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Fusarium head blight (scab) is a devastating disease of wheat and barley throughout the world. The disease has been reported worldwide wherever cereals are grown, cutting across diverse ecological and geographical distribution. In addition to being pathogenic to plants, which may cause severe crop yield reduction, many Fusarium species are also capable of producing mycotoxins deleterious to human health as secondary metabolites. Fusarium toxins are commonly detected in wheat, barley, maize, rice and beer. Traditionally malted barley (Hordeum vulgare L.) is the principal ingredient in clear beer and Fusarium toxins incidences are of major concern. Moreover, the spent grain from the brewing industry is used as feed and presence of mycotoxins can lead to harmful effects on domestic animals and also find a way into the human food chain. Studies carried out in Kenya have revealed presence of various Fusarium species with ability to produce mycotoxins and presence of Fusarium toxins in wheat and maize and beer. Based on the ubiquitous nature of Fusarium mold and the fact that barley production takes place in maize and wheat growing areas, this study set out to investigate the occurrence of Fusarium molds in Equatorial barley grown in Kenya and the ability of the isolates to produce selected mycotoxins. Grain samples were obtained from newly delivered barley lots originating from two regions and stored grain awaiting malting after break of dormancy from Kenya Maltings Ltd., Nairobi. The Fusarium isolates were identified to species level based on cultural and morphological characteristics. Additionally, they were screened in-vitro on rice cultures for their ability to produce Type A trichothecenes (T-2 toxin, HT-2 toxin, Diacetoxyscirpenol), Type B trichothecenes (deoxynivalenol and nivalenol) and Zearalenone. Samples from all sources were contaminated with Fusarium, but at varying magnitudes - 50%, 33.3% and 25% for barley kernels originating from Timau, Olchoro and in-storage grain with no common history of origin, respectively. The distribution of the species showed some regional specificity. F. graminearum and F. poae predominated in kernels sourced from Olchoro region. All strains of F. graminearum produced both deoxynivalenol and zearalenone. F. poae strains and F. chlamydosporum did not produce detectable amounts of the screened mycotoxins. However, two inconclusively identified isolates of Fusarium spp. isolated from Timau samples produced deoxynivalenol only. The study revealed that a number of toxigenic Fusarium spp. do occur in Equatorial barley grown in Kenya. Key words: Barley, Fusarium, trichothecenes, zearalenone, Kenya., INTRODUCTION Most genera of cultivated plants are susceptible to diseases caused by Fusarium species including all members of the Gramineae family [1]. Fusarium head blight (scab) is a devastating disease [...]

Control of Fusarium head blight (FHB), incited primarily by Fusarium graminearum Schwabe [telomorph Gibberella zea (Schwein)], has been a challenge for barley (Hordeum vulgare L.) producers in the upper midwestern United States for more than a decade. This disease manifests primarily in the form of food safety issues and quality discounts due to the presence of deoxynivalenol (DON), a mycotoxin produced by the pathogen. Cultural practices and fungicides only provide partial control; genetic resistance to FHB and DON accumulation may add to control of the disease. Field studies in North Dakota and China were conducted in FHB disease nurseries using 118 double-haploid lines from the cross C93-3230-24/'Foster'. Foster is susceptible to FHB, and C93-3230-24 is moderately resistant. The quantitative trait locus (QTL) analysis revealed five putative QTL for Type I FHB resistance; however, only the one in bins 7 and 8 of chromosome 2H was detected in more than 70% of the environments. In addition, QTL associated with late days to heading and maturity and tall plant height also were located in the same region. Three putative QTL for increased DON accumulation were detected; however, none were detected in more than three of the eight environments where data were collected. On the basis of these results, it is unlikely that C93-3230-24 has a novel QTL for FHB resistance different from those identified in other resistant accessions. However, C93-3230-24 is unique in that it provides QTL for improved FHB resistance but not reduced DON accumulation. This is an atypical response compared with other sources of FHB resistance and provides evidence that FHB resistance and DON accumulation are under separate genetic control.

