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The use of fungi as biological nanofactories is an innovative and promising research field of nanoscience. It offers the potential to yield sustainable and cost-effective nanostructures with fine-tuned features. This study reports the trichogenic synthesis of silver oxide/copper oxide (Ag2O/CuO) nanoalloys, using the culture filtrate of the endophytic fungus Trichoderma virens. The filtrate of T. virens mediated the synthesis of the Ag2O/CuO nanoalloys, which was visually verified by the formation of dark bluish-greenish colloidal suspension upon reacting with AgNO3 and CuSO4·5H2O (1:1 v/v). X-ray diffraction revealed the crystalline nature of the trichogenic Ag2O/CuO nanoalloys, showing the characteristic diffraction peaks of cubic Ag2O (Pn-3/201) and tenorite monoclinic CuO (C2/c/15). X-ray photoelectron spectroscopy revealed spectra corresponding to the core levels of Ag 3d, Cu 2p, and O 1 s in the Ag2O/CuO nanoalloys. Fourier-transform infrared spectroscopy showed nitro and ester stretching vibrations in addition to characteristic interatomic metal–metal vibrations near 500 cm−1. Dynamic light scattering revealed that the Ag2O/CuO nanoalloys had an average particle size distribution of 124 nm, while zeta potential measurement was + 23.8 mV. Field-emission scanning electron microscopy (FESEM) revealed a mixture of hemispherical and plate-like Ag2O/CuO particles arranged in crystalline clusters. High-resolution transmission electron microscopy revealed the formation of quasi-spherical, circular, cuboid, and triangular Ag2O NPs along with the sharp-edged nanoplates of CuO particles. Energy-dispersive X-ray analysis showed prominent emission peaks of Ag (12.41%), Cu (17.68%), and O (69.91%). The antibacterial efficacy of the Ag2O/CuO nanoalloys was evaluated against six Gram-negative and three Gram-positive bacterial strains. The Ag2O/CuO nanoalloys exhibited significant antibacterial activity against streptomycin-sensitive Xanthomonas citri pv. citri (Xcc) (17.85 ± 0.735 mm), streptomycin-resistant Xcc (17.77 ± 0.720 mm), Clavibacter michiganensis subsp. michiganensis (17.44 ± 3.492 mm), and Escherichia coli (16.23 ± 2.468 mm), whereas the effectiveness of the Ag2O/CuO nanoalloys was moderate against Salmonella enterica serovar Typhimurium (12.74 ± 1.302 mm), Staphylococcus aureus (12.56 ± 0.667 mm), streptomycin-sensitive Pectobacterium carotovorum subsp. carotovorum (Pcc) (11.77 ± 0.431 mm), streptomycin-resistant Pcc (11.63 ± 0.741 mm), and C. michiganensis subsp. capsici (11.62 ± 0.958 mm). The bacterial cells exposed to the Ag2O/CuO nanoalloys exhibited notable morphological changes, such as cell membrane disruption, structural damage, compromised membrane integrity, shrinkage, and swelling. This research lays the groundwork for developing and using trichogenic Ag2O/CuO nanoalloys as highly effective nano-antimicrobial agents for a diverse array of applications. [ABSTRACT FROM AUTHOR]

Abstract Background The soil-borne fungi, Rhizoctonia solani and Sclerotium rolfsii, are major pathogens of Brassicae crops. This study was performed to clarify the relationship between the accumulation pattern of the genus Trichoderma and disease suppression in frequently inoculated soils with binucleate Rhizoctonia (BNR), Rhizoctonia solani and Sclerotium rolfsii. Results As compared to the control group, five Trichoderma virens strains isolated from soil inoculated with R. solani or BNR significantly reduced the severity of S. rolfsii (85.6–100% covering percentage) and R. solani (95.7–100% covering percentage). Similarly, five T. hamatum strains obtained from soil inoculated with R. solani were shown to be highly suppressive against S. rolfsii (83.9–97.1% covering percentages) and R. solani (60.2–96.2% covering percentages). Four out of five T. hamatum strains obtained from soil infected with S. rolfsii exhibited considerable suppression against S. rolfsii (63.7–91.2% covering percentages), while the SM5 strain did not. The phylogenetic analysis of the TEF and ITS regions of Trichoderma hamatum revealed that most isolates were classified into the same cluster with homology of 99–100%. Five strains of each T. virens and T. hamatum were isolated from the suppressive soil with high antagonistic potentials against R. solani and S. rolfsii. Suppression and antagonistic activity of T. hamatum isolated from soil frequently inoculated with sterile barley grains were negligible, whereas T. hamatum isolated from frequently inoculated soil with BNR and R. solani demonstrated considerable suppression of the pathogens and antagonistic activity. Accumulation and quantification of T. virens and T. hamatum were confirmed using loop-mediated isothermal amplification (LAMP). Conclusion In conclusion, disease suppression in frequently inoculated soil with BNR, R. solani and S. rolfsii was due to Trichoderma spp. accumulated selectively in each replicate of soil inoculation.

Background: The soil-borne fungi, Rhizoctonia solani and Sclerotium rolfsii, are major pathogens of Brassicae crops. This study was performed to clarify the relationship between the accumulation pattern of the genus Trichoderma and disease suppression in frequently inoculated soils with binucleate Rhizoctonia (BNR), Rhizoctonia solani and Sclerotium rolfsii. Results: As compared to the control group, five Trichoderma virens strains isolated from soil inoculated with R. solani or BNR significantly reduced the severity of S. rolfsii (85.6–100% covering percentage) and R. solani (95.7–100% covering percentage). Similarly, five T. hamatum strains obtained from soil inoculated with R. solani were shown to be highly suppressive against S. rolfsii (83.9–97.1% covering percentages) and R. solani (60.2–96.2% covering percentages). Four out of five T. hamatum strains obtained from soil infected with S. rolfsii exhibited considerable suppression against S. rolfsii (63.7–91.2% covering percentages), while the SM5 strain did not. The phylogenetic analysis of the TEF and ITS regions of Trichoderma hamatum revealed that most isolates were classified into the same cluster with homology of 99–100%. Five strains of each T. virens and T. hamatum were isolated from the suppressive soil with high antagonistic potentials against R. solani and S. rolfsii. Suppression and antagonistic activity of T. hamatum isolated from soil frequently inoculated with sterile barley grains were negligible, whereas T. hamatum isolated from frequently inoculated soil with BNR and R. solani demonstrated considerable suppression of the pathogens and antagonistic activity. Accumulation and quantification of T. virens and T. hamatum were confirmed using loop-mediated isothermal amplification (LAMP). Conclusion: In conclusion, disease suppression in frequently inoculated soil with BNR, R. solani and S. rolfsii was due to Trichoderma spp. accumulated selectively in each replicate of soil inoculation. [ABSTRACT FROM AUTHOR]

Trichoderma spp. widely used as biocontrol agents for controlling a wide range of plant diseases. Banana anthracnose caused by Colletotrichum musae is a prominent, widely distributed postharvest disease. This study was carried out to assess the effectiveness of locally isolated Trichoderma virens against C. musae and the mass production of T. virens using locally available, low-cost solid substrates. T. virens was tested for the inhibition of C. musae isolate in-vitro and in-vivo by dual plating and inoculating into two varieties of ripened banana: Kolikuttu and Cavendish, respectively. For mass production, T. virens was inoculated into different solid substrates including scraped coconut waste, sawdust, tea waste, seeds from rice, finger millet, and maize, dried pieces of water hyacinth plant, paddy straw, and Panicum maximum leaves; regularly taken spore counts (cfu/g) and checked for viability by plating after 12 weeks of storage. The pathogen inhibition percentage by T. virens was 74.10%. Disease severity was 0% in Kolikuttu and 19% in Cavendish after 5 days of T. virens spore application (1x107 spores/ml). Significantly higher (p≤0.05) mean spore production resulted in rice seeds (9.345x109 spores/g) compared to the other substrates and the least resulted in sawdust (1.808x109 spores/g) at the 8th week after T. virens inoculation. Spores of T. virens were viable in all the tested substrates throughout the study period. The results conclude that T. virens is capable of controlling banana anthracnose and can be efficiently mass-produced by using rice seeds, dried pieces of P. maximum leaves, and finger millet seeds as substrates. [ABSTRACT FROM AUTHOR]

