Association Mapping of Drought Tolerance Indices in Ethiopian Durum Wheat (Triticum turgidum ssp. durum).

Ethiopia is a major producer of durum wheat in sub-Saharan Africa. However, its production is prone to drought stress as it is fully dependent on rain, which is erratic and unpredictable. This study aimed to detect marker-trait associations (MTAs) and quantitative trait loci (QTLs) related to indices. Six drought tolerance indices, i.e., drought susceptibility index (DSI), geometric mean productivity (GMP), relative drought index (RDI), stress tolerance index (STI), tolerance index (TOL), and yield stability index (YSI) were calculated from least-square means (lsmeans) of grain yield (GY) and traits significantly (p < 0.001) correlated with grain yield (GY) under field drought stress (FDS) and field non-stress (FNS) conditions. GY, days to grain filling (DGF), soil plant analysis development (SPAD) chlorophyll meter, seeds per spike (SPS), harvest index (HI), and thousand kernel weight (TKW) were used to calculate DSI, GMP, RDI, STI, TOL, and YSI drought indices. Accessions, DW084, DW082, DZ004, C037, and DW092 were selected as the top five drought-tolerant based on DSI, RDI, TOL, and YSI combined ranking. Similarly, C010, DW033, DW080, DW124-2, and C011 were selected as stable accessions based on GMP and STI combined ranking. A total of 184 MTAs were detected linked with drought indices at -log10p ≥ 4.0,79 of which were significant at a false discovery rate (FDR) of 5%. Based on the linkage disequilibrium (LD, r 2 ≥ 0.2), six of the MTAs with a positive effect on GY-GMP were detected on chromosomes 2B, 3B, 4A, 5B, and 6B, explaining 14.72, 10.07, 26.61, 21.16, 21.91, and 22.21% of the phenotypic variance, respectively. The 184 MTAs were clustered into 102 QTLs. Chromosomes 1A, 2B, and 7A are QTL hotspots with 11 QTLs each. These chromosomes play a key role in drought tolerance and respective QTL may be exploited by marker-assisted selection for improving drought stress tolerance in wheat.

The genetic diversity of Ethiopian barley genotypes in relation to their geographical origin.

Ethiopia is recognized as a center of diversity for barley, and its landraces are known for the distinct genetic features compared to other barley collections. The genetic diversity of Ethiopian barley likely results from the highly diverse topography, altitude, climate conditions, soil types, and farming systems. To get detailed information on the genetic diversity a panel of 260 accessions, comprising 239 landraces and 21 barley breeding lines, obtained from the Ethiopian biodiversity institute (EBI) and the national barley improvement program, respectively were studied for their genetic diversity using the 50k iSelect single nucleotide polymorphism (SNP) array. A total of 983 highly informative SNP markers were used for structure and diversity analysis. Three genetically distinct clusters were obtained from the structure analysis comprising 80, 71, and 109 accessions, respectively. Analysis of molecular variance (AMOVA) revealed the presence of higher genetic variation (89%) within the clusters than between the clusters (11%), with moderate genetic differentiation (PhiPT = 0.11) and five accessions were detected as first-generation migrants using Monte Carlo resampling methods. The Mantel test revealed that the genetic distance between accessions is poorly associated with their geographical distance. Despite the observed weak correlation between geographic distance and genetic differentiation, for some regions like Gonder, Jimma, Gamo-Gofa, Shewa, and Welo, more than 50% of the landraces derived from these regions are assigned to one of the three clusters.

The barley HvSTP13GR mutant triggers resistance against biotrophic fungi.

High-yielding and stress-resistant crops are essential to ensure future food supply. Barley is an important crop to feed livestock and to produce malt, but the annual yield is threatened by pathogen infections. Pathogens can trigger an altered sugar partitioning in the host plant, which possibly leads to an advantage for the pathogen. Hampering these processes represents a promising strategy to potentially increase resistance. We analysed the response of the barley monosaccharide transporter HvSTP13 towards biotic stress and its potential use for plant protection. The expression of HvSTP13 increased on bacterial and fungal pathogen-associated molecular pattern (PAMP) application, suggesting a PAMP-triggered signalling that converged on the transcriptional induction of the gene. Promoter studies indicate a region that is probably targeted by transcription factors downstream of PAMP-triggered immunity pathways. We confirmed that the nonfunctional HvSTP13GR variant confers resistance against an economically relevant biotrophic rust fungus in barley. Our experimental setup provides basal prerequisites to further decode the role of HvSTP13 in response to biological stress. Moreover, in line with other studies, our experiments indicate that the alteration of sugar partitioning pathways, in a host-pathogen interaction, is a promising approach to achieve broad and durable resistance in plants.

