Low Sporulation Research Articles

胞子形成能の低い麹菌株の簡便な胞子形成法

胞子形成能の低い麹菌株の簡便な胞子形成法

Natural Isolates of Saccharomyces cerevisiae Display Complex Genetic Variation in Sporulation Efficiency

Sporulation is a well-studied process executed with varying efficiency by diverse yeast strains. We developed a high-throughput method to quantify yeast sporulation efficiency and used this technique to analyze a line cross between a high-efficiency oak tree isolate and a low-efficiency wine strain. We find that natural variation in sporulation efficiency mirrors natural variation in higher eukaryotes: it shows divergence between isolated populations, arises from loci of major effect, and exhibits epistasis. We show that the lower sporulation efficiency of the wine strain results from a failure to initiate sporulation, rather than from slower kinetics of meiosis and spore formation. The two strains differentially regulate many genes involved in aerobic respiration, an essential pathway for sporulation, such that the oak tree strain appears better poised to generate energy from this pathway. We also report that a polymorphism in RME1 that affects sporulation efficiency in laboratory strains also cosegregates with significant phenotypic differences in our cross of natural isolates. These results lay the groundwork for the study of variation in sporulation efficiency among natural isolates of yeast.

Physiological studies on a new isolate of the gut fungus, Smittium culisetae (Trichomycetes: Harpellales), from wetland mosquito larvae, Aedes vexans (Diptera: Culicidae)

Aedes vexans (Diptera: Culicidae) larvae collected from ephemeral lentic habitats in the Haskell-Baker Wetlands, south of Lawrence, Kansas, were infested with a symbiotic fungus. Two isolates of Smittium culisetae, KS-108-02 and KS-108-06, were axenically cultured from the hindguts of the mosquito larvae. Physiological studies were conducted on isolate KS-108-02 to investigate conditions suitable for successful development of the fungus. The effects of temperature, pH, and different carbon and nitrogen sources on the growth of this new isolate were studied in vitro. Growth was measured as mycelium dry weight for all studies. Asexual trichospore production was monitored in the temperature, pH, and nitrogen source studies. Higher temperatures of 30 C and 24 C produced higher growth rates and trichospore production in shorter periods of time compared to temperatures of 18°C and 12°C. Media with lower pH produced greater growth than media with higher pH. Trichospore production was highest at pH 5.0, reaching an average of 1.21 × 105 trichospores/ml, but above pH 5.0 trichospore production decreased. Glucose produced the greatest amount of growth followed by maltose, fructose, and soluble starch when the isolate was tested with various carbon sources. When tryptone concentrations in the culture medium were reduced to 0.1% in the nitrogen-source study, isolate KS-108-02 exhibited some reciprocal effects between growth and sporulation, wherein nitrogen sources that produced higher mycelium dry weight yielded lower sporulation and vice versa. Results of this study differed slightly from previous studies conducted on S. culisetae.

Isolation, Characterization and Preliminary Genetic Analysis of Laboratory Tricyclazole‐resistant Mutants of the Rice Blast Fungus, Magnaporthe grisea

AbstractThe minimum inhibitory concentration of tricyclazole for hyphal melanization (MIC‐H) was adopted to detect the sensitivity of 129 Magnaporthe grisea isolates collected in China in 2000. Results showed that the mean MIC‐H was 0.2 μg/ml and no isolate with a MIC‐H ≥1 μg/ml was detected. Therefore, 1 μg/ml was chosen as a discriminatory dose to identify resistant mutants generated by ultraviolet (UV) radiation. Only three low‐level resistant (R) mutants derived from the sensitive (S) isolate TH16 were obtained. In addition, fitness decrease was observed for all mutants, with lower sporulation ability and pathogenicity to rice than that of the wild strain TH16. Four crosses between S × R and S × S were tested to determine the inheritance mode of resistance during the process of sexual recombination by analysing the sensitivity of hybrid F1 progeny to tricyclazole. Progeny of crosses between a tricyclazole‐sensitive strain and tricyclazole‐resistant mutants segregated in a 1 : 1 (R : S) ratio and no segregation was found in the cross of S × S, indicating that each mutant contained a single gene for resistance. No nucleotide differences leading to amino acid changes in the coding sequences for 1,3,6,8‐tetrahydroxynaphthalene reductase (4HNR) and 1,3,8‐trihydroxynaphthalene reductase (3HNR) were found between resistant mutants and sensitive strains. Therefore, it is preliminarily concluded that tricyclazole resistance in M. grisea was conferred by a one‐locus mutation in a single Mendelian gene other than those encoding for 4HNR or 3HNR.

