Evaluation of Trichoderma harzianum strain T22 as biological control agent of Calonectria pauciramosa

The objective of this research was to evaluate Trichoderma harzianum strain T22 as a biocontrol agent of collar and root rot caused by different Calonectria pauciramosa isolates. Thus, the microsclerotia-forming ability and virulence of twenty C. pauciramosa isolates were assessed. Microsclerotia production varied partially among the isolates and dual culture with T22 on carnation leaf agar revealed isolates with both high and low microsclerotia-forming ability. Inoculation tests on red clover (Triflolium pratense) demonstrated its susceptibility to the pathogen. On red clover, the degree of virulence and T22 effects in controlling infections were highly variable among the isolates tested. A nursery trial performed on Feijoa sellowiana seedlings confirmed previous results, clearly indicating virulence variability among C. pauciramosa isolates. For three isolates tested in nursery trial, T22 effectiveness in controlling infection was inversely related to their degree of virulence. Overall, T. harzianum strain T22 showed good antagonist activity in reducing microsclerotia production on carnation leaf and the incidence and severity of collar and root rot on both selected hosts. This data could be crucial in developing integrated pest management strategies in ornamental plant nurseries.     

Testing the effect of biological control agents on the formation of vesicular arbuscular mycorrhiza

A container system for rapid infection of roots with pathogenic or mycorrhizal fungi was used to test the effect of the two commercial biological control agents, Trichoderma harzianum and Streptomyces griseoviridis, on the formation of vesicular-arbuscular mycorrhiza in soybean. In the presence of these biocontrol agents, mycorrhiza formation with Glomus mosseae was significantly depressed, particularly with S. griseoviridis. Infection by the root pathogen Rhizoctonia solani was not altered by these agents. Remarkably, not only R. solani but also T. harzianum induced accumulation of large amounts of the phytoalexin glyceollin in the roots. In contrast, roots inoculated with S. griseoviridis or with the mycorrhizal fungus G. mosseae did not accumulate glyceollin.     

Efficacy of Bacillus subtilis and Trichoderma harzianum combination on chickpea Fusarium wilt caused by F. oxysporum f. sp. ciceris

Fusarium wilt is caused by F. oxysporum Schlecht end. Fr. f. sp. ciceris (FOC) is a devastating disease of chickpea in Algeria. In this study, antagonistic effects of B. subtilis MF352017 (Bs1) and Trichoderma harzianum KX523899 (T5) isolated from the rhizosphere of chickpea were investigated separately and in combination for their efficacy in controlling the disease in vivo. The efficacy of the antagonistic biocontrol agents on Fusarium wilt was evaluated based on vegetative and root growth parameters of chickpea. Seed bacterisation with B. subtilis MF352017 (Bs1) and seed treatment with T. harzianum (T5) significantly protected chickpea seedlings from FOC as compared to untreated plants. Plant protection was more pronounced in T. harzianum-treated plants than in bacterised plants. The application of both antagonists effectively suppressed 93.67% of the disease and also enhanced plant growth leading to increased plant height, root length, fresh and dry weights of shoot and root. The mixture of antagonists increased the effectiveness of B. subtilis MF352017 (Bs1) isolate on Fusarium wilt and improved chickpea growth.     

Efficiency of different application methods of biocontrol agents and biocides in control of Fusarium root rot on some citrus rootstocks

Abstract

The efficiency of two biocontrol agents (Trichoderma harzianum NB and Bacillus subtilis NB) and two commercial biocides (Plant Guard and Rhizo-N) in controlling Fusarium root rot disease on some citrus rootstocks was evaluated under artificially infested soil in green house.

Fusrium root rot on citrus rootstocks seedlings i.e. sour orange (SO), volkamer lime (VL), rangpur lime (RP) and cleopatra mandarin (CL) was successfully controlled by dipping the root system of such seedlings in water suspensions of each biological treatment i.e. Trichoderma harzianum (spore suspension 5×10⁶ spore/ml), Bacillus subtilis (cell suspension 8×10⁷ cell/ml), Plant Guard (3 g/l) and Rhizo-N (4 g/l), then transplanted into artificially infested soil with Fusarium solani and drenched with enough water suspension of such biological treatments. Plant Guard (3 g/l) and Rhizo-N (4 g/l) were highly effective treatments in decreasing infection and severity of the disease, Fusarium density in rhizosphere soil and colonization of Fusarium solani in the roots of all tested seedlings.