QTL for induced resistance against leaf rust in barley.

Leaf rust caused by Puccinia hordei is one of the major diseases of barley (Hordeum vulgare L.) leading to yield losses up to 60%. Even though, resistance genes Rph1 to Rph28 are known, most of these are already overcome. In this context, priming may promote enhanced resistance to P. hordei. Several bacterial communities such as the soil bacterium Ensifer (syn. Sinorhizobium) meliloti are reported to induce resistance by priming. During quorum sensing in populations of gram negative bacteria, they produce N-acyl homoserine-lactones (AHL), which induce resistance in plants in a species- and genotype-specific manner. Therefore, the present study aims to detect genotypic differences in the response of barley to AHL, followed by the identification of genomic regions involved in priming efficiency of barley. A diverse set of 198 spring barley accessions was treated with a repaired E. meliloti natural mutant strain expR+ch producing a substantial amount of AHL and a transformed E. meliloti strain carrying the lactonase gene attM from Agrobacterium tumefaciens. For P. hordei resistance the diseased leaf area and the infection type were scored 12 dpi (days post-inoculation), and the corresponding relative infection and priming efficiency were calculated. Results revealed significant effects (p<0.001) of the bacterial treatment indicating a positive effect of priming on resistance to P. hordei. In a genome-wide association study (GWAS), based on the observed phenotypic differences and 493,846 filtered SNPs derived from the Illumina 9k iSelect chip, genotyping by sequencing (GBS), and exome capture data, 11 quantitative trait loci (QTL) were identified with a hot spot on the short arm of the barley chromosome 6H, associated to improved resistance to P. hordei after priming with E. meliloti expR+ch. Genes in these QTL regions represent promising candidates for future research on the mechanisms of plant-microbe interactions.

The treasure inside barley seeds: microbial diversity and plant beneficial bacteria.

BACKGROUND: Bacteria associated with plants can enhance the plants' growth and resistance against phytopathogens. Today, growers aim to reduce the use of mineral fertilizers and pesticides. Since phytopathogens cause severe yield losses in crop production systems, biological alternatives gain more attention. Plant and also seed endophytes have the potential to influence the plant, especially seed-borne bacteria may express their beneficiary impact at initial plant developmental stages. In the current study, we assessed the endophytic seed microbiome of seven genetically diverse barley accessions by 16S rRNA gene amplicon sequencing and verified the in vitro plant beneficial potential of isolated seed endophytes. Furthermore, we investigated the impact of the barley genotype and its seed microbiome on the rhizosphere microbiome at an early growth stage by 16S rRNA gene amplicon sequencing. RESULTS: The plant genotype displayed a significant impact on the microbiota in both barley seed and rhizosphere. Consequently, the microbial alpha- and beta-diversity of the endophytic seed microbiome was highly influenced by the genotype. Interestingly, no correlation was observed between the endophytic seed microbiome and the single nucleotide polymorphisms of the seven genotypes. Unclassified members of Enterobacteriaceae were by far most dominant. Other abundant genera in the seed microbiome belonged to Curtobacterium, Paenibacillus, Pantoea, Sanguibacter and Saccharibacillus. Endophytes isolated from barley seeds were affiliated to dominant genera of the core seed microbiome, based on their 16S rRNA gene sequence. Most of these endophytic isolates produced in vitro plant beneficial secondary metabolites known to induce plant resistance. CONCLUSION: Although barley accessions representing high genetic diversity displayed a genotype-dependent endophytic seed microbiome, a core seed microbiome with high relative abundances was identified. Endophytic isolates were affiliated to members of the core seed microbiome and many of them showed plant beneficial properties. We propose therefore that new breeding strategies should consider genotypes with high abundance of beneficial microbes.

Genetic diversity of Ethiopian durum wheat landraces.

