Molecular characterization of emmer (Triticum dicoccon Schrank) Italian landraces

Emmer (Triticum dicoccon Schrank,2n = 4x = 28) is a hulled wheatspecies [more] widely spread in the Mediterranean basin. In Italy it survives as acrop in a few marginal areas and peculiar ecological niches in different regionsof central and southern Italy. A renewed interest has occurred during the lastdecade toward local varieties belonging to this species. As a matter of fact,local varieties have the highest genetic variation and adaptation to the naturaland anthropological environment from where they originated. Results on thegenetic diversity within and relationships among 11 Italian local varieties ofemmer as assessed with 17 RAPD marker loci are here reported. The proportion ofthe among-local variety genetic diversity was as high as 48% (G_ST =0.479). Thus, about 52% of the total variation was within population. Localvarieties of emmer proved to be formed by a variable number of lines geneticallydistinguishable from each other, and the vast majority of individuals overpopulations proved to be different multilocus genotypes. Landraces of emmer fromcentral and southern Italy showed distinctive molecular traits. In particular,local varieties classified as «Central Italy» types were characterized by a common set of RAPD marker alleles and proved to bedistinguishable from both the «Southern Italy» and the«Garfagnana» accessions. The overall results confirm the highvariability that can be found within landrace populations, underlining thevalues of landraces as an irreplaceable bank of genetically diversified andhighly co-adapted genotypes. Information for an appropriate insitu conservation and management of this valuable source of emmergermplasm is discussed.     

Molecular genetic diversity analysis in emmer wheat (Triticum dicoccon Schrank) from India

Emmer wheat (Triticum dicoccon Schrank) is still largely cultivated in India, and highly appreciated for the preparation of traditional dishes. Moreover, its nutritional characteristics could justify a development of its cultivation. The perspective of genetic improvement however requires a good knowledge of the genetic diversity existing within the eco-geographic group of Indian emmer wheats. A set of 48 emmer wheat accessions from India including 28 from a local collection and 20 Indian accessions obtained from CIMMYT, Mexico, was assessed for genetic variability using 47 microsatellite (SSR) markers, distributed over all the 14 chromosomes. The number of alleles per locus ranged from 2 to 9, with an average of 3.87 alleles per locus. A total of 201 alleles were detected at 52 loci with average polymorphic information content of 0.35 per locus and a mean resolving power of 1. The pair-wise similarity coefficients calculated from binary data matrix based on presence or absence of alleles varied from 0.15 to 0.98, but was greater than 0.5 for most accessions, indicating a high level of similarity. A cluster analysis based on the similarity matrix identified nine distinct accessions and three clusters. All the recently developed commercial varieties were distinctly different from the clusters. Based on the analysis, it appears that Indian emmer wheats are not very diverse. Consequently, there is a need to increase the diversity within the Indian emmer wheat eco-geographic group, by introducing diversity from other eco-geographic groups, or even from other wheat species.     

Dimeric inhibitors of human salivary alpha-amylase from emmer (Triticum dicoccon Schrank) seeds

The proteins belonging to the cereal trypsin/alpha-amylase inhibitor family are abundant water/salt-soluble flour proteins active against alpha-amylases from several seed parasites and pests and inactive against endogenous alpha-amylases. Three alpha-amylase inhibitor families have been described in cereals that vary in size and are differently expressed among Triticeae seeds. The present work investigates the presence of human salivary alpha-amylase inhibitors in emmer (Triticum dicoccon Schrank) flour. The isolation was obtained by a series of chromatography steps, and the purification progress was monitored through the inhibition of human salivary alpha-amylase activity. The purified fraction was subjected to protein sequencing by tandem mass spectrometry (MSMS) of the tryptic digests obtained after the sample separation on 2-DE. MSMS data indicated that the emmer alpha-amylase inhibitory fraction was composed of two newly identified proteins [emmer dimeric inhibitor 1 (EDI-1) and emmer dimeric inhibitor 2 (EDI-2)] sharing very high identity levels with related proteins from Triticum aestivum.     