Meanwhile, root dipping or soil drenching with the same treatments individually gave the least effect in reducing root rot incidenceon all tested rootstocks compared with application of the two methods together.

It should be noted that using biocontrol agents and commercial biocides could be successfully used in controlling root rot pathogens on citrus in commercial greenhouses or under field conditions before transplanting in new reclaimed lands in the desert.     

Biological control of strawberry crown rot,root rot and grey mould by the beneficial fungus Aureobasidium pullulans

Utilization of biocontrol agents is a sustainable approach to reduce plant diseases caused by fungal pathogens. In the present study, we tested the effect of the candidate biocontrol fungus Aureobasidium pullulans (De Bary) G. Armaud on strawberry under in vitro and in vivo conditions to control crown rot, root rot and grey mould caused by Phytophthora cactorum (Lebert and Cohn) and Botrytis cinerea Pers, respectively. A dual plate confrontation assay showed that mycelial growth of P. cactorum and B. cinerea was reduced by 33–48% when challenged by A. pullulans as compared with control treatments. Likewise, detached leaf and fruit assays showed that A. pullulans significantly reduced necrotic lesion size on leaves and disease severity on fruits caused by P. cactorum and B. cinerea. In addition, greenhouse experiments with whole plants revealed enhanced biocontrol efficacy against root rot and grey mould when treated with A. pullulans either in combination with the pathogen or pre-treated with A. pullulans followed by inoculation of the pathogens. Our results demonstrate that A. pullulans is an effective biocontrol agent to control strawberry diseases caused by fungal pathogens and can be an effective alternative to chemical-based fungicides.

     

Biocontrol of onion white rot by application of Trichoderma species formulated on wheat bran powder

In vitro, Trichoderma album, Trichoderma harzianum, Trichoderma koningii, Trichoderma viride and Trichoderma virens showed antagonistic effect against the most pathogenic isolate (Sc2) of Sclerotium cepivorum, the cause of onion white rot disease. Five Trichoderma preparations of each Trichoderma sp. were prepared on wheat bran powder to be used for controlling white rot disease of onion. Greenhouse and field experiments followed the same trend where T. harzianum and T. koningii were the most effective in reducing the incidence and severity of white rot disease compared with the control. Trichoderma species preparations caused promotion to vegetative parameters of onion plants in pots and increase bulb productivity in filed. In this regard, T. harzianum and T. koningii were the most effective. A positive correlation was found between the biocontrol activity of Trichoderma species preparations and enhancement of peroxidase, polyphenoloxidase and chitinase enzymes in onion plants to resist infection with S. cepivorum.     

Biological Control of Phytophthora Root Rot of Pepper Using Trichoderma harzianum and Streptomyces rochei in Combination

A combination of two compatible micro‐organisms, Trichoderma harzianum and Streptomyces rochei, both antagonistic to the pathogen Phytophthora capsici, was used to control root rot in pepper. The population of the pathogen in soil was reduced by 75% as a result. Vegetative growth of the mycelium of P. capsici was inhibited in vitro on the second day after P. capsici and T. harzianum were placed on the opposite sides of the same Petri plate. Trichoderma harzianum was capable of not only arresting the spread of the pathogen from a distance, but also after invading the whole surface of the pathogen colony, sporulating over it. Scanning electron microscopy showed the hyphae of P. capsici surrounded by those of T. harzianum, their subsequent disintegration, and the eventual suppression of the pathogen's growth. Streptomyces rochei produced a zone of inhibition, from which was obtained a compound with antioomycete property secreted by the bacteria. When purified by high‐pressure liquid chromatography, this compound was identified as 1‐propanone, 1‐(4‐chlorophenyl), which seems to be one of the principal compounds involved in the antagonism. A formulation was prepared that maintained the compound's capacity to inhibit growth of the pathogen for up to 2 years when stored at room temperature in the laboratory on a mixture of plantation soil and vermiculite. The two antagonists, added as a compound formulation, were effective at pH from 3.5 to 5.6 at 23–30°C. The optimal dose of the antagonists in the compound formulation was 3.5 × 10⁸ spores/ml of T. harzianum and 1.0 × 10⁹ FCU/ml of S. rochei. This is the first report of a compound biocontrol formulation of these two antagonists with a potential to control root rot caused by P. capsici.     

