Endophytic Trichoderma Research Articles

Simplex-lattice design and decision tree optimization of endophytic Trichoderma-multi-walled carbon nanotube composite for enhanced methylene blue removal

This study investigates a novel approach for enhancing methylene blue (MB) removal from water using a composite of endophytic Trichoderma mate and multi-walled carbon nanotubes (MWCNTs). For the first time, a unique combination of simplex-lattice design and decision tree learning algorithm was employed to optimize MB removal. This innovative approach effectively identified the optimal composite ratio of hyphal mate (0.5354 g/L) and MWCNTs (0.4646 g/L) for maximizing MB removal, which achieved remarkable removal efficiency ranging from 63.50 to 95.78 % depending on the combination used. The DT model further demonstrated promising potential for predicting MB removal efficiency. SEM revealed a unique hybrid material formed by the intertwining or entrapment of MWCNTs within the hyphal network of Trichoderma mate. FT-IR analysis confirmed the presence of novel functional groups on the MWCNTs' surface at 2438.79 and 528.25 cm−1, likely due to interactions with the endophytic fungi's biomolecules. These functional groups presumably act as reducing and stabilizing agents, promoting efficient MB adsorption. This research paves the way for utilizing the combined biological and chemical approach (fungal biomass and MWCNTs) in bioremediation applications. The findings suggest significant potential for practical applications in wastewater treatment, providing an eco-friendly and cost-effective method for dye removal. Furthermore, the proposed method shows promise for scaling up to industrial wastewater treatment and applicability in resource-limited settings, offering a sustainable solution for global water pollution challenges. Further investigations with larger datasets incorporating additional influencing factors are necessary to refine the predictive power of the DT model for practical applications.

Optimization, purification and characterization of laccase from a new endophytic Trichoderma harzianum AUMC14897 isolated from Opuntia ficus-indica and its applications in dye decolorization and wastewater treatment.

Hazardous synthetic dye wastes have become a growing threat to the environment and public health. Fungal enzymes are eco-friendly, compatible and cost-effective approachfor diversity of applications. Therefore, this study aimed to screen, optimize fermentation conditions, and characterize laccase from fungal endophyte with elucidating its ability to decolorize several wastewater dyes. A new fungal endophyte capable of laccase-producing was firstly isolated from cladodes of Opuntia ficus-indica and identified as T. harzianum AUMC14897 using ITS-rRNA sequencing analysis. Furthermore, the response surface methodology (RSM) was utilized to optimize several fermentation parameters that increase laccase production. The isolated laccase was purified to 13.79-fold. GFC, SDS-PAGE revealed laccase molecular weight at 72kDa and zymogram analysis elucidated a single band without any isozymes. The peak activity of the pure laccase was detected at 50°C, pH 4.5, with thermal stability up to 50°C and half life span for 4h even after 24h retained 30% of its activity. The Km and Vmax values were 0.1mM, 22.22µmol/min and activation energy (Ea) equal to 5.71kcal/mol. Furthermore, the purified laccase effectively decolorized various synthetic and real wastewater dyes. Subsequently, the new endophytic strain produces high laccase activity that possesses a unique characteristic, it could be an appealing candidate for both environmental and industrial applications.

Antagonistic and molecular characterization of endophytic Trichoderma from sugarcane

Antagonistic and molecular characterization of endophytic Trichoderma from sugarcane

Potential biological control agents of Geosmithia morbida restrict fungal pathogen growth via mycoparasitism and antibiosis

