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Bipolaris sorokiniana (Cochliobolus sativus) is a fungal pathogen that causes spot blotch, common root rot, and kernel blight in barley and wheat. Four pathotypes (0, 1, 2, and 7) of the fungus were previously identified based on their virulence on three barley differential lines (Bowman, ND 5883, and NDB 112). Although several genome assemblies have been reported for B. sorokiniana, a telomere-to-telomere genome assembly is still lacking for this fungus. In this study, we assembled the genome of a pathotype 0 isolate (ND93-1) of B. sorokiniana using both PacBio HiFi reads and ultralong Oxford Nanopore Technology reads corrected with Illumina paired-end 100-bp short reads. The combined genome assembly of ND93-1 has an estimated size of 35.7 Mb and consists of 16 scaffolds, each having two telomeres. A total of 11,564 protein-coding genes were predicted, including 1,633 genes encoding for secretory proteins and 473 genes for effectors. This telomere-to-telomere genome assembly provides an important resource for comparative genomics and understanding molecular biology of B. sorokiniana and related Bipolaris species. [Figure: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Two genes (TaHRC and Tsn1) conferring susceptibility to Fusarium head blight and tan spot, Septoria nodorum blotch, and spot blotch in wheat were targeted through wide hybridization with maize expressing Cas9 and guide RNA (gRNA). For each gene, two target sites were selected and corresponding gRNA expression cassettes were synthesized and cloned into a binary vector carrying the CRISPR/Cas9-mediated genome editing machinery. The constructed binary vectors were used to transform the hybrid maize Hi-II through an Agrobacterium-mediated approach to generate T0 and T1 plants, which were used to cross with wheat variety Dayn for targeting Tsn1 or the susceptible allele (TaHRC-S) of TaHRC as well as with the near-isogenic line (Day-Fhb1) of Dayn for targeting the resistant allele (TaHRC-R) of TaHRC. Haploid embryos were rescued in vitro from the wide crosses to generate haploid plants. PCR amplification and sequencing indicated that 15 to 33% of the haploid plants contained the target gene with mutations at the target sites. This wheat × maize hybridization combined with genome editing approach provides a useful alternative tool, not only for targeting susceptibility genes to improve disease resistance without regulatory issues, but also for understanding gene function in wheat. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The fungus Pyrenophora tritici-repentis causes tan spot, an important foliar disease of wheat worldwide. The fungal pathogen produces three necrotrophic effectors, namely Ptr ToxA, Ptr ToxB, and Ptr ToxC to induce necrosis or chlorosis in wheat. Both Ptr ToxA and Ptr ToxB are proteins, and their encoding genes have been cloned. Ptr ToxC was characterized as a low–molecular weight molecule 20 years ago but the one or more genes controlling its production in P. tritici-repentis are unknown. Here, we report the genetic mapping, molecular cloning, and functional analysis of a fungal gene that is required for Ptr ToxC production. The genetic locus controlling the production of Ptr ToxC, termed ToxC, was mapped to a subtelomeric region using segregating biparental populations, genome sequencing, and association analysis. Additional marker analysis further delimited ToxC to a 173-kb region. The predicted genes in the region were examined for presence/absence polymorphism in different races and isolates leading to the identification of a single candidate gene. Functional validation showed that this gene was required but not sufficient for Ptr ToxC production, thus it is designated as ToxC1. ToxC1 encoded a conserved hypothetical protein likely located on the vacuole membrane. The gene was highly expressed during infection, and only one haplotype was identified among 120 isolates sequenced. Our work suggests that Ptr ToxC is not a protein and is likely produced through a cascade of biosynthetic pathway. The identification of ToxC1 is a major step toward revealing the Ptr ToxC biosynthetic pathway and studying its molecular interactions with host factors.[Graphic: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Abstract Hexaploid‐derived resistance genes exhibit complex inheritance and expression patterns in tetraploid backgrounds. This study aimed to characterize the inheritance patterns and genomic compatibilities of hexaploid‐derived Fusarium head blight (FHB) resistance genes in tetraploid durum wheat (Triticum durum Desf.). Evaluation of FHB resistance for F1 hybrids of hexaploid ‘Sumai 3’ crossed with tetraploid and hexaploid wheats indicated that Sumai 3‐derived FHB resistance genes exhibit a dominant phenotypic effect seen only in hexaploid hybrids. Alternately, the hexaploid‐derived FHB resistance genes from PI 277012 exhibited complete dominance in the crosses with both tetraploid and hexaploid wheat. FHB evaluation of the F1 hybrids of Sumai 3 and PI 277012 crossed with ‘Langdon’ (LDN)–‘Chinese Spring’ D‐genome substitution lines suggested that chromosomes 2B, 3B, 4B, 5B, 6B, 3A, 4A, 6A, and 7A contain genes that suppress expression of the Sumai 3‐derived FHB resistance, whereas chromosomes 4A, 6A, and 6B contain genes required for expression of PI 277012‐derived FHB resistance. A wide range of segregation for FHB severity (10–90%) was observed in the F2 generation of Sumai 3 crossed with durum cultivars LDN and ‘Divide’, but the distribution of F3 families derived from the most resistant F2 segregants was skewed towards susceptibility. Similar segregation trends were observed in the crosses of PI 277012 with other durum wheats, whereby FHB resistance became slightly diluted over successive generations. These results suggest tetraploid durum wheat contains the unique alleles at multiple gene loci on different chromosomes that positively and/or negatively regulate the expression of hexaploid‐derived FHB resistance genes, which complicate efforts to deploy these genes in durum breeding programs.