Unmasking Novel Sporulation Genes in Bacillus subtilis

The Bacillus subtilis transcription factor sigma(E) directs the expression of a regulon of 262 genes, but null mutations in only a small fraction of these genes severely impair sporulation. We have previously reported that mutations in seven sigma(E)-controlled genes cause a mild (2- to 10-fold) defect in sporulation. In this study, we found that pairwise combinations of some of these seven mutations led to strong synthetic sporulation phenotypes, especially those involving the ytrHI operon and ybaN. Double mutants of ybaN and ytrH and of ybaN and ytrI had >10,000-fold lower sporulation efficiencies than the wild type. Thin-section electron microscopy revealed a block in cortex formation for the ybaN ytrH double mutant and coat defects for the ybaN single and ybaN ytrI double mutants. Sporulating cells of a ybaN ytrI double mutant and of a ybaN ytrHI triple mutant exhibited a pronounced loss of dipicolinic acid (DPA) between hours 8 and 24 of sporulation, in contrast to the constant levels seen for the wild type. An analysis of the spore cortex peptidoglycans of the ybaN ytrI and ybaN ytrHI mutants showed striking decreases in the levels of total muramic acid by hour 24 of sporulation. These data, along with the loss of DPA in the mutants, suggest that the developing spores were unstable and that the cortex underwent degradation late in sporulation. The existence of otherwise hidden sporulation pathways indicates that functional redundancy may mask the role of hitherto unrecognized sporulation genes.

Evaluation of components of resistance to powdery mildew in field pea

Field trials were conducted to evaluate 12 field pea (Pisum sativum L.) cultivars and lines (AC Melfort, AC Tamor, Highlight, Tara, 267-PS210713, 89-2910, JI100, JI1543, JI95, JI96, PI273605, PI311112) for resistance to disease progression and components of resistance to powdery mildew (Erysiphe pisi DC.), in comparison to the susceptible check cultivar Radley in 1998 and 1999. The disease progression was measured as area under the disease progress curve (AUDPC) and the components of resistance were leaf area with symptoms (LAS), lesion diameter (LD), lesion numbers (LN), and sporulation (SP). All 12 lines had significantly lower AUDPC, LAS, LN, and SP than the susceptible check, but only five (AC Tamor, Tara, 267-PS210713, 89-2910, JI1543) showed a smaller LD. A greater range of variability was observed for AUDPC, LAS, and LN than for SP among the 12 lines, suggesting that AUDPC, LAS, and LN are more useful for measuring varietal resistance. Since AUDPC, LAS, and LN were highly correlated (P < 0.01), only one component would be required in screening pea lines for powdery mildew resistance. Key words: Powdery mildew, Erysiphe pisi, field pea, Pisum sativum, resistance

Efficient Selection of Hybrids by Protoplast Fusion Using Drug Resistance Markers and Reporter Genes in Saccharomyces cerevisiae

We have developed a selection system for hybrids by protoplast fusion using dominant selective drug resistance markers, Tn 601 ( 903 ) against geneticin and AUR1-C against aureobasidin A, and reporter genes, ADH1 p- PHO5 - ADH1 t and CLN2 p- CYC1 - lacZ , in Saccharomyces cerevisiae . To examine the effectiveness of this system, plasmids with each marker and reporter gene were introduced into auxotrophic sake yeasts. From the resulting transformants, eight colonies were screened by protoplast fusion in combination with the drug resistance markers and the reporter genes. Among them, seven strains were judged as hybrids between parental strains by analysis of growth on a minimal medium. This selection system was applied to wine yeasts having no genetic markers. Six strains were regarded as hybrids between parental strains by polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) analysis of the MET2 gene and by karyotype analysis using a contour-clamped homogeneous electric field (CHEF). We propose that the plotoplast fusion using dominant selective geneticin- and aureobasidin A-resistance markers and reporter genes is useful for the selection of hybrids from wine yeasts, which are homothallic and have low sporulation ability.

