Effect of mixed function oxidase inhibitors on the toxicity of chlortoluron and isoproturon to wheat

Growth responses of wheat plants to combined treatments of four mixed function oxidase (MFO) inhibitors and chlortoluron were determined. Analysis of interactions showed that piperonyl-butoxide and especially ABT (1-aminobenzotriazole) increased the toxicity of chlortoluron. Metyrapone and 2,4-dichlorophenoxypropyne were phytotoxic and did not exert any clear interaction. ABT also increased the toxicity of isoproturon to wheat. Our results suggest that ABT Strongly inhibits the breakdown of chlortoluron and isoproturon in wheat. Since ABT is known to act as a suicide substrate for plant cytochrome P-450, wheat enzymes involved in the metabolism of these two herbicides are likely to belong to this class. It thus appears that compounds designed to inhibit plant cytochrome P-450 enzymes may interact with herbicide metabolism and are potential synergists of herbicide activity.     

黑龙江和云南葡萄产区灰霉病菌对多菌灵和异菌脲的抗性检测

为明确黑龙江省和云南省葡萄灰霉病菌对多菌灵和异菌脲的抗性频率,从两省分别采集、分离到葡萄灰霉病菌Botrytis cinerea 39株和38株,利用菌丝生长速率法测定其对多菌灵和异菌脲的敏感性,并分别检测上述菌株两种药剂靶标基因β-tubulin和Bos1的突变情况.结果表明,黑龙江省葡萄灰霉病菌对多菌灵和异菌脲的抗...     

Antagonism of Trichoderma harzianum ETS 323 on Botrytis cinerea Mycelium in Culture Conditions

ABSTRACT Previous studies have shown that the extracellular proteins of Trichoderma harzianum ETS 323 grown in the presence of deactivated Botrytis cinerea in culture include a putative l-amino acid oxidase and have suggested the involvement of this enzyme in the antagonistic mechanism. Here, we hypothesized that the mycoparasitic process of Trichoderma spp. against B. cinerea involves two steps; that is, an initial hyphal coiling stage and a subsequent hyphal coiling stage, with different coiling rates. The two-step antagonism of T. harzianum ETS 323 against B. cinerea during the mycoparasitic process in culture was evaluated using a biexponential equation. In addition, an l-amino acid oxidase (Th-l-AAO) was identified from T. harzianum ETS 323. The secretion of Th-l-AAO was increased when T. harzianum ETS 323 was grown with deactivated hyphae of B. cinerea. Moreover, in vitro assays indicated that Th-l-AAO effectively inhibited B. cinerea hyphal growth, caused cytosolic vacuolization in the hyphae, and led to hyphal lysis. Th-l-AAO also showed disease control against the development of B. cinerea on postharvest apple fruit and tobacco leaves. Furthermore, an apoptosis-like response, including the generation of reactive oxygen species, was observed in B. cinerea after treatment with Th-l-AAO, suggesting that Th-l-AAO triggers programmed cell death in B. cinerea. This may be associated with the two-step antagonism of T. harzianum ETS 323 against B. cinerea.     

A comparison of the mechanisms of action of vinclozolin,procymidone, iprodione and prochloraz against Botrytis cinerea

The mechanisms of action against Botrytis cinerea of the dicarboximide fungicides vinclozolin ( I ), procyrnidone ( II ), iprodione ( III ) and the less closely related compound prochloraza ( IV ) have been compared. They all inhibited mycelial growth much more than spore germination. None of the compounds affected respiration, membrane permeability or RNA production but III inhibited DNA synthesis and IV inhibited protein synthesis. Although chitin biosynthesis was inhibited by all the fungicides it was barely affected at the ED₅₀ concentrations (the concentration required to reduce the growth or germination of the test species by 50%) and is thus unlikely to be the primary target. The fungicides altered fungal lipid composition. I and II inhibited triglyceride production but III had no effect on it; III and IV reduced sterol biosynthesis. No common primary mode of action was found.     

