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Identification and characterization of Trichoderma species aggressive to Pleurotus in Italy

浙江大学学报(农业与生命科学版) 2004年第4期30卷 469-470页

作者： Woo S L Di Benedetto P Senatore m Abadi K Gigante S Soriente I Ferraioli S Scala F lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale La b. di Lotta Biologica Università di Napoli "Federico II" Via Università 1 00 80055 Portici (NA)Italy.

In the late 1980’s the deve lop ment of a severe epidemic of green mold caused by Trichoderma spp. was not ed in the commercial production of Agaricus bisporus (champignon) in the U nited Kingdom, North America, Spain and Holland, which caused extensive economic losses. The parasitic fungi isolated from the edible mushroom belonged to four biotypes, Th1, Th2, Th3 and Th4 of T. harzianum. However, among these biotypes, only Th2 (since c lassified as T. aggressivum f. europaeum) and Th4 (T. aggressivum f. aggressivum) were identified as the fungi causing problems in Agaricus production. In general, mushroom compost hosts both aggressive and innocuous is olates of Trichoderma, which are not morphologically distinguishable. Abo ut four years ago, a problem with green mold became apparent in the production o f Pleurotus ostreatus in Northern Italy, which eventually developed to a c risis situation in the South two years later and threatened to seriously comprom ise the Pleurotus market. This study was initiated to: isolate and identif y the aggressive fungi, then morphologically, physiologically and genetically characterize the isolates, dete rmine the source and phases of infection, and study methods of control. Samples were obtained from different phases of compost preparation at the locality of a major producer and supplier of compost to the mushroom industry in Southern Ital y, and microbial counts were conducted. Although the presence of Trichoderma was detected in the initial stages of composting, this value was reduced to zero from the phase of pasteurization to seeding with Pleurotus. Trichoderma infestations were noted in the packaged Pleurotus bales at various time s during the incubation phase (7-15 days after seeding) and after shipping to th e mushroom greenhouses, where the pathogen infestations greatly reduced the qual ity and quantity of the mushroom yield, as well as the number of potential harvest cycles. Preliminary r esults from the morphological and

关键词： commercial mushrooms green mold T.aggressivum

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Effect of rare earth application on the growth of Trichoderma spp. and several plant pathogenic fungi

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 424-424页

作者： d'Aquino L Carboni m Woo S L morgana m Nardi L lorito m ENEA Centro di Ricerche della Trisaia S.S. Jonica 106 Km 4195 75026 Rotondella (Matera) Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" Via Università 100 80055 Portici (NA) Italy

Rare earth elements (REEs) enriched fertilisers are currently used in China for soil and foliar applications to crops, but little is known about the effect of REEs applications on the growth of beneficial and detrimental soilborne microorganisms. The growth of biological control agents Trichoderma atroviride strain P1, Trichoderma harzianum strain A6 and strain T22, plant pathogens Botrytis cinerea, Alternaria alternata, Fusarium solani, Rhizoctonia solani and Sclerotinia sclerotiorum was investigated in the presence of REEs. An in vitro assays was used to monitor the effect of different concentration levels of either a mix of REEs (La, Ce, Pr, Nd) nitrates or lanthanum alone in comparison to treatments conducted with potassium nitrate and water. Although all fungi were affected when the REEs mix or lanthanum were present at concentrations higher than 100 mm, the growth inhibition depended mainly upon the combination of compounds, the dose and the fungal species or strains tested. Trichoderma strains and B. cinerea were more sensitive than A. alternata, F. solani, R. solani or at higher concentrations. Differing growth responses of some fungi to treatments with REEs mix vs. lanthanum alone indicated that in given situations the effect of the REEs compounds may be caused by elements other than lanthanum or by element mixtures. Further investigations are in progress to determine the effect of REEs on important interactions in the soil community between beneficial fungi, pathogenic fungi and/or the plant. REEs are naturally present in the environment and in biological systems but accumulation in soil can take place following successive applications. Therefore, it would be useful to achieve a better understanding of the effect of REEs accumulation on the activity of rhizosphere microorganisms given the widespread use in some regions of rare earths as fertilizers and their presence as fertilizer contaminants.