Efficient selection of hybrids by protoplast fusion using drug resistance markers and reporter genes in Saccharomyces cerevisiae

We have developed a selection system for hybrids by protoplast fusion using dominant selective drug resistance markers, Tn 601( 903) against geneticin and AUR1-C against aureobasidin A, and reporter genes, ADH1p- PHO5- ADH1t and CLN2p- CYC1- lacZ, in Saccharomyces cerevisiae. To examine the effectiveness of this system, plasmids with each marker and reporter gene were introduced into auxotrophic sake yeasts. From the resulting transformants, eight colonies were screened by protoplast fusion in combination with the drug resistance markers and the reporter genes. Among them, seven strains were judged as hybrids between parental strains by analysis of growth on a minimal medium. This selection system was applied to wine yeasts having no genetic markers. Six strains were regarded as hybrids between parental strains by polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) analysis of the MET2 gene and by karyotype analysis using a contour-clamped homogeneous electric field (CHEF). We propose that the plotoplast fusion using dominant selective geneticin- and aureobasidin A-resistance markers and reporter genes is useful for the selection of hybrids from wine yeasts, which are homothallic and have low sporulation ability.

In vitro fungicide sensitivity of Alternaria species pathogenic to crucifers and identification of Alternaria brassicicola field isolates highly resistant to both dicarboximides and phenylpyrroles

Blackspot, caused by Alternaria spp. is one of the most important diseases in crucifers and its management mainly relies on fungicide applications. However, growers report increasing prevalence of this disease and decline in its control that might be related to the development of resistance in Alternaria species through over-use of fungicides. To test this hypothesis, field isolates of Alternaria species pathogenic to crucifers were evaluated in vitro for their sensitivity to different fungicides. Toxicity responses were expressed as effective concentration, which inhibits mycelial growth by 50% (EC 50). On this basis, three sterol biosynthesis inhibiting fungicides, two of the triazole group (flutriafol and difenoconazole) and one of the imidazole group (prochloraz), performed equally well in reducing the mycelial growth of A. brassicae, A. brassicicola, and A. japonica. Of the two selected dicarboximides, iprodione proved to be more effective than procymidone. Most of the tested isolates were also very sensitive to fludioxonil (phenylpyrroles family). However, in vitro tests for mycelial growth, conidial germination and germ-tube elongation revealed the existence of A. brassicicola isolates highly resistant (EC 50>100 mg/l) to both dicarboximides (e.g. iprodione and procymidone) and phenylpyrroles (e.g. fludioxonil). These resistant isolates did not exhibit lower sporulation capacities or reduced aggressiveness towards host-plants compared to sensitive A. brassicicola isolates. To our knowledge, this is the first report of fungal field isolates highly cross-resistant to dicarboximides and phenylpyrroles.

Sporulation of Metarhizium anisopliae and Beauveria bassiana on Coptotermes formosanus and in vitro

The sporulation of 22 total isolates of Metarhizium anisopliae and Beauveria bassiana was quantified on cadavers of the Formosan subterranean termite, Coptotermes formosanus. Conidial production increased significantly over 11 days post-death. Effects of isolates of M. anisopliae and B. bassiana on in vivo sporulation were significant. Although the overall effects of fungal species on in vivo sporulation were not significant, the interactions between fungal species and certain times post-death were significant, indicating different sporulation patterns between the two fungal species. B. bassiana isolates could be categorized into a group with high total sporulation (day 11) and low quick sporulation (on days 2 and 3), while M. anisopliae isolates fell into another group with high quick sporulation and low total sporulation. This could give M. anisopliae an advantage over B. bassiana in termite microbial control due to termite defensive social behaviors. Conidial production was significantly higher in vitro than in vivo. In vitro and in vivo sporulation differed by as much as 89x and 232x among the selected isolates of B. bassiana and M. anisopliae, respectively. Correlation between in vivo and in vitro conidial production was positive and significant. This may allow preliminary in vitro screening of a large number of isolates for high in vivo sporulation.