Influence of bio-inoculants on the control of root rot and wilt disease of pyrethrum caused by Rhizoctonia solani

The effect of four bio-inoculants namely, Glomus aggregatum, Streptomyces sp., Bacillus subtilis and Trichoderma harzianum and a fungicide, Ridomil-mancozeb was evaluated on the biomass production and control of root rot and wilt disease under glasshouse conditions. Results showed that 21-day- prior inoculation with G. aggregatum was most effective where none of the treated plants produced disease symptoms and interestingly their growth was increased by 39.4%. The colonisation by G. aggregatum (>80%) also increased P concentration in shoot. While, similar treatment with Streptomyces sp., B. subtilis, T. harzianum and Ridomil-mancozeb individually failed to produce any significant effect over Rhizoctonia solani inoculated control, where all inoculated plant died, prematurely. The simultaneous and 3-day-post treatments of G. aggregatum were non-effective but simultaneous treatment with Streptomyces sp. produced 70% disease control, while B. subtilis and T. harzianum individually provided 50% control. Their effects were either better or at par when compared with the simultaneous treatment of Ridomil-mancozeb. In 3-day-post treatment, Streptomyces, B. subtilis and Ridomil-mancozeb individually provided 50% disease control, whereas T. harzianum was least effective as it could protect only 25% plants against infection. The results reveals that 15 days prior treatment of G. aggregatum can significantly controls the root rot and wilt disease of pyrethrum. Further, treatment of Streptomyces can also serve the next effective post infection control method.     

Evaluation of biological seed treatments in combination with management practices for the control of Fusarium dry rot of potato

Seed-borne diseases of potato represent a significant constraint to potato production in the US. The use of an effective fungicide in combination with good management practices during cutting and storage, prior to planting, is essential to reducing disease. The efficacy of two biocontrol agents (Bacillus subtilis and Trichoderma harzianum), and a commercially formulated mixture of the chemicals fludioxonil plus mancozeb, applied as seed treatments in combination with different management practices, were evaluated over two years for the control of seed piece decay and sprout rot caused by Fusarium sambucinum. Treatments were made 10 days prior to planting and at planting, and tubers were re-stored at either 18 °C and 95% RH with forced air ventilation at 5950 l min⁻¹ (optimal conditions), at 25 °C in the dark without ventilation (sub-optimal), or not stored at all prior to planting. Seed piece and sprout health were evaluated in vitro and agronomic impacts evaluated in field experiments. Results showed that the biological control agents B. subtilis and T. harzianum provided good control of sprout rot and seed piece decay caused by F. sambucinum, when seed was re-stored under optimal conditions or not re-stored at all. Under optimal conditions, treatment with B. subtilis reduced sprout rot and seed piece decay on average by 66% and 84%, respectively. Treatment with T. harzianum reduced sprout rot and seed piece decay on average by 70% and 81%, respectively. Treatment with fludioxonil + mancozeb reduced sprout rot and seed piece decay under both re-storage regimes. Under optimal conditions, disease incidence and severity was reduced on average by 81% and 97%, respectively. Neither biological control agent reduced seed piece decay incidence under either re-storage regime compared to the untreated controls.     

Efficacy of formulation and storage on rice straw waste on the activation of bioagents against root-rot diseases of bean plants

Trichoderma harzianum and Trichoderma viride, which were grown on modified media of rice straw waste, were used as suitable natural media for determining the population of the bioagents and stimulating the production of antimicrobial substances, i.e. toxins, enzymes at different periods (1, 2, 3, 4, 5 and 6 weeks). Also, the evaluation of synergistic effect between biocontrol fungi and bacteria may play a key role in the natural process of biocontrol. Synergism can occur when different agents are applied together and cell wall degrading enzymes produced by fungi can increase the efficacy of bacteria. The evaluation of the bioagents formulated on rice straw waste has influenced the percentage of infection of root rot on bean plants under green house and field conditions. The formulation of the Trichoderma spp. to reduce the incidence of the diseases caused by soil-borne fungi in the field has great importance in the biocontrol of the diseases. This work was aimed to determine and improve the efficacy of application and formulation of Trichoderma spp. on the rice straw waste against root-rot disease of bean under green house and field conditions.     