Thousand cankers disease of Juglans (walnut) and Pterocarya (wingnut) spp. (Fagales: Juglandaceae) is caused by the fungal pathogen Geosmithia morbida Kolarík, Freeland, Utley, and Tisserat (Hypocreales: Bionectriaceae) and bark beetle pest/vector, Pityophthorus juglandis Blackman (Coleoptera: Curculionidae). To further the development of biological management strategies for thousand cankers disease, we assessed the ability of 14 endophytic Trichoderma (Hypocreales: Hypocreaceae) isolates and the commercially available isolate T. afroharzianum strain KRL-AG2 to inhibit the in vitro growth of three different G. morbida isolates via mycoparasitism and antibiosis. To identify factors that may affect field success of candidate biological control agents, we quantified the growth responses of Trichoderma spp. and the commercially available entomopathogenic fungus, Beauveria bassiana (Bals.-Criv.) Vuill. (Hypocreales: Cordycipitaceae) strain GHA, to the plant secondary metabolite and antimicrobial compound, juglone in vitro. A total of 12 Trichoderma isolates (from six different Trichoderma species) demonstrated antagonistic activity towards G. morbida in dual-plate assays. Juglone consistently reduced the growth of B. bassiana strain GHA and 14 of the 15 screened Trichoderma isolates in vitro. Additionally, one metabolite-producing Trichoderma isolate, TN4-47, completely inhibited the growth of all three G. morbida isolates across all tested metabolite concentrations and had comparatively greater tolerance to juglone compared to other Trichoderma isolates. Future lines of research should focus on characterizing the active antagonistic compound present in the metabolite filtrates, determine the mode of action of the active component(s), and elucidate how abiotic and biotic factors may influence the growth, persistence, and antagonistic activity of candidate biological control agents in planta.

Endophytic Trichoderma: Potential and Prospects for Plant Health Management.

The fungus Trichoderma is widely regarded as the most common fungal biocontrol agent for plant health management. More than 25 Trichoderma species have been extensively studied and have demonstrated significant potential in inhibiting not only phytopathogen growth but also insect pest infestations. In addition to their use as biopesticides, there is increasing evidence that several Trichoderma species can function as fungal endophytes by colonizing the tissues of specific plants. This colonization enhances a plant's growth and improves its tolerance to abiotic and biotic stresses. In recent decades, there has been a proliferation of literature on the role of Trichoderma endophytes in crop protection. Although the mechanisms underlying plant-fungal endophyte interactions are not yet fully understood, several studies have suggested their potential application in agriculture, particularly in the mitigation of plant pests and diseases. This review focuses on the diversity of Trichoderma endophytic strains and their potential use in controlling specific diseases and pests of crop plants. Trichoderma endophytes are considered a potential solution to reduce production costs and environmental impact by decreasing reliance on agrochemicals.

Biological control of Corynespora leaf fall disease in rubber by endophytic Trichoderma spp. under field conditions

Biological control of Corynespora leaf fall disease in rubber by endophytic Trichoderma spp. under field conditions

Evaluation of the potential of endophytic Trichoderma sp. isolated from medicinal plant Ampelopsis japonica against MRSA and bioassay-guided separation of the anti-MRSA compound.

Endophytic fungi have been recognized as a valuable source for the production of biologically active compounds with potential applications in various domains. This study aimed to isolate endophytic fungi from Ampelopsis japonica (Thunb.) Makino and assess their anti-MRSA activity. Meanwhile, chromatographic separation techniques were applied to analyze the constituents of endophytic fungal secondary metabolites. The isolate BLR24, which exhibited strong inhibition activity against MRSA, was identified as Trichoderma virens based on morphological characteristics and ITS sequence analyses. The ethyl acetate extract of BLR24 (EA-BLR24) showed good anti-MRSA activity with the MIC and MBC values of 25 μg/mL and 50 μg/mL, separately. The inhibition of biofilm formation was up to 34.67% under MIC concentration treatment. Meanwhile, EA-BLR24 could significantly reduce the expression of biofilm-related genes (icaA, sarA, and agrA) of MRSA. Based on LC-MS/MS analysis, twenty compounds in EA-BLR24 could be annotated using the GNPS platform, mainly diketopiperazines. The anti-MRSA compound (Fr.1.1) was obtained from EA-BLR24 by bioassay-guided fractionation and determined as gliotoxin. The results indicated that endophytic Trichoderma virens BLR24 isolated from the medical plant A. japonica roots could be a promising source of natural anti-MRSA agents. Endophytic fungal secondary metabolites are abundant in biologically active compounds. Endophytic fungi from medicinal plants could be a source yielding bioactive metabolites of pharmaceutical importance.

