In the search for natural products with potent activity against plant diseases, we employed the one strain-many compounds (OSMAC) strategy to the marine-derived fungus Trichoderma virens B211 by supplementing cultures with halogen salts such as sodium bromide (NaBr). Both solid and liquid cultures grown under NaBr-supplemented conditions exhibited markedly darker pigmentation of B211, a diversified secondary metabolite profile, and enhanced antibacterial activity against Erwinia amylovora. Bioassay-guided fractionation of these extracts led to the isolation of fourteen compounds (1-14), including two new polycyclic aromatic terpenoids (1 and 5) belonging to the viridin family. In vitro antimicrobial assays demonstrated that viridin (3) and β-viridin (4), together with gliotoxin (8), displayed broad-spectrum activity against diverse plant pathogenic fungi and bacteria. Notably, viridin derivatives 1 and 2, which are structural isomers, exhibited distinct bioactivity profiles: compound 1 exhibited antifungal activity, whereas compound 2 showed antibacterial activity. Among the isolated metabolites, only gliotoxin (8) and bisdethiobis(methylthio)gliotoxin (9) exhibited direct antibacterial activity against E. amylovora. Furthermore, NaBr-induced crude extracts in which compounds 8 and 9 were the major constituents, as well as the purified compounds themselves, effectively suppressed fire blight symptoms. Collectively, these findings highlight the effectiveness of the OSMAC approach in expanding the chemical diversity of metabolites produced by T. virens B211 and enhancing its potential as a biocontrol agent for plant disease management.

Fungi of the genus Trichoderma have attracted attention because of their potential activity against phytophagous mites; however, information regarding their non-target effects on predatory mites remains limited. This study evaluated the effects of spore suspensions and secondary metabolites of Trichoderma afroharzianum Tr132 and Trichoderma virens Tvr2 on three predatory mite species widely used in biological control programs: Phytoseiulus persimilis, Neoseiulus californicus, and Amblyseius swirskii. Predator egg hatchability and adult mortality were assessed under laboratory conditions. Spore suspension treatments did not significantly affect egg hatchability, which remained high (97-99%) across all predator species. In contrast, secondary metabolites slightly reduced egg hatchability to 94-96%, compared with 99.5% in the control. Exposure to spore suspensions caused moderate mortality in adult predatory mites, increasing from 10 to 13% at 3 days post-application (dpa) to 15-19% at 6 dpa. Secondary metabolites produced higher mortality that increased over time, reaching 9-11% at 1 dpa, 17-18% at 3 dpa, and up to 22-25% at 6 dpa. Mortality responses were similar among predator species. Overall, Trichoderma applications had minimal effects on predator egg hatchability but caused moderate mortality in adult predatory mites, particularly following exposure to secondary metabolites. These findings highlight the importance of evaluating the compatibility of Trichoderma-based products with beneficial predatory mites before their integration into IPM programs.

This study was conducted to determine the practical feasibility of using Trichoderma virens as a potential biological control agent against Paramarasmius palmivorus . The pathogen was isolated from symptomatic cowpea plants collected in the research field at the Faculty of Agriculture, Rajarata University of Sri Lanka, Anuradhapura, Sri Lanka. The isolated pathogen was initially identified based on morphological characteristics and subsequently confirmed using molecular biological tools (PV216709). In dual culture plate assays, T . virens exhibited an 89.74% inhibition of mycelial growth against P. palmivorus . The efficacy of T . virens formulation was tested in a pot experiment under shade house conditions. Plant growth parameters and disease incidence were monitored. Plant growth parameters were analyzed using a repeated mixed effect model. Colony‐forming units were analyzed using logistic regression in SAS Studio version 9.0 software. The application of T . virens exhibited zero disease incidence under shade house conditions (28°C ± 1°C and 70% RH). Treatment with T . virens significantly enhanced ( p < 0.05) plant height, flower number, pod number, and root length compared to the untreated control. Results revealed that the spores of T . virens were viable and multiplied throughout the experimental period.

