Identifying and genetically characterizing new sources of resistance in barley (Hordeum vulgare) effective against the virulent wheat stem rust (Puccinia graminis f. sp. tritici [Pgt]) population in the Pacific Northwest (PNW) is critical. Isolates from this population, including Pgt isolate Lsp21, were virulent on barley stem rust resistance (R) genes Rpg1, Rpg2, Rpg3, rpg4, Rpg5, and rpg8. Notably, 10% of the Pgt isolates from the population were virulent on barley line Q21861, which contains Rpg1 and rpg4/5 stacked together. Virulence on these 2 broad and effective stem rust R-genes/loci, when combined, is unprecedented Pgt virulence on barley. To discover novel resistance, 277 wild barley (H. vulgare subsp. spontaneum) accessions from the Wild Barley Diversity Collection (WBDC) were screened with Pgt isolate Lsp21. Twelve percent showed moderate resistance, with WBDC-94 and WBDC-238 from Jordan exhibiting exceptional resistance, likely conferred by Rpg7, previously reported in both. To genetically characterize resistance in the WBDC, a genome-wide association study was conducted using disease reactions to Lsp21 and 37,338 genotyping-by-sequencing SNPs. Twelve resistance-associated loci were identified on chromosomes 1H, 2H, 3H, 5H, 6H, and 7H. Rpg7 was not detected due to its low allele frequency in the panel. Importantly, 7 novel resistance loci, WQRpg-2H01, WQRpg-2H02, WQRpg-3H01, WQRpg-5H01, WQRpg-5H03, WQRpg-7H02, and WQRpg-7H03, were discovered. These new sources of resistance can be integrated into cultivated barley, and the associated SNPs will aid in tracking resistance loci in prebreeding lines, enhancing breeding efforts against the virulent PNW Pgt population.

To exploit allelic variation in Hordeum vulgare subsp. spontaneum, the Wild Barley Diversity Collection was subjected to paired-end Illumina sequencing at ∼9 × depth and evaluated for several agronomic traits. We discovered 240.2 million single nucleotide polymorphisms (SNPs) after alignment to the Morex V3 assembly and 24.4 million short (1 to 50 bp) insertions and deletions. A genome-wide association study of lemma color identified one marker-trait association (MTA) on chromosome 1H close to HvBlp, the cloned gene controlling black lemma. Four MTAs were identified for seedling stem rust resistance, including 2 novel loci on chromosomes 1H and 6H and one co-locating to the complex RMRL1-RMRL2 locus on 5H. The whole-genome sequence data described herein will facilitate the identification and utilization of new alleles for barley improvement.

The introduction of invasive microbes compromises the structure, biodiversity, and function of naïve ecosystems. Sphaerulina musiva, a hemibiotrophic pathogen that causes leaf spot and stem cankers in Populus species, exemplifies an invasive fungal pathogen spread by human activities. However, the genetic mechanisms of pathogenicity and virulence are poorly understood, impeding mitigation strategies. We utilized RNA sequencing to identify fungal effectors linked to stem canker formation, informing the development of future strategies for effective disease management. Our analysis revealed 70 genes differentially expressed at 2 weeks and 110 genes at 3 weeks between inoculated trees and controls. Notably, the gene with the highest expression at 2 weeks and the second highest at 3 weeks was homologous to Extracellular protein 2 (Ecp2). Complementary genome-wide association studies linked sequence polymorphisms in this locus to phenotypic variation in disease severity. Infiltration of S. musiva Ecp2 into Populus trichocarpa leaves induced necrosis in susceptible genotypes. Gene disruption using a CRISPR-Cas9 RNP system resulted in a genotype-dependent reduction of stem canker and disease severity. Tracing the evolutionary history of this effector across the fungal kingdom, we uncovered clade-specific gene-family expansions and orthologs in new species. These findings raise questions about the function and adaptive significance of these gene families in fungal lifestyles. Our study provides the first tractable target for breeding resistant poplar genotypes, addressing the challenges of managing S. musiva and uncovering mechanisms that drive its virulence, and provides deeper insights into the evolutionary dynamics of a conserved small-secreted protein with a diversity of functions. IMPORTANCE:Populus species, key feedstocks in the bioeconomy, are severely impacted by leaf spot and stem canker caused by Sphaerulina musiva. This disease diminishes biomass, reduces wood quality, and increases tree mortality, jeopardizing industrial sustainability. Invasion of S. musiva into naïve ecosystems exacerbates these challenges by disrupting ecosystem processes. Breeding resistant poplar genotypes has been the primary strategy to combat this pathogen, but it has remained unclear which molecular drivers of infection breeders should target. Our study makes a significant advance by identifying a key necrotrophic effector that increases S. musiva virulence in specific Populus genotypes. We identify clade-specific gene-family expansions of this effector that raise questions about the function of closely related genes. Our research elucidates the ecology and evolution of a small-secreted protein across the fungal kingdom while offering insights that enable host-breeding efforts to reduce the economic impact of S. musiva.