Fusarium temperatum (Scaufl. & Munaut) is one of the most important fungal pathogens that cause ear and stalk rots in maize. In this study, we sequenced genomes of two F. temperatum isolates (KFI615 and KFI660) isolated from corn ears in Poland. A total of 110.3 and 116.3 million 100-nucleotide paired-end clean reads were obtained for KFI615 and KFI660, which were assembled into 20 and 18 scaffolds with an estimated genome size of 45.21 and 45.00 Mb, respectively. These genome sequences provide important resources for understanding pathogenicity and biology of the pathogens within the Fusarium fujikuroi complex.

Fusarium head blight (FHB) is a devastating disease in wheat. The use of resistant germplasm from diverse sources can significantly improve resistance to the disease. “Surpresa” is a Brazilian spring wheat cultivar with moderate FHB resistance, different from currently used sources. In this study, we aimed to identify and map the genetic loci for FHB resistance in Surpresa. A mapping population consisting of 187 recombinant inbred lines (RILs) was developed from a cross between Surpresa and a susceptible spring wheat cultivar, “Wheaton.” The population was evaluated for FHB by the point-inoculation method in three greenhouse experiments and four field trials between 2016 and 2018. Mean disease severity for Surpresa and Wheaton was 41.2 and 84.9% across the 3 years of experiments, ranging from 30.3 to 59.1% and 74.3 to 91.4%, respectively. The mean FHB severity of the NILs was 57%, with an overall range from 7 to 100%, suggesting transgressive segregation in the population. The population was genotyped using a two-enzyme genotyping-by-sequencing approach, and a genetic map was constructed with 5,431 single nucleotide polymorphism (SNP) markers. Four QTL for type II resistance were detected on chromosomes 3A, 5A, 6A, and 7A, explaining 10.4–14.4% of the total phenotypic variation. The largest effect QTL was mapped on chromosome 7A and explained 14.4% of the phenotypic variation; however, it co-localized with a QTL governing the days to anthesis trait. A QTL for mycotoxin accumulation was also detected on chromosome 1B, explaining 18.8% of the total phenotypic variation. The QTL for FHB resistance identified in the study may diversify the FHB resistance gene pool and increase overall resistance to the disease in wheat.

Fusarium head blight (FHB) is one of the most destructive diseases in wheat worldwide. Breeding for FHB resistance is hampered by its complex genetic architecture, large genotype by environment interaction, and high cost of phenotype screening. Genomic selection (GS) is a powerful tool to enhance improvement of complex traits such as FHB resistance. The objectives of this study were to (1) investigate the genetic architecture of FHB resistance in a North Dakota State University (NDSU) hard red spring wheat breeding population, (2) test if the major QTL Fhb1 and Fhb5 play an important role in this breeding population; and (3) assess the potential of GS to enhance breeding efficiency of FHB resistance. A total of 439 elite spring wheat breeding lines from six breeding cycles were genotyped using genotyping-by-sequencing (GBS) and 102,147 SNP markers were obtained. Evaluation of FHB severity was conducted in 10 unbalanced field trials across multiple years and locations. One QTL for FHB resistance was identified and located on chromosome arm 1AL, explaining 5.3% of total phenotypic variation. The major type II resistance QTL Fhb1 only explained 3.1% of total phenotypic variation and the QTL Fhb5 was not significantly associated with FHB resistance in this breeding population. Our results suggest that integration of many genes with medium/minor effects in this breeding population should provide stable FHB resistance. Genomic prediction accuracies of 0.22–0.44 were obtained when predicting over breeding cycles in this study, indicating the potential of GS to enhance the improvement of FHB resistance.

Synthetic hexaploid wheat (SHW) can serve as a bridge for the transfer of useful genes from Aegilops tauschii and tetraploid wheat (Triticum turgidum) into common wheat (T. aestivum). The objective of this study was to evaluate 149 SHW lines and their 74 tetraploid parents for their genetic diversity, breeding values and inter-genomic interactions for resistance to Fusarium head blight (FHB). The genetic diversity analysis was performed based on the population structure established using 4,674 and 3,330 polymorphic SNP markers among the SHW lines and tetraploid parents, respectively. The results showed that all T. carthlicum and most T. dicoccum accessions formed different clusters and subpopulations, respectively, whereas all the T. durum, T. polonicum, T. turgidum, and T. turanicum accessions were clustered together, suggesting that T. durum was more closely related to T. polonicum, T. turgidum, and T. turanicum than to T. dicoccum. The genetic diversity of the SHW lines mainly reflected that of the tetraploid parents. The SHW lines and their tetraploid parents were evaluated for reactions to FHB in two greenhouse seasons and at two field nurseries for 2 years. As expected, most of the SHW lines were more resistant than their tetraploid parents in all environments. The FHB severities of the SHW lines varied greatly depending on the Ae. tauschii and tetraploid genotypes involved. Most of the SHW lines with a high level of FHB resistance were generally derived from the tetraploid accessions with a high level of FHB resistance. Among the 149 SHW lines, 140 were developed by using three Ae. tauschii accessions CIae 26, PI 268210, and RL 5286. These SHW lines showed FHB severities reduced by 21.7%, 17.3%, and 11.5%, respectively, with an average reduction of 18.3%, as compared to the tetraploid parents, suggesting that the D genome may play a major role in reducing disease severity in the SHW lines. Thirteen SHW lines consistently showed a high level of FHB resistance compared to the resistant check, Sumai 3, in each environment. These SHW lines will be useful for the development of FHB-resistant wheat germplasm and populations for discovery of novel FHB resistance genes.