Basal rot is a soil borne disease caused by Fusarium oxysporum f. sp. cepae. It is known to affect garlic production throughout the world. Various strategies such as crop rotation and raising of resistant varieties have been tried to limit the damage caused due to this fungus. Biocontrol is another effective option that can be used to control the spread of disease. Biocontrol process is generally harmless to humans, a non-polluting, biodegradable and selective mode of action. Moreover, pathogens cannot develop resistance easily, no harm is caused to other beneficial microorganisms, improve soil health and help in achieving agricultural sustainability. Efficacy of fungal and bacterial bioagents were checked against the basal rot of garlic. The potential of six species of fungal species viz. Trichoderma harzianum, Trichoderma viridae, Trichoderma virens, Trichoderma hamatum, bacterial species viz. Pseudomonas fluorescens and Bacillus subtilis were evaluated against F. oxysporum under in vitro conditions. Dual culture and streak plate method was used evaluation of fungal and bacterial antagonists. Among the fungal antagonists maximum inhibition of mycelial growth was noted in Trichoderma viridae (74 %), Pseudomonas fluorescens proved to be the most effective bacteria in reducing (30 %) the mycelial growth.

Microbial community in a soilless cultivation system with organic fertilization, biofertilizers and cover crop rotation.

This study investigates the antifungal potential of hydrothermal liquefaction (HTL) wastewater against Verticillium dahliae and its effects on the growth of pepper plants (Capsicum annuum L.). The HTL process generates wastewater containing various antimicrobial compounds, which can offer a sustainable alternative for plant protection. In this research, the biological control agent Trichoderma virens and HTL wastewater were applied both individually and in combination to assess their impact on plant growth and pathogen suppression. The results demonstrated that specific HTL wastewater treatments significantly enhanced root and shoot growth, biomass, and chlorophyll content in pepper plants. Plant growth observed in pathogen-free conditions may be related to the stimulation of plant physiology by biologically active components contained in wastewater, indicating a biostimulatory effect. Notably, the 4th wastewater mixture (wm) exhibited the highest efficacy, promoting plant development and mitigating the negative effects of V. dahliae. The combination of T. virens and wastewater also showed synergistic effects, reducing disease severity by up to 64% and improving plant biomass and structural parameters. Statistical analysis revealed significant differences among treatments, highlighting the potential of HTL wastewater as a natural and sustainable strategy for managing soil-borne pathogens. These findings suggest that integrating HTL wastewater with biocontrol agents could offer a promising approach to sustainable agriculture.

Groundnut (Arachis hypogaea L.) is an important oilseed crop which is primarily grown in tropical and subtropical regions. Dry root rot caused by soil borne fungus Macrophomina phaseolina is the most destructive and widespread disease worldwide. For better management of this disease, rhizosphere antagonists and organic amendments were employed both in vitro and in vivo studies. Seven rhizosphere antagonists (Trichoderma sp. - Tsp 1, Tsp 2, Tsp 3, Tsp 4, Bacillus sp. - Bsp 1, Bsp 2 and Pseudomonas sp.) were isolated from rhizosphere regions of healthy groundnut plants and tested in vitro against M. phaseolina by dual culture method. Among the antagonists tested, Tsp1 recorded maximum mycelial inhibition of 67.81 %. The best performing antagonists Tsp1 was molecularly characterised and confirmed as Trichoderma virens. Among the seven different organic amendments tested, neem cake showed maximum mycelial inhibition of 52.21 % and 52.50 % at 10 % and 15 % concentrations respectively. The effective rhizosphere antagonists and organic amendments were tested against M. phaseolina under pot culture and field condition. In pot culture experiment, soil application of T. virens (5 g/pot) + neem cake (5 g/pot) was effective in managing the dry root rot disease with lowest disease incidence (22.21 %) against inoculated control (88.88 %). In field experiment, soil application of T. virens (2.5 kg/ha) + neem cake (150 kg/ha) recorded lowest disease incidence of 10.32 % and highest yield of 3200 kg/ha against untreated control (58.57 %, 900 kg/ha respectively).

