trichoderma spp

In this study, inhibitory effects of five plant essential oils (EOs), including Zataria multiflora, Nepeta sp., Lavandula stoechas, L. officinalis and Artemisia dracunculus, on Fusarium graminearum UM29, F. graminearum UM89, F. verticillioides, F. brasilicum, F. oxysporum f. sp. lentis, and F. oxysporum f. sp. lycopersici and two antagonistic fungi Trichoderma harzianum and T. viride at five concentrations (75, 150, 300, 600 and 1000 μl/l) by mixing with PDA medium was studied. The chemical composition of EOs was explored by GC-MS. Also, the inhibitory effect of Trichoderma (T. harzianum T447 and T. viride T125) was investigated by dual culture assay and the effect of volatile compounds methods. Based on GC-MS results, thymol and carvacrol were identified as main constituents of Z. multiflora EO, and Nepetalactone and 1,8-cineole were the major components of Nepeta sp. EO. Zataria multiflora EO at concentration of 300 μl/l completely inhibited mycelial growth of the majority of Fusarium species evaluated in this study. The results of dual culture assay revaled the highest percentage of inhibition in the interaction between T. viride and F. brasilicum with a mean of 64.78% and the lowest in the interaction between T. harzianum and F. oxysporum f. sp. lycopersici with a mean of 36.52%. According to our results, Z. multiflora EO and T. harzianum T447 and T. viride T125had high mycelial growth inhibitory potential against Fusarium spp. Greenhouse and field assays are necessary to further evualte their efficacy in control of these plant pathogens.

The rice blast disease caused by Pyricularia oryzae is one of the most important fungal diseases in Iran and worldwide. From 150 samples of healthy and infected rice seedlings, leaves and spikes collected from paddy fields of Guilan province, 68 fungal isolates were obtained. Eleven isolates identified based on morphological and molecular methods that were not able to cause disease on rice in pathogenicity test including Ulocladium alternariae, Ulocladium cf. alternariae, Ulocladium cf. consortiale, Ulocladium sp., Curvularia pallescens, Preussia sp., Trichoderma harzianum, Trichoderma virens, Trichoderma viride, Alternaria citriand Epicoccum sp. were selected for rice blast disease biocontrol studies. In the laboratory, we useddual culture, culture filtrate, slide culture and volatile metabolites methods and based on results, T. harzianum, T. virens, T. viride, A. citri and U. cf. consortialeisolates had highest percentage of mycelial growth inhibition of P. oryzae. These fungal isolates were inoculated in a four–leaf stage on rice plant of Hashemi cultivar under greenhouse conditions. All inoculated fungi except U. cf. alternariae, reduced leaf blast disease rating, among them T. harzianum with 27.36% reduction in disease rating was the most effective antagonist in greenhouse studies followed by T. virens, T. viride and A. citri. Comparison of data average in least significant difference (LSD) method showed that there was significant difference betweenbiocontrol fungi (T. harzianum, U. cf. alternariae and Preussia sp.)in terms of efficiency in rice blast control. Also, there was a statistically significant difference between the fungi in terms of effect on height, fresh weight and dry weight of rice plants in the greenhouse andamong these fungi, T. harzianum showed the highest efficiency in improving these traits.The results of the present study showed that there are different fungi in microflora of rice plants which are considered as potential biological control of rice blast disease.