Cultivated emmer wheat (Triticum dicoccon Schrank), an old crop with promising future: a review

Cultivated emmer wheat, Triticum dicoccon Schrank, a tetraploid species with hulled grain, has been largely cultivated during seven millennia in the Middle-East, Central and West Asia, and Europe. It has been largely replaced by hulless species and is now a minor crop, with the exception of some countries like India, Ethiopia and Yemen, where its grain is used for preparing traditional foods. Nutritional qualities and specific taste and flavor of emmer wheat products have led to a recent development of the cultivation in some European countries. Emmer wheat also possesses valuable traits of resistance to pests and diseases and tolerance to abiotic stresses and is increasingly used as a reservoir of useful genes in wheat breeding. In the present article, a review concerning taxonomy, diversity and history of cultivation of emmer wheat is reported. Grain characteristics and valuable agronomic traits are described. Some successful examples of emmer wheat utilization for the development of durum or bread wheat cultivars are examined, and the perspectives in using emmer wheat as health food and for the development of new breeding germplasm are discussed.     

Emmer (Triticum dicoccon Schrank) in Oman†

Emmer (Triticum dicoccon ) was collected recently in northern Oman. The material was analyzed morphologically and phenologically. It belongs to the Asiatic emmers (subsp. asiaticum) and not to the Ethiopian ones (subsp. abyssinicum), distributed in Ethiopia and Yemen, as originally expected. The determination of the material resulted in var. haussknechtianum and var. aeruginosum.     

Kinetics of carotenoids degradation during the storage of einkorn (Triticum monococcum L. ssp. monococcum) and bread wheat (Triticum aestivum L. ssp. aestivum) flours

To evaluate the effect of storage temperature, the degradation kinetics of carotenoids in wholemeal and white flour of einkorn cv. Monlis and bread wheat cv. Serio, stored at -20, 5, 20, 30, and 38 degrees C, was assessed by normal-phase high-performance liquid chromatography. In Monlis, the carotenoids content (8.1 and 9.8 mg/kg for wholemeal and white flour, respectively) was 8-fold higher than in Serio (1.0 and 1.1 mg/kg). Only lutein and zeaxanthin were detected in bread wheat, while significant quantities of (alpha and beta)-carotene and beta-cryptoxanthin were observed in einkorn. Carotenoids degradation was influenced by temperature and time, following first-order kinetics. The degradation rate was similar in wholemeal and white flour; however, loss of lutein and total carotenoids was faster in Serio than in Monlis. The activation energy E(a) ranged from 35.2 to 52.5 kJ/mol. Temperatures not exceeding 20 degrees C better preserve carotenoids content and are recommended for long-term storage.     

Cultivated einkorn wheat (Triticum monococcum L. subsp. monococcum): the long life of a founder crop of agriculture

The first cultivated wheat, cultivated einkorn (Triticum monococcum L. subsp. monococcum), was domesticated in South-East Turkey during the Pre-Pottery Neolithic period. It then spread to the Middle-East, the Balkans and Caucasus, Turkmenistan, Central and Mediterranean Europe, North-Africa, and finally to Western and Northern Europe. In all these regions, it played an important role in the development of agriculture and was cultivated for several centuries before being replaced by free-threshing wheats. Today, cultivated einkorn is only present in isolated, mountainous areas of a few countries. However, there is renewed interest for this crop due to the nutritional qualities of its grain, its adaptation to low-input agriculture and high level of resistance to pests and diseases that represent advantages for organic farming. Cultivated einkorn is also a valuable reservoir of genes for wheat improvement. Its utilization, limited by its hulledness, low yield and especially by a poor knowledge of its diversity and its low crossability with bread and durum wheat, is expected to increase in the future, particularly with the need for wheat breeding to face newly emerging diseases through the use of genetic resistances. Considering these perspectives, the present review attempts to analyse the current and historical importance of einkorn cultivation and utilization in wheat breeding, tracing back to its origin and diffusion. The main traits of resistance to pest and diseases, and the nutritional qualities and technological characteristics of the grain are described. Einkorn genetic resources diversity exploration is reviewed and successful examples of introgression of useful einkorn traits into cultivated wheat are reported. Lastly, perspectives of einkorn cultivation development in low-input agriculture and use for wheat enhancement are discussed.     