This study evaluated the efficacy of Trichoderma virens in controlling Agroathelia rolfsii, the causal agent of cowpea crown rot disease through in vitro, in vivo experiments. Following isolation and confirmation of the pathogen, dual plate assay was conducted in vitro. Pot and field trials were conducted under shade house and in both sterilized and non-sterilized field conditions. Agroathelia rolfsii was inoculated onto the crown area of cowpea seedlings using agar disc method, with one agar disc per plant. Trichoderma virens was applied as a spore suspension at planting and three-week intervals. Plant growth parameters, yield components, CFU, disease incidence were monitored throughout the study. According to the results, morphological, molecular identification confirmed the pathogen as A. rolfsii. Pathogenicity was verified through Koch’s postulate. In the dual plate assay, T. virens inhibited A. rolfsii by 86.02%. The application of T. virens significantly improved plant height, leaf number, flower number, pod number, root length, and seed weight, resulting in zero disease incidence. Furthermore, viability of T. virens in soil was ensured by showing fluctuations in CFU throughout the experimental period. In conclusion, T. virens exhibits significant potential as a biological alternative to chemical fungicides for controlling A. rolfsii, while enhancing cowpea growth.

Peat swamp forests are critical ecosystems that serve as significant carbon stores. However, they are increasingly threatened by deforestation and land-use changes. The peat swamp forests in Narathiwat, southern Thailand, represent the last remaining primary peat swamp ecosystems in the country, yet studies on microfungi in these habitats remain limited and mostly lack molecular data. This study focuses on identifying and characterizing fungi associated with palms, particularly Eleiodoxa conferta, in the peat swamp forests of Narathiwat, Thailand. Based on detailed morphological comparisons and multi-gene phylogenetic studies, we introduce one new genus, Narathiwatomyces, along with 25 new species, viz., Cancellidium narathiwatense, Chaetosphaeria narathiwatensis, Chloridium narathiwatense, Conioscypha narathiwatensis, Daldinia narathiwatensis, Gongromerizella palmicola, Helicoma narathiwatense, H. eleiodoxae, Javarisimilis narathiwatensis, Lentistoma narathiwatense, Longivarius narathiwatensis, Narathiwatomyces confertae, Nawawia narathiwatensis, Neohelicosporium narathiwatense, Ne. arecacearum, Neoleptodontidium narathiwatense, Pseudosaprodesmium narathiwatense, Savoryella narathiwatensis, Stanjehughesia narathiwatensis, Strossmayeria narathiwatensis, Tamhinispora narathiwatensis, Terriera narathiwatensis, Tubeufia narathiwatensis, Vamsapriya narathiwatensis and Yunnanomyces narathiwatensis. Furthermore, we report 16 new records based on host, geographical, or habitat associations. These include 11 previously described species — Ernakulamia cochinensis, Lasiodiplodia brasiliensis, La. theobromae, Linocarpon appendiculatum, Longicorpus striataspora, Megacapitula villosa, Neohelicosporium fusisporum, Nigrospora chinensis, Polyancora globosa, Pseudodactylaria longidenticulata, Trichoderma virens — as well as Conioscypha narathiwatensis, Longivarius narathiwatensis, Neoleptodontidium narathiwatense, Terriera narathiwatensis, and Vamsapriya narathiwatensis, which represent the first records of their respective genera on the host family Arecaceae. Each taxon is provided with detailed descriptions and illustrations, along with a concise summary for each family and genus. This study enhances our understanding of fungal diversity in peat swamp forests and validates the identification of species introduced in previous studies, which relied solely on morphological analysis. By incorporating molecular data, it ensures more accurate taxonomic placement

The adoption of biological control strategies plays a crucial role in ensuring the sustainability of organic agricultural practices. A field experiment was conducted in 2023 and 2024 to evaluate the impact of biological treatments using lactic acid bacteria (LAB) Lactiplantibacillus plantarum and mycoparasitic fungus (MPF) Trichoderma virens applied through seed treatment and foliar application separately and in combination on agronomic characteristics and pea yield in organic cultivation. Seed treatment with LAB and MPF resulted in a notable improvement in shoot length and root dry weight, while an increase in root nodule number was observed exclusively with LAB. The combined application of MPF as a seed treatment and LAB as a foliar application at the flowering stage significantly enhanced pod weight per plant, seed number per pod and per plant, and seed weight compared to treatments with LAB applied as either a foliar or seed treatment separately, as well as the untreated control. However, the yield responses to individual and combined treatments under field conditions demonstrated variability and inconsistency. Protein content ranged from 21.24% to 21.61%, and no significant differences observed between treatments. This is the first field report directly comparing the effectiveness of treatments on organic pea production. The findings offer promising avenues for assessing the long-term impacts of these treatments on the sustainable intensification of pea cultivation.

