Selectivity of SMY 1500 (4-amino-6-tert-butyl-3-ethylthio-l,2,4,triazin-5(4H)-one) in Triticum durum

Susceptibility to SMY 1500 (4-amino-6-tert-butyl-3-ethylthio-1.2.4-triazin-5(4H)-one) was studied in durum wheat (Triticum durum Desf.) cultivars in growth-chamber assays. Weight reduction, photosynthesis inhibition and herbicide levels were determined at various times after a 24-h herbicide treatment in two cultivars of durum wheat, one cultivar of spring wheat (Triticum aestivum L). and in the weed Bromus diandrus (Roth). Measurements of weight reduction distinguished between plants tolerant and plants susceptible to SMY 1500. Tolerant cultivars showed a lower herbicide content and photosynthesis inhibition than susceptible plants after the 24-h treatment. Thereafter, herbicide levels and photosystem II inhibition decreased faster in tolerant plants. Comparison with previous work showed that plants tolerated doses of SMY 1500 about four times higher than of its analogue, metribuzin. Herbicide content in leaves just after treatment, and rate of detoxification may explain the selectivity of SMY 1500 in these cereals.     

Deamination of metribuzin in tolerant and susceptible soybean (Glycine max) cultivars

The deamination of metribuzin was studied in vitro in peroxisomes isolated from the leaves of soybean cultivars which were either metribuzin tolerant, intermediate, or sensitive. The deamination rate observed with peroxisomes from tolerant leaves was about twice the rate observed with peroxisomes from sensitive leaves. The intermediate group was also intermediate with respect to the in-vitro deamination rate. Tolerant and sensitive intact soybean plants were pulse-labeled with [¹⁴C]metribuzin via the roots for 5 h. The extractable radioactivity in roots, stems and leaves was measured and separated into metabolites after the 5 h pulse and after an additional 24 h growth in water. The level of DA (deaminated metribuzin) was always significantly higher in the stems and leaves of tolerant soybean plants (4.8–10.0% of the extracted radioactivity) than in sensitive stems and leaves (1.8–2.9%). Conjugates were rapidly formed in tolerant as well as in sensitive soybean tissues. More conjugates were found in the tolerant cultivars, especially after the 5 + 24 h incubation time. Labeled [¹⁴C]DA fed to soybean plants via the roots was conjugated two to four times faster than [¹⁴C]metribuzin. Tolerant soybean tissue conjugated [¹⁴C] DA two to three times faster than sensitive tissue. The results are interpreted as showing that, in tolerant soybean plants, metribuzin is metabolized via deamination and subsequent conjugation, in addition to the well-known direct conjugation of metribuzin parent compound.     

Metribuzin metabolism as the basis for tolerance in barley (Hordeum vulgare L.)*

Metabolism of ¹⁴C-5 (ring)-metribuzin was studied in Steptoe (tolerant) and Morex (susceptible) barley (Hordeum vulgare L.) cultivars, 1, 4, and 8 days following a single application to roots. Both cultivars contained similar ether-soluble metribuzin metabolites and five water-soluble metabolites. Water-soluble compounds increased from 12 to 53% of the total ¹⁴C recovered for Steptoe and 5–17% for Morex between 1 day and 8 days, respectively, whereas the percentage of ether-soluble metabolites decreased. Ninhydrin reacting compounds were the major water-soluble metabolites in the leaf blades 8 days after treatment. On a d.p.m. mg^?1 dry weight basis, Steptoe leaves had five times more water-soluble material than Morex leaves and half the quantity of ether-soluble compounds. Metribuzin comprised 83 and 89% of the ether-soluble compounds in leaves of Morex and Steptoe, respectively, at 8 days. Terminal radioactivity comprised between 19% and 26% of total radioactivity for both cultivars as early as 1 day after application, with little change over 8 days. Rapid metabolism of metribuzin to non-phytotoxic water-soluble conjugates and terminal residues was the major mechanism responsible for the differential tolerance between these two barley cultivars.     

