Endophytic Trichoderma Research Articles

Biodegradation of ochratoxin A by endophytic Trichoderma koningii strains

Ochratoxin A (OTA) is a mycotoxin produced by Aspergillus spp. and Penicillium spp. that causes a threat to food safety and human health. Fungal biodegradation might be a promising strategy for reducing the OTA contamination in the future. In this study, the ability of Trichoderma koningii strains to degrade OTA produced by Aspergillus niger T2 (MW513392.1) isolated from tomato seeds was investigated. Among T. koningii strains tested, three strains; AUMC11519, AUMC11520 and AUMC11521 completely eliminated OTA from the culture medium, while AUMC11522 strain eliminated only 41.82% of OTA. OTα-amide, 3-phenylpropionic acid, OTα and phenylalanine were assayed as degradation products by FTIR analysis and LC–MS/MS spectra. Carboxypeptidase A (CPA) was found responsible for OTA degradation when a metal ion chelator, EDTA, was added to cell free supernatants of the three effective strains. OTA detoxification by T. koningii could present new prospective strategies for a possible application in food commodities intoxicated with ochratoxin.

The Biocontrol Potential of Endophytic Trichoderma Fungi Isolated from Hungarian Grapevines, Part II, Grapevine Stimulation.

In the first part of this two-piece publication, the isolation, identification and in vitro characterization of ten endophytic Trichoderma isolates were reported. Here we report the ability of two different mixes of some of these isolates (Trichoderma simmonsii, Trichoderma orientale and Trichoderma gamsii as well as of Trichoderma afroharzianum and T. simmonsii) to colonize and stimulate the growth of grapevines. Two commercial vineyards about 400 km away from the site of isolation were used as experimental fields, from which the strains of three Trichoderma species were re-isolated up to four years after rootstock soaking treatment with conidiospores, performed before planting. The treatments decreased the overall percentage of lost plants of about 30%, although a low number of lost plants (about 5%) were observed also in the control plot. For all cultivars and clones, the Trichoderma treatments significantly increased both the bud burst ratio and bud burst vigor index. In addition, the grape must parameters such as the Brix degrees, as well as the extract, the D-glucose and the D-fructose concentrations all appeared to be improved, suggesting a potentially higher ethanol content of the produced wine. We conclude that grapevine-endophytic Trichoderma isolates promote plant growth, which could be a useful feature for sustainable agriculture in general and integrated plant production in particular.

Potential and fungicidal compatibility of antagonist endophytic Trichoderma spp. from rice leaves in controlling dirty panicle disease in intensive rice farming

Potential and fungicidal compatibility of antagonist endophytic Trichoderma spp. from rice leaves in controlling dirty panicle disease in intensive rice farming

Endophytic Trichoderma spp. can protect strawberry and privet plants from infection by the fungus Armillaria mellea.

Armillaria mellea is an important fungal pathogen worldwide, affecting a large number of hosts in the horticulture and forestry industries. Controlling A. mellea infection is expensive, labour intensive and time-consuming, so a new, environmentally friendly management solution is required. To this effect, endophytic Trichoderma species were studied as a potential protective agent for Armillaria root rot (ARR) in strawberry and privet plants. A collection of forty endophytic Trichoderma isolates were inoculated into strawberry (Fragaria × ananassa) plants and plant growth was monitored for two months, during which time Trichoderma treatment had no apparent effect. Trichoderma-colonised strawberry plants were then inoculated with A. mellea and after three months plants were assessed for A. mellea infection. There was considerable variation in ARR disease levels between plants inoculated with different Trichoderma spp. isolates, but seven isolates reduced ARR below the level of positive controls. These isolates were further tested for protective potential in Trichoderma-colonized privet (Ligustrum vulgare) plants where five Trichoderma spp. isolates, including two highly effective Trichoderma atrobrunneum isolates, were able to significantly reduce levels of disease. This study highlights the potential of plants pre-colonised with T. atrobrunneum for effective protection against A. mellea in two hosts from different plant families.

Biological control agents potential and molecular identification of endophytic Trichoderma isolates originated from oil palm tissue against Ganoderma boninense Pat.