Rice foot rot disease caused by Fusarium fujikuroi, is one of the most important rice diseases in the world and in Iran. Integrated management with different methods is very important for the disease. In this research, to find appropriate fungal antagonistic isolates for the biological control of foot rot, the effect of four isolates of Trichoderma spp. and three isolates of Aspergillus spp. were studied on F. fujikuroi in vitro using dual culture, slide culture, volatile metabolites and non–volatile metabolites. The results of research showed that T. harzianum isolate with 81.81% was the most effective isolate in suppressing the mycelium growth of F. fujikuroi. In the volatile metabolites method, T. harzianum and T. virens isolates were the most effective with 73.33% in suppressing the mycelium growth of the pathogen. In the non–volatile metabolites method, A. awamori, T. viride and T. koningi isolates showed the highest percentage of F. fujikuroi inhibition. In the slide culture method, all isolates of Trichoderma spp. and Aspergillus spp. proved to be effective in inhibition of the mycelia of F. fujikuroi. These fungal isolates were inoculated on the rice plant under greenhouse conditions. All inoculated fungi were able to reduce the severity of the disease in the treated plants. The isolates of T. harzianum and T. virens were the most effective fungi in reducing the disease by reducing the severity by 83% and 79%, respectively. Also, application of all the studied fungi in the greenhouse increased the height, fresh weight and dry weight of shoots and roots in the presence of pathogen compared to the control plants (without pathogen). Analysis of variance and comparison of the mean of the treatments showed a significant difference between the fungi except in the dry weight trait. According to the results obtained from biocontrol studies in laboratory and greenhouse, T. harzianum, T. virens and T. viride isolates were the most effective antagonists in controlling rice foot rot disease.

This experiment was designed to investigate the antagonistic effect of six isolates of two Trichoderma species, i.e. Trichoderma harzianum and Trichoderma viride on growth and eliciting plant defense responses against Rhizoctonia solani in basil (Ocimum basilicum L.). The basil seeds treated with two Trichoderma species were planted in infested soil with R. solani and plant growth and severity of damping off measurement 40 days after planting. Peroxidase activity, total phenolic and malondialdehyde compounds levels were evaluated by colorimetic assay. The results show that eed treatments with two Trichoderma species significantly increased height and dry weight of seedling shoots and root dry weight, but their effects were not significantly different on stem diameter. Two Trichoderma species significantly reduced the severity of damping off disease where the percentages disease reduced from 71.25% in infected plants to 8.75-15.75% in plants treated with fungal anthagonists. The best protection to damping off disease was obtained by T. harzianum. Trichoderma species and Rhizoctonia solani individually increased peroxidase activity and total phenolic compounds levels, but they are more pronounced in the case of applying of combination of two fungi (Trichoderma and Rhizoctonia) in comparision to the control. Experiments indicated that maximum level of total phenolic compounds is induced by T. viride. Changes in malondialdehyde concentration occurred only in infected plants without antagonists treatments. Findings indicate that the growth and defense responses of basil significantly enhanced by seed coated with a Trichoderma different species.

Among crop culture, tomato (Lycopersicum esculentum L.) is an important vegetable crop that grown throughout the world in open as well as protected cultivation. Iran ranks sixth among tomato producing countries. One of the biggest challenges in modern agriculture worldwide, is the management of soil-borne plant pathogens, including plant-parasitic nematodes. Among plant parasite nematodes, root-knot nematode genus Meloidogyne spp. (RKN) is one of the most important nematodes in a wide spectrum of crops that causes serious damage in most agricultural crops worldwide. The eggshell of root-knot nematode is containing 50% protein, 30% chitin and 20% lipid and Species of Trichoderma due to the production of the high level of many hydrolytic enzymes, are used as \ biocontrol agents of plant pathogens.

In the current study, extracellular protease and lipase activity of 15 isolates of four species of Trichoderma spp. including T. harzianum, T. virens, T. konigii and T. saturnisporum were investigated to their biocontrol effects on tomato root-knot nematode in the greenhouse. These Trichoderma isolates were cultured on Potato Dextrose Agar (PDA) media for 7 days and were used to prepare inoculation of Trichoderma. In order to prepare Trichoderma inoculum, we used sterilized wheat as the substrate contains Trichoderma fresh culture and after that incubated in 28ºC for 12 days. The Meloidogyne javanica pure population was prepared and tomato seedlings were inoculated with 2000 of the fresh egg of nematode and 1×107 different Trichoderma strain conidia and were kept in greenhouse conditions. The enzyme activity was assessed based on the colorimetric method using a special substrate as the only energy source for Trichoderma isolates. We used casein as the substrate for protease and extra-pure olive oil as the substrate for lipase activity assessment.