Introduction: Pear Valsa canker, caused by Valsa pyri (V. pyri), poses a major threat to pear production. We aimed to assess the effectiveness of the cell-free supernatant (CFS) produced by Trichoderma virens (T. virens) to control the development of pear Valsa canker and reveal the inhibitory mechanism against the pathogenic fungi. Results: Using morphological characteristics and phylogenetic analysis, the pathogen G1H was identified as V. pyri, and the biocontrol fungus WJ561 was identified as Trichoderma virens. CFS derived from WJ561 exhibited strong inhibition of mycelial growth and was capable of reducing the pathogenicity of V. pyri on pear leaves and twigs. Scanning electron microscopy (SEM) observations revealed deformations and shrinkages in the fungal hyphae treated with CFS. The CFS also destroyed the hyphal membranes leading to the leakage of cellular contents and an increase in the malondialdehyde (MDA) content. Additionally, CFS significantly inhibited the activities of catalase (CAT) and superoxide dismutase (SOD), and downregulated the expression of antioxidant defense-related genes in V. pyri, causing the accumulation of reactive oxygen species (ROS). Artesunate, identified as the main component in CFS by liquid chromatograph-mass spectrometry (LC--MS), exhibited antifungal activity against V. pyri. Conclusion: Our findings demonstrate the promising potential of T. virens and its CFS in controlling pear Valsa canker. The primary inhibitory mechanism of CFS involves multiple processes, including membrane damage and negatively affecting enzymatic detoxification pathways, consequently leading to hyphal oxidative damage of V. pyri. This study lays a theoretical foundation for the utilization of T. virens to control V. pyri in practical production. [ABSTRACT FROM AUTHOR]

Soil-borne Trichoderma spp. have been extensively studied for their biocontrol activities against pathogens and growth promotion ability in plants. However, the beneficial effect of Trichoderma on inducing resistance against insect herbivores has been underexplored. Among diverse Trichoderma species, consistent with previous reports, we showed that root colonization by T. virens triggered induced systemic resistance (ISR) to the leaf-infecting hemibiotrophic fungal pathogens Colletotrichum graminicola. Whether T. virens induces ISR to insect pests has not been tested before. In this study, we investigated whether T. virens affects jasmonic acid (JA) biosynthesis and defense against fall armyworm (FAW) and western corn rootworm (WCR). Unexpectedly, the results showed that T. virens colonization of maize seedlings grown in autoclaved soil suppressed wound-induced production of JA, resulting in reduced resistance to FAW. Similarly, the bacterial endophyte Pseudomonas chlororaphis 30-84 was found to suppress systemic resistance to FAW due to reduced JA. Further comparative analyses of the systemic effects of these endophytes when applied in sterile or non-sterile field soil showed that both T. virens and P. chlororaphis 30-84 triggered ISR against C. graminicola in both soil conditions, but only suppressed JA production and resistance to FAW in sterile soil, while no significant impact was observed when applied in non-sterile soil. In contrast to the effect on FAW defense, T. virens colonization of maize roots suppressed WCR larvae survival and weight gain. This is the first report suggesting the potential role of T. virens as a biocontrol agent against WCR. [ABSTRACT FROM AUTHOR]

Volatile organic compounds (VOCs) emitted by fungi play a key role in locating and selecting hosts for fungivorous arthropods. However, the ecological functions of many common VOC classes, such as sesquiterpenes, remain unknown. Mutants of Trichoderma virens, defective in the emission of most sesquiterpenes owing to the deletion of the terpene cyclase vir4, were used to evaluate the role of this compound class in the food preference and fitness of the soil Collembola Folsomia candida. Choice experiments with and without direct contact with fungal mycelium revealed that Collembola were preferentially attracted to Δvir4 mutants impaired in sesquiterpene synthesis compared to wild-type T. virens. Grazing by F. candida on the sesquiterpene deficient T. virens strain had no effect on Collembola survival, reproduction and growth compared to wild-type T. virens. The results suggest that sesquiterpenes play an important role in fungal defence as repellents, but not as deterrents or toxins, against fungivorous Collembola. Our research contributes to the understanding of ecological interactions between fungi and fungivorous arthropods, providing insights into the specific ecological functions of sesquiterpenes. The study has implications for chemical ecology and the dynamics of multitrophic interactions in soil ecosystems.

Squalene is a triterpenoid compound and widely used in various industries such as medicine and cosmetics due to its strong antioxidant and anticancer properties. The purpose of this study is to increase the accumulation of squalene in filamentous fungi using exogeneous butenafine hydrochloride, which is an inhibitor for squalene epoxidase. The detailed settings achieved that the filamentous fungi, Trichoderma virens PS1-7, produced squalene up to 429.93 ± 51.60 mg/L after culturing for 7 days in the medium consisting of potato infusion with glucose at pH 4.0, in the presence of 200 μm butenafine. On the other hand, no squalene accumulation was observed without butenafine. This result indicated that squalene was biosynthesized in the filamentous fungi PS1-7, which can be used as a novel source of squalene. In addition, we successfully obtained highly 13C-enriched squalene by using [U-13C6]-glucose as a carbon source replacing normal glucose. [ABSTRACT FROM AUTHOR]

Sanghuangporus vaninii is a medicinal macrofungus that is increasingly cultivated in China. During cultivation, it was found that the fruiting body of S. vaninii was susceptible to pathogenic fungi, resulting in significant economic losses to the industry. The symptoms of the disease occur in the initial stage of fruiting body development. The isolate YZB-1 was obtained from the junction of the diseased and healthy areas of the fruiting body. In order to verify the pathogenicity of YZB-1, its purified spore suspension was inoculated into the exposed area nearby the developing fruiting body of S. vaninii. After 10 days, the same disease symptoms appeared in the inoculated area. Morphological identification and molecular analysis of rDNA ITS region confirmed that the isolate YZB-1 was identified as Trichoderma virens. The temperature stability assay revealed that the mycelia of YZB-1 grew the fastest at 25 °C, with growth slowing down gradually as the temperature increased or decreased. Dual-culture tests of T. virens and S. vaninii showed that the inhibition rate of T. virens on S. vaninii mycelium was the highest (79.01 ± 2.79%) at 25 °C, and more green spores were produced at the intersection of T. virens and S. vaninii. [ABSTRACT FROM AUTHOR]

Exploration of epiphytic microorganisms from different plant parts and their exploitation for the management of plant pathogens is a relevant approach in view of greater awareness of pollution free environment. The aim of the present study was to analyze the microbial communities with special focus on antagonists isolated from the fructosphere of cowpea (Vigna unguiculata L. Walp.) and the use of organic preparations such as panchagavya, jeevamruth, compost tea, vermiwash and fish amino acid for suppression of Choanephora cucurbitarum, the pathogen inciting pod rot in cowpea. A collection of six isolates of bacteria and fungi were isolated through serial dilution technique, and their efficacy in suppressing the pathogen were tested under in vitro conditions. Among the six isolates, the bacteria and fungi with maximum inhibitory activity against the targeted pathogen were selected for further identification and in vivo assay. Based on the cultural, morphological and biochemical characters, the bacterial and fungal antagonists were identified as Pseudomonas fluorescens and Trichoderma virens, respectively. In vitro assay of the organic preparations revealed that vermiwash (5% and 10%), jeevamurth (10%) and panchagavya (10%) completely inhibited the growth of pathogen. Application of effective dose of organic preparations and the selected antagonists on the excised cowpea pods revealed that, among organic preparations jeevamurth (10%) exhibited maximum suppression of pod rot by 60.64%, however the selected bacterial antagonist, i.e., P. fluorescens gave complete suppression of the pathogen. Under in vivo conditions, jeevamurth (10%), vermiwash (5%), T. virens (106cfu/ml), and P. fluorescens (106cfu/ml) showed the maximum suppression of the pathogen and the percentage suppression was recorded as 87.33, 75.22, 75.27 and 72.31% respectively. Therefore, the present study revealed that the organic preparations such as jeevamurth (10%), vermiwash (5%), and the indigenous species of Pseudomonas fluorescens and Trichoderma virens obtained from the fructosphere can be used in integrated disease management strategies against Choanephora pod rot of cowpea. [ABSTRACT FROM AUTHOR]