Effects of isolate virulence and host susceptibility on development of early blight (Alternaria solani) on tomato*

Early blight of tomato (Lycopersicon esculentum) caused by Alternaria solani has the potential to become one of the most serious diseases throughout the tomato‐producing regions of Greece. Controlled environment experiments were conducted to study the virulence of A. solani isolates and the susceptibility of commercial tomato cultivars and hybrids to early blight. The isolates used, derived from naturally infected tomato plants during the period 1997/1998, differed significantly (P>0.05) in the rate of mycelial growth as well as in their ability to sporulate in vitro. No correlation (R²= 0.33) was found between mycelial growth and conidia production. Isolates of A. solani were virulent to young tomato plants (cv. Ace 55VF), although they differed significantly (P>0.05) in the intensity of symptoms produced on leaves, stems, petioles and flowers. Defoliation was linearly related (R²= 0.87) to the percentage of leaf area with symptoms. Twenty‐three tomato cvs. or F₁ hybrids were evaluated for their susceptibility to early blight. The cultivars or hybrids were arbitrarily categorized as immune, highly tolerant, tolerant, moderately tolerant, susceptible and highly susceptible based on a percent disease index range: 0%, 1‐9%, 10‐24%, 25‐49%, 50‐74% and 75% or more respectively. None of the cultivars or hybrids tested was immune or tolerant to A. solani infection.     

Nature of resistance to Striga hermonthica (Del.) Benth. parasitism in some Sorghum vulgare (Pers.) cultivars

Ten Sorghum vulgare (Pers.) cultivars varying in tolerance to Striga hermonthica (Del.) Benth. parasitism were grown with or without Striga infection. Endodermal thickening, pericycle lignification and silica crystal deposition were studied microscopically and measured for infected and non-infected sorghum cultivars. Although differences in the root character measurements were statistically significant they were not closely related to the response of the plant to infection. Low stimulant producing cultivars showed low or medium root cell thickening. The cv. Framida had both low stimulant production and high root cell thickening and was the best of the tolerant cultivars. High stimulant producing, tolerant cultivars generally showed heavy or intermediate cell thickening. The high stimulant producing, susceptible cultivar Debaikri also showed intermediate root cell thickening.‘Antibiosis', measured by the content of phenolic compounds in the plant, was then studied. Varietal differences in quality and quantity of phenolic substances in the roots and shoots of sorghum cultivars infected or non-infected with Striga were observed. Infection increased total phenolic contents in both shoot and root extracts. Differences in the total phenolic content in the shoot of non-infected cultivars did not reflect tolerance to Striga infection. The total phenolic acid content of the root extracts was closely related to the response of the host plant to Striga infection, tolerant cultivars having greater total phenolic acid content than susceptible ones.     

Adzuki bean leaf infection by <Emphasis Type="Italic">Phytophthora vignae</Emphasis> f. sp. <Emphasis Type="Italic">adzukicola</Emphasis> and resistance evaluation using detached leaves inoculated with zoospores

We examined the response of adzuki bean leaves to infection by Phytophthora vignae f. sp. adzukicola and determined whether inoculated leaves can be used to evaluate cultivar resistance. Detached adzuki bean leaves were inoculated with zoospores, and the resulting symptoms were diagnosed. Resistant reactions were characterized by dark brown, speckled lesions or a lack of symptoms, while susceptible reactions were characterized by water-soaked spreading lesions. In an inoculation experiment using a combination of three differential cultivars and three races, the response of 10-day-old primary leaves accurately differentiate between race-specific resistance and susceptibility of adzuki cultivars.     

Rapid chlorophyll fluorescence technique for the study of penetration of photosynthetically active herbicides into leaf tissue

A rapid and sensitive technique, involving analysis of chlorophyll a fluorescence induction kinetics, for the study of the penetration of photosynthetically active herbicides into leaves is described. A range of chlorophyll fluorescence parameters are examined for wheat leaves treated with diuron in order to determine the most appropriate parameter to use in herbicide penetration studies. The usefulness of the technique is demonstrated by examination of the effect of a range of commercially available additives on diuron penetration in wheat leaves.     