Fusarium graminearum is the major causal agent of Fusarium head blight (FHB) in barley and wheat in North America. The fungus not only causes yield loss of the crops but also produces harmful trichothecene mycotoxins [Deoxynivalenol (DON) and its derivatives-3-acetyldeoxynivalenol (3ADON) and 15-acetyldeoxynivalenol (15ADON), and nivalenol (NIV)] that contaminate grains. Previous studies showed a dramatic increase of 3ADON-producing isolates with higher aggressiveness and DON production than the 15ADON-producing isolates in North America. However, the genetic and molecular basis of differences between the two types of isolates is unclear. In this study, we compared transcriptomes of the 3ADON and 15ADON isolates in vitro (in culture media) and in planta (during infection on the susceptible wheat cultivar 'Briggs') using RNA-sequencing. The in vitro gene expression comparison identified 479 up-regulated and 801 down-regulated genes in the 3ADON isolates; the up-regulated genes were mainly involved in C-compound and carbohydrate metabolism (18.6%), polysaccharide metabolism (7.7%) or were of unknown functions (57.6%). The in planta gene expression analysis revealed that 185, 89, and 62 genes were up-regulated in the 3ADON population at 48, 96, and 144 hours after inoculation (HAI), respectively. The up-regulated genes were significantly enriched in functions for cellular import, C-compound and carbohydrate metabolism, allantoin and allantoate transport at 48 HAI, for detoxification and virulence at 96 HAI, and for metabolism of acetic acid derivatives, detoxification, and cellular import at 144 HAI. Comparative analyses of in planta versus in vitro gene expression further revealed 2,159, 1,981 and 2,095 genes up-regulated in the 3ADON isolates, and 2,415, 2,059 and 1,777 genes up-regulated in the 15ADON isolates at the three time points after inoculation. Collectively, our data provides a foundation for further understanding of molecular mechanisms involved in aggressiveness and DON production of the two chemotype isolates of F. graminearum.

Mitogen-activated protein kinases (MAPKs) have been demonstrated to be involved in fungal development, sexual reproduction, pathogenicity and/or virulence in many filamentous plant pathogenic fungi, but genes for MAPKs in the fungal cereal pathogen Bipolaris sorokiniana have not been characterized. In this study, orthologues of three MAPK genes (CsSLT2, CsHOG1 and CsFUS3) and one MAPK kinase kinase (MAPKKK) gene (CsSTE11) were identified in the whole genome sequence of the B. sorokiniana isolate ND90Pr, and knockout mutants were generated for each of them. The ∆Csfus3 and ∆Csste11 mutants were defective in conidiation and formation of appressoria-like structures, showed hypersensitivity to oxidative stress and lost pathogenicity on non-wounded leaves of barley cv. Bowman. When inoculated on wounded leaves of Bowman, the ∆Csfus3 and ∆Csste11 mutants were reduced in virulence compared to the wild type. No morphological changes were observed in the ∆Cshog1 mutants in comparison with the wild type; however, they were slightly reduced in growth under oxidative stress and were hypersensitive to hyperosmotic stress. The ∆Cshog1 mutants formed normal appressoria-like structures but were reduced in virulence when inoculated on Bowman leaves. The ∆Csslt2 mutants produced more vegetative hyphae, had lighter pigmentation, were more sensitive to cell wall degrading enzymes, and were reduced in virulence on Bowman leaves, although they formed normal appressoria like the wild type. Root infection assays indicated that the ∆Cshog1 and ∆Csslt2 mutants were able to infect barley roots while the ∆Csfus3 and ∆Csste11 failed to cause any symptoms. However, no significant difference in virulence was observed for ∆Cshog1 mutants while ∆Csslt2 mutants showed significantly reduced virulence on barley roots in comparison with the wild type. Our results indicated that all of these MAPK and MAPKKK genes are involved in the regulation of fungal development under normal and stress conditions and required for full virulence on barley plants.

A molecular genetic map was constructed and an electrophoretic karyotype was resolved for Cochliobolus sativus, the causal agent of spot blotch of barley and wheat. The genetic map consists of 27 linkage groups with 97 amplified fragment length polymorphism (AFLP) markers, 31 restriction fragment length polymorphism (RFLP) markers, two polymerase chain reaction amplified markers, the mating type locus (CsMAT), and a gene (VHv1) conditioning high virulence on barley cv. Bowman. These linkage groups covered a map distance of 849 cM. The virulence gene VHv1 cosegregated with six AFLP markers and was mapped on one of the major linkage groups. Fifteen chromosome-sized DNAs were resolved in C. sativus isolates ND93-1 and ND90Pr with contour-clamped homogeneous electric field (CHEF) electrophoresis combined with telo-mere probe analysis of comigrating chromosome-sized DNAs. The chromosome sizes ranged from 1.25 to 3.80 Mbp, and the genome size of the fungus was estimated to be approximately 33 Mbp. By hybridizing genetically mapped RFLP and AFLP markers to CHEF blots, 25 of the 27 linkage groups were assigned to specific chromosomes. The barley-specific virulence locus VHv1 was localized on a chromosome of 2.80 Mbp from isolate ND90Pr in the CHEF gel. The total map length of the fungus was estimated to be at least 1,329 cM based on the map distance covered by the linked markers and the estimated gaps. Therefore, the physical to genetic distance ratio is approximately 25 kb/cM. Construction of a high-resolution map around target loci will facilitate the cloning of the genes conferring virulence and other characters in C. sativus by a map-based cloning strategy.