Soil-borne Trichoderma spp. have been extensively studied for their biocontrol activities against pathogens and growth promotion ability in plants. However, the beneficial effect of Trichoderma on inducing resistance against insect herbivores has been underexplored. Among diverse Trichoderma species, consistent with previous reports, we showed that root colonization by T. virens triggered induced systemic resistance (ISR) to the leaf-infecting hemibiotrophic fungal pathogens Colletotrichum graminicola. Whether T. virens induces ISR to insect pests has not been tested before. In this study, we investigated whether T. virens affects jasmonic acid (JA) biosynthesis and defense against fall armyworm (FAW) and western corn rootworm (WCR). Unexpectedly, the results showed that T. virens colonization of maize seedlings grown in autoclaved soil suppressed wound-induced production of JA, resulting in reduced resistance to FAW. Similarly, the bacterial endophyte Pseudomonas chlororaphis 30-84 was found to suppress systemic resistance to FAW due to reduced JA. Further comparative analyses of the systemic effects of these endophytes when applied in sterile or non-sterile field soil showed that both T. virens and P. chlororaphis 30-84 triggered ISR against C. graminicola in both soil conditions, but only suppressed JA production and resistance to FAW in sterile soil, while no significant impact was observed when applied in non-sterile soil. In contrast to the effect on FAW defense, T. virens colonization of maize roots suppressed WCR larvae survival and weight gain. This is the first report suggesting the potential role of T. virens as a biocontrol agent against WCR.

A new carotane sesquiterpene, trichocarotin N (1), was isolated from a culture of the marine-derived fungus Trichoderma virens QD-11. The structure of this compound was established by detailed analysis of 1D/2D NMR and HRESIMS data. Trichocarotin N (1) exhibited moderate cytotoxicity against HeLa and MCF-7 cancer cell lines, with IC50 values of 32.4 and 41.6 μM, respectively. A new carotane sesquiterpene, trichocarotin N (1), was isolated from a culture of the marine-derived fungus Trichoderma virens QD-11. The structure of this compound was established by detailed analysis of 1D/2D NMR and HRESIMS data. Trichocarotin N (1) exhibited moderate cytotoxicity against HeLa and MCF-7 cancer cell lines, with IC50 values of 32.4 and 41.6 μM, respectively. [ABSTRACT FROM AUTHOR]

Abstract Background N-acetylglucosaminidase (NAGase) could liberate N-acetylglucosamine (GlcNAc) from GlcNAc-containing oligosaccharides. Trichoderma spp. is an important source of chitinase, particularly NAGase for industrial use. nag1 and nag2 genes encoding NAGase, are found in the genome in Trichoderma spp. The deduced Nag1 and Nag2 shares ~ 55% homology in Trichoderma virens. Most studies were focus on Nag1 and nag1 previously. Results The native NAGase (TvmNAG2) was purified to homogeneity with molecular mass of ~ 68 kDa on SDS-PAGE analysis, and identified as Nag2 by MALDI/MS analysis from an isolate T. virens strain mango. RT-PCR analyses revealed that only nag2 gene was expressed in liquid culture of T. virens, while both of nag1 and nag2 were expressed in T. virens cultured on the plates. TvmNAG2 was thermally stable up to 60 °C for 2 h, and the optimal pH and temperature were 5.0 and 60–65 °C, respectively, using p-nitrophenyl-N-acetyl-β-D-glucosaminide (pNP-NAG) as substrate. The hydrolytic product of colloidal chitin by TvmNAG2 was suggested to be GlcNAc based on TLC analyses. Moreover, TvmNAG2 possesses antifungal activity, inhibiting the mycelium growth of Sclerotium rolfsii. And it was resistant to the proteolysis by papain and trypsin. Conclusions The native Nag2, TvmNAG2 was purified and identified from T. virens strain mango, as well as enzymatic properties. To our knowledge, it is the first report with the properties of native Trichoderma Nag2.

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A new strain of Trichoderma virens was isolated from mushroom compost which causes green mold disease from the mushroom production unit of Lovely Professional University, Punjab, India. Based on 5.8S rRNA, partial 18S, and 28S rRNA genes, ITS1, ITS4 sequence analysis, and morphological characteristics, the fungal isolate was identified as T. virens. The universal primers (ITS-1 and ITS-4) were used to amplify the 28S rRNA gene fragment that produced a sharp band of about 600 bp on the agarose gel. The amplified gene fragment was then sequenced and showed 100% similarity with T. virens. GC/MS analysis of the strain revealed the presence of several antimicrobial compounds, such as n-nonadecanol-, propanoic acid, 2-methyl-butanoic acid, and 2-methyl-dodecane. The current study confirmed the potent antimicrobial and chitinolytic activity against Fusarium and its potential as a bio-control agent for managing plant diseases. [ABSTRACT FROM AUTHOR]

Trichoderma is internationally recognized as a biocontrol fungus for its broad-spectrum antimicrobial activity. Intriguingly, the crosstalk mechanism between the plant and Trichoderma is dynamic, depending on the Trichoderma strains and the plant species. In our previous study, the Trichoderma virens 192-45 strain showed better pathogen inhibition through the secretive non-volatile and volatile substrates. Therefore, we studied transcriptional and metabolic responses altered in creeping bentgrass (Agrostis stolonifera L.) with T. virens colonization prior to a challenge with Clarireedia homoeocarpa. This fungal pathogen causes dollar spot on various turfgrasses. When the pathogen is deficient, the importance of T. virens to the enhancement of plant growth can be seen in hormonal production and microbe signaling, such as indole-3-acrylic acid. Therefore, these substrates secreted by T. virens and induced genes related to plant growth can be the 'pre-defense' for ensuing pathogen attacks. During C. homoeocarpa infection, the Trichoderma–plant interaction activates defense responses through the SA- and/or JA-dependent pathway, induced by T. virens and its respective exudates, such as oleic, citric, and stearic acid. Thus, we will anticipate a combination of genetic engineering and exogenous application targeting these genes and metabolites, which could make creeping bentgrass more resistant to dollar spot and other pathogens. [ABSTRACT FROM AUTHOR]

Aquaponics has the potential to produce sustainable and accessible quality food through the integration of hydroponics and aquaculture. Plants take up dissolved nutrients in fish wastewater, allowing water reuse for fish. However, the simultaneous presence of fish and plants in the same water loop has made phytosanitary treatments of diseases such as powdery mildew problematic due to risks of toxicity for fish and beneficial bacteria, limiting its commercialization. Entomopathogenic and mycoparasitic fungi have been identified as safe biological control agents for a broad range of pests. This study aimed to investigate the efficacy of entomopathogenic fungi, Lecanicillium attenuatum (LLA), Isaria fumosorosea (IFR), and mycoparasitic fungus Trichoderma virens (TVI) against Podosphaera xanthii. Also, we investigated the possible harmful effects of the three fungal biocontrol agents in aquaponics by inoculating them in aquaponics water and monitoring their survival and growth. The findings showed that the three biocontrol agents significantly suppressed the powdery mildew at 107 CFU/ml concentration. Under greenhouse conditions (65-73% relative humidity (RH)), a significant disease reduction percentage of 85% was recorded in L. attenuatum-pretreated leaves. IFR-treated leaves had the least AUDPC (area under disease progress curve) of ~434.2 and disease severity of 32% under 65-73% RH. In addition, L. attenuatum spores were the most persistent on the leaves, the spores population increased to 9.54 × 103 CFUmm-2 from the initial 7.3 CFUmm-2 under 65-73%. In contrast, in hydroponics water, the LLA, IFR, and TVI spores significantly reduced by more than 99% after 96 hrs. Initial spore concentrations of LLA of 107 CFU/ml spores were reduced to 4 x 103 CFU after 96 hrs. Though the results from this study were intended for aquaponics systems, relevance of the results to other cultivation systems are discussed.