Assessment of genetic diversity in emmer (Triticum dicoccon Schrank) × durum wheat (Triticum durum Desf.) derived lines and their parents using mapped and unmapped molecular markers

In the last few years, the renewed interest for emmer wheat (Triticum dicoccon Schrank) in Italy has stimulated breeding programs for this crop releasing improved genotypes obtained not only by selection from landraces, but even by crosses with durum wheat (Triticum durum Desf.) varieties. The purpose of this work has been to uncover the genetic make-up of some emmer × durum derivatives, specifically by comparing the differences from their parents. Genetic diversity of advanced breeding lines and varieties derived from a durum × emmer cross has been evaluated on the basis of AFLP and SSR markers in comparison with the corresponding emmer and durum wheat parent for addressing the seminal question of how much ‘wild’ variation remains after selection for agronomic type.     

NsLTP1 and NsLTP2 isoforms in soft wheat (Triticum aestivum Cv. Centauro) and farro (Triticum dicoccon Schrank) bran

Isoforms of nonspecific lipid-transfer protein 1 (nsLTP1) and nonspecific lipid-transfer protein 2 (nsLTP2) were investigated in bran tissues isolated from caryopses of two cereal crops quite relevant for the Italian market, the cultivar Centauro of soft wheat (Triticum aestivum) and Italian emmer or farro (Triticum dicoccon Schrank). By sequential separation of the bran extracts on cation-exchange and gel filtration chromatographies, fractions containing only proteins belonging to the nsLTP1 and nsLTP2 classes were obtained. The proteins were roughly identified by SDS-PAGE and by immunoreactions in Western blotting experiments. By MALDI-MS and RP-HPLC/ESI-MS analyses we were able to show the presence of several LTP1 and LTP2 isoforms in the investigated species. Bioinformatic searches based on the determined Mr indicated that (i) two nsLTP1s already identified in T. aestivum have Mr and number of Cys residues identical to that of a 9.6 kDa protein present both in soft wheat cv. Centauro and in farro; (ii) two isoforms of nsLTP2 detected in T. aestivum have the same Mr and number of Cys residues of two 7 kDa proteins found in Centauro; and (iii) a nsLTP1 detected in Ambrosia artemisiifolia has Mr and number of Cys residues coincident to that of a 9.9 kDa protein found both in soft wheat cv. Centauro and in farro.     

Patterns of morphological diversity in tetraploid wheat (Triticum turgidum L.) landraces from Ethiopia

Thirty‐four tetraploid (2n = 4x = 28) wheat (Triticum turgidum L.) landrace populations collected from four regions in Ethiopia, and consisting of 4099 entries were characterized for glume colour, glume pubescence, beak awn, seed colour and spike density. The main objective was to analyse the diversity and distribution of these traits on the basis of administrative regions and altitudinal gradients. The Shannon‐Weaver Diversity Index (H') was used to estimate the magnitude of diversity. With the exception of spike density, all characters were polymorphic in all regions and most altitude groups. However, clinal patterns were observed in only a few cases. At the population level, the mean H’ varied from completely monomorphic (H’ = 0.00) to highly polymorphic (H’ = 0.80 ± 0.07). The highest diversity was found in regions with relatively better climatic conditions and in optimal altitude ranges. On country basis, seed colour (H’ = 0.98) and spike density (H’ = 0.43) displayed the highest and the lowest diversity indices, respectively. Spike density was the only character that exhibited significant differences (P<0.01) both “between regions”; and “between altitude groups”;. Across characters, most of the variance was due to the lowest hierarchy, the “within‐population”; component. The “between region”; and “between altitude group”; variances were significant, but spike density alone was responsible for the differences. The overall mean H’ for Ethiopia was 0.77 + 0.09 and did not seem to have reduced appreciably within the past 25 years or so, when compared to previous estimates.     