This research aimed to determine the potential of endophytic Trichoderma sp originated from oil palm plant tissue as a biological control agent and its molecular identification. The potency assay was carried out using a completely randomized design consisting of three isolates of Trichoderma sp. from different plant tissues (TR01 from root tissue, TS01 from stem tissue, and TM01 from midrib tissue) with inhibitory ability parameters. The inhibition of the growth of G. boninense was performed at the concentration of 15% and 20%, and the molecular identification of Trichoderma sp. endophyte isolates was analyzed using BLAST software. The results showed that Trichoderma sp. TM01 isolate had larger inhibitory than the other isolates on days 3 and 4 by 60% and 68%. TM01 isolate also showed inhibition to the growth of the G. boninense at 15% and 20%, with the percentage of inhibition reaching 71.79% and 82.05%. Based on phylogenetic analysis, the three endophytic isolates of Trichoderma sp. were closely related to Trichoderma virens.

Genome sequencing and evolutionary analysis of a new endophytic Trichoderma species isolated from orchid roots with reduced repertoire of protein-coding genes

Genome sequencing and evolutionary analysis of a new endophytic Trichoderma species isolated from orchid roots with reduced repertoire of protein-coding genes

Effect of Endophytic Trichoderma sp. Strains on the Agronomic Characteristics of Ecotypes of Theobroma cacao L. under Nursery Conditions in Peru

Peru is one of the main producers of fine aroma native cacao, expanding its areas considerably in recent years, which makes it necessary to seek adequate management alternatives to obtain advantageous yields. The present work had the objective of testing the influence of Trichoderma sp. endophytic strains on the agronomic characteristics of ecotypes of Theobroma cacao L. under nursery conditions, Cajaruro district, Utcubamba, Amazonas, Peru. The Trichoderma strains evaluated were Trichoderma breve; T. harzianum; T. longibrachatum; T. afrojarzianum, and Trichoderma sp. which were inoculated on cocoa seedlings of ecotypes CCN51; TCHS565; and Nativo fino de aroma. The variables evaluated were chlorophyll indices (at 30, 45, 60, and 70 days after planting), percentage of endophyte colonization, root hair development, trichomes on stems, and agronomic characteristics (plant height, number of leaves, stem diameter, root dry and fresh weight, root size, etc.). The results showed that the application of Trichoderma breve in the fine aroma native cocoa ecotype (T12) presented the highest chlorophyll index at 75 days after planting (DAP) with 43.53 ± 1.59 and 49.77 ± 2.42 for the apex and leaf base, respectively, with a percentage of colonization in the root hairs of 66.67%, and with better characteristics for the number of leaves with 12.00 ± 3.46. T12 showed positive influences for plant height, leaf number, and chlorophyll index. Treatments based on T. harzianum + CCN51 (T5) and T. afroharzianum + TSHS565 (T6) showed 100% colonization of the root hairs and trichomes on stems. Endophytic Trichoderma fungi are an alternative for organic production of fine aroma cocoa in Peru, improving the agronomic characteristics of the crop.

Endophytic Trichoderma strains isolated from forest species of the Cerrado-Caatinga ecotone are potential biocontrol agents against crop pathogenic fungi.

The indiscriminate use of chemical pesticides increasingly harms the health of living beings and the environment. Thus, biological control carried out by microorganisms has gained prominence, since it consists of an environmentally friendly alternative to the use of pesticides for controlling plant diseases. Herein, we evaluated the potential role of endophytic Trichoderma strains isolated from forest species of the Cerrado-Caatinga ecotone as biological control agents of crop pathogenic fungi. Nineteen Trichoderma strains were used to assess the antagonistic activity by in vitro bioassays against the plant pathogens Colletotrichum truncatum, Lasiodiplodia theobromae, Macrophomina phaseolina, and Sclerotium delphinii isolated from soybean, cacao, fava bean, and black pepper crops, respectively. All Trichoderma strains demonstrated inhibitory activity on pathogen mycelial growth, with maximum percent inhibition of 70% against C. truncatum, 78% against L. theobromae, 78% against M. phaseolina, and 69% against S. delphinii. Crude methanol extracts (0.5 to 2.0 mg mL-1) of Trichoderma strains were able to inhibit the growth of C. truncatum, except Trichoderma sp. T3 (UFPIT06) and T. orientale (UFPIT09 and UFPIT17) at 0.5 mg mL-1, indicating that the endophytes employ a biocontrol mechanism related to antibiosis, together with multiple mechanisms. Discriminant metabolites of Trichoderma extracts were unveiled by liquid chromatography-tandem mass spectrometry-based metabolomics combined with principal component analysis (PCA), which included antifungal metabolites and molecules with other bioactivities. These results highlight the biocontrol potential of Trichoderma strains isolated from the Cerrado-Caatinga ecotone against crop pathogenic fungi, providing support for ongoing research on disease control in agriculture.