High density lipoprotein (HDL) cholesterol levels are inversely related to the risk of developing coronary heart disease. Apolipoprotein (apo) A-II is the second most abundant HDL apolipoprotein and apoA-II knockout mice show a 70% reduction in HDL cholesterol levels. There is also evidence, using human apoA-II transgenic mice, that apoA-II can prevent hepatic lipase-mediated HDL triglyceride hydrolysis and reduction in HDL size. These observations suggest the hypothesis that apoA-II maintains HDL levels, at least in part, by inhibiting hepatic lipase. To evaluate this, apoA-II knockout mice were crossbred with hepatic lipase knockout mice. Compared to apoA-II-deficient mice, in double knockout mice there were increased HDL cholesterol levels (57% in males and 60% in females), increased HDL size, and decreased HDL cholesteryl ester fractional catabolic rate. In vitro incubation studies of plasma from apoA-II knockout mice, which contains largely apoA-I HDL particles, showed active lipolysis of HDL triglyceride, whereas similar studies of plasma from apoA-I knockout mice, which contains largely apoA-II particles, did not. In summary, these results strongly suggest that apoA-II is a physiological inhibitor of hepatic lipase and that this is at least part of the mechanism whereby apoA-II maintains HDL cholesterol levels.—Weng, W., N. A. Brandenburg, S. Zhong, J. Halkias, L. Wu, X-c. Jiang, A. Tall, and J. L. Breslow. ApoA-II maintains HDL levels in part by inhibition of hepatic lipase: studies in apoA-II and hepatic lipase double knockout mice. J. Lipid Res. 1999. 40: 1064–1070.

The genomes of five Cochliobolus heterostrophus strains, two Cochliobolus sativus strains, three additional Cochliobolus species (Cochliobolus victoriae, Cochliobolus carbonum, Cochliobolus miyabeanus), and closely related Setosphaeria turcica were sequenced at the Joint Genome Institute (JGI). The datasets were used to identify SNPs between strains and species, unique genomic regions, core secondary metabolism genes, and small secreted protein (SSP) candidate effector encoding genes with a view towards pinpointing structural elements and gene content associated with specificity of these closely related fungi to different cereal hosts. Whole-genome alignment shows that three to five percent of each genome differs between strains of the same species, while a quarter of each genome differs between species. On average, SNP counts among field isolates of the same C. heterostrophus species are more than 25× higher than those between inbred lines and 50× lower than SNPs between Cochliobolus species. The suites of nonribosomal peptide synthetase (NRPS), polyketide synthase (PKS), and SSP-encoding genes are astoundingly diverse among species but remarkably conserved among isolates of the same species, whether inbred or field strains, except for defining examples that map to unique genomic regions. Functional analysis of several strain-unique PKSs and NRPSs reveal a strong correlation with a role in virulence.

The class Dothideomycetes is one of the largest groups of fungi with a high level of ecological diversity including many plant pathogens infecting a broad range of hosts. Here, we compare genome features of 18 members of this class, including 6 necrotrophs, 9 (hemi)biotrophs and 3 saprotrophs, to analyze genome structure, evolution, and the diverse strategies of pathogenesis. The Dothideomycetes most likely evolved from a common ancestor more than 280 million years ago. The 18 genome sequences differ dramatically in size due to variation in repetitive content, but show much less variation in number of (core) genes. Gene order appears to have been rearranged mostly within chromosomal boundaries by multiple inversions, in extant genomes frequently demarcated by adjacent simple repeats. Several Dothideomycetes contain one or more gene-poor, transposable element (TE)-rich putatively dispensable chromosomes of unknown function. The 18 Dothideomycetes offer an extensive catalogue of genes involved in cellulose degradation, proteolysis, secondary metabolism, and cysteine-rich small secreted proteins. Ancestors of the two major orders of plant pathogens in the Dothideomycetes, the Capnodiales and Pleosporales, may have had different modes of pathogenesis, with the former having fewer of these genes than the latter. Many of these genes are enriched in proximity to transposable elements, suggesting faster evolution because of the effects of repeat induced point (RIP) mutations. A syntenic block of genes, including oxidoreductases, is conserved in most Dothideomycetes and upregulated during infection in L. maculans, suggesting a possible function in response to oxidative stress.