Emmer wheat, a potential new host of Tilletia indica

Two representative Italian emmer wheat (Triticum dicoccum) landraces, two selected lines and three improved emmer wheat cultivars, derived from crosses with durum wheat (Molisano landrace × ‘Simeto’), were tested for their susceptibility to Tilletia indica, the cause of Karnal bunt of wheat. Plants of emmer wheat were inoculated by injecting allantoid sporidial suspensions into the boot cavity of plants, just prior to ear emergence. A highly susceptible Indian spring wheat cultivar (Triticum aestivum) was used as a comparative control. At maturity of the plants, the seeds were harvested and assessed for incidence and severity of disease. All emmer wheat genotypes tested were infected but showed differing levels of susceptibility. The percentage of infected seeds for individual genotypes ranged from 5.4 to 75.0% compared with 99.1% for WL-711. The severity of infection was less in the old landraces, but it was higher in all the improved emmer wheat cultivars. In conclusion, Italian cultivars of emmer wheat were found to be highly susceptible to T. indica, and are potentially able to support the establishment of the pathogen.Authors L. Riccioni and M. Valvassori contributed equally to this work and should both be considered as first author.     

Histochemical and chemical evidence for lignin accumulation during the expression of resistance to leaf rust fungi in wheat

Levels of individual phenolic acids were examined in primary leaves of wheat (Triticum aestivum) after inoculation with avirulent and virulent strains of the leaf rust fungus (Puccinia recondita f. sp. tritici) at stages when previous work had shown fungal and host cells to be affected by expression of the Lr20 or Lr28 alleles for resistance. The predominant phenolic acid, ferulic acid, as well as p-coumaric and syringic acids were detected in primary leaves in both unbound and bound forms. They were not detected in germinating urediniospores of either rust strain. Levels of unbound phenolic acids changed little in response to infection. In Lr28-bearing leaves inoculated with an avirulent strain, increased concentrations of bound phenolic acids and three other unidentified compounds were observed about 4 h after many single or small groups of cells had undergone hypersensitive collapse. In an Lr20-bearing cultivar, levels of bound phenolic acids fell in leaves inoculated with either a virulent or avirulent rust strain. Coniferyl aldehyde and coniferyl alcohol were not detected in healthy or inoculated leaves of either wheat cultivar. Attempts to affect expression of resistance by application of inhibitors of phenylalanine ammonia-lyase were not successful and both wheat cultivars remained resistant to avirulent rust strains. The bound phenolic acids which accumulate in cells undergoing a hypersensitive response may play a role in resistance of Lr28-bearing wheat to the leaf rust fungus.     

Metribuzin metabolism in tomato: Isolation and identification of N-glucoside conjugates

Metribuzin [4-amino-6-tert-butyl-3-(methylthio)-1,2,4-triazin-5(4H)-one] metabolism was studied in tomato (Lycopersicon esculentum Mill. “Sheyenne”). Pulse-treatment studies with seedlings and excised leaves showed that [5-¹⁴C]metribuzin was rapidly absorbed, translocated (acropetal), and metabolized to more polar products. Foliar tissues of 19-day-old seedlings metabolized 96% of the root-absorbed [¹⁴C]metribuzin in 120 hr. Excised mature leaves metabolized 85–90% of the petiole-absorbed [¹⁴C]metrubuzin in 48 hr. Polar metabolites were isolated by solvent partitioning, and purified by adsorption, thin-layer, and high-performance liquid chromatography. A minor intermediate metabolite (I) was identified as the polar β-d-(N-glucoside) conjugate of metribuzin. The biosynthesis of (I) was demonstrated with a partially purified UDP-glucose: metribuzin N-glucosyltransferase from tomato leaves. A possible correlation between foliar UDP-glucose: metribuzin N-glucosyltransferase activity levels and differences in the tolerance of selected tomato seedling cultivars to metribuzin was suggested. The major polar metabolite (II) was identified as the malonyl β-d-(N-glucoside) conjugate of metribuzin.     