Two new acorane-type sesquiterpenoids from an endophytic Trichoderma harzianum associated with Paeonia lactiflora Pall

Two new acorane-type sesquiterpenoids, harzianes A and B (1 and 2), together with two known cyclonerodiol-type sesquiterpenoids (3–4) and four known sterols (5–8) were isolated from the endophytic Trichoderma harzianum, associated with the medicinal plant Paeonia lactiflora Pall. Compounds 1 and 2 were identified as a pair of heterotropic isomers by spectroscopic analysis (HR-ESI-MS, 1D and 2D NMR), and their absolute configurations were determined by ECD calculations. All compounds were tested for anti-inflammatory activity, however, none demonstrated such activity.

Endophyte genomes support greater metabolic gene cluster diversity compared with non-endophytes in Trichoderma.

Trichoderma is a cosmopolitan genus with diverse lifestyles and nutritional modes, including mycotrophy, saprophytism, and endophytism. Previous research has reported greater metabolic gene repertoires in endophytic fungal species compared to closely-related non-endophytes. However, the extent of this ecological trend and its underlying mechanisms are unclear. Some endophytic fungi may also be mycotrophs and have one or more mycoparasitism mechanisms. Mycotrophic endophytes are prominent in certain genera like Trichoderma, therefore, the mechanisms that enable these fungi to colonize both living plants and fungi may be the result of expanded metabolic gene repertoires. Our objective was to determine what, if any, genomic features are overrepresented in endophytic fungi genomes in order to undercover the genomic underpinning of the fungal endophytic lifestyle. Here we compared metabolic gene cluster and mycoparasitism gene diversity across a dataset of thirty-eight Trichoderma genomes representing the full breadth of environmental Trichoderma's diverse lifestyles and nutritional modes. We generated four new Trichoderma endophyticum genomes to improve the sampling of endophytic isolates from this genus. As predicted, endophytic Trichoderma genomes contained, on average, more total biosynthetic and degradative gene clusters than non-endophytic isolates, suggesting that the ability to create/modify a diversity of metabolites potential is beneficial or necessary to the endophytic fungi. Still, once the phylogenetic signal was taken in consideration, no particular class of metabolic gene cluster was independently associated with the Trichoderma endophytic lifestyle. Several mycoparasitism genes, but no chitinase genes, were associated with endophytic Trichoderma genomes. Most genomic differences between Trichoderma lifestyles and nutritional modes are difficult to disentangle from phylogenetic divergences among species, suggesting that Trichoderma genomes maybe particularly well-equipped for lifestyle plasticity. We also consider the role of endophytism in diversifying secondary metabolism after identifying the horizontal transfer of the ergot alkaloid gene cluster to Trichoderma.

Characterization and antimicrobial activities of bioactive compounds from endophytic Trichoderma asperellum isolated from Dendrobium orchids

Characterization and antimicrobial activities of bioactive compounds from endophytic Trichoderma asperellum isolated from Dendrobium orchids

New endophytic strains of Trichoderma promote growth and reduce clubroot severity of rapeseed (Brassica napus).

Rapeseed (Brassica napus L.) is the world's third most important edible oilseed crop after soybean and palm. The clubroot disease caused by Plasmodiophora brassicae poses a significant risk and causes substantial yield losses in rapeseed. In this study, 13 endophytic fungal strains were isolated from the healthy roots of rapeseed (B. napus) grown in a clubroot-infested field and molecularly identified. Based on germination inhibition of resting spores of P. brassicae, two endophytic fungal antagonists, Trichoderma spp. ReTk1 and ReTv2 were selected to evaluate their potential for plant growth promotion and biocontrol of P. brassicae. The Trichoderma isolates were applied as a soil drench (1×107 spore/g soil) to a planting mix and field soil, in which plants were grown under non-infested and P. brassicae-infested (2×106 spore/g soil) conditions. The endophytic fungi were able to promote plant growth, significantly increasing shoot and root length, leaf diameter, and biomass production (shoots and root weight) both in the absence or presence of P. brassicae. The single and dual treatments with the endophytes were equally effective in significantly decreasing the root-hair infection, root index, and clubroot severity index. Both ReTk1 and ReTv2 inhibited the germination of resting spores of P. brassicae in root exudates. Moreover, the endophytic fungi colonized the roots of rapeseed extensively and possibly induced host resistance by up-regulated expression of defense-related genes involved in jasmonate (BnOPR2), ethylene (BnACO and BnSAM3), phenylpropanoid (BnOPCL and BnCCR), auxin (BnAAO1) and salicylic acid (BnPR2) pathways. Based on these findings, it is evident that the rapeseed root endophytes Trichoderma spp. ReTk1 and ReTv2 could suppress the gall formation on rapeseed roots via antibiosis, induced systemic resistance (ISR), and/or systemic acquired resistance (SAR). According to our knowledge, this is the first report of the endophytic Trichoderma spp. isolated from root tissues of healthy rapeseed plants (B. napus.), promoting plant growth and reducing clubroot severity.