Crescimento e esporulação de isolados de Verticillium lecanii sob diferentes condições nutricionais

O objetivo deste trabalho foi determinar as melhores condições de crescimento e esporulação de dois isolados de Verticillium lecanii (Zimm.) Viégas, cultivados em vários meios de cultura, fontes de C, fontes orgânicas e inorgânicas de N e relações C:N. Após 20 dias de cultivo, BDA e Meio Completo mostraram ser os melhores meios de cultura, e a lactose e o amido foram as fontes de C que mais favoreceram o crescimento dos isolados JAB 02 e JAB 45, respectivamente; baixa esporulação foi apresentada por JAB 02, enquanto JAB 45 obteve boa esporulação em glicose e maltose. JAB 02 desenvolveu melhor quando a fonte orgânica de N era casitona; JAB 45 obteve bom desenvolvimento em presença de casitona e peptona, mas houve pouca esporulação em presença de triptona e caseína hidrolisada. As fontes inorgânicas de N que favoreceram o melhor desenvolvimento dos isolados foram (NH4)2HPO4 e NaNO3, porém, JAB 02 apresentou baixa esporulação. Entre as relações C:N analisadas, JAB 02 obteve melhor crescimento na relação 60:1 e JAB 45 nas relações 5:1, 10:1 e 20:1; baixa esporulação foi obtida por JAB 02 nas relações testadas, mas não se verificou efeito da relação C:N do meio sobre a esporulação dos isolados.

The E1beta and E2 subunits of the Bacillus subtilis pyruvate dehydrogenase complex are involved in regulation of sporulation.

The pdhABCD operon of Bacillus subtilis encodes the pyruvate decarboxylase (E1alpha and E1beta), dihydrolipoamide acetyltransferase (E2), and dihydrolipoamide dehydrogenase (E3) subunits of the pyruvate dehydrogenase multienzyme complex (PDH). There are two promoters: one for the entire operon and an internal one in front of the pdhC gene. The latter may serve to ensure adequate quantities of the E2 and E3 subunits, which are needed in greater amounts than E1alpha and E1beta. Disruptions of the pdhB, pdhC, and pdhD genes were isolated, but attempts to construct a pdhA mutant were unsuccessful, suggesting that E1alpha is essential. The three mutants lacked PDH activity, were unable to grow on glucose and grew poorly in an enriched medium. The pdhB and pdhC mutants sporulated to only 5% of the wild-type level, whereas the pdhD mutant strain sporulated to 55% of the wild-type level. This difference indicated that the sporulation defect of the pdhB and pdhC mutant strains was due to a function(s) of these subunits independent of enzymatic activity. Growth, but not low sporulation, was enhanced by the addition of acetate, glutamate, succinate, and divalent cations. Results from the expression of various spo-lacZ fusions revealed that the pdhB mutant was defective in the late stages of engulfment or membrane fusion (stage II), whereas the pdhC mutant was blocked after the completion of engulfment (stage III). This analysis was confirmed by fluorescent membrane staining. The E1beta and E2 subunits which are present in the soluble fraction of sporulating cells appear to function independently of enzymatic activity as checkpoints for stage II-III of sporulation.

Components of resistance to late leaf spot and rust among interspecific derivatives and their significance in a foliar disease resistance breeding in groundnut ( Arachis hypogaea L.)