According to the enzyme activity results, the highest protease activity was in the isolates T. harzianum BI, T. virens T65 and T. virens T64 and the lowest protease activity was in isolates T. harzianum T16A, T. saturnisporum T12 and T. saturnisporum T12N, respectively. The highest lipase activity was in T. harzianum T19, T. virens T6 and T. virens T65 and the lowest lipase activity were in T. harzianum T14N, T. harzianum T16A and T. harzianum T16. Based on the greenhouse results, T.BI, T65 and T6 isolates were the highest effective isolates and T12N, T12 and T16 reported as the lowest effective isolates in the biocontrol of root-knot nematode on infected tomato plants. Also, according to the results of correlation between protease and lipase activity and nematode reproduction factor, the correlation coefficient between enzyme activity and nematode reproduction factor for protease activity measured R2=0.83 and for lipase activity measured R2=0.42 which given the constituents of the root-knot eggshell and larvae cuticle that contains high amounts of protein, this is understandable. There are many reports about the effects of protease and chitinase on the induction of resistance against plant pathogens in plants. An investigation on the effect of Trichoderma isolates extracellular protease level on the biocontrol ability of these isolates against tomato root-knot nematode showed that the higher number of protease-encoding genes copies increases the biocontrol ability of these isolates.

Root-knot nematode is one of the most important damaging agents on tomato plants. It is difficult to control this endo-parasite nematode because of its wide host range, short life cycle, and high reproductive potential. The activation of the plant's own defense system through biotic and abiotic agents, called elicitors, has been considered as a focus of research only in recent years for the control of plant pathogens. The resulting elevated resistance due to an inducing agent upon infection by a pathogen is called induced systemic resistance (ISR) or systemic acquired resistance (SAR). Several abiotic and biotic agents have been reported to induce resistance against plant-parasitic nematodes. So far, many pieces of research have done on the use of different species of Trichoderma as biological control agents of plant pathogens. The ability to produce high amounts of chitinase, protease and glucanase enzymes made this isolates as a biocontrol agent of a wide range of plant pathogens. Overall, there is no sure whether or not a biocontrol agent is successful in controlling a particular pathogen, and many variables influence the success rate of biocontrol agents in natural environments. 

The deficiency of macro-nutrients such as phosphorus and potassium is very important due to vital roles of these elements. Although the total amount of phosphorus and potassium in the soil is high, the formation of insoluble forms of phosphorus, as well as the stabilization of potassium in silicates, has led to the shortage of these essential elements. The use of microorganisms, having the ability to dissolve insoluble phosphate forms and potassium fixed in silicates, can be effective in reducing the deficiency of these elements for the plant. In this research, 7 species of Trichoderma fungis were selected and their effects on the release of phosphorus and potassium were evaluated in Pikovskaya’s, Alexandrov’s and modified Pikovskaya’s media. The results showed that in Pikovskaya’s broth medium, phosphorus release rate by different species of fungi was consistent with decreasing pH. Trichoderma koningii, T.harzianum, T.citrinoviride and T.viridescens had the most phosphate solubilizing ability and increased soluble phosphorus by 244, 205, 191 and 190%, respectively. In both Aleksandrov’s and modified pikovskaya’s media, which contain both insoluble inorganic phosphate and potassium as biotite, it was observed that the dissolution rate of tri-calcium phosphate was lower than that of pikovskaya’s medium which has available potassium. T.koningii in Aleksandrov’s medium and T.harzianum in modified Pikovskaya’s medium had the highest ability to release potassium from biotite. These species increased potassium in solution by 123 and 20% compared to control, respectively. In general, the results showed that Trichoderma fungi has the ability to solubilize phosphate from tri-calcium phosphate and release potassium from biotite under in-vitro conditions.