Construction of a Streptomyces lydicus A01 transformant with a chit42 gene from Trichoderma harzianum P1 and evaluation of its biocontrol activity against Botrytis cinerea

Streptomyces lydicus A01 and Trichoderma harzianum P1 are potential biocontrol agents of fungal diseases in plants. S. lydicus A01 produces natamycin to bind the ergosterol of the fungal cell membrane and inhibits the growth of Botrytis cinerea. T. harzianum P1, on the other hand, features high chitinase activity and decomposes the chitin in the cell wall of B. cinerea. To obtain the synergistic biocontrol effects of chitinase and natamycin on Botrytis cinerea, this study transformed the chit42 gene from T. harzianum P1 to S. lydicus A01. The conjugal transformant (CT) of S. lydicus A01 with the chit42 gene was detected using polymerase chain reaction (PCR). Associated chitinase activity and natamycin production were examined using the 3, 5-dinitrosalicylic acid (DNS) method and ultraviolet spectrophotometry, respectively. The S. lydicus A01-chit42 CT showed substantially higher chitinase activity and natamycin production than its wild type strain (WT). Consequently, the biocontrol effects of S. lydicus A01-chit42 CT on B. cinerea, including inhibition to spore germination and mycelial growth, were highly improved compared with those of the WT. Our research indicates that the biocontrol effect of Streptomyces can be highly improved by transforming the exogenous resistance gene, i.e. chit42 from Trichoderma, which not only enhances the production of antibiotics, but also provides a supplementary function by degrading the cell walls of the pathogens.     

Streptomyces sp. S160: a potential antagonist against chickpea charcoal root rot caused by Macrophomina phaseolina (Tassi) Goid

Biological control of charcoal root rot disease caused by Macrophomina phaseolina in chickpea was studied by using Streptomyces sp. S160. This biocontrol agent (BCA) inhibited the mycelial growth of M. phaseolina by 50 % in vitro and significantly reduced charcoal rot incidence in the greenhouse by 33.3 %. The greenhouse experiment revealed that seed treatment along with soil application supported the highest germination (88.6 %), vigor index (7326.91) and reduced root rot incidence (12.5 %) in comparison to seed treatment and soil application alone. BCA enhanced the growth and helped in inducing resistance against charcoal rot disease of chickpea caused by M. phaseolina by increasing activity of defense-related enzymes in chickpea plants, leading to the synthesis of defense chemicals in plants. BCA (Streptomyces sp. S160) was also characterized and identified by using polyphasic approaches including 16S rDNA sequencing.     

Biological efficacy of <Emphasis Type="Italic">Trichoderma harzianum</Emphasis> isolate to control some fungal pathogens of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis>) in Turkey

Gaeumannomyces graminis var. tritici, Fusarium culmorum and F. moniliforme are highly important and widespread pathogens of wheat in Turkey. Trichoderma isolates have been used as biocontrol agents to protect plants against soilborne diseases in several crops. The present work was carried out to evaluate the potential of Trichoderma harzianum isolate T1 as biocontrol agents for G. graminis, F. culmorum and F. moniliforme under field conditions in 2001 and 2002. Quantitative differences were found in microbial number in soil. T. harzianum T1 had considerable effect on population densities of the tested pathogens. The total number of G. graminis, F. culmorum and F. moniliforme were lower in the T. harzianum T1 application made to seed. T. harzianum T1 application to seed had increasing affect on the yield components of wheat through better control over pathogens. The greatest counts of T. harzianum T1 were detected on root segments. Seed application by T. harzianum T1 had increasing effect on yield components of wheat.     