This study aimed to elucidate the effect of D-genome chromosomes and their homoeologs in A and B genome on Fusarium head blight (FHB) resistance in durum wheat. Evaluation of the 'Langdon' (LDN) durum-'Chinese Spring' (CS) D-genome disomic substitution lines for reaction to FHB indicated that chromosome 5A could affect resistance to FHB by generating variation in the 5AQ gene-controlled spike structure. Also, we found that CS chromosome 6D might contain genes for FHB susceptibility and/or suppression of FHB resistance. Langdon chromosomes 2B and possibly 6A and 6B may contain the genes that enhance FHB resistance. In addition, we added the entire D genome of Aegilops tauschii Coss. RL5286 (2n = 2x = 14, DD) to LDN by producing LDN-RL5286 amphiploid (2n = 6x = 42, AABBDD) and dissected the A. tauschii D genome in LDN background by producing LDN-A. tauschii D genome addition lines. The A. tauschii chromosomes 1D and 5D were found to carry the genes that conditioned FHB resistance when they were individually or concurrently added to LDN. The genes for FHB resistance on these two chromosomes seemed to act additively in LDN. The other A. tauschii chromosomes (i.e., 2D, 3D, 4D, 6D, and possibly 7D) might contain the genes that downregulated or suppressed the FHB resistance-related genes on chromosome 1D or 5D. Apparently, there are complex interactions among the genes on the A. tauschii chromosomes, as well as A- and B-genome chromosomes, for FHB resistance in the durum background. Proper manipulation of the critical chromosomes may lead to the improvement of FHB resistance in durum wheat. [ABSTRACT FROM AUTHOR]

Cochliobolus sativus (anamorph: Bipolaris sorokiniana) causes spot blotch, common root rot, and kernel blight or black point in barley and wheat. However, little is known about the molecular mechanisms underlying the pathogenicity of C. sativus or the molecular basis of resistance and susceptibility in the hosts. This study aims to establish the model grass Brachypodium distachyon as a new model for studying plant-fungus interactions in cereal crops. Six B. distachyon lines were inoculated with five C. sativus isolates. The results indicated that all six B. distachyon lines were infected by the C. sativus isolates, with their levels of resistance varying depending on the fungal isolates used. Responses ranging from hypersensitive response-mediated resistance to complete susceptibility were observed in a large collection of B. distachyon (2n = 2x = 10) and B. hybridum (2n = 4x = 30) accessions inoculated with four of the C. sativus isolates. Evaluation of an F2 population derived from the cross between two of the B. distachyon lines, Bd 1-1 and Bd3-1, with isolate Cs07-47-l showed quantitative and transgressive segregation for resistance to C. sativus, suggesting that the resistance may be governed by quantitative trait loci from both parents. The availability of whole-genome sequences of both the host (.B. distachyon) and the pathogen (C. sativus) makes this pathosystem an attractive model for studying this important disease of cereal crops. [ABSTRACT FROM AUTHOR]

In the tribe Triticeae, several Thinopyrum species have been used as sources of resistance to stem rust (caused by Puccinia graminis Pers.:Pers. f. sp. tritici Eriks. & E. Henn., abbreviated as Pgt) and other wheat (Triticum aestivum L.) diseases. To identify novel sources of resistance to Pgt race TTKSK (Ug99), we evaluated and characterized the stem rust resistance of 242 accessions belonging to five Thinopyrum species, including beach wheatgrass [Th. bessarabicum (Savul. & Rayss) A. Löve], diploid tall wheatgrass [Th. elongatum (Host) D.R. Dewey], intermediate wheatgrass [Th. intermedium (Host) Barkworth & D. R. Dewey], sand cough or sea wheatgrass [Th. junceum (L.) A. Löve], and decaploid tall wheatgrass [Th. ponticum (Podp.) Barkworth & D.R. Dewey]. These accessions were evaluated for seedling reactions to nine Pgt races (TTKSK, TTTTF, TRTTF, RTQQC, QFCSC, TCMJC, TPMKC, TMLKC, and TPP KC), genotyped with molecular markers linked to four stem rust resistance genes (Sr24, Sr25, Sr26, and Sr43) derived from Thinopyrum species, and examined for ploidy levels. All accessions but one (Th. elongatum PI 531718) were resistant to all or most races. Most of the Th. elongatum and Th. ponticum accessions showed near-immunity to all of the races while the accessions of the other three species (Th. bessarabicum, Th. intermedium, and Th. junceum) had varied levels of resistance ranging from near immunity to moderate resistance. Molecular marker analysis showed that most of the markers appeared to be species- or genusspecific rather than linked to a gene of interest, and thus genotyping analysis was of limited value. Comparisons of infection types of accessions based on ploidy level suggested that higher ploidy level was associated with higher levels of stem rust resistance. The results from this study substantiate that the Thinopyrum species are a rich source of stem rust resistance. [ABSTRACT FROM AUTHOR]

Fusarium head blight (FHB) caused by Fusarium graminearum Schwabe causes extensive losses in durum wheat [Triticum turgidum subsp. durum (Desf.) Husn.]. The only verified resistance contributed by a tetraploid wheat species comes from T. turgidum L. subsp. dicoccoides (Körn. ex Asch. & Graebn.) Thell. This research was conducted to determine whether five Tunisian durum lines exhibiting Type II FHB resistance carry the same resistance alleles as the hexaploid wheat 'Frontana', 'Sumai 3', and 'Wangshuibai' and subsp. dicoccoides genotype (Israel A). One hundred thirty-one polymerase chain reaction (PCR)-based markers were used to determine the genetic similarity among 20 genotypes including resistant and susceptible Tunisian lines and adapted tetraploid and hexaploid cultivars. Phenotypic evaluation was conducted in two greenhouse seasons. Cluster analysis of the Tunisian lines based on the genetic markers showed they are genetically distant from known sources of resistance. Tunisian 7, the most resistant line in this study, was distinctly placed on a separate branch from all the other Tunisian lines, indicating the presence of some genetic diversity. Nineteen of the genotypes were haplotyped with PCR-based markers for nine loci on six chromosomes associated with known Type II FHB resistance. The Tunisian lines amplified different fragment sizes from the known resistant hexaploid wheat cultivars and subsp. dicoccoides. Results from this study indicate that these Tunisian lines may contain a novel source of Type II FHB resistance. [ABSTRACT FROM AUTHOR]