Molecular marker technology shows potential to select and combine favorable alleles via genotypic selection and can be used to develop improved crop cultivars with considerable resource savings. In the present investigation, the utility of molecular markers for net blotch resistance identified in Australian breeding material was investigated for Canadian breeding material. Screening 42 Canadian, Australian, and international barley (Hordeum vulgare L.) lines with 10 Canadian Pyrenophora teres Drechs. isolates identified three lines resistant to all isolates evaluated. Net-form net blotch isolates were more virulent than spot-form net blotch isolates. The strong agreement between seedling and adult-plant reactions indicated that seedling screening could be useful for selection for adult-plant resistance. Evaluation of the inheritance of resistance in four Australian populations against Canadian P. teres isolates revealed one to three resistance genes, depending on the isolate used. The majority of Australian barley mapping population parents were susceptible to Canadian P. teres isolates, suggesting markers linked to their resistance may not be useful for Canadian breeding; however, some molecular markers identified by Australian workers could be used for Canadian barley breeding. This study demonstrates that the differences in germplasm and fungal pathogen strains used in different countries result in molecular markers that may not be applicable in all breeding programs for that crop.

Breeding for resistance to Fusarium head blight {FHB; caused by Fusarium graminearum Schwabe [teleomorph Gibberella zeae (Schwein) Petch]} is complicated because there is no accurate method for quantifying the disease organism. Most breeders rely on visual scoring for FHB and deoxynivalenol (DON) analysis to assess disease severity, but both DON and visual scoring are subject to error. The objective of this study was to compare a Fusarium-specific quantifiable enzyme-linked immunosorbent assay (ELISA) to DON and visual assessment of FHB. A doubled-haploid mapping population was grown in two environments and breeding lines in the North American Barley Scab Evaluation Nursery (NABSEN) were grown at four locations. Both experiments used a randomized complete block design and were analyzed for Fusarium by ELISA, DON, and visually scored for FHB. ELISA values for the doubled-haploid lines were consistent over years, and lines that were low in ELISA were also Iow in DON. Broad sense heritability was greater for ELISA (0.48) than DON (0.19) or visual scores of disease (0.29) in the NABSEN study. Numbers of locations and replications necessary to screen for disease were calculated for ELISA, DON, ancl FHB. ELISA required one-third to one-fourth as many replications and locations as did DON or FHB. Temperature and osmotic potential had little effect on mycelial antigen used for ELISA in vitro, but both affected DON. ELISA is a practical alternative to combined visual scores and DON analysis for breeders interested in improving FHB resistance.

Inheritance studies on the nonhost resistance of plants would normally require interspecific crosses that suffer from sterility and abnormal segregation. Therefore, we developed the barley-Puccinia rust model system to study, using forward genetics, the specificity, number, and diversity of genes involved in nonhost resistance. We developed two mapping populations by crossing the line SusPtrit, with exceptional susceptibility to heterologous rust species, with the immune barley cultivars Vada and Cebada Capa. These two mapping populations along with the Oregon Wolfe Barley population, which showed unexpected segregation for resistance to heterologous rusts, were phenotyped with four heterologous rust fungal species. Positions of QTL conferring nonhost resistance in the three mapping populations were compared using an integrated consensus map. The results confirmed that nonhost resistance in barley to heterologous rust species is controlled by QTL with different and overlapping specificities and by an occasional contribution of an R-gene for hypersensitivity. In each population, different sets of loci were implicated in resistance. Few genes were common between the populations, suggesting a high diversity of genes conferring nonhost resistance to heterologous pathogens. These loci were significantly associated with QTL for partial resistance to the pathogen Puccinia hordei and with defense-related genes.