Increasing germination percentage and growth indices is important by using methods such as biopriming. The present study aims to investigate effect of biopriming with a number of selected species of native Trichoderma fungi of Iran on yield, germination, vegetative and morphological components of Talash vriety bean seedlings. Also, the effect of induced-mutation by gamma irradiation on the Trichoderma genome and probebly increase in the efficiency of biopriming via using mutated Trichoderma isolates was studied. Experiments with three treatments: control (non-inoculation), Inoculated with wild type Trichoderma based bio-formulation and inoculation with mutants Trichoderma, at the Nuclear Agriculture Research Institute in the form of a completely randomized design with four replications were done. Evaluation of indices (Germination percentage; seedlings, stems and roots length ; their fresh and dry weight and vigor index) based on the ISTA standard method and yield in greenhouse was implemented. Results showed, bean seeds biopriming method with bio-formulations of Trichoderma species (wild type and mutant), plant's yield increased significantly compared to control. Germination percentage of seeds was significantly higher than control in fungal treatments. The important trait of dry weight has also increased compared to control and also had significant difference with control. The length of seedlings of bioprimed with Trichoderma was shorter than the control. Most of the indicators were significantly higher in the one percent level than control. Overally, the results of this study shows, in bean plants, the use of bio-priming seeds with Trichoderma, improves plant growth parameters especially in establishment and its early growth.

In order to investigate the effect of biopriming with selected species of Trichoderma on the morphological and germination characteristics of Canola (Brassica napus) seedlings, and soybean (Glycine max) cultivar Williams, an experiment was conducted in a randomized complete block design with three replications. The effect of gamma-mutation on Trichoderma genome and its effectiveness was investigated. Experiment with three levels of inoculation: control (non-inoculation), Inoculation with formulations from spores of wild type Trichoderma and inoculation with mutant isolates of the same species, at the Nuclear Agriculture Research Institute was carried out. Evaluation of germination indices (Germination percentage, stem length and root and seedlings and their fresh and dry weight and vigor index) based on ISTA standard implemented. Soybean bioprimed seeds with Trichoderma in most of measured indicators had significant difference with other treatments and growth components, especially in initial development and early growth stages were improved. In Canola seeds, all germination indices were decreased by biopriming with Trichoderma and in the most of the indices, there were no significant differences between wild type and mutant Trichoderma treatments. Overall, the results of this study showed, using seed biopriming with Trichoderma had different effects on plant growth parameters, and use of this technique needs case study for each plant genome.

Three isolates of Trichoderma spp. and two isolates of bacillus bacterial were evaluated in biological control of Fusarium oxysporum f. sp melonis the laboratory and greenhouse condition. Dual culture, antimicrobial metabolites, and volatile metabolites were in invitro assay. Fungal pathogen colony area was recorded, compared with control and induced inhibition of growth was determined. The experiment displayed the antagonistic ability using dual culture simultaneous and dual culture un-simultaneous assays against fungal pathogen. Results showed that Trichoderma harzianum 6 whit 70% and T. viride10 whit 52.9 highest percentage of growth inhibition. Results of dual culture highest percentage of growth inhibition about 30.5 percent of isolates whit the Bacillus spp62 percent growth inhibit the growth factor. The volatile compounds Bacillus spp15 whit 12.37 percent of the precipitation was prevented from growing. In the treatment Bacillus spp62 and mixed two spice tin treatments at Trichoderma 14g1 respectively 84.93 and 84.95 were showed most effects on length growth of stem but most effects of increased fresh weight from up ground organ of plant, was mixed treatment as spices T. longibrachiatum and bacteria Bacillus spp.15. Most effects on root fresh weight were showed by Trichoderma harzianum. Most effect on stem dry weight T. viridae with T. harzianum. Most effects on increased dry weight were showed by Bacillus spp.15and T. harzianum. Results of the experiment show treatments have significantly different activity pathogen. Test average correspond were meant significant at 1% level by LSD test.