Phytotoxicity and detoxification of metribuzin in dark-grown suspension cultures of soybean

Differential resistance of soybean to metribuzin (4-amino-6-t-butyl 3-(methylthio)-as-triazin-5(4H)-one) was demonstrated in cell suspensions from resistant and susceptible cultivars. Whereas the herbicide had been reported to inhibit photosynthesis, the observations on achlorophyllous, dark-grown cultures indicated that phytotoxicity was not restricted to photosynthesis. The sites of light-independent herbicide activity were not identified, but the presence of these sites was demonstrated in both the herbicide-susceptible and herbicide-resistant cells. The mechanism to protect these sites was present in all cultures. The mechanism, enzymatic detoxification of the herbicide, was inoperative in the susceptible cultures due to accumulation of a substance which inhibited the enzyme. Resistant cultures metabolized that substance to an ineffectual form. Metribuzin selectivity was therefore determined according to the accessibility of the inhibitor to the enzyme responsible for resistance.     

The contact activity of difenzoquat in two United Kingdom spring wheat cultivars

The contact action of difenzoquat was investigated on seedlings of spring wheat (Triticum aestivum L.) cultivars tolerant (Maris Butler) and susceptible (Sicco) to the translocated etTect of the herbicide on the apical meristem. Applied as a spray the chemical caused an increase in electrolyte leakage before the development of visible damage. Chlorophyll levels were also reduced and both effects were greater on Maris Butler than Sicco. Droplets of difenzoquat caused similar ultrastructural effects on the two cultivars. These included swelling of the chloroplasts with subsequent disorganization of the thylakoid system accompanied by disruption of other cellular components. Similar effects have been reported with paraquat and it is suggested that the contact action of difenzoquat is due to a weak paraquat type activity.     

Microscopic detection of reactive oxygen species generation in the compatible and incompatible interactions of Alternaria alternata Japanese pear pathotype and host plants

 Reactive oxygen species (ROS) generation was examined in the interaction of Alternaria alternata Japanese pear pathotype and host plants using three methods: nitro blue tetrazolium (NBT) method for microscopic detection of O₂ ⁻, diaminobenzidine (DAB) methods for microscopic detection of H₂O₂, and cerium chloride methods for ultrastructural detection of H₂O₂. ROS generation was detected by NBT and DAB methods at appressoria on leaves of susceptible cultivars and heat-shocked leaves of resistant cultivars but not in leaves of resistant cultivars. Ultrastructural detection by the cerium chloride method identified ROS generation at cell walls of appressoria and penetration pegs in susceptible, resistant leaves and heat-shocked leaves. These differences in the ultrastructural and microscopic data in resistant areas were due to the restriction of ROS generation in limited areas, the side facing the plant surface, of appressoria and penetration pegs. Therefore, ROS generation was apparently induced regardless of the resistance or susceptibility of the cultivar with the difference being in the volumes generated. After evaluating the pathological role of ROS generation in fungal structures, such generation was found to be associated with early penetration of cell walls in pear plants. Additionally, ROS generation in plants was also found in degrading pectin layers near infected hyphae and in plasma membrane modification sites in susceptible leaves but not in resistant leaves. ROS generation in susceptible leaves might be accompanied with plasma membrane damage, although the role of ROS generation in the pectin layers is not clear. ROS generation in both fungal and plant cells during their interaction was likely associated with the expression of susceptibility. Received: June 3, 2002 / Accepted: July 31, 2002     

