Pythium periplocum, an aggressive mycoparasite of Botrytis cinerea causing the gray mould disease of grape-vine

Pythium periplocum Dreschler has been found to be an aggressive mycoparasite of Botrytis cinerea, the causal agent of the gray mould disease of the grape-vine. When grown together, the former enters the latter's mycelium, branches freely within, coagulates its cytoplasm and finally tears its hyphae apart, bringing about widespread destruction of the grape-vine pathogen. Extensive coiling around the host, as reported in the case of other mycoparasites belonging to the genus Pythium, has not been observed here. The infected mycelium of B. cinerea fails to infect the grape-vine and does not induce the characteristic gray mould symptoms. Since P. periplocum is not a grape-vine parasite, it could be useful for the biological control of B. cinerea. A brief account of this mycoparasitism is discussed in this article.     

Presence of a vanadium-dependent haloperoxidase in Botrytis cinerea

The presence of a haloperoxidase in the mycelium of Botrytis cinerea, extractable with buffer, is demonstrated. A low level of extracellular enzyme activity was also detected. The haloperoxidase from the fungus is a vanadium-dependent glycoprotein, with a pH optimum of about 5.5. Native gel electrophoresis indicates that it is a high molecular mass protein. It appears to react with antibodies against haloperoxidase from Caldariomyces fumago. Enzyme activity is increased 3.5-fold and 15-fold by culture of the fungus in the presence of NaCl or vanadium, respectively. Activity is partly reduced by removal of vanadium and activity can be restored by the addition of vanadium to the enzyme. The possible function of this haloperoxidase is discussed.     

拟南芥新型脂转移蛋白AtDHyPRP1的亚细胞定位及其对灰霉菌的抗性

通过遗传转化技术研究了拟南芥脂转移蛋白AtDHyPRP1在细胞中的定位及其对真菌病原体的抗性。采用PCR方法从拟南芥Ws生态型克隆了AtDHyPRP1基因,构建产生pRI101-AN-AtDHyPRP1植物双元表达载体和pCAMBIA1302-AtDHyPRP1-GFP融合表达载体,经农杆菌介导的叶盘和浸花法得到烟草和拟南芥转基因植株。AtDHyPRP1基因能够明显增加烟草对灰霉菌的抗性,转AtDHyPRP1烟草叶片的被侵染部位有大量H2O2积累,激光共聚焦显微观察发现AtDHyPRP1蛋白定位于细胞表面。说明AtDHyPRP1蛋白在合成后被分泌到细胞外执行特殊的功能,与植物抗病防御机制有关。     

Root inoculation with a forest soil streptomycete leads to locally and systemically increased resistance against phytopathogens in Norway spruce

Soil streptomycetes are commonly antagonistic against plant pathogens. However, interactions involving increased defense responses in the host plant, leading to suppression of plant disease development, have not yet been detailed. Here, the mechanisms were studied of disease suppression by Streptomyces sp. GB 4-2 against Heterobasidion root and butt rot in Norway spruce (Picea abies) seedlings. GB 4-2 promoted mycelial growth of the phytopathogenic fungus, germination rate of fungal spores, extension of germ tubes and early colonization of outer cortical layers of the plant root. Reduced colonization of the inner cortical cell layers was accompanied by the induction of cell wall appositions, and increased xylem formation in the vascular cylinder emerged after bacterium-fungus coinoculation. Bacterial treatment led to decreased water content in roots and needles and increased photosynthetic yield (F(v)/F(m)) and peroxidase activities in needles. The infection of needles by Botrytis cinerea was reduced by bacterial pretreatment. Complex interactions of GB 4-2 with Norway spruce and Heterobasidion abietinum were discovered. The bacterium promoted the growth of the phytopathogenic fungus but induced plant defense responses. Host responses indicate that GB 4-2 induces both local and systemic defense responses in Norway spruce.     