Genetic diversity and intra-specific phylogeny of Triticum turgidum L. subsp. dicoccon (Schrank) Thell. revealed by RFLPs and SSRs

Today, emmer wheat, T. turgidum subsp. dicoccon, widely grown in the past is a candidate crop for sustainable agriculture in Italy. As part of a research project aimed at the enhanced use of the hulled wheat germplasm, molecular characterization was carried out to understand the genetic structure of the crop and to identify accessions of interest. A collection of 194 accessions was analyzed with 15 microsatellite loci (SSRs), while only a sample of 38 accessions was tested with 19 RFLP probes. The marker loci were selected on the basis of their independent genomic distribution. Genetic distances and allelic frequencies were calculated for each marker class. The genetic relationships were visualized with dendrograms. RFLP loci were, on average, less polymorphic than SSRs. An average Dice's genetic distance of 0.22 for RFLPs vs 0.38 for SSRs was detected, while an expected average heterozygosity per locus of 0.23 for RFLPs vs 0.26 for SSRs was also estimated. With a least number of 10 loci per marker class it was possible to identify each genotype. The most diverse accessions had different geographic origins. Germplasms from Italy and Ethiopia appear to belong to a more primitive genepool, given that a group of accessions from these countries were genetically differentiated from a Russian-Iranian group.     

The effect of soil-climate conditions on yielding parameters,chemical composition and baking quality of ancient wheat species Triticum monococcum L., Triticum dicoccum Schrank and Triticum spelt L. in comparison with modern Triticum aestivum L.

Triticum monococcum L., Triticum dicoccum Schrank and Triticum spelt L. nowadays offer an alternative to Triticum aestivum L. We analyzed grain and straw yield, yielding parameters, chemical composition and bakery quality of these species and compared them with modern T. aestivum at three sites with different soil-climate conditions. The average grain yield varied from 0.41 t ha^?1 (T. monococcum) to 5.17 t ha^?1 (T. aestivum), straw yield varied from 1.50 t ha^?1 (T. dicoccum) to 5.83 t ha^?1 (T. aestivum). The yielding parameters and chemical composition of the grain were significantly influenced by soil-climate conditions and wheat species. The highest average crude protein content was recorded in T. spelta (20.55%), while the lowest in T. aestivum (11.20%). The Zeleny’s sedimentation test ranged from 9.0 ml (T. monococcum) to 34.5 ml (T. aestivum) and the value of the Gluten index varied from 7.45 (T. dicoccum) to 89.75 (T. aestivum). According to the results, ancient wheat species provides lower grain and straw yields, higher protein content and mineral concentrations. Concentration of proteins and grain’s baking quality strongly depends on wheat species and soil-climate conditions.     

Capillary Electrophoresis of Gliadins as a Tool in the Discrimination and Characterization of Hulled Wheats (Triticum dicoccon Schrank and T. spelta L.)

Twenty‐two lines of emmer (T. dicoccon Schrank) and 10 of spelt (T. spelta L.) were analyzed using capillary electrophoresis for their gliadins. These proteins were separated on an uncoated fused‐silica capillary (30 cm long, 22 cm to detector, 50 μm i.d.) using the isoelectric buffer 40 mM aspartic acid, 4M urea, 0.5% (w/v) HEC, and 20% (v/v) acetonitrile. Samples were run for 20 min at 22kV and 42°C. By using these conditions, gliadins were separated into 21–30 components (peaks and shoulders). The major peaks eluted between 4.5 and 8.5 min. Electrophoregrams of tested lines showed qualitative and quantitative differences, including number of peaks, presence or absence of some major peaks, and areas of peaks. Lines belonging to the same species can be discriminated mainly on the basis of β‐ and ω‐gliadin patterns. The γ‐and ω‐gliadins seem to be more useful in the differentiation of emmer from spelt. The comparison of electrophoregrams relative to hulled and unhulled species evidenced the high similarity between species with the same genome composition (durum wheat‐emmer, and common wheat‐spelt).     