Effects of biological treatments on disease intensity reduction of wheat brown rust caused by Puccinia reconditeRob. ex. Desem f.sp. tritici was studied in a split-split plot design, conducted based on completely randomized design in a glasshouse with three replications. Chamran and Boolani cultivars planted in main plots and three Trichoderma species including Trichoderma harzianum, T. citrinoviride and T. longibrachiatum were located in first subplots and leaf and seed treatments inoculation methods were used in second subplots. Changes in peroxidase enzyme level were also determined by Hemeda and Kelin method. The results showed a significant difference between chamran and boolani cultivars (P

Grape, like other horticultural products due to high humidity, low relative pH and wealth of nutrients, is subjected to attack by pathogenic fungi during the harvest and storage stages. In this study, at first, the some post-harvest pathogenic fungi of grape were isolated in Meshgin-shahr and two fungal species including Botrytis cinerea and Cladosporium cladosporioides were identified as the domonant species. Aspergillus tubingensis, causal agent of fruit rot and toxigenic species on grape and raisin was selected as well. In order to evaluate their biocontrol possibility, the effects of Trichoderma harzianum T447 and T. hamatum T622 and four medicinal plant essential oils (EOs) including mentha, spearmint, fennel and yarrow were tested in vitro. Results revealed that both of anthagonistic fungi were prospering than pathogenic fungi in nutrient competition and their volatile metabolites showed the highest inhibition to C. cladosporioides. 10% filtrate culture extract of T. harzianum T447 could effectively (≥85%) prevented the fungal mycelial growth. Also, our results showed that fungal species, EO type and its concentration play a critical role (P≤0.01) in fungal mycelia inhibition. Mentha and fennel EOs were known respectively as the most effective and weakest treatments. GC-MS analysis demonstrated that piperitoneoxid and Cis piperitone oxide, L-Menthone, dimethyl-4(E),6-octadien-2-ol and trans-Anethole were the main components of the essential oils of Mentha longifolia, M. spicata, Foeniculum vulgare and Achillea sp., respectively. Due to the significant inhibition of essential oils and biocontrol fungi, additional researches about their use for grape post-harvest diseases control are recommended.

Different Trichoderma species play role as suitable alternative for chemical fertilizers and successful biological tool in stable agriculture. These microorganisms show rivalry in food and place sources. They establish in the rhizosphere of most agricultural and non-agricultural plants via producing spores. They also could induce resistance in plants. These properties may lead to pathogen reduction as well as plant growth incitation. Favorite effect of Trichoderma species on the plant growth depends on type of species, strains and inoculation methods. Therefore this research has been tried to compare growth effects of four different Trichoderma species using three methods including seed coating, soil application and shoot spraying on tomato growth factors.
Four Trichoderma spp. including Trichoderma sp (T1), Trichoderm virens (T2), Trichoderma harzianum (T3, T4) and Trichoderma koningi (T5) were evaluated on tomato growth under completely randomized design based on factorial mmethod with four replications in greenhouse conditions. Three methods, consisting of seed coating, soil application and shoot spraying were applied under pot experiments and the performance of all three methods was evaluated according to the isolates. Fungal inoculation suspension was prepared for each method separately. For instance in seed coating method, 3×106 Trichoderma spore on seed surface was estimated. Approximately 107 propagle per gram of soil was calculated for soil application method and in shoot spraying 107 spores per ml was prepared. Statistical analysis was carried out based on averages comparison by LSD method (P=0.05).
Based on obtained results, soil application method showed meaningful promotion on tomato growth traits especially on dry root and foliar weight in comparison to other techniques. Overall, T1, T2 and T5 selected as the best candidates in trial conditions specifically in soil and seed application methods based on their positive effect on tomato growth factors. Isolate T1 increased root and shoot length by 54 and 45.5 percent respectively over controls in soil application treatment. In addition, the means comparisons using LSD method shows no significant difference between T1 and T2 isolates at 5% possibility level. So there is no difference between these isolates as a bio-fertilizer.
The soil application method was suggested as preferred technique in current study. In addition, it seems that application of T1 (Trichoderma sp.) and T2 (T. virens) isolates in the form of individual or combined formulation which can be used in the field level and could improve tomato growth and yields significantly.