Recently, conserving irrigation water via application of deficit irrigation is a main priority, particularly in aridity and semi- aridity conditions. However, water deficiency is one of the major issues that hinder crop production worldwide. Also, knowledge regarding the physiological efficiency of microbial inoculation (MI) to shrink drought impacts in broccoli is unclear. We hypothesized that AMF and TRI fungi may have different mechanisms to modulate the physiological state and growth of broccoli to be more tolerant to drought stress. A field experiment in two seasons of 2023-24 and 2024-25 was conducted to assess the influence of MI on drought tolerance in broccoli. A mycorrhiza inoculum (AMF) and Trichoderma (TRI) inoculum were applied under 100, 80, 60 and 40% of irrigation requirements, IR (referred to as IR100, IR80, IR60 and IR40, respectively). Firstly, the MI was applied in the nursery after sowing the seeds in the seed trays, further, MI was applied two times, 28 and 38 days after transplanting (DAT) with the drip irrigation system, while the irrigation regimes started on 27 DAT. The treatments were designed in a strip-plot system in a complete randomized block with three replicates (trial unit size was 12 m2). At 70 DAT, soil plant analysis development (SPAD), membrane stability index (MSI), relative water content (RWC), proline content, enzymatic and non-enzymatic antioxidants and total antioxidant activity of broccoli leaves were assessed, while at 85 DAT the fresh and dry weight of shoot and root, leaf area and head yield were estimated. Findings clarified that application of AMF and TRI inoculations under different levels of drought improved significantly (p < 0.05) SPAD, MSI and RWC compared to the corresponding control treatments, except AMF with IR60 for SPAD and TRI with IR60 for SPAD and MSI and with IR40 for SPAD. Under IR60 regime, AMF achieved the maximum improvements (p < 0.05) in peroxidase, polyphenol oxidase and superoxide dismutase, significantly equaling (p > 0.05) TRI for peroxidase. AMF×IR60 was the effective interaction for achieving the maximal values of total phenolic, total flavonoids and total antioxidant activity in broccoli, equaling (p > 0.05) TRI×IR60 combination in total phenolic. Compared to the counterpart control (CK) treatments the increases in proline content due to AMF or TRI applications amounted to 26.4 and 30.0% with IR80, 25.0 and 15.6% with IR60 and 36.6 and 32.5% with IR40, respectively. It is worth to observe that IR80×AMF combination achieved shoot dry weight and head yield values similar (p > 0.05) to IR100×AMF combination. Briefly, it can be concluded that microbial inoculations, specifically AMF, can relieve the injuries of drought. Practically, broccoli growers can save irrigation water by 20% with inoculating plants by mycorrhiza to maintain crop productivity and quality under water deficiency circumstances.

Chickpea (Cicer arietinum L.) is a legume of global importance, especially in arid and semi-arid regions, due to its nutritional value and adaptability. However, root-knot nematodes (Meloidogyne spp.) pose significant threats to chickpea cultivation by impairing root function and reducing yield. This review aimed to synthesize recent scientific findings on sustainable strategies for managing Meloidogyne spp. in chickpea. A systematic literature search was conducted in databases such as PubMed, Scopus, and Web of Science, selecting peer-reviewed articles published between 1985 and 2025. The study highlights biological control agents like Trichoderma virens, Pseudomonas fluorescens, and Purpureocillium lilacinum, as well as organic amendments, botanical extracts, and crop rotation as effective tools. Advances in genetic resistance and emerging technologies such as CRISPR-Cas9 and nanotechnology are also discussed. The review concludes that integrating these approaches is essential to reduce nematode pressure, ensure food security, and promote the sustainable cultivation of chickpea.