Ferulic acid from aleurone determines the antioxidant potency of wheat grain (Triticum aestivum L.)

Grain is an important source of phytochemicals, which have potent antioxidant capacity. They have been implicated in the beneficial health effect of whole grains in reducing cardiovascular disease and type 2 diabetes. The aim of the present study was to identify the most important antioxidant fractions of wheat grain. It was found that the aleurone content of these fractions was highly correlated with the antioxidant capacity of the fractions (r = 0.96, p < 0.0001). Ferulic acid appeared to be the major contributor to the antioxidant capacity in fractions with higher antioxidant capacity. The contribution of protein was rather limited. It was concluded that the antioxidant potency of wheat grain fractions is predominantly determined by aleurone content, which can be attributed to the presence of relatively large amounts of phenolic compounds, primarily ferulic acid.     

Molecular cloning and characterization of five novel low molecular weight glutenin subunit genes from Tibetan wheat landraces (Triticum aestivum L.)

The low molecular weight glutenin subunits (LMW-GS) are the major components of glutenins and are important for the end-use quality of wheat. Five novel LMW-GS genes (designated as LMW-Jiachazharen, LMW-Bangdadongmai-3, LMW-Jiachaaigan, LMW-Maoyintumai and LMW-Rikezehongmai) were isolated from Tibetan wheat landraces. The coding regions of LMW-Jiachazharen, LMW-Bangdadongmai-3, LMW-Jiachaaigan, LMW-Maoyintumai and LMW-Rikezehongmai were 912, 897, 915, 927 and 906 bp in length, which encoded 302, 297, 303, 307 and 300 amino acid residues, respectively. Analysis of the deduced amino acid sequences showed that the five novel genes were classified as LMW-m type, with the predicted molecular weights of 32,013.97, 31,622.89, 32,107.07, 32,939.41 and 31,731.64 Da, respectively. The LMW-Jiachazharen, LMW-Jiachaaigan, LMW-Maoyintumai possessed seven cysteine residues, which resulted from a single-nucleotide polymorphism (SNP) of the G–A transition. However, except for eight conserved cysteine residues, LMW-Bangdadongmai-3 contained an extra one, as the result of a SNP of the T–C transition. In addition, the corresponding five LMW-GS were identified and confirmed by sodium SDS-PAGE and MALDI-TOF-MS, respectively. Phylogenetic analysis indicated that the five novel genes were glutenin-like proteins and designated as LMW-m type genes. The five novel genes may be new candidate LMW-GS genes with potential value for wheat quality improvement.     

Combined use of gliadins and SSRs to analyse the genetic variability of the Spanish collection of cultivated diploid wheat (Triticum monococcum L. ssp. monococcum)

This work studied the combined use of gliadins and SSRs to analyse inter- and intra-accession variability of the Spanish collection of cultivated einkorn (Triticum monococcum L. ssp. monococcum) maintained at the CRF-INIA. In general, gliadin loci presented higher discrimination power than SSRs, reflecting the high variability of the gliadins. The loci on chromosome 6A were the most polymorphic with similar PIC values for both marker systems, showing that these markers are very useful for genetic variability studies in wheat. The gliadin results indicated that the Spanish einkorn collection possessed high genetic diversity, being the differentiation large between varieties and small within them. Some associations between gliadin alleles and geographical and agro-morphological data were found. Agro-morphological relations were also observed in the clusters of the SSRs dendrogram. A high concordance was found between gliadins and SSRs for genotype identification. In addition, both systems provide complementary information to resolve the different cases of intra-accession variability not detected at the agro-morphological level, and to identify separately all the genotypes analysed. The combined use of both genetic markers is an excellent tool for genetic resource evaluation in addition to agro-morphological evaluation.