Comparison of Fungal Genera Isolated from Cucumber Plants and Rhizosphere Soil by Using Various Cultural Media.

Plant endophytic fungi and rhizosphere soil fungi are often reported as biocontrol agents against plant pathogens or with plant growth promotion potential. Four treatments were performed in field and greenhouse experiments where cucumber plants were inoculated with Trichoderma harzianum and Fusarium oxysporum in 2022. The roots, stems and leaves of cucumber plants and their rhizosphere soil were collected twice individually from the field and greenhouse for isolation of cucumber endophytic and rhizosphere soil fungi. All fungal strains were identified through sequence similarity of the ITS1-5.8s-ITS2 rDNA region. The potato dextrose agar (PDA) media yielded the highest number of genera isolated from cucumber plants, rhizosphere soil and both compared to other media. There were no significant differences among the four media for the isolation of all cucumber endophytic fungi. However, in the roots, the number of endophytic fungi isolated by MRBA was significantly higher than that isolated on malt extract agar (MEA), while in the stems, the number of fungi isolated with PDA was significantly higher than that isolated with Martin's rose bengal agar medium (MRBA). PDA had significantly higher isolation efficiency for the rhizosphere soil fungi than MRBA. The 28 fungal genera had high isolation efficiency, and the endophytic Trichoderma strains were significantly more isolated by MEA than those of MRBA. It is suggested that PDA can be used as a basic medium, and different cultural media can be considered for specific fungal genera.

Application of Endophytic Trichoderma harzianum to Alleviate the Adverse Impact of Chromium in Lupinus albus L

The present study was conducted to understand the role of interactive usage of Trichoderma harzianum to alleviate the adverse impact of chromium (Cr) stress for sustainable production of the legume plants (Lupinus albus L.) with special reference to physiological mechanism. The exogenous application of T. harzianum significantly enhanced seed germination and plant growth, along with all physiological and biochemical attributes examined in lupin plant. The germination percentage of lupin seeds treated with conidia suspension of T. harzianum was 100% with 37% increase as compared to the control. T. harzianum inoculation increased seedling height (41.3%) and root length (75.6%). Treatment of T. harzianum also resulted in significant enhancement in the photosynthetic attributes like chlorophyll pigment synthesis (13.75% increase as compares to the control), reflecting a significant enhancement in the net photosynthetic rate. Results of our study demonstrated that L. albus have improved tolerance to inhibitory concentrations of Cr when inoculated with T. harzianum. So, we suggest the application of T. harzianum, which could be used to improve the quality and yield of lupin (L. albus) plants in Cr-contaminated areas and in fertile soils.

Trichodermic acids from an endophytic Trichoderma sp. and their antifungal activity against the phytopathogen Botrytis cinerea

Phytopathogen Botrytis cinerea is a necrotrophic pathogen with a wide host range, which causes considerable economic losses in agriculture. Consequently, it is necessary to find safe and effective antifungal agents against B. cinerea from natural sources. In this study, eight octahydronaphthalene derivatives (1−8) including four new natural products named trichodermic acids E − H (1−4), were isolated from an endophytic fungus Trichoderma sp. Their structures were determined by analysis of their nuclear magnetic resonance (NMR) and high-resolution electrospray ionization mass spectra (HR-ESI-MS) and by electronic circular dichroism (ECD) data. All compounds showed some extent of antifungal activities against B. cinerea with minimum inhibitory concentration (MIC) values ranging from 4 to 64 μg/mL. Notably, compound 8 showed significant inhibition with an MIC of 4 μg/mL, which was visually confirmed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). These findings provide a potential application of trichodermic acids as natural “hit” fungicides in agriculture.