During survey of potato Rhizoctonia disease in 2006، 50 isolates with characters of Rhizoctonia solani fungus from stems and roots and eight isolates from sclerotia، on potatoes surface، were obtained from fields and storages of Hamadan and Kurdistan Provinces. After nuclear staining and physiological surveys، all of the isolates identified as R. solani. For determining anastomosis groups of isolates، anastomosis carried out with tester isolates. Finally، 56 isolates belonged to AG-3، one isolate belonged to AG-4 and one isolate did not anastomoses with any available tester anastomosis groups in this study. Pathogenicity test of 25 isolates shown they are virulence. Biocontrol potential of four species of Trichoderma، including T. brevicompactum، T. koningiopsis، T. andinensis، and T. virens were studied on two pathogen isolates (6 and 16)، that had highest pathogenicity in pathogenicity test، in laboratory. Results showed that except T. andinensis، other Trichoderma spp. developed، establish and sporulated on pathogen colony after stopping its growth and destroyed pathogen hypha as coiling them. Volatile compounds of all Trichoderma spp. had significant effects on reducing pathogen isolates growth and among them T. virens، T. koningiopsis and T. brevicompactum had a higher inhibition، respectively.

One of the most destructive soil-borne diseases of carnation is fusarium wilt caused by Fusarium oxysporumf.sp. dianthi. In This study, antagonistic potential of different isolotes of three Trichoderma species on growth of the pathogen in laboratory and control of the disease in greenhouse was investigated. Isolates of the antagonist were Ta1, Ta2 and Ta3 belonging to Trichoderma atroviride, sixteen isolates Th1-Th16 belonging to T. harzianum and one isolate Tl1 belonging to T. longibrachiatum. Dual culture and colonization tests were carried out in a completely randomized design with three replications. Isolates  Ta3, Ta2 and Ta1 which completely covered colonies of the pathogen after five days, were the mosts successful colonizers. Inhibitory effect of volatile metabolites of Ta3 isolate in 72h and 96h cultures of the pathogen and antagonis (60% and 72%, respectively) was higher than those of the other isolates. In greenhouse, soil treatment with isolates of Trichoderma, to study the biocontrol effects the isolates was carried out in an experiment with 21 treatments in three replications. The results showed that different isolates could control the disease in different levels. Isolate Ta3 had the highest effect on disease control and reduction of cuttings death, followed by isolates Ta1, Ta2, Ta3 and Th2. In general, the antagonist isolates had less effects on growth of the plant, but reduced disease severity.

Trichoderma spp. is considered as promising biological control agents. Antagonistic Trichoderma species control numerous of plant pathogenic fungi. The highest value of degrading enzymes production (chitinases and glucanases) between known micro- organisms belongs to this genus. Hence that chitinases have effective role in myco-parasitism mechanism of plant pathogenic fungi، therefore superior bio-control potential may then be found in isolates which produce a high capacity of these enzymes. Application of genetic engineering and direct mutation in fungal genome in order to increase T. harzianum antagonistic potential via enhancement in chitinase activity has some advantages. In this study، we use 250 Gry gamma irradiation dose to induce mutation in T. harzianum. Then، chitinase activity of 20 mutants have been evaluated and 15 mutants showed higher enzyme activity than wild type isolate. Thus، induced mutation by gamma irradiation in order to improve antagonistic capacity of native iranian Trichoderma spp. to obtain effective and useful bio- control agents is advisable.