Simple Summary: Rice, together with wheat and corn, is among the most important food crops for mankind. Half of the world's population consumes rice, mainly in Asia, southern Europe, tropical America, and parts of Africa. Rice sheath blight, caused by Rhizoctonia solani, is one of the main diseases in rice production. The control of this pathogen is difficult due to its ecological behavior, broad host range, and the high survival rate of sclerotia under various environmental conditions. In this research, after morphological and molecular identification of fungal isolates, five superior isolates, including Trichoderma virens, Trichoderma harzianum, Curvularia lunata, Aspergillus fumigatus, and Aspergillus awamori were studied in the in vitro and greenhouse trials, towards their potential to inhibit R. solani. The statistical analysis showed a significant difference between the effectiveness of fungi used in the volatile metabolites assay and in terms of height and fresh weight of plants in the greenhouse. It can be concluded that T. virens and A. fumigatus are the most effective antagonists in rice sheath blight disease control. The rice sheath blight disease, caused by Rhizoctonia solani J.G. Kühn fungus, is a major disease of Oryza sativa L. occurring all over the world. Therefore, efforts need to be undertaken to limit the spread of this pathogen, preferably by using environmentally friendly methods. In the present study, 57 fungal isolates were recovered by surface sterilization technique from 120 rice samples collected from paddy fields in Guilan province, Iran. Biological characterizations of the isolated taxa were performed in vitro, in the dual culture, volatile metabolites, and slide culture methods. Among the studied isolates, Trichoderma virens (J. H. Miller, Giddens and A. A. Foster) Arx was most effective in inhibiting the mycelial growth of R. solani in the dual culture (44.16% inhibition level), while Aspergillus fumigatus Fresen and T. virens had a 62.50–68.75% inhibition efficiency by volatile metabolites. In the slide culture, all of the isolates, except for T. harzianum Rifai, were effective in inhibiting the hyphae growth of R. solani. Under greenhouse conditions, rice plants inoculated with these potential antagonistic fungi showed a reduction in disease severity by even 41.4% as in the case of T. virens. Moreover, phenotypic properties of rice, such as plant height, fresh weight, and dry weight were increased in the plants inoculated with all antagonistic fungi tested, compared to the infected plants, except for the fresh weight of plants inoculated with Curnularia lunata (Wakker) Boedijn. The present in vivo and in vitro studies revealed that T. virens and A. fumigatus are the most effective antagonists in rice sheath blight disease control and could be applied in agricultural practice. [ABSTRACT FROM AUTHOR]

Peanut stem white rot caused by Sclerotium rolfsii Sacc. is a soil-borne disease that is widely prevailing across peanut farms, leading to serious economic losses. Screening for biocontrol agents against this pathogen is urgent. In this research, 166 fungal isolates including 136 isolates of S. rolfsii and 30 isolates of antagonistic endophytic fungi were obtained from a total of 220 samples collected from peanut farms in Guilan province, Iran. After morphological and molecular identification, six superior endophytic isolates were finally selected for the in vitro and greenhouse trials, including four isolates from Trichoderma viride, Trichoderma virens, Penicillium decaturense, and Aspergillus flavus and two isolates from Penicillium rubens. Four methods of biocontrol were used during the in vitro phase, i.e., dual culture, volatile metabolites assay, non-volatile metabolites assay (culture extract), and slide culture. It was found that T. virens had the highest capability of suppressing the mycelial growth of S. rolfsii in the dual culture method (90.98%). As for the volatile metabolites assay, the most effective isolates in inhibiting the pathogen's mycelial growth were P. rubens (MN395854.1) and A. flavus (84.30% and 73.50% inhibition, respectively). In the non-volatile metabolites method, the isolates that performed the best in suppressing the mycelial growth of S. rolfsii were T. viride and P. rubens (MN395854.1) with 91.80% and 90.20% inhibitory effects, respectively. On the other hand, in the slide culture method, all isolates, except for T. virens and T. viride, successfully controlled the development of S. rolfsii hyphae. The greenhouse trials also supported the effectiveness of endophytic fungi in controlling the pathogen on the host plants. According to the results, T. viride, A. flavus, and P. rubens (MN395854.1) were 44%, 42%, and 38% effective in alleviating the disease incidence and severity. Moreover, the application of these antagonistic fungi in the greenhouse conditions increased the height, fresh weight, and dry weight of the Arachis hypogaea plants infected with the disease causal agent compared to the plants treated only with the pathogen. The results of the in vitro and greenhouse experiments revealed that the endophytic fungi occurring in the natural microbiota of peanut are capable of bio-controlling S. rolfsii, the causal agent of peanut stem white rot disease. These findings shed new insights into the possible resistance induction in A. hypogaea plants through biological protection. [ABSTRACT FROM AUTHOR]

Fusarium oxysporum f. sp. cubense (Foc) tropical race 4 (TR4) is threatening banana production because of its increasing spread. Biological control approaches have been widely studied and constitute interesting complementary measures to integrated disease management strategies. They have been based mainly on the use of single biological control agents (BCAs). In this study, we moved a step forward by designing a synthetic microbial community (SynCom) for the control of Fusarium wilt of banana (FWB). Ninety-six isolates of Pseudomonas spp., Bacillus spp., Streptomyces spp., and Trichoderma spp. were obtained from the banana rhizosphere and selected in vitro for the antagonism against Foc TR4. In pot experiments, a large community such as SynCom 1.0 (44 isolates with moderate to high antagonistic activity) or a small one such as SynCom 1.1 (seven highly effective isolates) provided similar disease control (35% symptom severity reduction). An in vitro study of the interactions among SynCom 1.1 isolates and between them and Foc revealed that beneficial microorganisms not only antagonized the pathogen but also some of the SynCom constituents. Furthermore, Foc defended itself by antagonizing the beneficial microbes. We also demonstrated that fusaric acid, known as one of the secondary metabolites of Fusarium species, might be involved in such an interaction. With this knowledge, SynCom 1.2 was then designed with three isolates: Pseudomonas chlororaphis subsp. piscium PS5, Bacillus velezensis BN8.2, and Trichoderma virens T2C1.4. A non-simultaneous soil application of these isolates (to diminish cross-inhibition) delayed FWB progress over time, with significant reductions in incidence and severity. SynCom 1.2 also performed better than two commercial BCAs, BioPak R and T-Gro. Eventually, SynCom 1.2 isolates were characterized for several biocontrol traits and their genome was sequenced. Our data showed that assembling a SynCom for biocontrol is not an easy task. The mere mixtures of antagonists (e.g., SynCom 1.0 and 1.1) might provide effective biocontrol, but an accurate investigation of the interactions among beneficial microorganisms is needed to improve the results (e.g., SynCom 1.2). SynCom 1.2 is a valuable tool to be further developed for the biological control of FWB. [ABSTRACT FROM AUTHOR]

Background: Rhipicephalus microplus and Amblyomma mixtum are the most important tick species affecting livestock in tropical areas. They can have a direct harmful effect on cattle and humans due to transmission of pathogens of tick-borne diseases (TBD). Entomopathogenic fungi have an important role in crop pest control. However, data concerning the efficacy of use of entomopathogenic fungicombinations are scarce. Objective: For this reason, in this study, the efficacy of commercial fungal combinations was assessed: Trishok® (Trichoderma harzianaum and Trichoderma virens) and Esporomax® (Beauveria bassiana, Metarhizium anisopliae, Paecelomyces fumosoroseus). Methodology: Adult immersion tests were performed with six different concentrations of fungal solutions. Results: In this study, Trishok® and Esporomax® showed potential efficacy against R. microplus. Different concentrations of fungal application tested on A. mixtum did not show any effect. The observed mortality of R. microplus was 85. 0 ± 2.0 and 65.0 ± 2.0 for Trishok® and Esporomax®, respectively. The estimated LC50 for R. microplus immersed in solutions of Trishok® and Esporomax® thereof were: 6.5x104 and3.9x105 conidia mL-1, respectively. Implications: These results should be considered in the design and implementation of alternatives based on the biological control of ticks. Conclusion: We can conclude that combined commercial strains of entomopathogenic fungi have in vitro acaricidal activity against R. microplus.