ABSTRACT The remarkable ability of Trichoderma virens (J.H. Mill., Giddens & A.A. Foster) Arx to promote plant growth presents a promising solution for agriculture. This study evaluated the efficacy of T. virens for rice growth promotion under in-vitro and semi in-vivo conditions (pot trial and artificial filed contidions). A series of trials, including a small-scale cup trial, a pot trial, and an artificial field trial, were conducted to explore the potential benefits of T. virens. In-vivo trials were arranged as follows: T1: negative control, T2: T. virens only (positive control), T3: Department of Agriculture, Sri Lanka (DoA) fertiliser recommendation only, T4: full DoA fertiliser recommendation + T. virens, T5: ½ DoA fertiliser recommendation only, and T6: ½ DoA fertiliser recommendation + T. virens. In all three experiments, Trichoderma virens (2.5 × 106 spores/ml) were applied as the initial inoculum. Germinated seedling length, number of leaves, and number of tillers were measured weekly until the growth stages were complete. Soil multiplication of T. virens was monitored weekly through CFU counts. Plant growth parameter data were analysed using a repeated mixed-effect model, and CFU counts were analysed using a logistic regression model in SAS Studio version 9.4. The results of the in-vitro experiment revealed that the treated rice plants with T. virens had significant plant growth. The pot trial data revealed a significant effect of the T. virens formulation on increasing all plant growth parameters. The highest plant height (37.34 cm ± 0.25) was observed in the T4 treatment, while the negative control exhibited the lowest plant height (28.89 cm ± 0.17). The viable spores of T. virens multiplied throughout the experimental period in both pot and field trials. The results demonstrate that T. virens significantly enhances rice plant growth, offering a hopeful outlook for its potential application in agriculture.

Secondary Metabolites of the Endophytic Fungi Trichoderma virens Cultivated in Potato Medium and Their Bioactivity

Our previous study shows that Coptotermes formosanus (Blattodea: Rhinotermitidae) preferred to stay on filter paper treated with ethyl 2,4-dioxovalerate, a metabolite in the soil fungus Trichoderma virens. Here, we hypothesized that adding ethyl 2,4-dioxovalerate in sand could trigger aggregation and tunneling preferences of C. formosanus and improve the effectiveness of liquid termiticide. In aggregation-choice tests, significantly more termites were found on/in sand blocks containing ethyl 2,4-dioxovalerate (250 µg/g) than untreated blocks throughout the 24-h experiments. In the tunneling-choice tests, termites also excavated significantly more tunnels in the sand treated with ethyl 2,4-dioxovalerate (2.5, 25, or 250 µg/g) than untreated sand. However, in no-choice tests, ethyl 2,4-dioxovalerate (2.5, 25, or 250 µg/g) did not significantly affect tunneling activities, termite survival, wood consumption, or activities of detoxification enzymes (peroxidase, superoxide dismutase, and catalase) compared to controls. Interestingly, in aggregation- and tunneling-choice tests, termites preferred to stay and made more tunnels in sand treated with both ethyl 2,4-dioxovalerate (250 µg/g) and fipronil (1 µg/g) than untreated sand. In addition, in choice tests, sand treated with the combination of ethyl 2,4-dioxovalerate (250 µg/g) and fipronil (1 µg/g) caused significantly higher termite mortality than the sand treated with only fipronil (1 µg/g). Our study showed that ethyl 2,4-dioxovalerate may enhance the effectiveness of fipronil (1 µg/g in sand) by triggering aggregation and tunneling preferences of termites, thereby increasing the contact between termites and fipronil.

Fusarium sacchari is a pathogenic fungus that causes the pokkah boeng disease in sugarcane plants. The symptoms of pokkah boeng disease are systemic, spreading throughout all parts of the plant. The control measures implemented so far still rely on synthetic pesticides but have not yielded optimal results and are not environmentally friendly. One of the environmentally friendly control methods is the use of endophytic fungi. Endophytic fungi live within plant tissues without causing disease symptoms. This research aims to discover endophytic fungi derived from sugarcane and evaluate their ability to suppress the growth of the pathogen F. sacchari. The research began with the isolation of sugarcane tissue, followed by the morphological and molecular characterization of endophytic fungi and testing their inhibitory capacity against the pathogen F. sacchari. Morphological identification was based on colony and microscopic characteristics, while molecular identification was conducted using PCR and sequencing, with similarities based on GenBank. Inhibition tests were conducted through dual culture and volatile compound testing, as well as enzyme production. The research results obtained 38 endophytic fungal isolates from sugarcane plants that have the ability to inhibit the pathogen F. sacchari. Five of these isolates showed the highest inhibition against F. sacchari and produced cellulase, protease, and chitinase enzymes. Molecular identification results indicated that these five-sugarcane endophytic fungal isolates are Daldinia eschscholdzii, Hypoxylon pulicicidum, Trichoderma virens, Trichoderma harzianum, and Diaporthe phaseolorum. The endophytic fungus T. harzianum provides the highest inhibition rate of 83.33% in dual culture and 55.5% inhibition through the production of volatile compounds. Keywords: Endophytic fungi, inhibitory power, Fusarium sacchari, morphological characteristics, molecular characteristics, pokkah boeng disease, sugarcane plants.