The stem-rust-susceptible barley cv. Golden Promise was transformed by Agrobacterium-mediated transformation of immature zygotic embryos with the Rpg1 genomic clone of cv. Morex containing a 520-bp 5' promoter region, 4,919-bp gene region, and 547-bp 3' nontranscribed sequence. Representatives of 42 transgenic barley lines obtained were characterized for their seedling infection response to pathotype Pgt-MCC of the stem rust fungus Puccinia graminis f. sp. tritici. Golden Promise was converted from a highly susceptible cultivar into a highly resistant one by transformation with the dominant Rpg1 gene. A single copy of the gene was sufficient to confer resistance against stem rust, and progenies from several transformants segregated in a 3:1 ratio for resistance/susceptibility as expected for Mendelian inheritance. These results unequivocally demonstrate that the DNA segment isolated by map-based cloning is the functional Rpg1 gene for stem rust, resistance. One of the remarkable aspects about the transformants is that they exhibit a higher level of resistance than the original sources of Rpg1 (cvs. Chevron and Peatland). In most cases, the Golden Promise transformants exhibited a highly resistant reaction where no visible sign of infection was evident. Hypersensitive necrotic 'fleck' reactions were also observed, but less frequently. With both infection types, pathogen sporulation was prevented. Southern blot and RT-PCR analysis revealed that neither Rpg1 gene copy number nor expression levels could account for the increased resistance observed in Golden Promise transformants. Nevertheless, this research demonstrates that stem-rust-susceptible barley can be made resistant by transformation with the cloned Rpg1 gene.

Stem rust caused by Puccinia graminis f. sp. tritici was among the most devastating diseases of barley in the northern Great Plains of the U.S. and Canada before the deployment of the stem rust-resistance gene Rpg1 in 1942. Since then, Rpg1 has provided durable protection against stem rust losses in widely grown barley cultivars (cvs.). Extensive efforts to clone Rpg1 by synteny with rice provided excellent flanking markers but failed to yield the gene because it does not seem to exist in rice. Here we report the map-based cloning and characterization of Rpg1. A high-resolution genetic map constructed with 8,518 gametes and a 330-kb bacterial artificial chromosome contig physical map positioned the gene between two crossovers [approximately equal to] 0.21 centimorgan and 110 kb apart. The region including Rpg1 was searched for potential candidate genes by sequencing low-copy probes. Two receptor kinase-like genes were identified. The candidate gene alleles were sequenced from resistant and susceptible cvs. Only one of the candidate genes showed a pattern of apparently functional gene structure in the resistant cvs. and defective gene structure in the susceptible cvs. identifying it as the Rpg1 gene. Rpg1 encodes a receptor kinase-like protein with two tandem protein kinase domains, a novel structure for a plant disease-resistance gene. Thus, it may represent a new class of plant resistance genes.

Screening individual barley plants for resistance to stem rust is difficult and time consuming. The objective of this work was to develop PCR-based markers for assisted selection of stem rust resistance in barley. Near isogenic barley lines (backcrossed 29 times) differing at the Rpg1 locus (where the gene that confers resistance to the stem rust pathogen Puccinia graminis f. sp. tritici is located) were screened for random amplified polymorphic DNAs (RAPDs) with 10 base primers. A total of 540 different primers were screened, and one was identified which amplified a DNA fragment closely linked to Rpg1. The DNA fragment was cloned, mapped, and sequenced. Specifically amplifying primers (SAPs) were designed to amplify related polymorphic DNA in other barley cultivars. The SAPs act as a co-dominant marker suitable for marker assisted selection of the Rpg1 gene. Three different amplification patterns were observed in the various cultivars tested. Surprisingly, no specific banding pattern is diagnostic for the presence or absence of Rpg1, despite the demonstrated linkage. Of the three amplification patterns, only one (the 500-bp banding pattern) was present in recently developed resistant cultivars. A similar amplification pattern is also found in many of the susceptible lines that have the cultivar Betzes in their pedigree. RFLP analysis can be used to differentiate the resistant and susceptible cultivars with this amplification pattern.

Analysis of the inheritance of Puccinia hordei resistance in wild and cultivated barley using leaf rust isolates reveals the presence of Riph3 resistance gene in the cultivars. Allelism tests indicate the high frequency origin of Rph3 allele from barley growing in the Mediterranean and Egypt. The accessions of Hordecum spontaneum exhibit higher diversity than those of H. vulgare. The Rph3 gene is recessive for ND89-3 resistance and dominant for ND8702 resistance.