Antagonism of Nutrient-Activated Conidia of Trichoderma harzianum (atroviride) P1 Against Botrytis cinerea

ABSTRACT The effect of preliminary nutrient activation on the ability of conidia of the antagonist Trichoderma harzianum (atroviride) P1 to suppress Botrytis cinerea was investigated in laboratory, greenhouse, and field trials. Preliminary nutrient activation at 21 degrees C accelerated subsequent germination of the antagonist at temperatures from 9 to 21 degrees C; at >/=18 degrees C, the germination time of preactivated T. harzianum P1 conidia did not differ significantly from that of B. cinerea. When coinoculated with B. cinerea, concentrated inocula of preactivated but ungerminated T. harzianum P1 conidia reduced in vitro germination of the pathogen by >/=87% at 12 to 25 degrees C; initially quiescent conidia achieved this level of suppression only at 25 degrees C. Application of quiescent T. harzianum P1 conidia to detached strawberry flowers in moist chambers reduced infection by B. cinerea by >/=85% at 24 degrees C, but only by 35% at 12 degrees C. Preactivated conidia reduced infection by >/=60% at 12 degrees C. Both quiescent and preactivated conidia significantly reduced latent infection in greenhouse-grown strawberries at a mean temperature of 19 degrees C, whereas only preactivated conidia were effective in the field at a mean temperature of 14 degrees C on the day of treatment application. An antagonistic mechanism based on initiation of germination in sufficiently concentrated inocula suggests that at suboptimal temperatures the efficacy of Trichoderma antagonists might be improved by conidia activation prior to application.     

Effect of Water Potential on Conidial Germination and Antagonism of Ulocladium atrum Against Botrytis cinerea

ABSTRACT The saprophytic fungus Ulocladium atrum was selected for its ability to competitively exclude Botrytis spp. from aboveground necrotic plant tissues which can play a crucial role in the epidemiology of diseases caused by necrotrophic Botrytis spp. Fungal growth in necrotic aboveground tissues can be hampered by fluctuating water availability. Adaptation to these adverse conditions is a key factor for the successful establishment of an antagonist population in this niche. Conidia of U. atrum germinated at water potentials between -1 and -7 MPa on water agar. Germinated conidia survived one to three interruptions of moist incubation by periods at -10 or -42 MPa. The speed of germination was significantly slower with interruption of the moist period as compared with the control, which had continuously moist incubation. However, the maximum germination percentage was the same for conidia incubated with or without interruption of the moist incubation. In bioassays with necrotic cyclamen leaves at -1, -3, and -7 MPa, U. atrum significantly reduced the sporulation of B. cinerea by more than 80%. The results demonstrate that U. atrum tolerates water stress during competitive substrate colonization with B. cinerea. The antagonist is, therefore, an attractive candidate for field applications on aboveground tissues.     

Spotting of tomato fruits caused by Botrytis cinerea

Botrytis spot or ghost spot on tomato fruits occurs after penetration of germ tubes ofB. cinerea into epidermal cells. A few days after the penetration a halo appears around the infected necrotic cells. These symptoms can be reproduced by inoculating young fruits with a few dry conidia. When many conidia alight on the epidermis of the fruit, scab-like symptoms develop, while under conditions of high humidity, blisters can be formed on the fruit surface before the fungus spreads through the fruit parenchyma. Under conditions of low humidity, necrotic areas are formed.In the necrotic cells, developed after inoculation with many or with a few conidia, no mycelium could be found by the histological methods so far used. However,B. cinera can be reisolated, implying that theBotrytis spot is a latent infection by the fungus. No renewed growth takes place when the fruit is fully ripe.Samenvatting Botrytis-stip op tomatevruchten ontstaat na binnendringen van kiembuizen vanB. cinerea in epidermiscellen van een jonge vrucht. Enkele dagen na de infectie treden celdelingen op in het onderliggende parenchymweefsel. De geïnfecteerde cel en enkele aangrenzende cellen sterven af, terwijl rondom dit necrotische plekje een zilverwit gekleurde ring ontstaat. Worden veel conidiën bijeen op een vruchtwand gebracht, dan krijgt de epidermis binnen 24 uur een schurftig uiterlijk. Blijft de luchtvochtigheid na de inoculatie hoog, dan kunnen daarentegen kleinere en grotere blaasjes in de vruchtwand ontstaan. Deze barsten na enkele dagen open, terwojl mycelium zich door de vrucht verbreidt. Daalt de luchtvochtigheid circa 16 uur na de inoculatie, dan blijft de aantasting beperkt tot de epidermis. Hoewel in het necrotische weefsel geen mycelium vanB. cinerea kon worden aangetoond, blijkt herisolatie van de schimmel gemakkelijk te zijn. Ook uitBotrytis-stippen van uit de praktijk afkomstige vruchten kanB. cinerea geïsoleerd worden. Dit betekent, dat het hier om een latente infectie gaat, waarbij evenwel geen hernieuwde groei plaats vindt als de vrucht geheel rijp is.     