Trichoderma asperellum integrated with Fusarium decemcellulare and Lasiodioplodia pseudotheobromae applications increase its colonisation in cacao seedling tissues and suppression of leaf anthracnose disease

Fusarium and Lasiodiplodia are fungal endophytes found frequently in cacao tissues. Their presence has suggested an increase endophytic Trichoderma asperellum’s ability to inhibit primary diseases. The research aims to evaluate the occurrence of T. asperellum in root, stem, and leaf cacao tissues after the plants were inoculated by F. decemcellulare, L. pseudotheobromae, and T. asperellum. Also, at the same time to observe their impact on inhibiting leaf anthracnose caused by Colletotrichum gloeosporioides. In plants treated with the combination of T. asperellum and F. decemcellulare at 30 days post-inoculation, colonisation of T. asperellum was four to 68 times higher than that of T. asperellum alone. Whereas when combined with L. pseudotheobromae, the colonisation was 4 to 68 times higher. T. asperellum, F. decemcellulare, and L. theobromae self-applied could decrease the incidence and severity of anthracnose disease. However, the highest decrease was observed in plants treated notably by the combination of T. asperellum and F. decemcellulare. This data demonstrated that the presence of T. asperellum in plant tissues was supported by other endophytic fungi, which increased its capability to suppress the disease of cacao.

Endophytic Trichoderma species from rubber trees native to the Brazilian Amazon, including four new species.

Fungi belonging to the genus Trichoderma have been widely recognized as efficient controllers of plant diseases. Although the majority of isolates currently deployed, thus far, have been isolated from soil, endophytic Trichoderma spp. is considered to be a promising option for application in biocontrol. In this study, 30 endophytic Trichoderma isolates-obtained from the leaves, stems, and roots of wild Hevea spp. in the Brazilian Amazon-were analyzed using specific DNA barcodes: sequences of internal transcribed spacers 1 and 2 of rDNA (ITS region), genes encoding translation elongation factor 1-α (TEF1-α), and the second largest subunit of RNA polymerase II (RPB2). The genealogical concordance phylogenetic species recognition (GCPSR) concept was used for species delimitation. A phylogenetic analysis showed the occurrence of Trichoderma species, such as T. erinaceum, T. ovalisporum, T. koningiopsis, T. sparsum, T. lentiforme, T. virens, and T. spirale. Molecular and morphological features resulted in the discovery of four new species, such as T. acreanum sp. nov., T. ararianum sp. nov., T. heveae sp. nov., and T. brasiliensis sp. nov. The BI and ML analyses shared a similar topology, providing high support to the final trees. The phylograms show three distinct subclades, namely, T. acreanum and T. ararianum being paraphyletic with T. koningiopsis; T. heveae with T. subviride; and T. brasiliensis with T. brevicompactum. This study adds to our knowledge of the diversity of endophytic Trichoderma species in Neotropical forests and reveals new potential biocontrol agents for the management of plant diseases.

Biocontrol efficacy of endophytic Trichoderma spp. in fresh and dry powder formulations in controlling northern corn leaf blight in sweet corn

Seventy-three endophytic Trichoderma spp. obtained from healthy corn leaves collected in the central and northeast parts of Thailand were evaluated for their biocontrol activity against northern leaf blight disease due to Exserohilum turcicum. Among them, T. asperellum KUFA0702, KUFA0703, T. hamatum KUFA0706, T. harzianum KUFA0710, and T. harzianum KUFA0713 showed promising biocontrol efficacy against the disease under laboratory and greenhouse conditions; thus, they were selected for control efficacy evaluation under field conditions using their fresh and dry powder formulations. The results found that the fresh formulation of each Trichoderma strain displayed better biocontrol effects than their dry formulation. In field trials, the application of fresh formulation of T. harzianum KUFA0710 displayed the best disease reduction at 55.70%, while its dry formulation caused 47.46% disease reduction followed by T. harzianum KUFA0713 at 49.92% and 43.84% disease reduction when applied as fresh and dry formulations, respectively. The antagonistic activities of Trichoderma spp. included fast growth with high competition for space and nutrients and production of antifungal crude extracts. Moreover, all Trichoderma strains showed compatibility fungicides at recommendation rate with azoxystrobin (1000 ppm) and difenoconazole + azoxystrobin (500 ppm). These fungi can serve as alternative fungicides or be combined with fungicides for integrated pest management.