The occurrence of a wheat stem rust (Puccinia graminis Pers.:Pers. f. sp. tritici Eriks. & E. Henn.) pathotype (Pgt-QCC) with virulence for the Rpg1 gene in barley (Hordeum vulgare L.) necessitated the search for resistant barley germplasm. From preliminary screenings of over 18 000 barley accessions, 13 lines were identified as possessing resistance to pathotype QCC: 'Diamond', 'Hietpas 5', Q21861, PC 11, PC 84, PC 249, PC 250, CI 5541, PI 452406, PI 452421, PI 477843, PI 477854, and PI 477860. This study was conducted to further characterize the reaction of the selected lines to pathotype QCC. The reaction was assessed by evaluating infection types at the seedling stage and infection responses at the adult plant stage in the greenhouse, and by evaluating disease severity and infection responses at the adult plant stage in the field compared to susceptible cultivars. Most lines exhibited low to intermediate infection types at the seedlings stage and moderately resistant to moderately susceptible infection responses at the adult plant stage in the greenhouse experiments. Among the selected lines, Q21861 exhibited the highest level of resistance at both the seedling and adult plant stages. These lines may provide an adequate level of resistance to pathotype QCC for cultivar development.

To investigate relationships between barley yellow dwarf virus (BYDV) titer and symptom expression in barley, enzyme-linked immunosorbent assay (ELISA) was used to compare the titers of specific virus isolates in artificially infected plants of cultivars previously assessed for symptom expression in field tests. Four Mexican isolates of BYDV were used: Mex-PAV, Mex-MAV, Mex RPV, and Mex-RMV. The near-isogenic lines 'CM67' (Yd2+) and 'California Mariout' (Yd2-) were compared in most detail. The titer of Mex-PAV was significantly lower in shoots and roots of the relatively tolerant line CM67 3 to 29 d after inoculation than in the relatively sensitive California Mariout. Similar differences were observed, but in roots only, for Mex-MAV infections. For the Mex-RPV and Mex-RMV isolates, no significant differences in virus content occurred between these lines. With each isolate and cultivar, maximum virus content occurred 11 d after inoculation. Assessments at this time indicated relatively low levels of some of the virus isolates used in the relatively tolerant cultivars 'Shyri', 'Laurel', 'Agave', and 'Zarza'. The results with other cultivars were less consistent, but in general their virus contents were relatively high whether they had shown tolerance or sensitivity in field tests. The trends noted remained the same when the data were compared using a 'susceptibility index' designed to provide a convenient comparison of cultivars across serotypes by compensating for differences in the reactivities of different antisera used for ELISA. The need for identification of the specific strains of BYDV involved in tests for tolerance and resistance is emphasized.

The barley yellow dwarf viruses (BYDVs) constitute an important group of plant viruses of cultivated cereal crops in which only tolerance and/or low levels of resistance are known. High levels of resistance, which we here define as low concentrations of virus in plant tissues, have been discovered in Thinopyrum spp. (formerly Agropyron). In this study we have characterized such resistance in [BC.sub.2] and [BC.sub.3] populations, derived by crossing cultivated wheat (Triticum aestivum L. emend. Thell.) with Th. ponticum (Podp.) Barkw. & D.R. Dewey Agropyron Elongatum [Syn (Host) P. Beauv.] and backcrossing to wheat. The objectives were to evaluate the levels of resistance genetically transferable to wheat lines using both enzyme-linked immunosorbent assay (ELISA) and cDNA dot-blot hybridization for the detection of low levels of virus in plant tissues. Uniformity in results using both techniques depends on procedural uniformity, but a high correlation (0.93) between ELISA values and densitometry scans was obtained. A range in resistance was observed among the BC[sub2] and BC[sub3] populations from highly resistant to moderately resistant to susceptible. Lines classified as resistant had densitometry absorbance values of 0.25 to 0.60 and ELISA values from 0.03 to 0.30. The low values .for both ELISA and dot-blot assays indicate that significant resistance can be maintained in more advanced generations. Such low virus levels also suggest that inhibition of virus replication or cell to cell movement may occur in resistant Thinopyrum spp.