We studied genetic variation in Puccinia psidii, a newly introduced rust fungus in Hawaii, using microsatellite markers. All 36 Hawaiian P. psidii isolates analyzed had the same genotype at 11 microsatellite loci, while three genotypes were identified among four isolates from Florida and 11 genotypes among 18 isolates from Brazil. One of the isolates from Florida had 95% similarity to the Hawaiian isolates. We conclude that the genetically homogeneous P. psidii isolates in Hawaii might have derived from the same strain originally introduced into the Islands and are closely related to one of the strains found in Florida. [ABSTRACT FROM AUTHOR]

Fusarium head blight (FHB) is primarily caused by Fusarium graminearum in North America. Isolates of F. graminearum can be identified as one of three chemotypes: 3-acetyl-deoxynivalenol (3ADON), 15-acetyl-deoxynivalenol (15ADON), and nivalenol (NIV). In this study, we characterized E graminearum isolates collected in 1980 to 2000 (old collection) and in 2008 (new collection) from Noah Dakota and found a 15-fold increase of 3ADON isolates in the new collection. Evaluation of randomly selected 3ADON isolates and 15ADON isolates on three spring wheat genotypes (Grandin, Steele-ND, and ND 2710) by single-floret inoculation indicated that the 3ADON population caused a higher disease severity and produced more DON at a significant level than the 15ADON population on Grandin (susceptible to FHB) and ND 2710 (with FHB resistance from Sumai 3). However, no significant differences in disease severity and DON production were observed between the two populations on Steele-ND (with moderate resistance from Triticum dicoccoides). The 3ADON isolates also exhibited a higher DON production in rice culture and produced more spores on agar media than the 15ADON isolates. suggesting a fitness advantage of the newly emerging 3ADON population over the prevalent 15ADON population. Population genetic analyses using DNA markers revealed a significant genetic differentiation between the two populations. The information obtained in this study could have an impact on development of FHB-resistant wheat cultivars and disease management. [ABSTRACT FROM AUTHOR]

The stem rust (Puccinia graminis Pers.:Pers. f.sp. tritici Eriks. and Henn.) resistance gene Sr39, which confers resistance to TTKSK (Ug99), has been incorporated into the wheat (Triticum aestivum L.) genome from Aegilops speltoides in the form of a chromosome translocation but it has not been deployed into adapted cultivars. In this study, we characterized translocation lines carrying Sr39 in four different wheat backgrounds with fluorescent genomic in situ hybridization (GISH) and simple sequence repeat (SSR) markers. The results indicated that RL5711 and RL6082 had translocation chromosomes of comparable structure. The translocation chromosome in PI 600683 has lost Ae. speltoides chromatin in the telomeric end of the 2S long arm. Six translocation lines derived from the cross PI 600683/3*HY438 had translocation chromosomes of comparable structure to the one found in PI 600683. However, one line (P9714-AM03C51), showed a substantial reduction in Ae. speltoides chromatin in the short arm of the translocated chromosome. The study demonstrated that it is apparently feasible to shorten Ae. speltoides chromatin in some wheat-Ae. speltoides translocation lines. These results and the identification of diagnostic SSR markers will be useful in guiding chromosome manipulation efforts to further shorten the Ae. speltoides chromosome segments in these materials. Greenhouse inoculation of translocation lines with stem rust indicated that the Sr39 gene conditions resistance to at least seven stem rust races. [ABSTRACT FROM AUTHOR]

Common root rot (CRR), primarily caused by Cochliobolus sativus, is one of the important soil borne diseases of wheat ( Triticum aestivum L). To gain information about resistance of hard red spring wheat (HRSW) to the disease, forty HRSW cultivars and advanced breeding lines were grown and evaluated in a completely randomized block design for reaction to CRR from 2000 to 2002 in a field plot with high natural inoculum of C. sativus at Williston, ND. Plant root samples during the soft to mid-dough stage were collected and CRR incidences and severities were scored using the subcrown internode index method. Disease severities (DS) varied from 24–61% in 2000 and 2002, but were relatively low in 2001 (19–40%). None of the wheat cultivars was immune to CRR. The genotypes Canon, Forge, Dandy, Ember, Butte 86, Oxen, AC Barrie, Parshall, and 2371 had higher DS (>45%) and exhibited susceptible disease reactions. However, the genotypes ND 722, AC Cadillac, HJ 98, Argent and Scholar had lower levels of DS throughout the study period, and these were better than, or similar to Amidon, the resistant check. The HRSW cultivars with consistent resistance to CRR during the three year evaluation could be valuable resistance sources for use in breeding programs. [ABSTRACT FROM AUTHOR]

