG/2GSH and ECySS/2Cys, respectively). Upon seed germination and early seedling growth, Cys and CySS concentrations significantly increased in both embryo and endosperm, progressively contributing to the cellular LMW thiol-disulphide redox environment (Ethiol-disulphide). The changes in ECySS/2Cys could be related to the mobilisation of storage proteins in the endosperm during early seedling growth. We suggest that EGSSG/2GSH and ECySS/2Cys can be used as markers of the physiological and developmental stage of embryo and endosperm. We also present a model of interaction between LMW thiols and disulphides with hydrogen peroxide (H2O2) in redox regulation of bread wheat germination and early seedling growth.

Next generation breeding.

The genomic revolution of the past decade has greatly improved our understanding of the genetic make-up of living organisms. The sequencing of crop genomes has completely changed our vision and interpretation of genome organization and evolution. Re-sequencing allows the identification of an unlimited number of markers as well as the analysis of germplasm allelic diversity based on allele mining approaches. High throughput marker technologies coupled with advanced phenotyping platforms provide new opportunities for discovering marker-trait associations which can sustain genomic-assisted breeding. The availability of genome sequencing information is enabling genome editing (site-specific mutagenesis), to obtain gene sequences desired by breeders. This review illustrates how next generation sequencing-derived information can be used to tailor genomic tools for different breeders' needs to revolutionize crop improvement.

Morpho-physiolological and qualitative traits of a bread wheat collection spanning a century of breeding in Italy.

Evaluation and characterization are crucial steps in the exploitation of germplasm collections. The Sant'Angelo Lodigiano unit of the Consiglio per la ricerca in agricoltura e l'analisi dell'economia agraria (CREA) maintains a broad collection of Triticum spp, including more than 4000 genotypes of T. aestivum. Such collection represents a wide source of genetic variability for many agronomic and qualitative traits, extremely useful in modern breeding programs. The collection size, however, makes very difficult its management as a whole. A reduced subset, representing the process of wheat breeding in Italy during the last hundred years, was hence identified for an in-depth characterization. The lines were cropped in two locations over two growing seasons, and analyzed using 16 morpho-agronomic and qualitative descriptors. Most of the analysed characters showed a broad variation throughout the collection, allowing to follow the plant ideotype changes across the breeding progress in Italy during the 20th century.

Diversity trends in bread wheat in Italy during the 20th century assessed by traditional and multivariate approaches.

A collection of 157 Triticum aestivum accessions, representative of wheat breeding in Italy during the 20(th) century, was assembled to describe the evolutionary trends of cultivated varieties throughout this period. The lines were cultivated in Italy, in two locations, over two growing seasons, and evaluated for several agronomical, morphological and qualitative traits. Analyses were conducted using the most common univariate approach on individual plant traits coupled with a correspondance multivariate approach. ANOVA showed a clear trend from old to new varieties, leading towards earliness, plant height reduction and denser spikes with smaller seeds. The average protein content gradually decreased over time; however this trend did not affect bread-making quality, because it was counterbalanced by a gradual increase of SDS sedimentation volume, achieved by the incorporation of favourable alleles into recent cultivars. Correspondence analysis allowed an overall view of the breeding activity. A clear-cut separation was observed between ancient lines and all the others, matched with a two-step gradient, the first, corresponding roughly to the period 1920-1940, which can be ascribed mostly to genetics, the second, from the 40s onward, which can be ascribed also to the farming practice innovations, such as improvement of mechanical devices and optimised use of fertilizers.

Expression of α-amylase inhibitors in diploid Triticum species.

The aim of the work was to characterize the expression of various α-amylase inhibitors (αAIs), well known anti-nutritional compounds, for the development of healthier diploid wheat-based functional foods. The salt-soluble protein fractions from the seeds of 53 accessions among Triticum monococcum subsp. monococcum (T.m.), T. monococcum subsp. boeoticum (T.b.) and Triticum urartu (T.u.) were analyzed by immunoblotting after SDS-PAGE and Urea-PAGE using polyclonal antibodies (PABs) raised against 0.19 and 0.28 αAIs expressed in bread-wheat. Reverse zymography with human saliva and Tenebrio molitor α-amylases was used to assay inhibition activity. A great variability of the expression of αAI-related proteins was observed among T.b. and T.u. PABs, and reverse zymography revealed different bands, often not correlating with those present in bread-wheat. Two-dimensional electrophoresis followed by immunoblotting and mass spectrometric analysis identified these proteins as αAIs. Interestingly, no signal was observed within T.m. accessions. This makes T.m. an important candidate for the production of novel functional foods.

A catalogue of Triticum monococcum genes encoding toxic and immunogenic peptides for celiac disease patients.

The celiac disease (CD) is an inflammatory condition characterized by injury to the lining of the small-intestine on exposure to the gluten of wheat, barley and rye. The involvement of gluten in the CD syndrome has been studied in detail in bread wheat, where a set of "toxic" and "immunogenic" peptides has been defined. For wheat diploid species, information on CD epitopes is poor. In the present paper, we have adopted a genomic approach in order to understand the potential CD danger represented by storage proteins in diploid wheat and sequenced a sufficiently large number of cDNA clones related to storage protein genes of Triticum monococcum. Four bona fide toxic peptides and 13 immunogenic peptides were found. All the classes of storage proteins were shown to contain harmful sequences. The major conclusion is that einkorn has the full potential to induce the CD syndrome, as already evident for polyploid wheats. In addition, a complete overview of the storage protein gene arsenal in T. monococcum is provided, including a full-length HMW x-type sequence and two partial HMW y-type sequences.

Impact of Eurygaster maura (Heteroptera: Scutelleridae) feeding on quality of bread wheat in relation to attack period.

Sunn pest (or cereal bug) (Heteroptera: Pentatomidae and Scutelleridae) infestations of wheat, Triticum aestivum L., in the grain filling stage have the potential to adversely affect the quality of harvested grain for bread making. In the absence of resistant wheat cultivars, producers must rely on chemical control to protect their crop from sunn pest infestations. To implement an efficient environment friendly control strategy, there is a need to pinpoint the relationships between the timing of the bug attack and gluten degradation. Recent outbreaks of Eurygaster maura (L.) in northwestern Italy have increased the local concern toward this problem. A 3-yr study was carried out by caging plants of two bread wheat cultivars, characterized by different seed texture and bread-making quality, and introducing adults of E. maura in four periods corresponding to different grain filling stages: heading, early milk-ripe, milk-ripe, and late milk-ripe. The degree of bread-making quality depletion was assessed by analytical and biochemical methods and related to the attack period. Using analysis of variance, significant differences were found in the quality traits of kernels attacked by E. maura in different grain filling stages, the maximum damage occurring with bug feeding at the late milk-ripe stage. Biochemical investigations on gluten confirmed analytical results; in grain samples infested at the late milk-ripe stage, SDS gel electrophoresis revealed the degradation of some components of the high-molecular-weight glutenins, and high-performance liquid chromatography analyses showed a breakdown of the first peak of the insoluble fraction, mainly containing polymeric proteins highly related to dough strength.