Wheat stem rust, caused by the obligate biotrophic fungal pathogen Puccinia graminis f. sp. tritici (Pgt), is an important disease of barley and wheat worldwide. Alarmingly, the Pacific Northwest contains a highly virulent Pgt population on barley. This population includes the Pgt isolate Lsp21, which is virulent on the barley stem rust resistance genes Rpg1, Rpg2, Rpg3, rpg4, Rpg5, and rpg8. The virulence on barley lines containing Rpg1 and the rpg4/Rpg5-mediated resistance locus, when stacked together, is a Pgt virulence profile on barley not previously reported. Thus, this population contains the most virulent Pgt isolates on barley resistance genes characterized worldwide. The Elliot line (PI 592261) was identified from the world barley core collection as containing effective seedling resistance to Pgt isolate Lsp21. To genetically characterize the resistance present in Elliot, 129 recombinant inbred lines were developed by advancing a population from the cross Elliot (resistant) x Palmer (susceptible) to the F6 generation. The population was phenotyped with Pgt isolate Lsp21 at the seedling stage and genotyped with the Illumina 50K bead express single-nucleotide polymorphism chip, resulting in 7,284 high-quality single-nucleotide polymorphism markers. Two significant resistance quantitative trait loci (QTLs) (EPRpg_4H-1 and EPRpg_5H-1) contributed by Elliot were identified on chromosomes 4H and 5H, respectively. The major QTL, EPRpg_4H-1, is novel, whereas EPRpg_5H-1 localized to a region approximately 9 Mbp distal of the rpg4/Rpg5-mediated resistance locus within a region of the barley genome that contains previously identified stem rust resistance loci. These QTLs should be useful in developing barley cultivars with resistance to the virulent Pacific Northwest Pgt population.

The genome-wide association study utilizing a diverse global collection of 318 barley accessions identified 44 loci for stripe rust resistance, including 14 potentially novel loci. Stripe rust is an important disease of barley in temperate regions worldwide. Identification and genetic characterization of stripe rust resistance are essential for development and deployment of durable resistance in barley cultivars. A total of 318 spring barley accessions from a global barley collection were evaluated for resistance to the stripe rust pathogen Puccinia striiformis f. sp. hordei (Psh) at the seedling stage in the greenhouse (4–20 °C) using five Psh races (PSH-33, PSH-48, PSH-72, PSH-117 and PSH-118) and at the adult-plant stage in the greenhouse under a high-temperature profile (15–25 °C) and in four field environments in Washington, USA. The frequencies of resistant accessions ranged from 15 to 54