Septoria speckled leaf blotch (SSLB), caused by Septoria passerinhi, has emerged as one of the most important foliar diseases of barley in the Upper Midwest region of the United States. To map and tag genes for SSLB resistance, we developed two populations derived from the resistant accessions CIho 4780 and CIho 10644 and the susceptible malting cv. Foster. Segregation analysis of F2 plants or F2:3 families from the Foster/CIho 4780 and Foster/CIho 10644 populations revealed that a single dominant gene conferred resistance at the seedling stage. Bulked segregant analysis identified an amplified fragment length polymorphism marker, E-ACT/M-CAA-170, that co-segregated with the SSLB resistance gene Rsp2 in the Foster/CIho 4780 F2 population. Southern hybridization analysis with DNA from the wheat/barley addition lines localized E-ACT/M-CAA-170 on the short arm of the barley chromosome 5(1H). Restriction fragment length polymorphism analysis with DNA clones previously mapped to the short arm of chromosome 5(1H) placed Rsp2 at a position flanked by the markers Act8 and ksuD14. A sequence-characterized amplified region (SCAR) marker (E-ACT/M-CAA-170a) was developed that co-segregated with not only Rsp2 in the Foster/CIho 4780 population but also resistance gene Rsp3 in the Foster/CIho 10644 population. This result indicates that Rsp3 is closely linked to Rsp2 on the short arm of chromosome 5(1H). The utility of SCAR marker E-ACT/M-CAA-170a for selecting Rsp2 in two different breeding populations was validated. [ABSTRACT FROM AUTHOR]

We used a combination of genomic techniques to monitor chromosomal evolution across hundreds of generations as Escherkhia coil adapted to growth-limiting concentrations of either lactulose, methyl-galactoside, or a 72:28 mixture of the two. DNA microarrays identified 8 unique duplications and 16 unique deletions among 42 evolvants from 23 chemostat experiments. Each mutation was confirmed by sequencing PCR-amplified flanking genomic DNA and, except for one deletion, an insertion sequence was found at the break point. vPCR of insertion sequences identified these same mutations and 16 additional insertions (all confirmed by sequencing). The pattern of genomic evolution is highly reproducible. Statistical analyses show that duplications at lac and mutations in mgl are adaptations specific to lactulose and to methyl-galactoside, respectively. Adaptation to mixed sugars is characterized by similar mutations, but lac duplications and mgl mutations usually arise in different backgrounds, producing ecological specialists for each sugar. This suggests that an antagonistic pleiotropic tradeoff between duplications at lac and mutations in mgl retards the evolution of generalists. Other mutations that repeatedly appear in replicate experiments are adaptations to the chemostat environment and are not specific to one or the other sugar. [ABSTRACT FROM AUTHOR]

Background: Insertion sequences (IS) are small DNA segments capable of transposing within and between prokaryotic genomes, often causing insertional mutations and chromosomal rearrangements. Although several methods are available for locating ISs in microbial genomes, they are either labor-intensive or inefficient. Here, we use vectorette PCR to identify and map the genomic positions of the eight insertion sequences (IS1, 2, 3, 4, 5, 30, 150, and 186) found in E. coli strain CGSC6300, a close relative of MG1655 whose genome has been sequenced. Results: Genomic DNA from strain CGSC6300 was digested with a four-base cutter Rsa I and the resulting restriction fragments ligated onto vectorette units. Using IS-specific primers directed outward from the extreme ends of each IS and a vectorette primer, flanking DNA fragments were amplified from all but one of the 37 IS elements identified in the genomic sequence of MG1655. Purification and sequencing of the PCR products confirmed that they are IS-associated flanking DNA fragments corresponding to the known IS locations in the MG1655 genome. Seven additional insertions were found in strain CGSC6300 indicating that very closely related isolates of the same laboratory strain (the K12 isolate) may differ in their IS complement. Two other E. coli K12 derivatives, TD2 and TD10, were also analyzed by vectorette PCR. They share 36 of the MG1655 IS sites as well as having 16 and 18 additional insertions, respectively. Conclusion: This study shows that vectorette PCR is a swift, efficient, reliable method for typing microbial strains and identifying and mapping IS insertion sites present in microbial genomes. Unlike Southern hybridization and inverse PCR, our approach involves only one genomic digest and one ligation step. Vectorette PCR is then used to simultaneously amplify all IS elements of a given type, making it a rapid and sensitive means to survey IS elements in genomes. The ability to rapidly identify the IS complements of microbial genomes should facilitate subtyping closely related pathogens during disease outbreaks. [ABSTRACT FROM AUTHOR]

We developed and characterized 15 polymorphic microsatellite markers present in the genome of the guava rust fungus, Puccinia psidii. The primers for these microsatellite markers were designed by sequencing clones from a genomic DNA library enriched for a simple sequence repeat (SSR) motif of (AG). All these 15 primer pairs successfully amplified DNA fragments from a sample of 22 P. psidii isolates, revealing a total of 71 alleles. The observed heterozygosity at the 15 loci ranged from 0.05 to 1.00. The SSR markers developed would be useful for population genetics study of the rust fungus. [ABSTRACT FROM AUTHOR]

Acacia koa (koa), a native tree in Hawaii, suffers from a dieback caused by Fusarium oxysporum. Pathogenicity tests, vegetative compatibility group (VCG) tests, and amplified fragment length polymorphisms (AFLP) analyses were conducted on Fusarium isolates recovered from diseased koa. Koa seedling mortality with individual strains ranged from 0 to 85%, with 42% of the strains killing no seedlings. Thus, strains of F. oxysporum recovered from dying koa trees may or may not be virulent. In addition to F. oxysporum, F. pseudocircinatum strains were isolated from diseased koa; however, they were either nonvirulent or had weak virulence. This is the first report of F. pseudocircinatum in Hawaii. The 46 strains of F oxysporum and F. pseudocircinatum were grouped into 16 VCGs, but 86% of the highly virulent strains belonged to VCG 2. In AFLP analyses, strains from the same VCG generally clustered with one another. Identification of the same set of strains using VCG, AFLP, and pathogenicity tests showed that the highly virulent strains are genetically close and that high virulence toward koa is not a property of all strains of F. oxysporum. Thus, VCG 2 with the corresponding AFLP data is a significant biological entity for which we propose the name F. oxysporum f. sp. acaciae to reflect its virulence on koa. [ABSTRACT FROM AUTHOR]