Abstract The objective of this work was to evaluate the chitinolytic activity and bioactivity of fungi isolated from the palm weevil (Rhynchophorus palmarum) and from soil samples from coconut (Cocos nucifera) crops against the insect itself. Initially, to determine the chitinolytic properties of the isolated fungi, their ability to hydrolyze chitin in a liquid culture medium was evaluated. Then, preliminary pathogenicity assays were carried out, using the bean weevil (Acanthoscelides obtectus) as a reference, to select the fungal isolates to be used in the experiments with the palm weevil. Fi n a l ly, the bioactivity of two selected entomopathogenic fungi on palm weevil larvae and adults was assessed. There was no direct correlation between chitinolytic activity and pathogenicity capacity on the bean weevil nor between the isolates and bioactivity on the palm weevil. Beauveria bassiana CSU9 shows the highest activity on palm weevil larvae and adults, with a median lethal time of 0.8 and 14.4 days, respectively.

Downy mildew (DM), caused by P. cubensis, is harmful to cucurbits including luffa, with increased shortcomings associated with its control through cultural practices, chemical fungicides, and resistant cultivars; there is a prompt need for an effective, eco-friendly, economical, and safe biocontrol approach. Current research is therefore dealt with the biocontrol of luffa DM1 through the endophytic fungi (EF) consortium. Results revealed that T. harzianum (ThM9) and T. virens (TvA1) showed pathogen-dependent inducible metabolic production of squalene and gliotoxins by higher gene expression induction of SQS1/ERG9 (squalene synthase) and GliP (non-ribosomal peptide synthetase). Gene expression of lytic enzymes of EF was also induced with subsequently higher enzyme activities upon confrontation with P. cubensis. EF-inoculated luffa seeds showed efficient germination with enhanced growth potential and vigor of seedlings. EF-inoculated plants showed an increased level of growth-promoting hormone GA with higher gene expression of GA2OX8. EF-pre-inoculated seedlings were resistant to DM and showed an increased GSH content and antioxidant enzyme activities (SOD, CAT, POD). The level of MDA, H2O2, REL, and disease severity was reduced by EF. ACC, JA, ABA, and SA were overproduced along with higher gene expression of LOX, ERF, NCED2, and PAL. Expression of defense-marker genes (PPO, CAT2, SOD, APX, PER5, LOX, NBS-LRR, PSY, CAS, Ubi, MLP43) was also modulated in EF-inoculated infected plants. Current research supported the use of EF inoculation to effectively escalate the systemic immunity against DM corresponding to the significant promotion of induced systemic resistance (ISR) and systemic acquired resistance (SAR) responses through initiating the defense mechanism by SA, ABA, ET, and JA biosynthesis and signaling pathways in luffa. [ABSTRACT FROM AUTHOR]

Background: N-acetylglucosaminidase (NAGase) could liberate N-acetylglucosamine (GlcNAc) from GlcNAc-containing oligosaccharides. Trichoderma spp. is an important source of chitinase, particularly NAGase for industrial use. nag1 and nag2 genes encoding NAGase, are found in the genome in Trichoderma spp. The deduced Nag1 and Nag2 shares ~ 55% homology in Trichoderma virens. Most studies were focus on Nag1 and nag1 previously. Results: The native NAGase (TvmNAG2) was purified to homogeneity with molecular mass of ~ 68 kDa on SDS-PAGE analysis, and identified as Nag2 by MALDI/MS analysis from an isolate T. virens strain mango. RT-PCR analyses revealed that only nag2 gene was expressed in liquid culture of T. virens, while both of nag1 and nag2 were expressed in T. virens cultured on the plates. TvmNAG2 was thermally stable up to 60 °C for 2 h, and the optimal pH and temperature were 5.0 and 60–65 °C, respectively, using p-nitrophenyl-N-acetyl-β-D-glucosaminide (pNP-NAG) as substrate. The hydrolytic product of colloidal chitin by TvmNAG2 was suggested to be GlcNAc based on TLC analyses. Moreover, TvmNAG2 possesses antifungal activity, inhibiting the mycelium growth of Sclerotium rolfsii. And it was resistant to the proteolysis by papain and trypsin. Conclusions: The native Nag2, TvmNAG2 was purified and identified from T. virens strain mango, as well as enzymatic properties. To our knowledge, it is the first report with the properties of native Trichoderma Nag2. [ABSTRACT FROM AUTHOR]

Trichoderma is a highly beneficial species for use in agriculture due to its ability to combat soilborne disease and increase growth in plants. This study investigated the effects of the biological control agent Trichoderma spp. on the incidence of charcoal rot and growth in strawberry plants. Sixteen Trichoderma harzianum (Thr) and three Trichoderma virens (Tvr) isolates were isolated from strawberry fields in different locations in Aydın province. The Trichoderma isolates were identified with molecular methods based on the rDNA internal transcribed spacer region sequences (ITS1/ITS4), and phylogenetic analysis. The Trichoderma spp. isolates growth was evaluated at temperatures ranging from 15 to 45 °C on Potato Dextrose Agar (PDA). The optimum growth was observed at 30 °C, and no isolates grew at 40 °C and 45 °C. Tvr4 demonstrated a high performance in inhibiting the mycelial growth of Macrophomina phaseolina by 66.4%, in dual culture assay. Volatile organic compounds antifungal activity Thr15 and Tvr4 suppressed the microsclerotia produced in M. phaseolina by 80.55% and 73%, respectively. A pot trial was carried out in climatic chambers with the following treatments: i) strawberry plant inoculated with Trichoderma spp. (Tr); ii) plant inoculated with Trichoderma spp. and M. phaseolina (Tr + Mp); iii) applying M. phaseolina 15 days after the treatment of Trichoderma (Tr + Mp15). Thr15 gave the maximum (281.95%) fresh weight increase compared to the other tested isolates. Plant mortality in Tr + Mp was significantly reduced with Thr15, Thr14, Thr13, and Tvr2. In the Tr + Mp15 treatment, Thr20 had the greatest reduction in plant mortality and the greatest increase in plant weight. [ABSTRACT FROM AUTHOR]

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.

Palu Valley shallot production is still very low namely 5.31 tonnes/ha when compared to national production which reaches 9.7 tonnes/ha, this is due to the attack of the base stem rot disease caused by Fusarium oxsyporum f.sp. cepae. This study aims to determine the effect of various types of organic fertilizer decomposed by Trichoderma virens on the incidence of stem rot disease and shallot yields. The research was conducted at the Laboratory of Pests and Plant Diseases, Faculty of Agriculture, Tadulako University, and shallot planting in Oloboju Village, Sigi Biromaru District, Sigi Regency, Central Sulawesi Province and lasted from March 2018 to August 2018. This research used a Randomized Block Design (RBD) with five treatments consisting of, without organic fertilizer (B0), cow manure 10 tons/ha + Trichoderma virens 100g/L (B1), chicken manure 10 tons/ha + T. virens 100g/L (B2), goat manure 10 tons/ha + T. virens 100g/L (B3), and petrogenic 5 tons/ha+ T. virens 100g/L (B4). The results showed that the use of 10 tons/ha cow manure decomposed by T. virens (B1) can reduce the intensity of stem rot disease on the Palu Valley shallot which is 5.61% to be 1.88% (3rd week) and 2.89% to be 0.98% (7th week), and increase shallot yield from 4.09 tons/ha to be 7.48 tons/ ha.