Late leaf spot (LLS) and rust cause substantial yield losses and reduce the fodder and seed quality in groundnut (Arachis hypogaea L.). Adoption of resistant cultivars by the semi-arid tropic farmers is the best option to overcome yield losses. Knowledge on components of resistance to these diseases should facilitate the development of groundnut cultivars with enhanced resistance to LLS and rust. The objectives of the experiments were to study the genetic variability and relationships among components of resistance to LLS and rust, and assess their significance in disease resistance breeding. Fifteen interspecific derivatives for LLS and 14 for rust and a susceptible control, TMV 2, were evaluated in a randomised complete block design with two or three replications under greenhouse conditions. The experiments were repeated twice. Genotypic differences were highly significant for all the traits studied. Resistance to LLS is due to longer incubation and latent periods, lesser lesions per leaf, smaller lesion diameter, lower sporulation index, and lesser leaf area damage and disease score. Selection based on components of resistance to LLS may not lead to plants with higher retained green leaf area. The remaining green leaf area on the plant should, therefore, be the major selection criteria for resistance to LLS in breeding programs. Resistance to rust is due to longer incubation and latent periods, fewer pustules per leaf, smaller pustule diameter, lower sporulation index, and lesser leaf area damage and disease score. Rust resistant components appear to work additively, therefore, selection based on resistance components together with green leaf area retained on the plant should be the basis of selecting for resistance to rust in breeding programs. ICGV 99005, 99003, 99012, and 99015 for rust and ICGV 99006, 99013, 99004, 99003, and 99001 for LLS are the better parents for use in resistance breeding programs.

Isolation and properties of genetically defined strains of the methylotrophic yeast Hansenula polymorpha CBS4732.

Genetically defined strains of the yeast Hansenula polymorpha were constructed from a clone of H. polymorpha CBS4732 with very low mating and sporulation abilities. Mating, spore viability, and the percentage of four-spore-containing asci were increased to a level at which tetrad analysis was possible. Auxotrophic mutations in 30 genes were isolated and used to construct strains with multiple markers for mapping studies, transformation with plasmid DNA, and mutant screening. Various other types of mutants were isolated and characterized, among them mutants that displayed an altered morphology, methanol-utilization deficient mutants and strains impaired in the biosynthesis of alcohol oxidase and catalase. Also, the mutability of H. polymorpha CBS4732 vs H. polymorpha NCYC495 was compared. The data revealed clear differences in frequencies of appearance and mutational spectra of some mutants isolated. Many of the mutants isolated had good mating abilities, and diploids resulting from their crossing displayed high sporulation frequencies and high spore viability. Most of the markers used revealed normal Mendelian segregation during meiosis. The frequency of tetratype spore formation was lower than in Saccharomyces cerevisiae suggesting a lower frequency of recombination during the second meiotic division. The properties of genetically defined strains of H. polymorpha CBS4732 as well as their advantages for genetics and molecular studies are discussed.

INFLUENCE OF DIFFERENT PHOSPHORUS MANAGEMENT STRATEGIES ON THE SPORULATION AND GROWTH OF AZOLLA

Phosphorus at the recommended dose of 10 kg P2O5 ha−1 divided between three equal applications, one on each of 0, 7 and 14 d after Azolla inoculation (DAI), increased biomass production but sporulation was invariably inhibited. Of the different options tested, lowering the phosphorus application rate to 4–8 kg P2O5 ha−1 significantly improved sporulation in A. microphylla (strain 202) but there was a substantial reduction in biomass production. On the other hand, changing the schedule of phosphorus application from 0, 7 and 14 DAI to 0, 3 and 6 DAI did not hamper biomass production and improved sporulation frequency and sporocarp number in A. microphylla (strain 202), A. caroliniana and A. pinnata. It was comparable to the no-P treatment for the number of sporocarps, with slightly lower sporulation frequency. In these species, the sporulation frequency and sporocarp number of Azolla enriched with 30 or 60 kg P2O5 ha−1 and then grown without any further added phosphorus were higher than those of unenriched Azolla grown with 10 kg P2O5 ha−1 and mostly comparable to those of Azolla grown without phosphorus. Foliar spray of 2.5 μg ml−1 gibberellic acid (GA) solution (7 DAI) along with the application of 10 kg P2O5 ha−1 (split between 0, 7 and 14 DAI) to unenriched Azolla increased the sporulation frequency and number of sporocarps in A. microphylla (strains 202 and 203), A. caroliniana and A. pinnata, not only over that of phosphorus application alone but also over the untreated control. Combining the use of P-enriched Azolla (A. microphylla strains 202 and 203) with application of GA was more effective for increasing sporulation than was the use of P-enriched Azolla without GA, or application of phosphorus plus GA to unenriched Azolla.