Genetic diversity and population structure assessment in crops is essential for marker trait association, marker assisted breeding and crop germplasm conservation. We analyzed a set of 285 durum wheat accessions comprising 215 Ethiopian durum wheat landraces, 10 released durum wheat varieties, 10 advanced durum wheat lines from Ethiopia, and 50 durum wheat lines from CIMMYT. We investigated the genetic diversity and population structure for the complete panel as well as for the 215 landraces, separately based on 11,919 SNP markers with known physical positions. The whole panel was clustered into two populations representing on the one hand mainly the landraces, and on the other hand mainly released, advanced and CIMMYT lines. Further population structure analysis of the landraces uncovered 4 subgroups emphasizing the high degree of genetic diversity within Ethiopian durum landraces. Population structure based AMOVA for both sets unveiled significant (P < 0.001) variation between populations and within populations. Total variation within population accessions (81%, 76%) was higher than total variation between populations (19%, 24%) for both sets. Population structure analysis based genetic differentiation (FST) and gene flow (Nm) for the whole set and the Ethiopian landraces were 0.19 and 0.24, 1.04, and 0.81, respectively indicating high genetic differentiation and limited gene flow. Diversity indices verify that the landrace panel was more diverse with (I = 0.7, He = 0.46, uHe = 0.46) than the advanced lines (I = 0.6, He = 0.42, uHe = 0.42). Similarly, differences within the landrace clusters were observed. In summary a high genetic diversity within Ethiopian durum wheat landraces was detected, which may be a target for national and international wheat improvement programs to exploit valuable traits for biotic and abiotic stresses.

Expression profiling of genes involved in drought stress and leaf senescence in juvenile barley.

BACKGROUND: Drought stress in juvenile stages of crop development and premature leaf senescence induced by drought stress have an impact on biomass production and yield formation of barley (Hordeum vulgare L.). Therefore, in order to get information of regulatory processes involved in the adaptation to drought stress and leaf senescence expression analyses of candidate genes were conducted on a set of 156 barley genotypes in early developmental stages, and expression quantitative trait loci (eQTL) were identified by a genome wide association study. RESULTS: Significant effects of genotype and treatment were detected for leaf colour measured at BBCH 25 as an indicator of leaf senescence and for the expression level of the genes analysed. Furthermore, significant correlations were detected within the group of genes involved in drought stress (r = 0.84) and those acting in leaf senescence (r = 0.64), as well as between leaf senescence genes and the leaf colour (r = 0.34). Based on these expression data and 3,212 polymorphic single nucleotide polymorphisms (SNP) with a minor allele frequency >5% derived from the Illumina 9 k iSelect SNP Chip, eight cis eQTL and seven trans eQTL were found. Out of these an eQTL located on chromosome 3H at 142.1 cM is of special interest harbouring two drought stress genes (GAD3 and P5CS2) and one leaf senescence gene (Contig7437), as well as an eQTL on chromosome 5H at 44.5 cM in which two genes (TRIUR3 and AVP1) were identified to be associated to drought stress tolerance in a previous study. CONCLUSION: With respect to the expression of genes involved in drought stress and early leaf senescence, genotypic differences exist in barley. Major eQTL for the expression of these genes are located on barley chromosome 3H and 5H. Respective markers may be used in future barley breeding programmes for improving tolerance to drought stress and leaf senescence.

Identification of genomic regions involved in tolerance to drought stress and drought stress induced leaf senescence in juvenile barley.

BACKGROUND: Premature leaf senescence induced by external stress conditions, e.g. drought stress, is a main factor for yield losses in barley. Research in drought stress tolerance has become more important as due to climate change the number of drought periods will increase and tolerance to drought stress has become a goal of high interest in barley breeding. Therefore, the aim is to identify quantitative trait loci (QTL) involved in drought stress induced leaf senescence and drought stress tolerance in early developmental stages of barley (Hordeum vulgare L.) by applying genome wide association studies (GWAS) on a set of 156 winter barley genotypes. RESULTS: After a four weeks stress period (BBCH 33) leaf colour as an indicator of leaf senescence, electron transport rate at photosystem II, content of free proline, content of soluble sugars, osmolality and the aboveground biomass indicative for drought stress response were determined in the control and stress variant in greenhouse pot experiments. Significant phenotypic variation was observed for all traits analysed. Heritabilities ranged between 0.27 for osmolality and 0.61 for leaf colour in stress treatment and significant effects of genotype, treatment and genotype x treatment were estimated for most traits analysed. Based on these phenotypic data and 3,212 polymorphic single nucleotide polymorphisms (SNP) with a minor allele frequency >5% derived from the Illumina 9 k iSelect SNP Chip, 181 QTL were detected for all traits analysed. Major QTLs for drought stress and leaf senescence were located on chromosome 5H and 2H. BlastX search for associated marker sequences revealed that respective SNPs are in some cases located in proteins related to drought stress or leaf senescence, e.g. nucleotide pyrophosphatase (AVP1) or serine/ threonin protein kinase (SAPK9). CONCLUSIONS: GWAS resulted in the identification of many QTLs involved in drought stress and leaf senescence of which two major QTLs for drought stress and leaf senescence were located on chromosome 5H and 2H. Results may be the basis to incorporate breeding for tolerance to drought stress or leaf senescence in barley breeding via marker based selection procedures.