Incidence of benomyl and iprodione resistance in isolates of Botrytis cinerea in tomato crops in England and Wales in 1986

In 1986, mass-mycelial isolates of Botrytis cinerea from 67 tomato crops in England and Wales were examined for benomyl and iprodione resistance. Of the 706 isolates obtained, 62.7% were resistant to benomyl at 2μg/ml and 43.2% were resistant to iprodione at 2 μg/ml. Iprodione resistance persisted in the absence of a dicarboximide spray programme. The incidence of benomyl resistance has not decreased since the last survey in 1984 in spite of a considerable reduction in the use of benzimidazole fungicides. There were no clear indications that the use of dichlofluanid influenced the incidence of benomyl or iprodione resistance. Disease control was poorer in crops with a higher incidence of iprodione resistance.     

Virulence factors of Botrytis cinerea

Botrytis cinerea is responsible for gray mold disease in more than 200 host plant species. The infection of host plants is mediated by numerous extracellular enzymes, proteins and metabolites. Each of these compounds may play a role in different stages of the infection process. Cell wall-degrading enzymes may facilitate the penetration into the host surface, while toxins, oxalic acid and reactive oxygen species may contribute to killing of the host cells. Cell wall-degrading enzymes contribute to the conversion of host tissue into fungal biomass. On the other hand, B. cinerea infection induces biosynthesis of phytoalexins. Therefore, the ability to overcome a wide spectrum of phytoalexins contributes to the pathogenicity of the fungus with a broad host range. The cloning of the corresponding genes has facilitated studies on gene expression and targeted mutagenesis. This review gives an overview of the research performed on virulence factors that play the roles in pathogenesis.     

Infection of plum and nectarine flowers by Botrytis cinerea

The ability of Botrytis cinerea to infect plum and nectarine flowers was studied in moist chambers and in the orchard. Within 36 h of inoculation, the pathogen penetrated and colonized the stamens, styles and petals on shoots placed in moist chambers, causing blossom blight. Similar lesion development was observed following inoculation with dry and wet conidia, Hyphae were usually distorted in stylar tissue, but grew normally in petals and filaments. Growth of the fungus through filaments into the sepals or floral tubes, or through the transmitting tissue of the style into the ovary, was never observed. Symptoms of blossom blight were not observed on inoculated shoots in the orchard. The floral tube, bearing the sepals and stamens, dehisced within 14 days of fruit set and infected floral parts did not remain attached to young developing fruit. No relation was found between post-harvest decay and flower infection. Losses following post-harvest decay might have been caused by direct penetration of ripening fruit and not by flower infection. The importance of infected floral parts as a source of secondary inoculum on ripening fruit is discussed.     