The interaction of biostimulant species of Trichoderma may have benefits to increase the production of metabolites involved in promoting plant growth; combining two organisms with a higher degree of antagonism could enhance their plant growth promoter effects. The objective of this study was to select in vitro native species of Trichoderma in a dual cultivation, and to evaluate their growth-promoting effect on Capsicum chinense. The selection of isolates in vitro was done by dual confrontations with the following characteristics: proportional growth of both isolates in the Petri dish; presence of mycelial barrier; and colour of the culture medium. The selected confrontations and their individual isolates were evaluated in plants of C. chinense cv. Mayapan. Also, two controls without fungal inoculation were assessed, one as control with 100% of the chemical fertilization (250N-200P-300K) recommended for the culture; and the second as control at 50%, with half of the indicated fertilization. Three applications of suspensions of 106 conidia mL-1 were made in the seedbed stage and another three during the growth of the culture. The effect of the treatments was evaluated through plant growth and crop production 145 days after sowing. The interaction of Trichoderma virens (Th05-02)/Trichoderma koningiopsis (Th41-11) was equal (p>0.05) to the 100% fertilized control in the variables evaluated. In addition, increases of 14.53% were recorded in plant height, 11.20% in stem diameter, 28.54% in foliar dry biomass, 12.97% in dry root biomass, 33.13% in a number of fruits, and 25.77% in the weight of fruits, compared to the control with 50% chemical fertilization. The species interaction of T. virens (Th05-02)/T. koningiopsis (Th41-11) exerted a plant growth promoting effect with 50% reduction in the recommended chemical fertilization for habanero pepper cultivation. [ABSTRACT FROM AUTHOR]

(1) Background: Ralstonia solanacearum causes tomato bacterial wilt disease, one of the most serious tomato diseases. As the combination of Trichoderma virens (Tvien6) and Bacillus velezensis (X5) was more effective at controlling tomato bacterial wilt disease than a single agent, we investigated the synergistic effect of Tvien6 and X5 in controlling this disease; (2) Methods: The disease incidence, plant heights and weights, relative chlorophyll content (SPAD values), defensive enzymes (PPO, POD, and SOD) activities, and metabolome were estimated among four treatment groups (BR treatment, X5 + R. solanacearum (RS-15); TR treatment, Tvien6+ RS-15; TBR treatment, Tvien6 + X5 + RS-15; and R treatment, RS-15); (3) Results: The R treatment group had the highest disease incidence and lowest plant heights, plant weights, SPAD values, defensive enzyme activities, and D-fructose and D-glucose contents; the TBR treatment group had the lowest disease incidence and highest plant heights, plant weights, SPAD values, defensive enzyme activities, and D-fructose and D-glucose contents; (4) Conclusions: The results revealed that Tvien6 and X5 can both individually promote tomato plant growth, increase leaf chlorophyll content, enhance defensive enzyme activities, and induce the accumulation of D-fructose and D-glucose; however, they were more effective when combined. [ABSTRACT FROM AUTHOR]

Trichoderma is internationally recognized as a biocontrol fungus for its broad-spectrum antimicrobial activity. Intriguingly, the crosstalk mechanism between the plant and Trichoderma is dynamic, depending on the Trichoderma strains and the plant species. In our previous study, the Trichoderma virens 192-45 strain showed better pathogen inhibition through the secretive non-volatile and volatile substrates. Therefore, we studied transcriptional and metabolic responses altered in creeping bentgrass (Agrostis stolonifera L.) with T. virens colonization prior to a challenge with Clarireedia homoeocarpa. This fungal pathogen causes dollar spot on various turfgrasses. When the pathogen is deficient, the importance of T. virens to the enhancement of plant growth can be seen in hormonal production and microbe signaling, such as indole-3-acrylic acid. Therefore, these substrates secreted by T. virens and induced genes related to plant growth can be the ‘pre-defense’ for ensuing pathogen attacks. During C. homoeocarpa infection, the Trichoderma–plant interaction activates defense responses through the SA- and/or JA-dependent pathway, induced by T. virens and its respective exudates, such as oleic, citric, and stearic acid. Thus, we will anticipate a combination of genetic engineering and exogenous application targeting these genes and metabolites, which could make creeping bentgrass more resistant to dollar spot and other pathogens.

Fusarium oxysporum Schlecht. (Hypocreales: Nectriaceae) is one of the most devastating plant pathogens worldwide, causing vascular wilt in several crops. Management of this disease primarily relies on chemical fungicides and resistant cultivars in high value crops. However, due to the limited efficacy of these methods, alternative control methods are needed. Biological control is a sustainable, safe, and effective alternative, but the use of a single biological control agent (BCA) usually has inconsistent results. The consistency of biocontrol could be enhanced using microbial consortia. In this context, the aim of this work was to select an effective microbial consortium against vascular wilt in cape gooseberry (Physalis peruviana L.) caused by Fusarium oxysporum f. sp. physali, from a mixture of four strains of Trichoderma spp. Pers. (Hypocreales: Hypocraceae) and Bacillus velezensis (Bacillales: Bacillaceae) Bs006. The calculated synergy factor was used as a selection criterion. Then the selected consortium was evaluated in the field and compared to carbendazim. The Trichoderma virens Gl006 and B. velezensis Bs006 consortium showed synergistic activity against vascular wilt under greenhouse and field conditions and efficacy similar to chemical control. These results suggest that Gl006 and Bs006 have a higher potential in controlling Fusarium wilt in cape gooseberry when applied as a consortium compared to separate, single strains. [ABSTRACT FROM AUTHOR]

Gliotoxin is a potent antibiotic showing antifungal activity against various phytopathogenic fungi. It is produced by Q strains of Trichoderma virens and gliotoxin non-producing strains of T. virens are designated as P strains. There is no detailed study on effect of gliotoxin on suppression of damping off of tomato caused by Pythium aphanidermatum and its phytotoxicity effect on tomato plants. Thus, the present study was carried out to assess the effect of gliotoxin on inhibition of mycelial growth of P. aphanidermatum, its phytotoxicity effect on tomato and its role on the suppression of damping off of tomato. Culture filtrates of Q strains of T. virens containing gliotoxin highly inhibited the mycelial growth of P. aphanidermatum compared to that of P strains of T. virens. Purified gliotoxin but not bis-thiomethyl gliotoxin effectively inhibited the mycelial growth of P. aphanidermatum. Tomato seeds treated with purified gliotoxin did not inhibit the germination of seeds, its root and shoot length even at higher concentration that is at 1000 ppm (fivefold inhibitory concentration against P. aphanidermatum). Foliar spray of gliotoxin on tomato plants did not show any phytotoxic effect at lower concentration but showed scorching effect at higher concentration. Seed treatment with gliotoxin producing Q strains of T. virens showed greater suppression of damping-off tomato compared to P strains of T. virens. This study clearly showed that gliotoxin producing T. virens could be used in suppression of damping-off disease incidence in tomato. [ABSTRACT FROM AUTHOR]

Abstract Background Trichoderma spp. are majorly composed of plant-beneficial symbionts widely used in agriculture as bio-control agents. Studying the mechanisms behind Trichoderma-derived plant benefits has yielded tangible bio-industrial products. To better take advantage of this fungal-plant symbiosis it is necessary to obtain detailed knowledge of which genes Trichoderma utilizes during interaction with its plant host. In this study, we explored the transcriptional activity undergone by T. virens during two phases of symbiosis with maize; recognition of roots and after ingress into the root cortex. Results We present a model of T. virens – maize interaction wherein T. virens experiences global repression of transcription upon recognition of maize roots and then induces expression of a broad spectrum of genes during colonization of maize roots. The genes expressed indicate that, during colonization of maize roots, T. virens modulates biosynthesis of phytohormone-like compounds, secretes a plant-environment specific array of cell wall degrading enzymes and secondary metabolites, remodels both actin-based and cell membrane structures, and shifts metabolic activity. We also highlight transcription factors and signal transduction genes important in future research seeking to unravel the molecular mechanisms of T. virens activity in maize roots. Conclusions T. virens displays distinctly different transcriptional profiles between recognizing the presence of maize roots and active colonization of these roots. A though understanding of these processes will allow development of T. virens as a bio-control agent. Further, the publication of these datasets will target future research endeavors specifically to genes of interest when considering T. virens – maize symbiosis.