Identification of Botrytis spp. on Plants Grown in Iran

A total of 363 isolates were collected from all over Iran. They were isolated from apple, arum lily, briar rose, bride wort, broad bean, camellia, canola, carnation, cucumber, egg plant, feijoa, geranium, gerbera, gladiolus, grape, guilder rose, hibiscus, iris, kiwifruit, oleander, onion, orange, pear, pomegranate, primrose, quince, redbud, robinia, rose, rubber plant, sow thiste, spathe flower, strawberry, tomato, violet, wall flower and wheat. To identify the species, morphological characters such as conidiophore length, conidial and sclerotial dimensions were measured. According to morphological and cultural characters, eight Botrytis species were identified: B. aclada sensu lato, B. cinerea, B. fabae, B. convoluta, B. gladiolorum, B. paeoniae, B. pelargonii and B. porri. As far as we are aware, this is the first report of the last five species from Iran. These species were examined by polymerase chain reaction (PCR) using necrosis and ethylene‐inducing protein (NEP2) and C729 primers. A 835 bp band was amplified in B. cinerea, B. fabae and B. pelargonii, using NEP2, but not in others. However, C729 primers amplified a 700 bp band in B. cinerea and B. pelargonii and a 600 bp in B. fabae.     

Control of preharvest and postharvest fruit rot in Strawberry by Metschnikowia fructicola

The yeast Metschnikowia fructicola was tested as a preharvest treatment to control preharvest and postharvest rots of strawberry fruit in Turkey and Israel. In greenhouse trials, the efficacy of the yeast antagonist against preharvest rots was equal to that of a chemical control (fenhexamid) in two growing seasons. In an open-field experiment, the yeast reduced the incidence of rot to commercially acceptable levels. M. fructicola reduced the incidence of fruit rot by 56-69% in greenhouses, open-field culture, and in low plastic tunnels. The yeast suppressed postharvest incidence of fruit rot significantly better than fenhexamid. Among fruit from greenhouses, open-field culture, or tunnels, M. fructicola treatment reduced the incidence of fruit rot during postharvest storage by 70, 64, and 72%, respectively. When applied weekly in the greenhouse or in the field, the population density of M. fructicola was about 1×10⁵ cfu/fruit. Similar population density of the antagonist was also observed during storage of the fruit at 0° C.     

Ability of Clonostachys rosea to Establish and Suppress Sporulation Potential of Botrytis cinerea in Deleafed Stems of Hydroponic Greenhouse Tomatoes

The ability of Clonostachys rosea to establish and persist in deleafed tomato stems and to suppress sporulation potential of Botrytis cinerea was investigated in plots of hydroponic tomatoes in commercial greenhouses. Leaves near lower fruit clusters were removed according to standard practice and deleafed portions of the stems were treated with C. rosea , iprodione or water. Inoculum of B. cinerea was from natural infections. Stem lesions were not produced by the pathogen during the trials. Development of C. rosea and B. cinerea in stems was estimated indirectly by quantifying sporulation on excised stem tissues that were incubated on an agar medium containing paraquat. Incidence and area of sporulation of C. rosea on tissue pieces were high (76-99%) and moderately high (33-79%), respectively, when stems were treated with the agent at 0, 6, 24 or 48 h after deleafing and sampled 11 to 75 days later. In various instances, the agent also sporulated on tissues from water controls and iprodione treatments, apparently after interplot transmission. In most instances, incidence and area of sporulation of B. cinerea on tissue pieces were high (83-100%) and moderate to high (35-76%), respectively, in the water controls, but moderate (31-44%) and moderate to low (5-34%), respectively, for stems treated with C. rosea at 0 to 48 h after deleafing and sampled after 11-75 days. Without exception, C. rosea suppressed B. cinerea as or more effectively than iprodione. Correlations between inoculum density of C. rosea (0-10 6 conidia mL -1 ) and sporulation potential of B. cinerea in deleafed stems were strongly negative in each of three tests ( r = -0.95 to -0.99). Conidial suspensions and a talc formulation of C. rosea were of similar effectiveness against B. cinerea . We conclude that C. rosea persisted and suppressed sporulation potential of B. cinerea in deleafed tomato stems for at least 11 weeks after application.     