Acetolactate synthase activity and chlorsulfuron sensitivity of wheat cultivars

Differences were observed in the sensitivity of three wheat (Triticum aestivum L.) cultivars to chlorsulfuron. Shoot dry weight was reduced by foliar applications of chlorsulfuron to a greater extent in cv. Rongotea than in cvs Lancer and Kotare. There was no difference between these cultivars in the specific activity of acetolactate synthase (ALS) enzyme extracted from leaves or roots. Moreover, chlorsulfuron inhibited ALS from the wheat cultivars to the same extent in?vitro. ALS measurement in vivo showed that after 15 h of incubating excised leaf tissues with chlorsulfuron, there was a greater reduction in ALS enzyme activity in Rongotea than in the other cultivars. Furthermore, 1 day after a foliar application of chlorsulfuron, in vitro ALS activity in leaves was reduced more in Rongotea than in Lancer or Kotare. Recovery of the enzyme activity following chlorsulfuron application was quicker in Kotare than in Lancer or Rongotea. It is concluded that differences in sensitivity of these wheat cultivars to chlorsulfuron are not due to differential ALS sensitivity or level, but may be due to differential rates of herbicide metabolism.     

Effect of host factors on the susceptibility of Rhododendron to Phytophthora ramorum

Phytophthora ramorum causes sudden oak death (SOD) in western coastal forests of the USA. In Europe, the pathogen is mainly present in the nursery industry, particularly on Rhododendron. Because of the primary role of Rhododendron as a host and potentially as a vector, the effect of Rhododendron host factors on P. ramorum susceptibility and sporulation was investigated. Inoculation methods using either wounded or non‐wounded detached leaves were applied to 59 Rhododendron cultivars and 22 botanical species, replicated in three separate years. All Rhododendron species and cultivars were susceptible when using wounded leaves, but not when using non‐wounded leaves, suggesting a resistance mechanism operating at the level of leaf penetration. Using a regression tree analysis, the cultivars and species were split into four susceptibility classes. Young leaves were more susceptible than mature leaves when wounded, but less susceptible when non‐wounded. This effect was not correlated with leaf hydrophobicity or the number of leaf hairs. The presence or the type of rootstock did not affect the cultivar susceptibility level. Sporangia and chlamydospore production in the leaf lesions varied widely among Rhododendron cultivars and was not correlated with the susceptibility level. The susceptibility to P. ramorum correlated well with the susceptibility to P. citricola and P. hedraiandra × cactorum, suggesting that the resistance mechanisms against these species are non‐specific. Susceptibility to P. kernoviae was low for most cultivars. These findings have implications for detection, spread and disease control, and are therefore important in pest risk assessment.     

The relative tolerances of wild and cultivated oats to infection byErysiphe graminisf.sp.avenae: I. The effects of infection on vegetative growth and yield

The effects of infection by the powdery mildew fungusErysiphe graminisf.spavenaewere studied in one line of wild oat (Avena fatua), and two cultivars, Lustre and Peniarth, of cultivated oat (A. sativa) to determine if the wild oat was more tolerant of infection than the two cultivated oats. Seven weeks after inoculation, when the plants were 10-weeks-old with fully expanded flag leaves, the fungus had colonized approx. 40% of the leaf surfaces of wild oat and cv. Lustre but only about 30% of the leaf surfaces of cv. Peniarth. The lower leaves of cv. Peniarth were at least as susceptible, if not more so, than those of the other two lines but the upper leaves, including the flag leaf, were much more resistant. Although cv. Peniarth supported the production of about half the number of mildew conidia as the wild oat and cv. Lustre its total dry weight and grain yield were reduced to the greatest extent. The wild oat was clearly much more tolerant of mildew infection than cv. Peniarth and slightly more tolerant than cv. Lustre. The greater tolerances of the wild oat and cv. Lustre compared to cv. Peniarth appeared to be due to the lower sensitivities of their metabolism to the activities of the mildew fungus.     