Bio-control of Stem Rot in Jerusalem Artichoke (Helianthus tuberosus L.) in Field Conditions

Stem rot is a serious disease in Jerusalem artichoke (JA). To reduce the impact of this disease on yield and quality farmers often use fungicides, but this control method can be expensive and leave chemical residues. The objective of this study was to evaluate the efficacy of two biological control agents, Trichoderma harzianum T9 and Bacillus firmus BSR032 for control of Sclerotium rolfsii under field conditions. Four accessions of JA (HEL246, HEL65, JA47, and JA12) were treated or notreated with T. harzianum T9 and B. firmus BSR032 in a 4 × 2 × 2 factorial experiment in two fields (environments), one unfertilized and one fertilized. Plants were inoculated with S. rolfsii and disease was evaluated at 3-day intervals for 46 days. T. harzianum T9 and B. firmus BSR032 reduced disease incidence by 48% and 49%, respectively, whereas T. harzianum T9 + B. firmus BSR032 reduced disease incidence by 37%. The efficacy of T. harzianum T9 and B. firmus BSR032 for control of S. rolfsii was dependent on environments and genotypes. The expression of host plant resistance also depended on the environment. However, HEL246 showed consistently low disease incidence and severity index in both environments (fertilized and unfertilized). Individually, T. harzianum T9, B. firmus BSR032, or host plant resistance control stem rot caused by S. rolfsii in JA. However, no combination of these treatments provided more effective control than each alone.     

Biological control of Botrytis cinerea on tomato using naturally occurring fungal antagonists

Abstract

In order to evaluate the potential of naturally occurring filamentous fungi having potential as biocontrol agents effective against grey mould and post-harvest fruit rot caused by Botrytis cinerea on tomato, fungal saprophytes were isolated. They were obtained from leaves, fruits and flowers belonging to different species of cultivated and spontaneous Solanaceous plants collected at the horticultural area of La Plata, Argentina. Of 300 isolates screened for inhibition of B. cinerea using the dual culture technique on agar plate, 12 strains inhibited strongly mycelial growth of the pathogen. Among the antagonists one isolate of Epicoccun nigrum (126), four of Trichoderma harzianum (110, 118, 248 and 252) and four isolates of Fusarium spp. decreased the spore germination of B. cinerea between 30 and 70%. These isolates were probed on tomato fruits to evaluate their biocontrol activity against post-harvest grey mould. In growth chamber tests, E. nigrum (27), F. equiseti (22, 105) and T. harzianum (118, 252) reduced the diameter of fruit lesions by 50 – 90% and were selected for further biocontrol assays of tomato plants in the greenhouse. Although there were not significant differences between the treatments and the control, F. equiseti (105), E. nigrum (27) and T. harzianum (118) reduced by 20, 22 and 22 respectively the disease on whole plants. The targeted application of isolates of E. nigrum, T. harzianum and F. equiseti provides a promising alternative to the use of fungicide spray to control B. cinerea on tomatoes.     

Effect of Trichoderma harzianum on microelement concentrations and increased growth of cucumber plants

The potential of the biocontrol agent Trichoderma harzianum strain T-203 to induce a growth response in cucumber plants was studied in soil and under axenic hydroponic growth conditions. When soil was amended with T. harzianum propagules, a 30% increase in seedling emergence was observed up to 8 days after sowing. On day 28, these plants exhibited a 95 and 75% increase in root area and cumulative root length, respectively, and a significant increase in dry weight (80%), shoot length (45%) and leaf area (80%). Similarly, an increase of 90 and 30% in P and Fe concentration respectively, was observed in T. harzianum inoculated plants. To better characterize the effect of T. harzianum during the early stages of root colonization, experiments were carried out in a gnotobiotic hydroponic system. An increased growth response was apparent as early as 5 days post-inoculation with T. harzianum, resulting in an increase of 25 and 40% in the dry weight of roots and shoots, respectively. Similarly a significant increase in the concentration of Cu, P, Fe, Zn, Mn and Na was observed in inoculated roots. In the shoots of these plants, the concentration of Zn, P and Mn increased by 25, 30 and 70%, respectively. Using the axenic hydroponic system, we showed that the improvement of plant nutritional level may be directly related to a general beneficial growth effect of the root system following T. harzianum inoculation. This phenomenon was evident from 5 days post-inoculation throughout the rest of the growth period, resulting in biomass accumulation in both roots and shoots.     