Wheat stem rust caused by the obligate biotrophic fungal pathogen Puccinia graminis f. sp. tritici (Pgt) is an important disease of barley and wheat worldwide. Alarmingly, the Pacific Northwest (PNW) contains a highly virulent Pgt population on barley. This population includes the Pgt isolate Lsp21 which is virulent on the barley stem rust resistance genes Rpg1, Rpg2, Rpg3, rpg4, Rpg5, and rpg8. The virulence on barley lines containing Rpg1 and the rpg4/Rpg5-mediated resistance locus (RMRL), when stacked together is a Pgt virulence profile on barley that had not been previously reported, thus, represents the most virulent Pgt isolates on barley R-genes characterized worldwide. The line Elliot (PI 592261) was identified from the world barley core collection as containing effective seedling resistance to Pgt isolate Lsp21. To genetically characterize the resistance present in Elliot, 129 recombinant inbred lines were developed by advancing a population from the cross Elliot (resistant) x Palmer (susceptible) to the F 6 generation. The population was phenotyped with Pgt isolate Lsp21 at the seedling stage and genotyped with the Illumina 50K bead express SNP chip, resulting in 7,284 high-quality SNP markers. Two significant resistance QTL (EPRpg_4H-1 and EPRpg_5H-1) contributed by Elliot were identified on chromosomes 4H and 5H, respectively. The major QTL, EPRpg_4H-1, is novel, while EPRpg_5H-1 localized to a region ~9 Mbp distal of RMRL within a region of the barley genome that contains previously identified stem rust resistance loci. These QTL should be useful in developing barley cultivars with resistance to the virulent PNW Pgt population.

Cereal rust diseases, including leaf, stem, and stripe rust, are some of the most devastating and economically important diseases of barley. However, host–pathogen genetic interaction research for each pathosystem is typically conducted independently and in isolation. Examples of host resistance/susceptibility genes functioning sympathetically to multiple pathogens or antagonistically to additional pathogens have been reported. Therefore, consolidation of loci that have been reported in multiple studies and across pathosystems is useful for variety development to maximize resistance to multiple pathogens and avoid inadvertent incorporation of susceptibility loci that act antagonistically to other pathogens. This review summarizes loci reported in three key biotrophic pathosystems of barley, including leaf, stem, and stripe rust. In conjunction with previously consolidated net blotch loci, this review lays the foundation for a wider barley rust resistance/susceptibility atlas. This review aims to inform breeders and researchers in rapidly identifying accessions and loci that need further characterization and which loci would be most useful to introgress into elite varieties.

Hybrid genotypes can provide significant yield gains over conventional inbred varieties due to heterosis or hybrid vigor. However, hybrids can also display unintended negative attributes or phenotypes such as extreme pathogen susceptibility. The necrotrophic pathogen Pyrenophora teres f. maculata (Ptm) causes spot form net blotch, which has caused significant yield losses to barley worldwide. Here, we report on a non-transgressive hybrid susceptibility locus in barley identified between the three parental lines CI5791, Tifang and Golden Promise that are resistant to Ptm isolate 13IM.3. However, F2 progeny from CI5791 × Tifang and CI5791 × Golden Promise crosses exhibited extreme susceptibility. The susceptible phenotype segregated in a ratio of 1 resistant:1 susceptible representing a genetic segregation ratio of 1 parental (res):2 heterozygous (sus):1 parental (res) suggesting a single hybrid susceptibility locus. Genetic mapping using a total of 715 CI5791 × Tifang F2 individuals (1430 recombinant gametes) and 149 targeted SNPs delimited the hybrid susceptibility locus designated Susceptibility to Pyrenophora teres 2 (Spt2) to an 198 kb region on chromosome 5H of the Morex V3 reference assembly. This single locus was independently mapped with 83 CI5791 × Golden Promise F2 individuals (166 recombinant gametes) and 180 genome wide SNPs that colocalized to the same Spt2 locus. The CI5791 genome was sequenced using PacBio Continuous Long Read technology and comparative analysis between CI5791 and the publicly available Golden Promise genome assembly determined that the delimited region contained a single high confidence Spt2 candidate gene predicted to encode a pentatricopeptide repeat-containing protein.