Barley yellow dwarf (BYD) is the most economically important and widespread virus disease of small grain cereals in the world. The North American bread wheat (Triticum aestivum L. emend. Thell.) cultivar Anza and several other International Maize and Wheat improvement Center (CIMMYT) wheats have shown tolerance to BYD. Anza and nine other tolerant wheats were studied to understand the genetic basis of tolerance. The tolerant wheats were intercrossed and crossed with the susceptible wheats |Bobwhite' and/or |Bagula'. Parents and the [F.sub.1], and [F.sub.2] generations were tested during 1990 in the field near Toluca (State of Mexico) with the MAV-Mex serotype of barley yellow dwarf virus. Seventy-two individual [F.sub.2]-plant-derived [F.sub.3] lines from each cross were classified in the field during 1991, together with the parents and [F.sub.1]'s, with the same serotype. The intercrosses among tolerant parents failed to segregate, and the observed distribution of [F.sub.3] lines in the crosses of tolerant and susceptible wheats was in ac. cordance with a monogenic segregation ratio. We conclude that the tolerance of Anza and the other nine wheats is due to a common, partially effective and partially dominant gene, designated as Bdv1. Bdv1 could have originated in the Brazilian cultivar Frontana and should be considered a durable source, because of its deployment in numerous CIMMYT wheats worldwide.

Barley yellow dwarf (BYD) is an important viral disease that reduces grain yield in small grains. Breeders usually do not screen for barley yellow dwarf virus (BYDV) tolerance prior to selection of homozygous lines from segregating populations. If early generations of segregating populations could be screened in the greenhouse to increase the frequency of BYDV-tolerant genotypes, the overall efficiency of selection would be improved. The objective was to determine if selection for BYDV tolerance in four F(sub 2) populations of spring oat (Avena sativa L.), which resulted from crosses of tolerant x tolerant parents and sensitive x tolerant parents, is effective in increasing the frequency of tolerant genotypes in the F(sub 4). F(sub 2) populations grown in the greenhouse were infested with viruliferous oat-bird cherry aphids (Rhopalosiphum padi L.) that had been reared on plants infected with the BYDV PAV-IL isolate. Response to selection was evaluated in the F(sub 4) in the field using the same PAV-IL isolate. Selection in the F(sub 2) generation was moderately effective in increasing the frequency of tolerant genotypes in the sensitive x tolerant crosses, but was not effective in changing the mean BYDV tolerance in tolerant x tolerant populations. Heritability estimates calculated in 1990 and 1991 were high (0.5-0.8 and 0.8-0.9, respectively) and confirm that selection based on inoculated hills in the field should be effective in eliminating sensitive genotypes.

A polyclonal antibody directed against antigens on the surface of the germinated conidia of Erysiphe graminis f.sp. hordei E.M. Marchal was used to determine the presence and location of these antigens in host (barley) tissues from 4 to 96 h after inoculation. These studies were carried out using protein A-gold immunolabeling followed by quantitative analysis of labeling density in the nuclei, chloroplasts, vacuoles, and walls of the host cell. Four hours after inoculation the level of labeling detected in mesophyll cells of infected tissues (which are not penetrated by the fungus) was similar to that found in control healthy tissues. However, at 24 h and especially at 96 h after inoculation, the level of labeling in infected tissues was significantly higher than in the healthy tissues. This increase in labeling in infected tissues may be a result of synthesis of co-antigens by the plant as a response to infection, or it may indicate that fungal antigens enter the mesophyll cells very early in the infection process (4 - 24 h) around the time that penetration of the epidermal cell by the infection peg occurs.