Spot blotch (SB) caused by Bipolaris sorokiniana and powdery mildew (PM) caused by Blumeria graminis f. sp. hordei are two important diseases of barley. To map genetic loci controlling susceptibility and resistance to these diseases, a mapping population consisting of 138 recombinant inbred lines (RILs) was developed from the cross between Bowman and ND5883. A genetic map was constructed for the population with 852 unique single nucleotide polymorphism markers generated by sequencing-based genotyping. Bowman and ND5883 showed distinct infection responses at the seedling stage to two isolates (ND90Pr and ND85F) of Bipolaris sorokiniana and one isolate (Race I) of Blumeria graminis f. sp. hordei. Genetic analysis of the RILs revealed that one major gene (Scs6) controls susceptibility to Bipolaris sorokiniana isolate ND90Pr, and another major gene (Mla8) confers resistance to Blumeria graminis f. sp. hordei isolate Race I, respectively. Scs6 was mapped on chromosome 1H of Bowman, as previously reported. Mla8 was also mapped to the short arm of 1H, which was tightly linked but not allelic to the Rcs6/Scs6 locus. Quantitative trait locus (QTL) analysis identified two QTLs, QSbs-1H-P1 and QSbs-7H-P1, responsible for susceptibility to spot blotch caused by Bipolaris sorokiniana isolate ND85F in ND5883, which are located on chromosome 1H and 7H, respectively. QSbs-7H-P1 was mapped to the same region as Rcs5, whereas QSbs-1H-P1 may represent a novel allele conferring seedling stage susceptibility to isolate ND85F. Identification and molecular mapping of the loci for SB susceptibility and PM resistance will facilitate development of barley cultivars with resistance to the diseases. [ABSTRACT FROM AUTHOR]

ND2710 is a hard red spring wheat line with a very high level of resistance to fusarium head blight (FHB). It was selected from the progeny of a cross between ND2603 (an advanced breeding line derived from the Sumai 3AVheaton cross) and Grandin (a spring wheat cultivar). The FHB resistance of ND2710 is presumably derived from Sumai 3 because the other parents (Grandin and Wheaton) are very susceptible to fHB. To identify and map the quantitative trait loci (QTL) for FHB resistance in ND2710, we developed a mapping population consisting of 233 recombinant inbred lines (RILs) from the cross between ND2710 and the spring wheat cultivar Bobwhite. These RILs along with their parents and checks were evaluated for reactions to fHB in three greenhouse experiments and one field experiment during 2013 to 2014. The population was also genotyped with the wheat 90K iSelect single-nucleotide polymorphism (SNP) assay, and a genetic linkage map was developed with 1,373 nonco segregating SNP markers, which were distributed on all 21 wheat chromosomes spanning 914.98 centimorgans of genetic distance. Genetic analyses using both phenotypic and genotypic data identified one major QTL (iQflib.ndwp-3B) on the short arm of chromosome 3B. and three minor QTL (Qjhb.ndwp-6B, Qfhb.ndwp-2A, and Qfhb.ndwp-6A) on 6B. 2A, and 6A, respectively. The major QTL on 3B was detected in all experiments and explained 5 to 20% of the phenotypic variation, while the three minor QTL on 6B, 2A, and 6A explained 5 to 12% phenotypic variation in at least two experiments, except for Qfhb.ndwp-2A, which was only detected in the field experiment. QJ'hb.ndwp-3B and Qfhb.ndwp-6B were mapped to the genomic regions containing Fhb1 and Fhb2, respectively, confirming that they originated from Sumai 3. The additive effect of the major and minor QTL may contribute to the high level of FHB resistance in ND2710. The SNP markers closely linked to the FHB resistance QTL will be useful for marker-assisted selection of FHB resistance in wheat breeding programs. [ABSTRACT FROM AUTHOR]

Spot blotch, caused by Cochliobolus sativus, is one of the important barley diseases in the northern Great Plains of the United States and the Prairie Provinces of Canada. The disease has been under control for almost five decades due to the use of durable spot blotch resistance derived from the barley line ND B112. However, the emergence of isolate ND4008 with virulence on ND B 112 prompted us to identity new sources of resistance to this new pathotype. In this study, we screened 2,062 barley accessions from the United States Department of Agriculture National Small Grains Collection for spot blotch resistance, and identified 40 barley accessions exhibiting a high level of resistance to isolate ND4008 at the seedling stage. In all, 24 of the barley accessions with seedling resistance also exhibited moderate to high adult plant resistance to ND4008 in greenhouse tests. Seven of the ND4008-resistant barley accessions showed seedling resistance to two other pathotypes (1 and 2) of the pathogen. Genetic study of resistant barley accessions PI 235186, PI 592275, and PI 643242 indicated that a single major dominant gene controls spot blotch resistance to ND4008 in each of these three accessions. These resistant sources are useful for developing barley cultivars with spot blotch resistance to all pathotypes of C. sativus. [ABSTRACT FROM AUTHOR]