Biocontrol potential of endophytic Trichoderma citrinoviride HT-1 against root rot of Rheum palmatum through both antagonistic effects and induced systemic resistance.

Some Trichoderma species have been used as biocontrol agents in agriculture. The effectiveness of T. citrinoviride HT-1, a beneficial endophyte isolated from Rheum palmatum root was explored for control of root rot and its mechanisms of induced systemic resistance. The results showed that the inhibition rate of F. oxysporum was 71.85% in dual culture. The fermentation metabolites (FM) of the T. citrinoviride HT-1 strain suppressed mycelial growth of F. oxysporum, recording an inhibition rate of 79.07%. Conidial suspensions of T. citrinoviride HT-1(1 × 107 spores/mL) can suppress the root rot of R. palmatum caused by F. oxysporum to a low disease index (17.60) and had significant control effects on root rot (72.53%). The activities of induced defense-related enzymes in R. palmatum plants were significantly increased following T. citrinoviride HT-1 treatment. The RT-PCR analysis of the defense-related genes showed that T. citrinoviride HT-1 can increase the defense response-related gene expression. This study has contributed to our understanding of the biocontrol potential of T. citrinoviride HT-1 and provided a theoretical basis for the application as a bio-fungicide.

In vitro and in silico investigation of the antifungal activity of endophytic fungi against phytopathogenic fungi of tomato

Plants are threatened by several diseases caused by phytopathogenic fungi. Melanin is an important pathogenicity factor in some fungal plant diseases. The enzyme 6,3,8-trihydroxynaphthalene reductase (3HNR) is implicated in the catalysis of the melanin biosynthesis in fungi. The chemical fungicide Phthalide acts by inhibiting this enzyme. But despite its efficacy, Phthalide can be detrimental to environmental health, hence the need to look for natural inhibitors to combat phytopathogenic fungi. This study aimed to screen the antifungal activity of some endophytic strains against phytopathogenic fungi of tomato. A total of 7 endophytic fungi were isolated and pre-identified from different parts of celery, parsley, mint, and coriander. On the other hand, five phytopathogenic fungal strains were isolated and pre-identified from tomatoes. The agar cylinder method showed that the endophytic fungi strains Fusarium and Trichoderma have significant inhibitory activity against four phytopathogenic fungi identified as Alternaria and Penicillium. Molecular docking was also used to study the inhibitory effect of some bioactive fungal compounds against the 3HNR enzyme. Drug-likeness and ADMET analyses were conducted on the selected chemicals to test their reliability and pharmaceutical efficacy. Phenylethyl alcohol interacts intensely with the binding site of the 3HNR receptor giving binding energy of -5.3 Kcal/mol, which is close to the co-crystallized ligand Phthalide. In addition, ADMET and pharmacokinetic analysis revealed that Phenylethyl alcohol verify the majority of the filters and pharmacokinetic properties necessary to select an effective antifungal molecule, including Lipinski’s and Veber’s rules.

Screening of endophytic Trichoderma isolates to improve the growth and health of Eucalyptus pellita seedlings

Since its first introduction to the Indonesian forest plantations, eucalyptus has been associated with pests and diseases. As a component of integrated disease management, some biocontrol agents have been developed to manage eucalyptus diseases. Application of endophytic Trichoderma is a critical option in this effort; it has been demonstrated in other pathosystems that the fungus can improve seedling growth and health. This study aims to screen and evaluate the effect of endophytic Trichoderma isolates on the growth and health of E. pellita seedlings. Field isolation resulted in 43 endophytic Trichoderma isolates. The isolates have the antagonistic ability with varied percentages of inhibition of radial growth (PIRG) against Rhizoctonia sp. (4.2–48.6%); Cylindrocladium sp. (4.8–43.5%); and Fusarium sp. (3.3–52.2%). Based on the Analytical Hierarchy Process on the variables of the growth rate of the isolates and their ability to inhibit several fungal pathogens, the best six isolates were selected for further tests. In general, the use of single and/or a consortium of the isolates increases seedling height and reduces the mortality rate of the seedlings. In summary, the tested isolates can improve plant vigor, which would later make the plant more resilient against root and foliar diseases in plantations.