Peanut stem white rot caused by Sclerotium rolfsii Sacc. is a soil-borne disease that is widely prevailing across peanut farms, leading to serious economic losses. Screening for biocontrol agents against this pathogen is urgent. In this research, 166 fungal isolates including 136 isolates of S. rolfsii and 30 isolates of antagonistic endophytic fungi were obtained from a total of 220 samples collected from peanut farms in Guilan province, Iran. After morphological and molecular identification, six superior endophytic isolates were finally selected for the in vitro and greenhouse trials, including four isolates from Trichoderma viride, Trichoderma virens, Penicillium decaturense, and Aspergillus flavus and two isolates from Penicillium rubens. Four methods of biocontrol were used during the in vitro phase, i.e., dual culture, volatile metabolites assay, non-volatile metabolites assay (culture extract), and slide culture. It was found that T. virens had the highest capability of suppressing the mycelial growth of S. rolfsii in the dual culture method (90.98%). As for the volatile metabolites assay, the most effective isolates in inhibiting the pathogen’s mycelial growth were P. rubens (MN395854.1) and A. flavus (84.30% and 73.50% inhibition, respectively). In the non-volatile metabolites method, the isolates that performed the best in suppressing the mycelial growth of S. rolfsii were T. viride and P. rubens (MN395854.1) with 91.80% and 90.20% inhibitory effects, respectively. On the other hand, in the slide culture method, all isolates, except for T. virens and T. viride, successfully controlled the development of S. rolfsii hyphae. The greenhouse trials also supported the effectiveness of endophytic fungi in controlling the pathogen on the host plants. According to the results, T. viride, A. flavus, and P. rubens (MN395854.1) were 44%, 42%, and 38% effective in alleviating the disease incidence and severity. Moreover, the application of these antagonistic fungi in the greenhouse conditions increased the height, fresh weight, and dry weight of the Arachis hypogaea plants infected with the disease causal agent compared to the plants treated only with the pathogen. The results of the in vitro and greenhouse experiments revealed that the endophytic fungi occurring in the natural microbiota of peanut are capable of bio-controlling S. rolfsii, the causal agent of peanut stem white rot disease. These findings shed new insights into the possible resistance induction in A. hypogaea plants through biological protection.

The rice sheath blight disease, caused by Rhizoctonia solani J.G. Kühn fungus, is a major disease of Oryza sativa L. occurring all over the world. Therefore, efforts need to be undertaken to limit the spread of this pathogen, preferably by using environmentally friendly methods. In the present study, 57 fungal isolates were recovered by surface sterilization technique from 120 rice samples collected from paddy fields in Guilan province, Iran. Biological characterizations of the isolated taxa were performed in vitro, in the dual culture, volatile metabolites, and slide culture methods. Among the studied isolates, Trichoderma virens (J. H. Miller, Giddens and A. A. Foster) Arx was most effective in inhibiting the mycelial growth of R. solani in the dual culture (44.16% inhibition level), while Aspergillus fumigatus Fresen and T. virens had a 62.50–68.75% inhibition efficiency by volatile metabolites. In the slide culture, all of the isolates, except for T. harzianum Rifai, were effective in inhibiting the hyphae growth of R. solani. Under greenhouse conditions, rice plants inoculated with these potential antagonistic fungi showed a reduction in disease severity by even 41.4% as in the case of T. virens. Moreover, phenotypic properties of rice, such as plant height, fresh weight, and dry weight were increased in the plants inoculated with all antagonistic fungi tested, compared to the infected plants, except for the fresh weight of plants inoculated with Curnularia lunata (Wakker) Boedijn. The present in vivo and in vitro studies revealed that T. virens and A. fumigatus are the most effective antagonists in rice sheath blight disease control and could be applied in agricultural practice.

Trichoderma spp. are widely used biocontrol agents which are antagonistic to a variety of plant pathogens. Chlamydospores are a type of propagules produced by many fungi that have thick walls and are highly resistant to adverse environmental conditions. Chlamydospore preparations of Trichoderma spp. can withstand various storage conditions, have a longer shelf life than conidial preparations and have better application potential. However, large-scale production of chlamydospores has proven difficult. To understand the molecular mechanisms governing chlamydospore formation (CF) in Trichoderma fungi, we performed a comprehensive analysis of transcriptome dynamics during CF across 8 different developmental time points, which were divided into 4 stages according to PCA analysis: the mycelium growth stage (S1), early and middle stage of CF (S2), flourishing stage of CF (S3), and late stage of CF and mycelia initial autolysis (S4). 2864, 3206, and 3630 DEGs were screened from S2 vs S1, S3 vs S2, and S4 vs S3, respectively. We then identified the pathways and genes that play important roles in each stage of CF by GO, KEGG, STC and WGCNA analysis. The results showed that DEGs in the S2 vs S1 were mainly enriched in organonitrogen compound metabolism, those in S3 vs S2 were mainly involved in secondary metabolite, cell cycle, and N-glycan biosynthesis, and DEGs in S4 vs S3 were mainly involved in lipid, glycogen, and chitin metabolic processes. We speculated that mycelial assimilation and absorption of exogenous nitrogen in the early growth stage (S1), resulted in subsequent nitrogen deficiency (S2). At the same time, secondary metabolites and active oxygen free radicals released during mycelial growth produced an adverse growth environment. The resulting nitrogen-deficient and toxin enriched medium may stimulate cell differentiation by initiating cell cycle regulation to induce morphological transformation of mycelia into chlamydospores. High expression of genes relating to glycogen, lipid, mannan, and chitin synthetic metabolic pathways during the flourishing (S3) and late stages (S4) of CF may be conducive to energy storage and cell wall construction in chlamydospores. For further verifying the functions of the amino sugar and nucleotide sugar metabolism (tre00520) pathway in the CF of T. virens GV29-8 strain, the chitin synthase gene (TRIVIDRAFT_90152), one key gene of the pathway, was deleted and resulted in the dysplasia of mycelia and an incapability to form normal chlamydospores, which illustrated the pathway affecting the CF of T. virens GV29-8 strain. Our results provide a new perspective for understanding the genetics of biochemical pathways involved in CF of Trichoderma spp.

Trichoderma spp. are widely used biocontrol agents which are antagonistic to a variety of plant pathogens. Chlamydospores are a type of propagules produced by many fungi that have thick walls and are highly resistant to adverse environmental conditions. Chlamydospore preparations of Trichoderma spp. can withstand various storage conditions, have a longer shelf life than conidial preparations and have better application potential. However, large-scale production of chlamydospores has proven difficult. To understand the molecular mechanisms governing chlamydospore formation (CF) in Trichoderma fungi, we performed a comprehensive analysis of transcriptome dynamics during CF across 8 different developmental time points, which were divided into 4 stages according to PCA analysis: the mycelium growth stage (S1), early and middle stage of CF (S2), flourishing stage of CF (S3), and late stage of CF and mycelia initial autolysis (S4). 2864, 3206, and 3630 DEGs were screened from S2 vs S1, S3 vs S2, and S4 vs S3, respectively. We then identified the pathways and genes that play important roles in each stage of CF by GO, KEGG, STC and WGCNA analysis. The results showed that DEGs in the S2 vs S1 were mainly enriched in organonitrogen compound metabolism, those in S3 vs S2 were mainly involved in secondary metabolite, cell cycle, and N-glycan biosynthesis, and DEGs in S4 vs S3 were mainly involved in lipid, glycogen, and chitin metabolic processes. We speculated that mycelial assimilation and absorption of exogenous nitrogen in the early growth stage (S1), resulted in subsequent nitrogen deficiency (S2). At the same time, secondary metabolites and active oxygen free radicals released during mycelial growth produced an adverse growth environment. The resulting nitrogen-deficient and toxin enriched medium may stimulate cell differentiation by initiating cell cycle regulation to induce morphological transformation of mycelia into chlamydospores. High expression of genes relating to glycogen, lipid, mannan, and chitin synthetic metabolic pathways during the flourishing (S3) and late stages (S4) of CF may be conducive to energy storage and cell wall construction in chlamydospores. For further verifying the functions of the amino sugar and nucleotide sugar metabolism (tre00520) pathway in the CF of T. virens GV29-8 strain, the chitin synthase gene (TRIVIDRAFT_90152), one key gene of the pathway, was deleted and resulted in the dysplasia of mycelia and an incapability to form normal chlamydospores, which illustrated the pathway affecting the CF of T. virens GV29-8 strain. Our results provide a new perspective for understanding the genetics of biochemical pathways involved in CF of Trichoderma spp. [ABSTRACT FROM AUTHOR]