Little is known about the inflience of genetic diversity in plant populations on the dynamics of plant viruses, particularly those transmitted by insects. For these viruses, plant genetic diversity may affect virus incidence through impacts on the population dynamics of the vector insects or through impacts on vector feeding behavior, which determines transmission of the virus. This study was designed to explore the influence of plant genetic diversity on virus dispersal by aphid vectors and to examine the biological mechanisms responsible for that influence. In a set of field experiments using the aphid-transmitted barley yellow dwarf virus, I examined the influence of genetic diversity in oat (Avena sativa L.) populations on the spread of the virus and on the population dynamics and movement behavior of aphid vectors of the virus. Only at relatively high aphid abundance were the densities of aphid vectors influenced by plant genetic diversity. In one year out of three, densities of the oat-bird cherry aphid, Rhopalosiphum padi (L.), were significantly lower in the genetically diverse stand than in the genetically homogeneous stands. In no year were densities of the English grain aphid, Sitobion avenae (F.), influenced by the host-plant population. Despite these weak or absent effects on vector abundance, the incidence of the virus was consistently lower in the genetically diverse oat populations. Disease reduction in the diverse populations appears to depend upon changes in aphid movement behavior that affect the efficiency of virus transmission. Mark-release experiments with S. avenae demonstrated that movement rates were significantly higher and plant tenure times were significantly lower in the genetically diverse oat populations. Because the barley yellow dwarf virus requires several hours of aphid feeding for effective transmission, these reduced tenure times and increased travel time among plants led to a reduction of virus transmission. While plant genotype can clearly influence herbivorous insects dramatically, this study suggests that the effects on insects of genetic diversity per se in the host-plant population are likely to be subtle and not easily detected using standard field sampling techniques, except at high insect densities. Yet even at low vector densities, behavioral responses to plant genetic diversity can lead to significant effects on the spread of pathogens.

A trichothecene biosynthesis gene (TRI)-based PCR assay is employed to analyze a collection of 336 isolates from Gibberella zeae from barley, wheat, potatoes, and sugar beets and determine the toxigenic potential of G. zeae. The findings from the studies might be applied to monitor mycotoxin contamination and to formulate Fusarium head blight (FHB) management strategies for these crops.

Powdery mildew of barley, caused by Erysiphe graminis f. sp. hordei, is a model system for investigating the mechanism of gene-for-gene interaction between large-genome cereals and obligate-fungal pathogens. A large number of loci that confer resistance to this disease are located on the short arm of chromosome 5(1H). The Mla resistance-gene cluster is positioned near the telomeric end of this chromosome arm. AFLP-, RAPD-, and RFLP-derived markers were used to saturate the Mla region in a high-resolution recombinant population segregating for the (Mla6 + Mla14) and (Mla13 + Ml-Ru3) resistance specificities. These tightly linked genetic markers were used to identify and develop a physical contig of YAC and BAC clones spanning the Mla cluster. Three distinct NBS-LRR resistance-gene homologue (RGH) families were revealed via computational analysis of low-pass and BAC-end sequence data derived from Mla-spanning clones. Genetic and physical mapping delimited the Mla-associated, NBS-LRR gene families to a 240-kb interval. Recombination within the RGH families was at least 10-fold less frequent than between markers directly adjacent to the Mla cluster.

PAV, one of the luteoviruses inducing the barley yellow dwarf disease, greatly reduces yield of cereal crops and has been reported to cause reddening in several varieties of maize (Zea mays L.). However, there have been no reports that the virus causes significant reductions in size or yield of irrigated maize. Trials to assess the effect of PAV on maize yields were conducted for 3 yr with two hybrids (`Dea' and `Nobilis') and two isolates of PAV (2t and L14). These two isolates differed in geographic origin and virulence on barley (Hordeum vulgare L.), but they multiplied similarly to wild isolates in leaves of maize hybrids. The virus was detected in maize leaves during the same period after inoculation in two out of three years. The reason for low levels of virus in leaves in the 1996 season may have been high temperatures ([is greater than] 30 ° C) following inoculation. Under these conditions, the virus was restricted to the roots for long periods. This blocking of virus migration and/or foliar multiplication was accompanied by less reddening, and there was almost no decrease in grain yield. High early leaf infection of the virus had little effect on the vegetative development of the hybrids ([is less than] 10% reduction in plant height), but grain yield was between 15 and 20% lower for a production of 10 to 12 t [ha.sup.-1]. This drop in yield was due to fewer kernels of maize per ear, while the 1000 kernel mass was unaffected. The loss in yield was not affected by either the virus isolate or the maize hybrid, although the intensity of symptoms differed between hybrids. The data show that there are potential losses incurred by PAV on maize yield. Therefore, these findings justify the establishment of a breeding program., BARLEY YELLOW DWARF VIRUSES (BYDVs) are frequently encountered in maize crops in temperate areas. Those viruses of similar biology designated GPV, MAV, PAV, RMV, RPV, and SGV have been detected [...]