Inheritance of Downy Mildew Resistance in Table Grapes

A study was conducted to determine how resistance to downy mildew [Plasmopara viticola (Bert. & Curt.) Berl. & de Toni] is inherited in germplasm (Vitis vinifera L., V. labrusca L., V. rupestris Scheele, and V. riparia Michx.) used for breeding table grapes. Crosses, including reciprocals, among parents possessing different levels of downy mildew resistance were evaluated in 1994 and 1995. The proportion of foliar tissue with sporulation, chlorosis, or necrosis was used to measure resistance. All genotypes were rated for these characters on two separate dates in 1994 and 1995. Hypersensitive flecking was also evaluated in the 1995 seedlings to determine its relationship with downy mildew resistance. Crosses with at least one resistant parent had a larger number of resistant offspring than crosses between two susceptible parents. General combining ability (GCA) effects were highly significant for 1994 and 1995. Specific combining ability effects were significant, but were relatively small compared to GCA, suggesting additive gene action was a primary influence on downy mildew resistance. Heritability estimates for sporulation, chlorosis, and necrosis were the highest at the second rating in 1994 (0.88, 0.74, and 0.57, respectively) and 1995 (0.50, 0.60, and 0.60, respectively). Reciprocal crosses indicated that maternal inheritance did not influence downy mildew resistance. A small percentage of progeny with hypersensitive flecking were identified from the germplasm. Seedlings with the flecking characteristic tended to have lower sporulation, chlorosis, and necrosis ratings earlier in the growing season.

Development of sorghum downy mildew on sequential plantings of maize in Uganda

Development of sorghum downy mildew (SDM), caused by Perenosclerospora sorghi (Weston and Uppal) C.G. Shaw, was monitored in three to five sequential plantings of six maize genotypes for three seasons in two locations in Uganda. Generally, incidence of SDM was not influenced by time of planting, suggesting that infection was initiated by oospores, the primary inoculum, and not conidia, the secondary inoculum. Genotype effects were highly significant (P<0.001); disease was high in Farida, Pool 16 and NZ 2, but low in Suwan 1. The six maize genotypes were grouped into three categories: resistant (Suwan 1), moderately resistant (102 and Longe 1), and susceptible (Farida, Pool 16 and NZ 2). Sporulation of P. sorghi varied significantly with the genotypes, being highest in the most susceptible genotype (Farida) and least on 102 and Longe 1. Suwan 1 was not tested for sporulation because of low infection. Thus, the resistant genotypes exhibited low incidence of SDM and low sporulation of P. sorghi .

Aggressiveness of Isolates of Phytophthora infestans from the Columbia Basin of Washington and Oregon

ABSTRACT The aggressiveness of 22 isolates of Phytophthora infestans collected from naturally infected potato plants in the Columbia Basin of Washington and Oregon was determined on detached potato leaflets at 18 degrees C in an incubator. Selected isolates were evaluated on whole plants in a greenhouse. Aggressiveness was measured by using the area under the lesion expansion curve (AULEC), incubation period, latent period, sporulation capacity, and lesion size on detached leaflets and the area under the disease progress curve and sporulation capacity on whole plants. The detached-leaflet assay was useful in that a large number of isolates were tested, several components of aggressiveness were studied, and significant differences were found among isolates. Significant variation for components of aggressiveness was found within and among isolates classified according to genotype. Significant interactions among isolates and cultivars were found for some components of aggressiveness, so results were pooled according to cultivar. On average, US-8 and US-11 isolates had higher AULEC scores, indicating aggressiveness higher than that of US-7, US-6, and US-1 genotypes. One US-8 genotype isolate had a higher standardized sporulation capacity than isolates of the other genotypes. US-6 genotype isolates were the least aggressive group, as indicated by low AULEC, sporulation capacity, and lesion size values. The replacement of the US-1 genotype by the US-8 genotype in the Columbia Basin may be partially explained by the increased aggressiveness of US-8 isolates. Additionally, potato growers may need to shorten intervals between fungicide applications and begin applications earlier.