Biological Control of Botrytis cinerea Pers. ex Fr. in Strawberry by Paenibacillus polymyxa (Isolate 18191)

Isolate 18191, obtained from mature strawberry fruit and determined as Paenibacillus polymyxa has shown an antagonistic potential against Botrytis cinerea , the causal agent of grey mould in strawberries. Germ tube growth of conidia of B. cinerea was strongly inhibited by the culture suspension of the antagonist in aqueous strawberry fruit pulp suspension (1%) but germination rate of conidia was not affected. The application of the culture suspension and the washed cells on detached strawberry leaf discs reduced conidiophore density of B. cinerea by 67 and 84%, respectively. The treatment of detached leaf discs with culture suspensions of different cell densities (1 × 10⁶, 1 × 10⁷, 1 × 10⁸) showed that the lowest density already reduced incidence of B. cinerea by 68% after 8 days incubation period. Investigating the influence of the temperature on the effectiveness of P. polymyxa it was observed that the antagonist was highly effective already at 10°C and reduced incidence and conidiophore density of B. cinerea by 53 and 58%, respectively. In 3-year field trials the effectiveness of P. polymyxa was in a range of 24–36% as compared to the water control.     

Mycelial elongation and sporulation of two fungi on amended media in light or dark

Botrytis allii andCollectotrichum dematium are onion pathogens which can infect in the field and cause decay in storage. Some phenolics can hinder development of these fungi, but the effect of cytokinins is not clear. Cytokinins (kinetin or 6-benzyladenine) or phenolics (caffeic or chlorogenic acids) were added to agar at concentrations of 0 to 10^–3 M. Cultures were continuously irradiated with fluorescent light or maintained in the dark for 6 days. On unamended media, final mycelial elongation was 45 or 17.8 mm and sporulation was 28 or 10.6 × 10⁴ spores/ml forBotrytis andColletotrichum, respectively. ForBotrytis, mycelial elongation was slightly (5%) but significantly increased and sporulation increased by 21% by incubation on phenolics as compared to cytokinins. Mycelial extension ofColletotrichum was not affected by amendment. Sporulation ofColletotrichum on kinetin was 16 to 28% greater than on the other amendments. As amendments concentration increased elongation of mycelia of both fungi decreased. Sporulation ofBotrytis increased by 60% as amendment concentration increased from 0 to 10^–5 M and then decreased 25% at 10^–3 M. As amendment concentration increased from 0 to 10^–3 M, sporulation ofColletotrichum increased by 45%. Incubation in light increased mycelial extension 3 to 17% forBotrytis andColletotrichum respectively, and sporulation was increased approximately 78% for both fungi. These compounds do not appear to inhibit development of theseBotrytis orColletotrichum species in culture.     

Fate of transformed Trichoderma harzianum in the phylloplane of tomato plants

A propiconazole-resistant Trichoderma harzianum strain with high phylloplane survival capability was transformed with the E. coli hygromycin B phosphotransferase gene (hph), coding for hygromycin B resistance. Four transformants were analysed for survival ability on the phylloplane of tomato plants grown under glasshouse conditions in comparison with their prototype and a yellow, hygromycin B-sensitive mutant. Over 2 weeks, the four transformants showed higher survival rates in comparison with the wild-type strain. The yellow mutant TF3/973 did not significantly differ in survival from the transformants. Both hygromycin B resistance and mitotic stability of transformants were evaluated during growth in vitro and after reisolation from tomato phylloplane. Hybridization patterns with the complete plasmid indicated that all four transformants were mitotically stable after several rounds of vegetative growth without selective pressure and during 2 weeks on tomato plants. None of the transformants had lost the ability to grow in the presence of both propiconazole and hygromycin B after growth under the same conditions. The results are discussed in relation to risk assessment of the release of transgenic fungi.