Viridin and viridiol, along with wortmannin, metabolized by filamentous fungus Trichoderma virens, are identical furanosteroids with high-potent inhibitory activity towards phosphatidylinositol 3-kinase (PI3K) that associates the growth of tumor cells. Therefore, structure–activity relationship study (SAR) of these furanosteroids contributes to the development of novel drugs. However, rational supply methods have not been established yet. In this study, we generated an efficient method to produce both viridin and viridiol by using a unique pH regulated T. virens culture manner. Besides, we successfully obtained β-viridin (epimer of viridin) crystal and X-ray structure, of which the CAS number was registered without stereochemistry. Furthermore, applying the original method to stable isotope study, we also obtained [U-13C]-viridn and [U-13C]-viridol by using [U-13C6]-glucose as a carbon source replacing normal glucose which can be used to elucidate the biosynthesis pathway of furanosteroids.

To improve the precision and operability of antifungal performance testing, this study systematically examines the cultivation time, spore suspension preparation, and inoculation protocols for fungal spores commonly used in mildew and rot resistance testing. Key optimisations were achieved in inoculum time, centrifugation conditions, and spray inoculation methods, enhancing experimental reliability. The findings indicate that Aspergillus brasiliensis and Trichoderma lignorum spores form abundantly within 5–7 days, while Chaetomium globosum requires 17–20 days. Penicillium funiculosum, Aureobasidium pullulans, Paecilomyces variotii, and Trichoderma virens reach sufficient spore production in 13–15 days. Effective spore separation was achieved via centrifugation at 6000 r/min for 10 minutes. Uniform and quantitative spore inoculation was ensured using 20–50 μm spray nozzles. Quantitative methods, including specific turbidity, hemocytometer counting, and the plate count method, were evaluated, with turbidity and hemocytometer methods offering efficiency and simplicity, while plate counts provided verification. These findings contribute to establishing standardised and reproducible antifungal testing procedures, addressing operational guidelines gaps and supporting the development of antifungal technologies.

BackgroundBanana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense is a soil-borne fungal disease. Especially, tropical Race 4 (Foc TR4) can infect almost Cavendish subgroup and has a fatal threat to banana industry. Use of antagonistic microbes to manage soil-borne pathogen is viewed as a promising strategy.ResultsStrain XZ11-1 isolated from tropical rainforest has the production ability of high siderophore. By the analysis of physiological and biochemical profiles, construction of phylogenetic tree, and comparative results from the NR database, strain XZ11-1 was identified as Trichoderma virens. A relative content of 79.45% siderophores was produced in the optimized fermentation solution, including hydroxamate and carboxylate-type siderophores. Siderophores were key for inhibiting the growth of Foc TR4 by competing for environmental iron. Similarly, T. virens XZ11-1 also had antagonistic activities against 10 phytopathogenic fungi. Pot experiments demonstrated that T. virens XZ11-1 could colonize in the root system of banana plants. The symbiotic interaction not only improve plant resistance to Foc TR4, but also enhance iron absorption of roots to promote plant growth by secreting siderophores.ConclusionsT. virens XZ11-1 with the high-yield siderophores was isolated and identified. The strain could effectively inhibit the infection of Foc TR4 in banana roots and promote plant growth. It is a promising biocontrol agent for controlling fungal disease.

Identification and expression of a hydrophobic carboxylic acid reductase from Trichoderma virens.