关键词： soilborne fungi enriched fertilizers lanthanum

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Factors that contribute to the mycoparasitism stimulus in Trichoderma atroviride. strain P1

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 421-421页

作者： Woo S L Formisano E Fogliano V Cosenza C mauro A Turrà D Soriente I Ferraioli S Scala F lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica and Dip. Di Scienza degli Alimenti Università di Napoli "Federico II" Via Università 100 80055 Portici (NA) Italy.

Trichoderma atroviride strain P1 has been used extensively to study the mycoparasitic mechanisms in the interaction between plant pathogenic host and beneficial antagonistic fungi. mutants of P1 containing the green fluorescent protein (gfp) or glucose oxidase (gox) reporter systems and different inducible promoters (from the exochitinase nag1 gene, or the endochitinase ech42 gene of P1) were used to determine the factors that activate the biocontrol gene expression cascade in the antagonist. The following compounds were tested singly and in various combinations: purified Trichoderma P1 enzymes (endochitinase, exochitinase, chitobiosidase, glucanase); antagonist culture filtrates (T. atroviride P1 wild-type and relative knock-out mutants, T. harzianum, T. reesei); pathogen culture filtrates (Botrytis, Pythium, Rhizoctonia); purified fungal cell walls (CWs) from Trichoderma, Botrytis, Pythium, Rhizoctonia; colloidal crab shell chitin; and plant extracts from cucumber leaves, stems or roots. Strong induction of mycoparasitism was found with the various digestion products produced by treating fungal CWs and colloidal chitin with purified enzymes or fungal culture filtrates. Filtrates from chitinase knock-out mutants, as well as CWs from Oomycetes fungi, were less active in producing the stimulus for mycoparasitism. The host CW digestion products were separated by molecular weight (mW) to determine which compounds were able to activate Trichoderma. micromolecules of mW less than 3 kDa were found to trigger mycoparasitism gene expression before physical contact with the host pathogen. These compounds stimulated mycelial growth and spore germination of the antagonist. Purification of these host-derived compounds was conducted by HPLC and in vivo assay. The obtained inducers were able to stimulate both the production of endochitinase and exochitinase enzymes, even under repressing conditions in the presence of glucose. Inducers stimulated the biocontrol effect of P1 in the

关键词： fungal cell walls hydrolytic enzymes mycoparasitism inducers

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Study of signal transduction factors involved in mycoparasitic response of Trichoderma atroviride

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 451-451页

作者： Scala V Zeilinger S Ambrosino P Brunner K Reithner B mach R L Woo S L Cristilli m Scala F lorito m Dipartimento di Arboricoltura Botanica e Patologia vegetale Lab. Lotta Biologica Facoltà di Agraria-Università degli Studi di Napoli "Federico II" via Università 100 80055 Portici (Napoli) Italy and Microbial Biochemistry and Gene Technology Department Institute for Chemical Engineering Technical University of Vienna Getreidemarkt 9/166-5 1060 Wien Austria.

Numerous Trichoderma spp. are mycoparasites and commercially applied as biological control agents against a large number of plant pathogenic fungi. The mycoparasitic interaction is host-specific and several research strategies have been applied to identify the main genes and compounds involved in the antagonist-plant-pathogen three-way interaction. During mycoparasitism, signals from the host fungus are recognised by Trichoderma, stimulating antifungal activities that are accompanied by morphological changes and the secretion of hydrolytic enzymes and antibiotics. Interestingly some morphological changes appeared highly conserved in the strategy of pathogenicity within the fungal world, i.e. the formation of appressoria as well as the secretion of hydrolytic enzymes seem to be general mechanisms of attack both for plant pathogens and mycoparasitic antagonists. This knowledge is being used to identify receptors and key components of signalling pathways involved in fungus-fungus interaction. For this purpose we have cloned the first genes (tmk1, tga1, tga3) from T. atroviride showing a high similarity to mAP kinase and G protein subunits (see abstract by Zeilinger et al.), which have been found to have an important role in pathogenicity by magnaporthe grisea. To identify the function and involvement of these factors in mycoparasitism by T. atroviride, tmk1, tga1, tga3 disruptant strains were produced. The knock-out mutants were tested by in vivo biocontrol assays for their ability to inhibit soil and foliar plant pathogens such as Rhizoctonia solani, Pythium ultimum and Botrytis cinerea . Disruption of these genes corresponded to a complete loss of biocontrol ability, suggesting a significant role in mycoparasitism. In particular, it has been suggested that tga3 regulates the expression of chitinase-encoding genes, the secretion of the corresponding enzymes and the process of conidiation. Comparative proteome analysis of wild type and disruptants supported this hypoth