AVALIAÇÃO DE GENÓTIPOS DE SERINGUEIRA (Hevea spp.) AO Microcyclus ulei VISANDO À IDENTIFICAÇÃO DE FONTES DE RESISTÊNCIA HORIZONTAL

Avaliou-se, sob condições controladas, a resistência de genótipos de seringueira ao mal-das-folhas. Empregaram-se dois isolados de Microcyclus ulei, pertencentes aos grupos I e II. Utilizaram-se mudas em sacos plásticos e inocularam-se folíolos nos estádios B1/B2 com uma suspensão de 2 χ 105conídios/ml. Incubaram-se as mudas em câmara úmida a 24° C por 24 horas. Determinaram-se o período latente (PL), diâmetro das lesões (DL), tipo de lesão (TL) e tipo de reação (TR). A maioria dos genótipos testados foi suscetível a pelo menos um isolado do patógeno. Os genótipos que apresentaram resistência moderada, com maior período latente e menor esporulação a um isolado, em geral, foram suscetíveis ao outro. Apenas o genotipo AM/86/271 exibiu resistência incompleta, caracterizada por maior PL, menor DL e baixa esporulação. Entretanto, o número de isolados testados não assegurou o caráter da resistência. Com base nos resultados obtidos concluiu-se que os genótipos avaliados não apresentaram perspectivas para utilização em programas de melhoramento visando resistência ao M.ulei.

Characterization of eds1, a mutation in Arabidopsis suppressing resistance to Peronospora parasitica specified by several different RPP genes.

The interaction between Arabidopsis and the biotrophic oomycete Peronospora parasitica (downy mildew) provides an attractive model pathosystem to identify molecular components of the host that are required for genotype-specific recognition of the parasite. These components are the so-called RPP genes (for resistance to P. parasitica). Mutational analysis of the ecotype Wassilewskija (Ws-0) revealed an RPP-nonspecific locus called EDS1 (for enhanced disease susceptibility) that is required for the function of RPP genes on chromosomes 3 (RPP1/RPP14 and RPP10) and 4 (RPP12). Genetic analyses demonstrated that the eds1 mutation is recessive and is not a defective allele of any known RPP gene, mapping to the bottom arm of chromosome 3 (approximately 13 centimorgans below RPP1/RPP14). Phenotypically, the Ws-eds1 mutant seedlings supported heavy sporulation by P. parasitica isolates that are each diagnostic for one of the RPP genes in wild-type Ws-0; none of the isolates is capable of sporulating on wild-type Ws-0. Ws-eds1 seedlings exhibited enhanced susceptibility to some P. parasitica isolates when compared with a compatible wild-type ecotype, Columbia, and the eds1 parental ecotype, Ws-0. This was observed as earlier initiation of sporulation and elevated production of conidiosporangia. Surprisingly, cotyledons of Ws-eds1 also supported low sporulation by five isolates of P. parasitica from Brassica oleracea. These isolates were unable to sporulate on > 100 ecotypes of Arabidopsis, including wild-type Ws-0. An isolate of Albugo candida (white blister) from B. oleracea also sporulated on Ws-eds1, but the mutant exhibited no alteration in phenotype when inoculated with several oomycete isolates from other host species. The bacterial resistance gene RPM1, conferring specific recognition of the avirulence gene avrB from Pseudomonas syringae pv glycinea, was not compromised in Ws-eds1 plants. The mutant also retained full responsiveness to the chemical inducer of systemic acquired resistance, 2,6-dichloroisonicotinic acid; Ws-eds1 seedlings treated with 2,6-dichloroisonicotinic acid became resistant to the Ws-0-compatible and Ws-0-incompatible P. parasitica isolates Emwa1 and Noco2, respectively. In summary, the EDS1 gene appears to be a necessary component of the resistance response specified by several RPP genes and is likely to function upstream from the convergence of disease resistance pathways in Arabidopsis.