Sensitivity of Botrytis cinerea from vegetable greenhouses to boscalid

Between 2004 and 2006, 228 isolates of Botrytis cinerea from two regions in China were characterized for baseline sensitivity to boscalid, a new active ingredient that interferes with succinate ubiquinone reductase in the electron transport chain. The isolates showed similar sensitivity in different years and regions. Baseline sensitivities were distributed as unimodal curves with mean EC₅₀ values of 1·07 (± 0·11) and 0·42 (± 0·05) mg L⁻¹ for inhibition of mycelial growth and conidial germination, respectively. Laboratory studies were conducted to evaluate the risk of development of resistance to boscalid. Boscalid-resistant mutants were obtained by UV-treatment at lower frequencies and with smaller resistance factors than pyrimethanil-resistant mutants. All boscalid-resistant mutants were also significantly more sensitive to Qo inhibitors than their wild-type parents and showed reduced sporulation in vitro and pathogenicity on aubergine leaves. The results suggested that the risk of resistance developing for boscalid was lower than for pyrimethanil. However, as B. cinerea is a high-risk pathogen, appropriate precautions against resistance development should be taken. Synergism between the activity of boscalid and that of kresoxim-methyl was observed.     

Some characteristics of Botrytis cinerea isolates tolerant to procymidone

Of 104 isolates of Botrytis cinerea collected from greenhouses in south-east Spain, 70 (67%) were tolerant to procymidone. The tolerant isolates appeared as often in greenhouses treated with dicarboximides for only 1 year as in those treated for several years, indicating a rapid development of tolerance under greenhouse conditions. Although the tolerant isolates grew more slowly on PDA than the sensitive ones, their sporulation per unit area was similar. The sporulation and the viability of conidia of tolerant isolates collected in greenhouses where dicarboximides were used for 3 years were enhanced by the presence of procymidone in the growth medium, features that may have important epidemiological implications.     

Biosuppression of Botrytis cinerea in grapes

There is increasing interest in the use of biological control agents (BCAs) and plant resistance stimulants to suppress botrytis bunch rot in grapes, caused by Botrytis cinerea . Numerous different filamentous fungi, bacteria and yeasts have been selected as potential BCAs for control of grey mould based upon demonstrated antagonism towards B. cinerea. Biological suppression of the pathogen arises via competition for nutrients and space, the production of inhibitory metabolites and/or parasitism. Preformed and inducible grapevine defence mechanisms also contribute to disease suppression by preventing or delaying pathogenic infection. Furthermore, various biotic and abiotic agents can stimulate grapevine defence mechanisms and so elevate resistance to B. cinerea infection. Biosuppression of B. cinerea in vineyards, using BCAs and resistance stimulants, has been inconsistent when compared with that observed in controlled glasshouse or laboratory conditions. This may be attributable, in part, to the innate variability of the field environment. Research to improve field efficacy has focused on formulation improvement, the use of BCA mixtures and combinational approaches involving BCAs and plant resistance stimulants with complementary modes of action.     

Electrophoretic karyotype analysis of Botrytis cinerea

The karyotypes of five strains ofBotrytis cinerea were analysed by pulsed field gel electrophoresis (PFGE). None of the five chromosome patterns were identical and all five strains contained one or several minichromosomes of variable length. Considering that some of the chromosomal bands represent unresolved doublets, the total genome size of the five strains was estimated to vary between at least 33.9 and 39.7 megabasepairs (Mbp). Hybridization of blots of pulsed field gels with rDNA and ß-tubulin probes revealed that these hybridize with the same chromosomal band. Chromosome-specific probes were isolated hybridizing to chromosomal bands 9 and 11 of strain SAS56. The latter two probes hybridized to only two of the five strains tested. The results are discussed in relation to the genome plasticity and variability of filamentous fungi.     

Expression of an oxalate oxidase gene in tomato and severity of disease caused by Botrytis cinerea and Sclerotinia sclerotiorum

A cDNA clone of wheat oxalate oxidase (OxO) under the control of the constitutive CAMV 35S promotor was expressed in tomato plants by Agrobacterium -mediated transformation. Twenty-six transgenic tomato lines were obtained and analysed. PCR experiments confirmed the incorporation of the OxO gene in all tested tomato lines. The transgenic tomato plants expressed a 124-kDa protein showing OxO activity, and were able to convert different oxalic acid (OA) concentrations in vitro . In a detached leaf assay, most of the transgenic lines showed reduced disease symptoms compared with controls, following inoculation with Botrytis cinerea . In addition, leaves of the line T15 showed a marked reduction in symptoms compared with the control following inoculation with Sclerotinia sclerotiorum .