关键词： pathogenicity mAP kinase G proteins

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Extracellular proteome of Trichoderma harzianum to identify proteins with biotechnological value

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浙江大学学报（农业与生命科学版） 2004年第4期30卷 452-452页

作者： Ambrosino P Scala V marra R Vinale F Soriente I Ferraioli S Carbone V Ruocco m Woo S L lorito m Dep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyISA-CNR Avellino ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples ItalyDep. Ar.Bo.Pa.Ve.-Plant Pathology University of Naples "Federico II"-80055 Portici Naples Italy

Trichoderma harzianum strain T22 parasitizes and controls many phytopatogenic fungi and is applied commercially as biological control agent. The production of hydrolitic enzymes appears to be a key factor in the parasitic process. We tested the endo-esochitinolitic and glucanolitic activities of culture filtrates of T22 grown under carbon and nitrogen starvation or in presence of biomass or cell walls of the phytopathogenic fungi Botrytis cinerea, Rhizoctonia solani and Pythium ultimum. The highest level of enzimatic activities was found in culture where the mycoparasite interacted with a phytopathogenic fungus. Therefore we used a proteomic approach to investigate changes in the complex mixture of extracellular proteins secreted by T. harzianum strain T22 in order to identify proteins of potential biotechnology value for commercial and industrial *** technology has greatly enhanced our ability to conduct functional genomics studies. Nevertheless only a few studies have been published so far on the fungal extracellular proteome. Sample preparation remains the most critical step in analyses based on two-dimensional gel electrophoresis (2-DE), and it requires to be optimized for each specific application. In this study, our first aim was to set up the extraction protocol of the extracellular proteins secreted by T . harzianum strain T22 when it was grown in vitro .The secreted proteins were analysed by two-dimensional electrophoresis (2-DE) and substantial changes in the extracellular proteome of the mycoparasite have been observed. Comparing the 2D maps of the fungus grown in minimal medium with glycerol as carbon source (used as control condition) with those obtained in inducing conditions, a lot of novel proteins appeared. The higher number of novel and up-regulated spots was obtained in the presence of Rhizoctonia solani biomass. Other spots were specifically up-regulated by the interaction with different plant pathogens. Differentially expressed proteins

关键词：木霉属真菌蛋白质鉴定生物技术植物病原体水解酶

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Use of Trichoderma spp. for biological control of the livestock feed contaminant fungus Fusarium proliferatum

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浙江大学学报（农业与生命科学版） 2004年第4期30卷 414-414页

作者： Ruocco m Pane F Ritieni A Lanzuise S Ambrosino P marra R Woo S L Ciliento R Soriente I Ferraioli S Scala F lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Uni

Fusarium spp. are pathogens of many important agricultural crops, and are often strong mycotoxin producers. Fusarium proliferatum, in particular, causes disease in cereals and secretes the toxin Beauvaricin that contaminates livestock feed and cereals, producing a variety of toxicity symptoms ranging from poor weight gain to mortality. Beauvaricin is a cyclodepsipeptide and acts as a potent mycotoxin known to have insecticidal properties. This compound is highly toxic to human cell lines, where it induces apoptosis and specifically inhibits cholesterol acetyltransferase. Nothing is known about the role of this mycotoxin during the interaction of F. proliferatum with other microorganisms, including the fungal antagonists Trichoderma spp. In vitro tests have demonstrated that the antagonistic and mycoparasitic activity of Trichoderma is not inhibited by the presence of Beauvaricin at concentrations up to 10 mg/kg in the substrate. In vivo biocontrol assays on barley and wheat with Trichoderma against F. proliferatum isolates, producing and non-producing Beauvaricin, confirmed the ability of the antagonist to control this pathogen in all cases. Also Trichoderma culture filtrates obtained in conditions that promote Cell Wall Degrading Enzyme (CWDE) secretion, were able to inhibit. spore. germination. of. different. F.. proliferatum. isolates.. These. results. suggest. the. possibility. of. using. Trichoderma. and/or. its. metabolites. to. control. contaminants. of. livestock. feed. by. mycotoxin-producing. Fusarium..