<i>In Vitro</i> Screening of Endophytic <i>Trichoderma</i> sp. Isolated from Oil Palm in FGV Plantation against <i>Ganoderma boninense</i>

The potential of several endophytic Trichoderma isolates, isolated from healthy oil palm roots was assessed against Ganoderma boninense (PER71), the causal agent of Basal Stem Rot (BSR) disease through in vitro screening. In the dual culture assay, all endophytic Trichoderma isolates were found to be capable of inhibiting the growth of PER71 mycelium, by causing more than 70% inhibition of the pathogen’s radial growth (PIRG). Trichoderma asperellum M103 has caused the highest inhibition at 93.14%. The volatile compound produced by M103 also suppressed 65% of the pathogen’s radial growth. On the other hand, T. harzianum, M108 was found to have the ability to produce non-volatile compounds that is toxic and it has effectively inhibited 98.18% of the growth of PER71. Collectively, T. asperellum M103 and T. harzianum M108 have the potential to inhibit the growth of G. boninense PER71. These isolates were shown to be viable options for the future management of G. boninense in the oil palm field.

Why Is the Correct Selection of Trichoderma Strains Important? The Case of Wheat Endophytic Strains of T. harzianum and T. simmonsii.

The search for endophytic fungi in the roots of healthy wheat plants from a non-irrigation field trial allowed us to select 4 out of a total of 54 cultivable isolates belonging to the genus Trichoderma, identified as T. harzianum T136 and T139, T. simmonsii T137, and T. afroharzianum T138. In vitro assays against the phytopathogenic fungus Fusarium graminearum showed that the T. harzianum strains had the highest biocontrol potential and that T136 exhibited the highest cellulase and chitinase activities. Production patterns of eight phytohormones varied among the Trichoderma strains. All four, when applied alone or in combination, colonized roots of other wheat cultivars and promoted seed germination, tillering, and plant growth under optimal irrigation conditions in the greenhouse. Apart from T136, the endophytic Trichoderma strains showed plant protection capacity against drought as they activated the antioxidant enzyme machinery of the wheat plants. However, T. simmonsii T137 gave the best plant size and spike weight performance in water-stressed plants at the end of the crop. This trait correlated with significantly increased production of indole acetic acid and abscisic acid and increased 1-aminocyclopropane-1-carboxylic acid deaminase activity by T137. This study shows the potential of Trichoderma endophytes and that their success in agricultural systems requires careful selection of suitable strains.

The Biocontrol Potential of Endophytic Trichoderma Fungi Isolated from Hungarian Grapevines. Part I. Isolation, Identification and In Vitro Studies.

This paper reports on the identification and in vitro characterization of several Trichoderma strains isolated from the Tokaj Wine Region in North-East Hungary. Ten isolates were analyzed and found to consist of six individual species—T. gamsii, T. orientale, T. simmonsii, T. afroharzianum, T. atrobrunneum and T. harzianum sensu stricto. The growth potential of the strains was assessed at a range of temperatures. We also report here on the in vitro biocontrol properties and fungicide tolerance of the most promising strains.

Scaling-up production of cost-effective and eco-friendly bio-fertilizer and its application on Barley green fodder via IoT hydroponic system