Abstract Background Wheat stem rust, caused by Puccinia graminis f. sp. tritici (Pgt), is an important disease of barley and wheat. A diverse sexual Pgt population from the Pacific Northwest (PNW) region of the US contains a high proportion of individuals with virulence on the barley stem rust resistance (R) gene, Rpg1. However, the evolutionary mechanisms of this virulence on Rpg1 are mysterious considering that Rpg1 had not been deployed in the region and the gene had remained remarkably durable in the Midwestern US and prairie provinces of Canada. Methods and results To identify AvrRpg1 effectors, genome wide association studies (GWAS) were performed using 113 Pgt isolates collected from the PNW (n = 89 isolates) and Midwest (n = 24 isolates) regions of the US. Disease phenotype data were generated on two barley lines Morex and the Golden Promise transgenic (H228.2c) that carry the Rpg1 gene. Genotype data was generated by whole genome sequencing (WGS) of 96 isolates (PNW = 89 isolates and Midwest = 7 isolates) and RNA sequencing (RNAseq) data from 17 Midwestern isolates. Utilizing ~1.2 million SNPs generated from WGS and phenotype data (n = 96 isolates) on the transgenic line H228.2c, 53 marker trait associations (MTAs) were identified. Utilizing ~140 K common SNPs generated from combined analysis of WGS and RNAseq data, two significant MTAs were identified using the cv Morex phenotyping data. The 55 MTAs defined two distinct avirulence loci, on supercontig 2.30 and supercontig 2.11 of the Pgt reference genome of Pgt isolate CRL 75-36-700-3. The major avirulence locus designated AvrRpg1A was identified with the GWAS using both barley lines and was delimited to a 35 kb interval on supercontig 2.30 containing four candidate genes (PGTG_10878, PGTG_10884, PGTG_10885, and PGTG_10886). The minor avirulence locus designated AvrRpg1B identified with cv Morex contained a single candidate gene (PGTG_05433). AvrRpg1A haplotype analysis provided strong evidence that a dominant avirulence gene underlies the locus. Conclusions The association analysis identified strong candidate AvrRpg1 genes. Further analysis to validate the AvrRpg1 genes will fill knowledge gaps in our understanding of rust effector biology and the evolution and mechanism/s of Pgt virulence on Rpg1.

Barley net form net blotch (NFNB) is a destructive foliar disease caused by Pyrenophora teres f. teres. Barley line CIho5791, which harbors the broadly effective chromosome 6H resistance gene Rpt5, displays dominant resistance to P. teres f. teres. To genetically characterize P. teres f. teres avirulence/virulence on the barley line CIho5791, we generated a P. teres f. teres mapping population using a cross between the Moroccan CIho5791-virulent isolate MorSM40-3 and the avirulent reference isolate 0-1. Full genome sequences were generated for 103 progenies. Saturated chromosome-level genetic maps were generated, and quantitative trait locus (QTL) mapping identified two major QTL associated with P. teres f. teres avirulence/virulence on CIho5791. The most significant QTL mapped to chromosome (Ch) 1, where the virulent allele was contributed by MorSM40-3. A second QTL mapped to Ch8; however, this virulent allele was contributed by the avirulent parent 0-1. The Ch1 and Ch8 loci accounted for 27 and 15% of the disease variation, respectively, and the avirulent allele at the Ch1 locus was epistatic over the virulent allele at the Ch8 locus. As a validation, we used a natural P. teres f. teres population in a genome-wide association study that identified the same Ch1 and Ch8 loci. We then generated a new reference quality genome assembly of parental isolate MorSM40-3 with annotation supported by deep transcriptome sequencing of infection time points. The annotation identified candidate genes predicted to encode small, secreted proteins, one or more of which are likely responsible for overcoming the CIho5791 resistance.

The foliar disease net form net blotch (NFNB), caused by the necrotrophic fungal pathogen Pyrenophora teres f. teres (Ptt), causes significant yield and quality losses of barley worldwide. Dominant resistance conferred by the Resistance to Pyrenophora teres 5 (Rpt5) gene from barley line CI5791 is the broadest and most effective resistance reported in this pathosystem. The Rpt5 locus was identified in multiple independent genetic studies utilizing diverse host populations and Ptt isolates, and harbors both dominant Rpt5 resistance and isolate-specific susceptibility genes/alleles that are dominant in the absence of Rpt5, designated susceptibility to Pyrenophora teres 1 (Spt1). Ptt virulence and avirulence effectors from diverse pathogen isolates genetically interact with the Rpt5/Spt1 locus, suggesting a complex locus with a function targeted by the evolution of a diversity of pathogen effectors. High-resolution mapping utilizing 1,920 recombinant gametes from a CI5791 x Tifang biparental population, identified 12 candidate genes in an ~4.6 Mb delimited region in the cv Morex V3 genome assembly, but is 1.1 − 2.2 Mb in the pangenome assemblies, containing 5−12 genes. Analysis revealed a strong correlation between the CI5791 allele of a receptor-like protein (RLP), provisionally designated Rpt5 candidate gene 1, (Rcg1), and broad Rpt5−mediated resistance. Two independent transformants of the CI5791 Rcg1 allele in the susceptible cv Golden Promise background showed significantly increased resistance when challenged with Rpt5 avirulent Ptt isolates 6A, 15A, and 0−1 compared to the Golden Promise wildtype. Thus, Rpt5, encodes an RLP and is the first net blotch resistance gene cloned in barley. ### Competing Interest Statement The authors have declared no competing interest.