Rice Plants’ Resistance to Sheath Blight Infection Is Increased by the Synergistic Effects of Trichoderma Inoculation with SRI Management

The capability of endophytic Trichoderma spp. to reduce sheath blight disease in rice caused by Rhizoctonia solani was assessed under the growth conditions established by practices of the System of Rice Intensification (SRI), compared to those of standard irrigated rice cultivation. Rice seeds inoculated with a local isolate of the fungus Trichoderma asperellum SL2 were grown under respective SRI and conventional conditions with the inoculated or uninoculated plants and then infected with the pathogen R. solani. It was seen that inoculation with this strain of Trichoderma protected rice plants against R. solani infection while enhancing plant growth, photosynthetic rate, and stomatal conductance. The biocontrol effectiveness of inoculation with a particular strain of Trichoderma was significantly greater under SRI management compared to conventional cultivation. This is the first report on how a crop management system, in this case, SRI, can influence the biocontrol effectiveness of Trichoderma spp.

Biodegradation of Aflatoxin B1 in Maize Grains and Suppression of Its Biosynthesis-Related Genes Using Endophytic Trichoderma harzianum AYM3

Aflatoxin B1 is one of the most deleterious types of mycotoxins. The application of an endophytic fungus for biodegradation or biosuppression of AFB1 production by Aspergillus flavus was investigated. About 10 endophytic fungal species, isolated from healthy maize plants, were screened for their in vitro AFs-degrading activity using coumarin medium. The highest degradation potential was recorded for Trichoderma sp. (76.8%). This endophyte was identified using the rDNA-ITS sequence as Trichoderma harzianum AYM3 and assigned an accession no. of ON203053. It caused a 65% inhibition in the growth of A. flavus AYM2 in vitro. HPLC analysis revealed that T. harzianum AYM3 had a biodegradation potential against AFB1. Co-culturing of T. harazianum AYM3 and A. flavus AYM2 on maize grains led to a significant suppression (67%) in AFB1 production. GC-MS analysis identified two AFB1-suppressing compounds, acetic acid and n-propyl acetate. Investigating effect on the transcriptional expression of five AFB1 biosynthesis-related genes in A. flavus AYM2 revealed the downregulating effects of T. harzianum AYM3 metabolites on expression of aflP and aflS genes. Using HepaRG cell line, the cytotoxicity assay indicated that T. harazianum AYM3 metabolites were safe. Based on these results, it can be concluded that T. harzianum AYM3 may be used to suppress AFB1 production in maize grains.

Potential of endophytic Trichoderma in controlling Phytophthora leaf fall disease in rubber (Hevea brasiliensis)

Fifty-nine endophytic Trichoderma spp., isolated from healthy rubber leaves collected in the eastern and southern parts of Thailand, were evaluated for their antagonistic activity against Phytophthora leaf fall disease, caused by Phytophthora palmivora. Three strains of T. asperellum, three strains of T. harzianum, three strains of T. hamatum, one strain of T. longibrachiatum, and two strains of T. virens were selected based on their potent antagonistic activity against P. palmivora when tested by dual culture. Among the endophytic Trichoderma strains tested, T. harzianum KUFA0436 and KUFA0437 showed potent biocontrol efficacy, suppressing the disease severity by 43 % and 48 %, respectively, under greenhouse conditions. Meanwhile, the other strains showed low to moderate suppression of the disease severity when tested in laboratory and under greenhouse conditions. Trichoderma harzianum KUFA0437 and KUFA0437 were selected to be tested under field conditions and were found to display a significant reduction in defoliation of 30–33 %; while metalaxyl (positive control) showed the best suppression of defoliation of up to 66 %. The antagonistic activities of Trichoderma spp. included fast growth, strong competition for natural resources, and production of antifungal crude extracts. Moreover, all Trichoderma strains showed compatibility with propineb (1,750 ppm), fosetyl-Al (1,600 ppm), and metalaxyl (500 ppm). To the best of our knowledge, this is the first report of endophytic Trichoderma spp. as potential biocontrol agents for controlling rubber diseases. These fungi can serve as alternative fungicides to reduce the use of synthetic fungicides or as part of an integrated pest management scheme in combination with fungicidal treatments.