BackgroundPlant-associated microbes (endophytes) have a significant relationship to enhance plant growth and crop productivity by producing proficient bioactive metabolites. Since endophytes promoted plant growth either directly by releasing active metabolites such as phytohormones or indirectly by suppressing the growth of phytopathogens, so, in this work, biomass yield of local endophytic Trichoderma harzianum was maximized at shake-flask scale and scaled up via 7-L Bioflo310 fermenter using continuous exponential fed-batch fermentation mode. Subsequently, the effect of these cells as bio-fertilizer was assessed using two-barley grain genotypes (Russian and Egyptian seeds) via an intelligent hydroponic system based on Internet of Things (IoT). ResultsTo reduce the cost of a biomass production line, agro-waste media containing potato, onion, garlic, pea, and cabbage peels were chosen as the culturing medium. The pea peel medium was found to be the best producer of biomass (2.2 g/L). The cultivation factors were evaluated to improve this biomass yield. The results showed that the maximum biomass production (4.9 g/L) was reported by adjusting the medium pH at 5.0 that inoculated with 10% of spore suspension, then incubated at 30°C, and 200 rpm. Then, this biomass yield was scaled up kinetically (505.4 g/L) by using exponential fed-batch fermentation mode via a 7-L bioreactor. The stimulation impacts of this endophytic T. harzianum on the growth of different barley genotypes (Russian and Egyptian seeds) were determined using a controlled hydroponic chamber. The total chlorophyll, carotenoid, and carbohydrate amounts in treated Russian showed the proficient stimulation percentage (81.05, 80, 40.8%) compared to the Egyptian barley groups (76.39, 73.5, 25.9%) respectively. Also, the maximum carbohydrate content (83.95 ± 1.7%) was recorded in the case of Russian barley. ConclusionVia this work, the optimal combination conditions for the cost-effective biomass production of endophytic T. harzianum were designed industrially via a fed-batch fermentation system using the cheapest culturing medium. Furthermore, by applying this promising bio-fertilizer, the total cost of barley production via an IoT hydroponic growing system was reduced. Besides, these animal diets (sprouted barley) could be produced in 3 cycles per month.

Use of Endophytic Trichoderma to Control Fruit Rot Phytophthora infestans, Growth and Yield in Tomato

Endophytic Trichoderma is a fungus found in plant tissue and has the potential as a plant biological control agent. The aim of this study was to study endophytic Trichoderma isolated from tomato roots, stems and leaves to control Phytophthora infestans fruit rot and growth and yield of tomato plants. The treatments that were tested were endophytic tricoderma isolated from the roots, stems and leaves of tomato plants and applied with doses of 50, 75, and 100 g per tomato plant, respectively. The experimental design used a completely randomized design with three replications. The observed variables were disease intensity, plant height, number of flowers formed, and number of fruits formed. The results showed that the endophytic Trichoderma used could control the pod rot disease of Phytophthora infestans as well as increase the growth and yield of tomato plants. Giving endophytic Trichodema from roots applied with a dose of 100 g is the best treatment because it can reduce disease intensity by 48.02%, while giving at a dose of 75 g per plant is the best treatment because it can increase plant height by 51.74%, the number of flowers formed amounted to 68.42% and the number of fruit formed was 74.70%.Keywords: Endophytic Tricoderma, Tomato, Phytophthora infestans.

Nursery evaluation of potential endophytic Trichoderma spp. from North Sumatra, Indonesia as a biocontrol agent against Ganoderma boninense

Basal stem rot (BSR) caused by the fungus Ganoderma boninense is regarded as the most destructive disease of oil palm (Elaeis guineensis Jaqc.) causes significant economic losses in the oil palm industry. An early study on the endophytic Trichoderma spp. showed they were promising potential in suppessing and preventing BSR. However, investigation on the effectiveness of endophytic Trichoderma from North Sumatra, Indonesia in preventing BSR disease in oil palms still scarce. The aim of this study was to evaluate the effectiveness of endophytic Trichoderma from North Sumatra, Indonesia in preventing BSR disease in the nursery. The active Trichoderma mixture (ATM) was prepared by first grown in PDA for six days before mixed and homogenized in 3 ml of sterile water. The mixture was later inoculated with a ratio of 1 ml to 150 g of mixture of cooked and sterilized broken corn and rice (3:4) medium. ATM was further dried and mixed homogeneously with zeolite (1:10) for application in the nursery. The potential endophytic Trichoderma spp. in suppressing BSR was monitored in the nursery for ten months. The external and internal symptoms of disease development were observed and disease incidence (DI) was also recorded. Total phenolic content (TPC) was analyzed using Folin-Ciocalteu method to determine the effect of endophytic Trichoderma in inducing oil palm seedling resistance to BSR disease. The result showed that application of endophytic Trichoderma spp. was found to be effective in suppressing G. boninense infection, where endophytic T. reesei ET501 to be the most effective in preventing disease incidence (DI) with only 13.4 % infection after ten months of observation followed by endophytic T. asperellum ET537 (48.4 %) and T. asperellum ET523 (71.8 %), respectively. Application of endophytic Trichoderma spp. in the nursery also exhibited the potential in inducing TPC amount and it helps in developing resistance in oil palm seedlings, thus protecting the seedlings from G. boninense infection.