Net form net blotch (NFNB), caused by Pyrenophora teres f. teres, is an important barley disease. The centromeric region of barley chromosome 6H has often been associated with resistance or susceptibility to NFNB, including the broadly effective dominant resistance gene Rpt5 derived from barley line CIho 5791. We characterized a population of Moroccan P. teres f. teres isolates that had overcome Rpt5 resistance and identified quantitative trait loci (QTL) that were effective against these isolates. Eight Moroccan P. teres f. teres isolates were phenotyped on barley lines CIho 5791 and Tifang. Six isolates were virulent on CIho 5791, and two were avirulent. A CIho 5791 x Tifang recombinant inbred line (RIL) population was phenotyped with all eight isolates and confirmed the defeat of the 6H resistance locus formerly mapped as Rpt5 in barley line CI9819. A major QTL on chromosome 3H with the resistance allele derived from Tifang, as well as minor QTL, was identified and provided resistance against these isolates. F2 segregation ratios supported dominant inheritance for both the 3H and 6H resistance. Furthermore, inoculation of progeny isolates derived from a cross of P. teres f. teres isolates 0-1 (virulent on Tifang/avirulent on CIho 5791) and MorSM 40-3 (avirulent on Tifang/virulent on CIho 5791) onto the RIL and F2 populations determined that recombination between isolates can generate novel genotypes that overcome both resistance genes. Markers linked to the QTL identified in this study can be used to incorporate both resistance loci into elite barley cultivars for durable resistance.

Stem rust, caused by the biotrophic fungal pathogen Puccinia graminis f. sp. tritici ( Pgt), is an important disease of wheat. However, the majority of Pgt virulence/avirulence loci and underlying genes remain uncharacterized due to the constraints of developing bi-parental populations with this obligate biotroph. Genome-wide association studies (GWAS) using a sexual Pgt population mainly collected from the Pacific Northwestern United States were used to identify candidate virulence/avirulence effector genes corresponding to the six wheat Sr genes: Sr5, Sr21, Sr8a, Sr17, Sr9a, and Sr9d. The Pgt isolates were genotyped using whole-genome shotgun sequencing that identified approximately 1.2 million single nucleotide polymorphisms (SNPs) and were phenotyped at the seedling stage on six Sr gene differential lines. Association mapping analyses identified 17 Pgt loci associated with virulence or avirulence phenotypes on six Pgt resistance genes. Among these loci, 16 interacted with a specific Sr gene, indicating Sr-gene specific interactions. However, one avirulence locus interacted with two separate Sr genes ( Sr9a and Sr17), suggesting two distinct Sr genes identifying a single avirulence effector. A total of 24 unique effector gene candidates were identified, and haplotype analysis suggests that within this population, AvrSr5, AvrSr21, AvrSr8a, AvrSr17, and AvrSr9a are dominant avirulence genes, while avrSr9d is a dominant virulence gene. The putative effector genes will be fundamental for future effector gene cloning efforts, allowing for further understanding of rust effector biology and the mechanisms underlying virulence evolution in Pgt with respect to race-specific R-genes. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