Genotype-specific responses to the effects of commercial Trichoderma formulations in lentil (Lens culinaris ssp. culinaris) in the presence and absence of the oomycete pathogen Aphanomyces euteiches

Members of the endophytic fungal genus Trichoderma have been established as plant-beneficial microbes and are most successful commercial biologicals in the form of bio-fertilisers, biocontrol agents, and growth stimulators. We report the variable interactions among different lentil genotypes and Trichoderma strains in both the presence and absence of biotic stress (root-rot pathogen Aphanomyces euteiches). Two commercial Trichoderma formulations, namely RootShield^® (RS) and RootShield^® Plus (RSP) based on T. harzianum T22 and T. virens G41, respectively, were evaluated for control of Aphanomyces root rot and plant growth promotion in 23 wild and cultivated lentil genotypes. No significant disease control was recorded with either formulation in any lentil genotype. Significant genotype-specific plant growth promotion was observed in terms of root and shoot development and leaf parameters in a genotype-specific manner. Genotypes of Lens culinaris and Lens tomentosus, both in the primary lentil gene pool, demonstrated the maximum response. The overall effect of Trichoderma treatment was markedly higher under biotically stressed conditions in comparison to unstressed conditions. In many cases, negative responses were recorded, particularly in the absence of root-rot disease. L. tomentosus PI 572390 exhibited positive responses for most of the tested parameters. Our findings clearly indicate that, in the case of lentil, plant genotype plays a major role in interactions among the tested Trichoderma strains and the plant. Moreover, the influence of Trichoderma was greater and more favourable under conditions of biotic stress vs. the absence of stress.     

Biocontrol of the Potato Blackleg and Soft Rot Diseases Caused by Dickeya dianthicola

Development of protection tools targeting Dickeya species is an important issue in the potato production. Here, we present the identification and the characterization of novel biocontrol agents. Successive screenings of 10,000 bacterial isolates led us to retain 58 strains that exhibited growth inhibition properties against several Dickeya sp. and/or Pectobacterium sp. pathogens. Most of them belonged to the Pseudomonas and Bacillus genera. In vitro assays revealed a fitness decrease of the tested Dickeya sp. and Pectobacterium sp. pathogens in the presence of the biocontrol agents. In addition, four independent greenhouse assays performed to evaluate the biocontrol bacteria effect on potato plants artificially contaminated with Dickeya dianthicola revealed that a mix of three biocontrol agents, namely, Pseudomonas putida PA14H7 and Pseudomonas fluorescens PA3G8 and PA4C2, repeatedly decreased the severity of blackleg symptoms as well as the transmission of D. dianthicola to the tuber progeny. This work highlights the use of a combination of biocontrol strains as a potential strategy to limit the soft rot and blackleg diseases caused by D. dianthicola on potato plants and tubers.     

Biological and Integrated Control of Botrytis Bunch Rot of Grape UsingTrichodermaspp.

Field trials were carried out in upstate New York in 1990, 1992, 1993, and 1994 and in Chile in 1992–1993 and 1993–1994 in order to evaluate the ability of various strains ofTrichodermaspp. to control bunch rot of grape, to assess the compatibility and possible additive effects of selected biocontrol fungi and dicarboximide fungicides, and to determine factors affecting biocontrol efficacy. In 1990, three strains ofTrichodermaspp. were evaluated for their biocontrol ability, and all provided significant control ofBotrytis cinerea.As few as two late applications of the biocontrol fungi were nearly as effective as up to five applications throughout bloom and fruit development. Trials in New York in 1992 and in Chile in 1992–1993 indicated thatTrichoderma harzianumcould replace some applications of iprodione or vinclozolin with little reduction in efficacy. In New York in 1993, we found that applications ofT. harzianumat bloom and early fruit development followed by a tank-mix application ofT. harzianumand half rates of iprodione gave extremely effective control of bunch rot. In 1994, less effective control was obtained than in earlier years. Addition of a nutritive adhesive (Pelgel, a mixture of carboxymethyl cellulose and gum arabic) applied with the biocontrol agent tended to improve results. Thus, biological control of bunch rot of grape withT. harzianumcan be an effective method of management of this disease.     

Vermicompost-based bioformulation for the management of sugarbeet root rot caused by Sclerotium rolfsii

Vermicompost-based bioformulations of bacterial and fungal biocontrol agents were examined against sugarbeet root rot caused by Sclerotium rolfsii. The result showed that the Pseudomonas fluorescens strain Pf1 in combination with either Trichoderma asperellum strain TTH1 or Bacillus subtilis strain EPCO-16 performed better in reducing disease next to the chemical difenoconazole. Similarly, enhanced yield was observed in the same combination treatments under both pot and field conditions.