Identification of native endophytic Trichoderma spp. for investigation of in vitro antagonism towards Armillaria mellea using synthetic- and plant-based substrates.

To isolate endophytic Trichoderma species and investigate the potential for biological control of the root rot pathogen Armillaria mellea. In all, 40 Trichoderma isolates were obtained from a range of host plants and identities were confirmed by ITS, rpb2 and tef1 sequence. When tested in dual culture assays for antagonism against A. mellea, Trichoderma isolates overgrew the A. mellea colonies within four days and by eightdays 38 Trichoderma isolates significantly reduced A. mellea colony size. Armillaria mellea was unable to be recovered from five of eight co-cultivations tested, suggesting Trichoderma had killed the A. mellea in these cases. Pre-colonized hazel disks were used to determine what happens in a more heterogeneous situation with A. mellea and a refined set of eight Trichoderma isolates. Similar to plate-based assays, Trichoderma quickly covered A. mellea stopping any further growth and two Trichoderma isolates were able to eradicate A. mellea. Of the Trichoderma spp. tested, endophytic isolates of Trichoderma virens and T. hamatum offered the greatest antagonism towards A. mellea. Using pre-colonized hazel disks was of great importance for this work to demonstrate the fungal interactions in plant material. Controlling Armillaria root rot is difficult with chemical treatments, thus an environmentally benign and cost-effective alternative is required. This study highlights the prospect of biological control as an effective, environmentally friendly alternative to chemicals.

An Endophytic Trichoderma Strain Promotes Growth of Its Hosts and Defends Against Pathogen Attack.

Plants host numerous endophytic microbes which promote plant performance, in particular under stress. A new endophytic fungus was isolated from the leaves of a deciduous wood tree Leucas aspera. Morphological inspection and multilocus phylogeny identified the fungus as a new Trichoderma strain. If applied to Arabidopsis thaliana and Nicotiana attenuata, it mainly colonizes their roots and strongly promotes initial growth of the plants on soil. The fungus grows on high NaCl or mannitol concentrations, and shows predatory capability on the pathogenic fungus Alternaria brassicicola. Colonized Arabidopsis plants tolerate higher salt stress and show lower A. brassicicola spread in roots and shoots, while arbuscular mycorrhiza formation in N. attenuata is not affected by the Trichoderma strain. These beneficial features of the novel Trichoderma strain are important prerequisites for agricultural applications.

Inhibition of plant pathogenic fungi by endophytic Trichoderma spp. through mycoparasitism and volatile organic compounds

Antagonism of plant pathogenic fungi by endophytic fungi is a well-known phenomenon. In plate assays, the antagonism could be due to mycoparasitism, competition for space or antibiosis, involving a chemical diffusate, or a volatile organic compound (VOC). In this study, we demonstrate that besides mycoparasitism, VOCs play a major role in antagonism of pathogenic fungi by four endophytic fungi belonging to the genus Trichoderma. Using a double-plate assay, we show that all the four endophytic Trichoderma species significantly inhibited mycelial growth of three of the four pathogens, (Sclerotinia sclerotiorum-TSS, Sclerotium rolfsii-CSR and Fusarium oxysporum-CFO), while that of Macrophomina phaseolina-CMP was not affected. GC–MS analysis of the pure cultures of one of the endophytic fungi studied, namely, Trichoderma longibrachiatum strain 2 (Acc. No. MK751758) and the pathogens, F. oxysporum-CFO and M. phaseolina-CMP revealed the presence of several VOCs including hydrocarbons, alcohols, ketones, aldehydes, esters, acids, ethers and different classes of terpenes. In mixed double plates, where the endophyte was grown along with either of the two plant pathogens, F. oxysporum-CFO or M. phaseolina-CMP, there was an induction of a number of new VOCs that were not detected in the pure cultures of either the endophyte or the pathogens. Several of these new VOCs are reported to possess antifungal and cytotoxic activity. We discuss these results and highlight the importance of such interactions in endophyte-pathogen interactions.