The Hessian fly (HF), Mayetiola destructor (Diptera: Cecidomyiidae), is one of the most devastating insect pests of cereals including wheat, barley, and rye. Although wheat is the preferred host for HF, this continuously evolving pest has been emerging as a threat to barley production. However, characterization and identification of genetic resistance to HF has not been conducted in barley. In the present study, we used a genome-wide association study (GWAS) to identify barley resistance loci to HF using a geographically diverse set of 234 barley accessions. The results showed that around 90% of barley lines were highly susceptible, indicating a significant vulnerability to HF in barley, and a total of 29 accessions were resistant, serving as potential resistance resources. GWAS with a mixed linear model revealed two marker-trait associations, both on chromosome 4H. The resistance loci and associated markers will facilitate barley improvement and development for breeders. In addition, our results are fundamental for genetic studies to understand the HF resistance mechanism in barley.

Hybrid genotypes can provide significant yield gains over conventional inbred varieties due to heterosis or hybrid vigor. However, hybrids can also display unintended negative attributes or phenotypes such as extreme pathogen susceptibility. The necrotrophic pathogen Pyrenophora teres f. maculata ( Ptm ) causes spot form net blotch, which has caused significant losses to barley worldwide. Here, we report on a non-transgressive hybrid susceptibility locus in barley initially recognized because the three parental lines CI5791, Tifang and Golden Promise are resistant to Ptm isolate 13IM.3, however F2 progeny from CI5791 × Tifang and CI5791 × Golden Promise crosses exhibited extreme susceptibility. The susceptible phenotype segregated in a ratio of 1 resistant:1 susceptible representing a genetic segregation ratio of 1 parental (res):2 heterozygous (sus):1 parental (res) suggesting a single hybrid susceptibility locus. Genetic mapping using a total of 715 CI5791 × Tifang F2 individuals (1430 recombinant gametes) and 149 targeted SNPs delimited the hybrid susceptibility locus designated Susceptibility to Pyrenophora teres 2 ( Spt2 ) to an ∼198 kb region on chromosome 5H of the Morex V3 reference assembly. This single locus was independently mapped with 83 CI5791 × Golden Promise F2 individuals (166 recombinant gametes) and 180 genome wide SNPs that colocalized to the same Spt2 locus. The CI5791 genome was sequenced using PacBio Continuous Long Read technology and comparative analysis between CI5791 and the publicly available Golden Promise genome assembly determined that the delimited region contained a single high confidence Spt2 candidate gene predicted to encode a pentatricopeptide repeat-containing protein. ### Competing Interest Statement The authors have declared no competing interest. * HR : Hypersensitive Response LOD : Logarithm of Odds NE : Necrotrophic Effector NFNB : Net Form Net Blotch PACE® : PCR Allele Competitive Extension® PCD : Programmed Cell Death PCR-GBS : Polymerase Chain Reaction – Genotyping-by-Sequencing Ptm : Pyrenophora teres f. maculata Ptt : Pyrenophora teres f. teres QTL : Quantitative Trait Loci R Gene/Protein : Resistance Gene/Protein RLK : Receptor-Like Kinase RLP : Receptor-Like Protein Rpt# : Resistance to Pyrenophora teres # SFNB : Spot Form Net Blotch SNP : Single Nucleotide Polymorphism Spt# : Susceptibility to Pyrenophora teres # V8-PDA : V8 Juice – Potato Dextrose Agar WRKY : Tryptophan (W)-Arginine (R)-Lysine (K)-Tyrosine (Y)

Barley net form net blotch (NFNB) is a foliar disease caused by Pyrenophora teres f. teres . Barley line CIho5791, which harbors the chromosome 6H broad spectrum resistance gene Rpt5 , displays dominant resistance to P. teres f. teres . To genetically characterize P. teres f. teres avirulence/virulence on the barley line CIho5791, we generated a P. teres f. teres mapping population using a cross between the Moroccan CIho5791-virulent isolate MorSM40-3, and the avirulent reference isolate 0-1. Genetic maps were generated for all 12 chromosomes (Ch) and quantitative trait locus (QTL) mapping identified two significant QTL associated with P. teres f. teres avirulence/virulence on CIho5791. The most significant QTL mapped to P. teres f. teres Ch1 where the virulent allele was contributed by MorSM40-3. A second QTL mapped to Ch8, however, this virulent allele was contributed by 0-1. The Ch1 and Ch8 loci accounted for 27 and 15% of the disease variation, respectively and the avirulent allele at the Ch1 locus was shown to be epistatic over the virulent allele at the Ch8 locus. Additionally, we used 177 sequenced P. teres f. teres isolates in a genome wide association study that identified the same Ch1 and Ch8 loci as the two most significant associations. Within the identified genomic regions, we identified several genes that encoded small secreted proteins, one or more of which may be responsible for overcoming the CIho5791 resistance. Results presented here demonstrate the complexity of avirulence/virulence in the P. teres f. teres - barley interaction.### Competing Interest StatementThe authors have declared no competing interest.