Laboratory Evaluation of Citrus Cultivars Susceptibility and Influence of Fruit Size on Fortune Mandarin to Infection by Alternaria alternata pv. citri

Young leaves of 62 citrus cultivars were inoculated with conidia of three Spanish isolates of Alternaria alternata pv. citri, the causal agent of brown spot of citrus. Hybrids with Dancy mandarin, King mandarin or their derivates as a parent, grapefruit cultivars and the mandarin cultivars Guillermina, Emperor, Clemenpons and Esbal were highly susceptible to the pathogen. Satsuma cultivar Clausellina and orange cultivars, with the exception of Sanguinelli, were slightly susceptible. Lemon and lime cultivars were not susceptible, with the exception of Mexican lime (Citrus aurantifolia), which was slightly susceptible. Although this study shows a range of potential hosts for this pathogen, to date the only affected cultivars in Spain are Fortune and Nova mandarins, and Minneola tangelo. The susceptibility of Fortune fruits decreased as diameter increased, being susceptible through the whole season. This was confirmed with field observations in autumn where fruit infections have been detected when the diameter reaches 6–7 cm.     

Screening tomato and cucurbit rootstocks for resistance to Verticillium dahliae*

Seedlings of tomato (19) and cucurbits (33), previously selected as rootstocks for commercial cultivars, were evaluated for their resistance to verticillium wilt under glasshouse conditions. Disease scoring was based on foliar symptoms, but in plants with no visual or ambiguous symptoms the presence of the pathogen was confirmed by isolations or PCR using specific DNA primers. Five of the cucurbit rootstocks showed resistance to infection, 11 were moderately infected, 11 were susceptible, while the remaining six were very susceptible. In general, Cucurbita pepo (courgette, pumpkin) and Lagenaria siceraria tolerated infection, Cucumis melo rootstocks were susceptible and Citrullus vulgaris was the most susceptible. When inoculated with race 1 of the pathogen, one of the tomato rootstocks was very resistant, three moderately resistant, nine tolerant, ten susceptible and one very susceptible. Pathogenicity tests with race 2 showed that none of the rootstocks exhibited high tolerance, but that there was some variation in susceptibility. Research is under way to evaluate the reaction of commercial cultivars to verticillium wilt when grafted onto resistant rootstocks.     

Responses of cucumber cultivars to induction of systemic resistance against anthracnose by plant growth promoting fungi

Initial experiment on the reactions of five Japanese cultivars of cucumber toColletotrichum orbiculare infection in the greenhouse revealed that cv Suyo and Gibai were susceptible and moderately susceptible, respectively, while cv Shogoin fushinari and Sagami hanjiro were resistant to infection byC. orbiculare; cv Ochiai fushinari was moderately resistant. The ability of 16 plant growth promoting fungi (some isolates belonged to species ofPhoma and some non-sporulating isolates) isolated from zoysiagrass rhizospheres to induce systemic resistance in the above five cucumber cultivars was tested by growing plants in potting medium infested with barley grain inocula of PGPF in the greenhouse. The second true leaves of 21-day-old plants were challenge inoculated withC. orbiculare and disease assessed. Nine, out of 16 isolates, caused significant reduction of disease caused byC. orbiculare in at least two cultivars.Phoma isolates (GS8-1 and GS8-2) and non-sporulating isolates (GU21-2, GU23-3, and GU24-3) significantly reduced the disease in all the five cultivars. The disease suppression in cucumber was due to the induction of systemic resistance, since the inducer(s) and the pathogen were separated spatially and that the inducer did not colonize aerial portions. The resistance induced by certain isolates in a susceptible cultivar was less than that in a resistant cultivar. Disease suppression caused by isolate GU21-2 was similar to theC. orbiculare induced control in certain cultivars. The average rate of expansion of lesion diameter on leaves due toC. orbiculare was slower due to induction with the selected plant growth promoting fungi compared to the uninduced control plants. Roots of four cultivars were colonized by only three isolates, however, roots of one cultivar (Suyo) was colonized by five isolates suggesting the cultivar-specific root colonization ability.Abbreviations cv cultivar(s) - PGPF plant growth promoting fungal isolates - PGPR plant growth promoting rhizobacteria