关键词：家畜霉菌饲料污染物镰刀霉木霉属生物防治

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Cloning of first abc transporter encoding gene from Trichoderma spp. and its expression during stress and mycoparasitism

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浙江大学学报（农业与生命科学版） 2004年第4期30卷 450-450页

作者： Lanzuise S Ruocco m Scala V Catapano L Woo S Ciliento R Ferraioli S Soriente I Vinale F Scala F Del Sorbo G lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 ItalyDip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Un

Trichoderma in its natural environment competes for nutrient uptake and is required to protect itself from adverse natural toxic compounds, such as those produced by plants and other microbes in the soil community, or synthetic toxic compounds released human activity. One of the most important metabolic pathways for drug resistance and substrate uptake, both in prokaryotes and eukaryotes, is ATP dependent. The role of ABC transporter proteins in the biology of Trichoderma is still not known. We present the cloning of the first four ABC transporter genes (TABC1, TABC2, TABC3, TABC4 ) in Trichoderma, and in particular T. atroviride P1, and the characterization of TABC2 The complete sequence of this gene is 6535 bp, which includes a promoter of 1624 bp, a terminator of 642 bp and a coding region of 4264 bp. The promoter contains many of the potential transcription factor binding sites found in the 5’ upstream region of the ech42 gene of T. atroviride P1. These included: heat shock factors (HSF), a nitrogen-regulating factor (Nit-2), a stress-response element (STRE), a GCR1 elements, and a Cre BP1 motif. Northern analysis and RT-PCR demonstrated that TABC2 is highly expressed when Trichoderma is subjected to nitrogen starvation, grown in the presence of culture filtrates of Botrytis cinerea, Rhizoctonia solani, and Pythium ultimum, or when N-acetylglucosamine is added to the substrate. TABC2 appears to be co-regulated with some CWDE-encoding genes, suggesting that this is the first ABC transporter encoding gene involved in mycoparasitic events. It’s role in the interaction of Trichoderma with fungal hosts or plants is being investigated by targeted gene disruption and overexpression.

关键词：真菌寄生现象应激反应木霉属真菌克隆病原体

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Proteomic study of three component interactions: plant, pathogens and antagonistic fungi

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 449-449页

作者： marra R Ambrosino P Scala V Romano C Vinale F Ferraioli S Ruocco m Carbone V Woo S L Turrà D Scala F lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" Via Università 100 80055 Portici (NA)Italy ISA-CNR Avellino.

The molecular factors involved in the three-way interaction between plant, pathogenic fungi and antagonistic/biocontrol fungi, such as Trichoderma, are still poorly understood, even if they represent a matter of interest for improving crop management and developing new strategies for plant diseases control. The aim of this work is to investigate the components involved in this interaction and, for this purpose, a proteomic approach was used. 2-D maps of the protein extracts from the single components in various interactions between plants (potato, bean, tobacco or tomato), pathogens (Botrytis cinerea, Rhizoctonia solani or Pythium ultimum) and biocontrol fungi (Trichoderma atroviride strain P1 or Trichoderma harzianum strain T22) were obtained. The proteome of each partner was collected separately and extracted by acetone precipitation in presence of trichloroacetic acid and a reducing agent (DTT). The extracted proteins were separated by isoelectrofocusing (IEF), using IPG (Immobilized pH gradient) strips, followed by SDS-PAGE. In order to improve resolution the separations were performed both on wide than narrow pH range and on different gel lengths. Differential spots were noted in the proteome of the three-way interaction when compared to each single component. These were further characterized by mass spectrometry and in silico analysis with the aim of identifying and cloning the relative genes. During the in vitro interaction of T. harzianum strain T22 with tomato and the culture filtrate or cell walls of pathogens, the spot number was higher than in the presence of pathogen biomass. In terms of Trichoderma differential proteins displayed on 2D gels, the most important changes were obtained in the presence of P. ultimum . During the in vivo interaction with tomato, the antagonist proteome changed much more in presence of soilborne fungi R. solani and P. ultimum than with the foliar fungus B. cinerea, both in terms of total and increased or novel spots. In silic

关键词： differential proteins in vivo interactions induced disease resistance

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The Trichoderma-plant interaction is mediated by avirulence proteins produced by this fungus