Downy mildew (DM), caused by P. cubensis, is harmful to cucurbits including luffa, with increased shortcomings associated with its control through cultural practices, chemical fungicides, and resistant cultivars; there is a prompt need for an effective, eco-friendly, economical, and safe biocontrol approach. Current research is therefore dealt with the biocontrol of luffa DM1 through the endophytic fungi (EF) consortium. Results revealed that T. harzianum (ThM9) and T. virens (TvA1) showed pathogen-dependent inducible metabolic production of squalene and gliotoxins by higher gene expression induction of SQS1/ERG9 (squalene synthase) and GliP (non-ribosomal peptide synthetase). Gene expression of lytic enzymes of EF was also induced with subsequently higher enzyme activities upon confrontation with P. cubensis. EF-inoculated luffa seeds showed efficient germination with enhanced growth potential and vigor of seedlings. EF-inoculated plants showed an increased level of growth-promoting hormone GA with higher gene expression of GA2OX8. EF-pre-inoculated seedlings were resistant to DM and showed an increased GSH content and antioxidant enzyme activities (SOD, CAT, POD). The level of MDA, H2O2, REL, and disease severity was reduced by EF. ACC, JA, ABA, and SA were overproduced along with higher gene expression of LOX, ERF, NCED2, and PAL. Expression of defense-marker genes (PPO, CAT2, SOD, APX, PER5, LOX, NBS-LRR, PSY, CAS, Ubi, MLP43) was also modulated in EF-inoculated infected plants. Current research supported the use of EF inoculation to effectively escalate the systemic immunity against DM corresponding to the significant promotion of induced systemic resistance (ISR) and systemic acquired resistance (SAR) responses through initiating the defense mechanism by SA, ABA, ET, and JA biosynthesis and signaling pathways in luffa.

Trichoderma virens is a commercial biofungicide used in agriculture. We have earlier isolated a mutant of T. virens using gamma ray-induced mutagenesis. This mutant, designated as M7, is defective in morphogenesis, secondary metabolism, and mycoparasitism. The mutant does not produce conidia, and the colony is hydrophilic. M7 cannot utilize cellulose and chitin as a sole carbon source and is unable to parasitize the plant pathogens Rhizoctonia solani and Pythium aphanidermatum in confrontation assay. Several volatile (germacrenes, beta-caryophyllene, alloaromadendrene, gamma-muurolene) and non-volatile (viridin, viridiol, gliovirin, heptelidic acid) metabolites are not detected in M7. In transcriptome analysis, many genes related to secondary metabolism, carbohydrate metabolism, hydrophobicity, and transportation, among others, were found to be downregulated in the mutant. Using whole genome sequencing, we identified five deletions in the mutant genome, totaling about 250 kb (encompassing 71 predicted ORFs), which was confirmed by PCR. This study provides novel insight into genetics of morphogenesis, secondary metabolism, and mycoparasitism and eventually could lead to the identification of novel regulators of beneficial traits in plant beneficial fungi Trichoderma spp. We also suggest that this mutant can be developed as a microbial cell factory for the production of secondary metabolites and proteins.

Cacao pod rot caused by Phytophthora palmivora, is an important disease and contributes significant disease losses to global cocoa production. This research objective was to determine the effect of Trichoderma harzianum and T. virens suspensions to cacao pod rot disease on the field. This research was carried out in Pulo Hagu village, Pidie Regency, Aceh, Indonesia from March to July 2017. The single pattern randomized block design was adopted to evaluate three treatments, i.e. without suspension (control), suspensions of T. Harzianum, and T. virens for eight replications. Each replication consisted of three of experimental units. The result showed that both of Trichoderma species contained only Alkaloid metabolite based on Phytochemical test. On the field, the application of T. harzianum suspension reduced the percentage of fruit infection and disease intensity for 48.57 %, 46.04 % at 12 weeks after application (WAA) respectively. Based on the percentage reduction in the area of the spot between the metabolites T. harzianum suspension and control and T. virens and control are 47.24 % and 27.46 % at 87 WAA respectively. In addition, T. virens suppressed the percentage of infected fruit and the intensity of infected fruit for 40.61 % and 38.02 % at 12 WAA.

The anticancer antibiotic heptelidic acid is a sesquiterpene lactone produced by the beneficial plant fungus Trichoderma virens. This species has been separated into two strains, referred to as P and Q, based on its biosynthesis of secondary metabolites; notably, only P-strains were reported to produce heptelidic acid. While characterizing a Q-strain of T. virens containing a directed mutation in the non-ribosomal peptide synthetase encoding gene Tex7 , the appearance of an unknown compound in anomalously large quantities was visualized by TLC. Using a combination of HPLC, LC-MS/MS, and NMR spectroscopy, this compound was identified as heptelidic acid. This discovery alters the strain classification structure of T. virens. Additionally, the Tex7 mutants inhibited growth of maize seedlings, while retaining the ability to induce systemic resistance against the foliar fungal pathogen, Cochliobolus heterostrophus. • TvTex7 deletion induces accumulation of heptelidic acid and viridin. • Identification of heptelidic acid in a Q-strain of T. virens alters the strain classification structure. • Maize treated with Tex7 deletion mutants are significantly smaller than control and WT treated plants. [ABSTRACT FROM AUTHOR]

Trichoderma virens is a commercial biofungicide used in agriculture. We have earlier isolated a mutant of T. virens using gamma ray-induced mutagenesis. This mutant, designated as M7, is defective in morphogenesis, secondary metabolism, and mycoparasitism. The mutant does not produce conidia, and the colony is hydrophilic. M7 cannot utilize cellulose and chitin as a sole carbon source and is unable to parasitize the plant pathogens Rhizoctonia solani and Pythium aphanidermatum in confrontation assay. Several volatile (germacrenes, beta-caryophyllene, alloaromadendrene, gamma-muurolene) and non-volatile (viridin, viridiol, gliovirin, heptelidic acid) metabolites are not detected in M7. In transcriptome analysis, many genes related to secondary metabolism, carbohydrate metabolism, hydrophobicity, and transportation, among others, were found to be downregulated in the mutant. Using whole genome sequencing, we identified five deletions in the mutant genome, totaling about 250 kb (encompassing 71 predicted ORFs), which was confirmed by PCR. This study provides novel insight into genetics of morphogenesis, secondary metabolism, and mycoparasitism and eventually could lead to the identification of novel regulators of beneficial traits in plant beneficial fungi Trichoderma spp. We also suggest that this mutant can be developed as a microbial cell factory for the production of secondary metabolites and proteins. [ABSTRACT FROM AUTHOR]

Small secreted cysteine‐rich proteins (SSCPs) from fungi play an important role in fungi–host interactions. The plant‐beneficial fungi Trichoderma spp. are in use worldwide as biocontrol agents and protect the host plant from soil‐borne as well as foliar pathogens. Recently, a novel SSCP, Tsp1, has been identified in the secreted protein pool of T. virens and is overinduced upon its interaction with the roots of the maize plant. The protein was observed to be well conserved in the Ascomycota division of fungi, and its homologs are present in many plant‐pathogenic fungi such as Fusarium oxysporum and Magnaporthe oryzae. However, none of these homologs have yet been characterized. Recombinant Tsp1 protein has been expressed and purified using an Escherichia coli expression system. The protein, with four conserved cysteines, forms a dimer in solution as observed by size‐exclusion chromatography. The dimerization, however, does not involve disulfide bonds. Circular‐dichroism data suggested that the protein has a β‐strand‐rich secondary structure that matched well with the secondary structure predicted using bioinformatics methods. The protein was crystallized using sodium malonate as a precipitant. The crystals diffracted X‐rays to 1.7 Å resolution and belonged to the orthorhombic space group P212121 (Rmeas = 5.4%), with unit‐cell parameters a = 46.3, b = 67.0, c = 173.2 Å. The Matthews coefficient (VM) of the crystal is 2.32 Å3 Da−1, which corresponds to nearly 47% solvent content with four subunits of Tsp1 protein in the asymmetric unit. This is the first report of the structural study of any homolog of the novel Tsp1 protein. These structural studies will help in understanding the classification and function of the protein. [ABSTRACT FROM AUTHOR]