Polygalacturonase-inhibiting proteins (PGIPs) are cell wall proteins that inhibit pathogen polygalacturonases (PGs). PGIPs, like other defense-related proteins, contain extracellular leucine-rich repeats (eLRRs), which are required for pathogen PG recognition. The importance of these PGIPs in plant defense has been well documented. This study focuses on chickpea (Cicer arietinum) PGIPs (CaPGIPs) owing to the limited information available on this important crop. This study identified two novel CaPGIPs (CaPGIP3 and CaPGIP4) and computationally characterized all four CaPGIPs in the gene family, including the previously reported CaPGIP1 and CaPGIP2. The findings suggest that CaPGIP1, CaPGIP3, and CaPGIP4 proteins possess N-terminal signal peptides, ten LRRs, theoretical molecular mass, and isoelectric points comparable to other legume PGIPs. Phylogenetic analysis and multiple sequence alignment revealed that the CaPGIP1, CaPGIP3, and CaPGIP4 amino acid sequences are similar to the other PGIPs reported in legumes. In addition, several cis-acting elements that are typical of pathogen response, tissue-specific activity, hormone response, and abiotic stress-related are present in the promoters of CaPGIP1, CaPGIP3, and CaPGIP4 genes. Localization experiments showed that CaPGIP1, CaPGIP3, and CaPGIP4 are located in the cell wall or membrane. Transcript levels of CaPGIP1, CaPGIP3, and CaPGIP4 genes analyzed at untreated conditions show varied expression patterns analogous to other defense-related gene families. Interestingly, CaPGIP2 lacked a signal peptide, more than half of the LRRs, and other characteristics of a typical PGIP and subcellular localization indicated it is not located in the cell wall or membrane. The study's findings demonstrate CaPGIP1, CaPGIP3, and CaPGIP4's similarity to other legume PGIPs and suggest they might possess the potential to combat chickpea pathogens.

Spring barley production and quality in the United States is severely impacted by Fusarium head blight (FHB), primarily caused by Fusarium graminearum. Management of FHB is best accomplished using an integrated approach including resistant varieties and fungicides. Field experiments were established from 2014 to 2019 to quantify the effects of host resistance, fungicide, and fungicide timing on management of FHB and deoxynivalenol (DON) and yield protection in spring barley. Three separate research objectives were investigated using multiyear and multilocation experiments. These included investigating the efficacy of prothioconazole + tebuconazole applied at either Feekes growth stage (FGS) 10.5 (full head) or 3 to 7 days after FGS 10.5; the efficacy of sequential fungicide applications occurring at both FGS 10.5 and 3 to 7 days after FGS 10.5 using prothioconazole + tebuconazole, metconazole, prothioconazole, or tebuconazole; and the efficacy and timing of pydiflumetofen + propiconazole. Pooled analysis results suggest that metconazole, prothioconazole + tebuconazole, and pydiflumetofen + propiconazole are all effective fungicides for reducing DON, reducing FHB, and protecting yield. The use of sequential fungicide applications with one occurring at FGS 10.5 and another at 3 to 7 days after FGS 10.5 provided greater suppression than a single application at FGS 10.5. The greatest DON reduction from pydiflumetofen + propiconazole occurred when applied at full head or 3 to 7 days after full head. Across all research objectives, the most effective single fungicide application timing to reduce FHB, reduce DON, and protect yield in barley is 3 to 7 days after FGS 10.5.