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 422-422页

作者： Ruocco m Lanzuise S Woo S L Ambrosino P marra R Turrà D Gigante S Formisano E Scala F Kip N P J G m de Wit lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy

The molecular basis of Trichoderma -plant interaction is very complex and still not completely understood. The colonization of the root system by rhizosphere competent strains of Trichoderma results in increased development of root/aerial systems, in improved yields and in plant disease control. Other beneficial effects, such as the induction of plant systemic resistance, have also been described. To understand the mechanisms involved we are using different approaches, including the making of transformants expressing genes that encode for compounds able to affect plant response to pathogens. Trichoderma transformants carrying the avirulence gene Avr4 from Cladosporium fulvum under the control of constitutive and inducible promoters were obtained and tested on tomato plants having the Cf4 resistance gene. Necrosis and suberification zones, similar to the symptoms appearing during Cladosporium-tomato interaction, were found when the roots of the Cf4 plants were treated with Avr4-Trichoderma. This demonstrates that selected Trichoderma strains are able to transfer to the plant molecules that may deeply affect metabolism, disease resistance etc. Therefore, these beneficial fungi can be regarded as biotechnological tools to provide a variety of crops with useful compounds. moreover, in in vitro competition assays the transformants were found to be more effective as antagonists against Alternaria alternata than the wild type. Trichoderma sends a variety of biochemical signals to the plants including avirulence molecules; therefore the presence of avr-like proteins in the fungus proteome was investigated. Proteome analysis has permitted us to isolate and sequence many proteins potentially having this function. From the extracellular protein extracts, we have purified and sequenced a protein with structural characteristics similar to Avr4 of C. fulvum. The protein, Hytra1, was found to be a hydrophobin with chitin binding activity, the typical 8 cysteine residues, and 4 dis

关键词： systemic resistance Avr4 biochemical signals

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Genetic improvement of Trichoderma ability to induce systemic resistance

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浙江大学学报(农业与生命科学版) 2004年第4期30卷 423-423页

作者： Ciliento R Woo S L Di Benedetto P Ruocco m Scala F Soriente I Ferraioli S Brunner K Zeilinger S mach R L lorito m Dip. Arboricoltura Botanica e Patologia Vegetale-Sez. Patologia Vegetale Lab. di Lotta Biologica Università di Napoli "Federico II" and CNR-IPP Via Università 100 Portici (NA) 80055 Italy and Microbial Biochemistry-Gene Technology Department Institute for Chemical Engineering Technical University of Vienna Getreidemarkt 9/166-5 1060 Wien Austria

The beneficial applications of Trichoderma spp. in agriculture include not only the control of plant pathogens, but also the improvement of plant growth, micronutrient availability, and plant tolerance to abiotic stress. In addition, it has been suggested that these fungi are able to increase plant disease resistance by activating induced systemic resistance (ISR) . The mode of action of these beneficial fungi in the Trichoderma -plant-pathogen interaction are many, complex and not completely understood. Numerous lytic enzymes have been characterized, the encoding genes (ech42 gluc78, nag1 from T. atroviride strain P1) cloned, and their role in biocontrol demonstrated. The corresponding biocontrol-related inducible promoters have been used in a reporter system based on the Aspergillus niger glucose oxidase gene (goxA) to monitor biocontrol activity. Glucose oxidase catalyzes the oxygen-dependent oxidation of D-glucose to D-glucono-1,5-lactone and hydrogen peroxide; this latter compound is known to have an antifungal effect and activate the plant defence cascade, thus increasing resistance to pathogen attack. T. atroviride P1 transformants with various promoters gox were tested as seed coating treatments on bean seeds planted in soil infested with a soilborne fungal pathogen. Successively, the emergent leaves were inoculated with a foliar pathogen to determine the effect of the GOX transformants on biocontrol and resistance to pathogen attack. Inoculations with the P1-GOX transformants not only reduced disease symptoms caused by a soil pathogen, but also the lesions of various foliar pathogens applied far from the Trichoderma colonization, thus activating ISR. A similar approach is being use to genetically improve T. harzianum T22, a rhizosphere competent and commercially marketed strain not transformed yet, by using four different gox gene constructs under the control of constitutive and inducible promoters. Plasmids have been introduced in Trichoderma by protoplast

关键词： glucose oxidase induced systemic resistance lytic enzymes

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