Screening and evaluation of the biocontrol efficacy of a Trichoderma brevicompactum strain and its metabolite trichodermin against banana Fusarium wilt / 生物工程学报

The banana Fusarium wilt (BFW) caused by Fusarium oxysporum f. sp. cubense tropical race4 (FocTR4) is difficult to control worldwide, which causes a huge economic losse to banana industry. The purpose of this study was to screen Trichoderma strains with antagonistic activity against FocTR4, to isolate and purify the active compound from the fermentation broth, so as to provide important biocontrol strains and active compound resources. In this work, Trichoderma strains were isolated and screened from the rhizosphere soil of crops, and the strains capable of efficiently inhibiting FocTR4 were screened by plate confrontation, and further confirmed by testing inhibition for the conidial germination and mycelial growth of FocTR4. The phylogenetic tree clarified the taxonomic status of the biocontrol strains. Moreover, the active components in the fermentation broth of the strains were separated and purified by column chromatography, the structure of the most active component was analyzed by nuclear magnetic resonance spectroscopy (NMR), the BFW control effect was tested by pot experiments. We obtained a strain JSHA-CD-1003 with antagonistic activity against FocTR4, and the inhibition rate from plate confrontation was 60.6%. The fermentation broth of JSHA-CD-1003 completely inhibited the germination of FocTR4 conidia within 24 hours. The inhibition rate of FocTR4 hyphae growth was 52.6% within 7 d. A phylogenetic tree was constructed based on the ITS and tef1-α gene tandem sequences, and JSHA-CD-1003 was identified as Trichoderma brevicompactum. Purification and NMR identification showed that the single active compound was trichodermin, and the minimum inhibitory concentration (MIC) was 25 μg/mL. Pot experiments showed that the fermentation broth of strain JSHA-CD-1003 was effective against BFW. The control rate of leaf yellowing was 47.4%, and the rate of bulb browning was 52.0%. Therefore, JSHA-CD-1003 effectively inhibited FocTR4 conidial germination and mycelium growth through producing trichodermin, and showed biocontrol effect on banana wilt caused by FocTR4, thus is a potential biocontrol strain.

Thirteen new peptaibols with antimicrobial activities from Trichoderma sp / 中国天然药物

From the fungus Trichoderma sp., we isolated seven novel 18-residue peptaibols, neoatroviridins E-K (1-7), and six new 14-residue peptaibols, harzianins NPDG J-O (8-13). Additionally, four previously characterized 18-residue peptaibols neoatroviridins A-D (14-17) were also identified. The structural configurations of the newly identified peptaibols (1-13) were determined by comprehensive nuclear magnetic resonance (NMR) and high-resolution electrospray ionization tandem mass spectrometry (HR-ESI-MS/MS) data. Their absolute configurations were further determined using Marfey's method. Notably, compounds 12 and 13 represent the first 14-residue peptaibols containing an acidic amino acid residue. In antimicrobial assessments, all 18-residue peptaibols (1-7, 14-17) exhibited moderate inhibitory activities against Staphylococcus aureus 209P, with minimum inhibitory concentration (MIC) values ranging from 8-32 μg·mL-1. Moreover, compound 9 exhibited moderate inhibitory effect on Candida albicans FIM709, with a MIC value of 16 μg·mL-1.

Study on alkaloids of endophytic Trichoderma atroviride B7 from Colquhounia coccinea var. mollis / 药学学报

Nine compounds were isolated from the crude extract of the solid culture of endophyte Trichoderma atroviride B7 of Colquhounia coccinea var. mollis by silica gel column chromatography, Sephadex LH-20 gel column chromatography, and HPLC. They were identified as atroviridanol (1), 3-oxo-3-[(2-phenylethyl) amino]-propanoic acid (2), N-(2′-phenylethyl)-acetamide (3), neoechinulin A (4), echinulin (5), gancidin W (6), N-isobutyl-3-methylbutanamide (7), 5-acetamido-1-pentanol (8), and N-2-methylpropyl-2-methylbutenamide (9) by NMR, HR-MS, and so on. Among them, compound 1 is a new compound, and compounds 2-9 are firstly isolated from Trichoderma spp.

Amazonian soil fungi are efficient degraders of glyphosate herbicide; novel isolates of Penicillium, Aspergillus, and Trichoderma / Os fungos do solo amazônico são degradadores eficientes do herbicida glifosato; novos isolados de Penicillium, Aspergillus e Trichoderma

Pesticide residues that contaminate the environment circulate within the hydrological cycle can accumulate within the food chain and cause problems to both environmental and human health. Microbes, however, are well known for their metabolic versatility and the ability to degrade chemically stable substances, including recalcitrant xenobiotics. The current study focused on bio-prospecting within Amazonian rainforest soils to find novel strains fungi capable of efficiently degrading the agriculturally and environmentally ubiquitous herbicide, glyphosate. Of 50 fungal strains isolated (using culture media supplemented with glyphosate as the sole carbon-substrate), the majority were Penicillium strains (60%) and the others were Aspergillus and Trichoderma strains (26 and 8%, respectively). All 50 fungal isolates could use glyphosate as a phosphorous source. Eight of these isolates grew better on glyphosate-supplemented media than on regular Czapek Dox medium. LC-MS revealed that glyphosate degradation by Penicillium 4A21 resulted in sarcosine and aminomethylphosphonic acid.

Resíduos de agrotóxicos que contaminam o meio ambiente circulam no ciclo hidrológico, podendo se acumular na cadeia alimentar e causar problemas tanto à saúde ambiental quanto humana. Por sua vez, microrganismos são bem conhecidos por sua versatilidade metabólica e capacidade de degradar substâncias quimicamente estáveis, incluindo xenobióticos recalcitrantes. O estudo atual se concentrou na bioprospecção nos solos da floresta amazônica para encontrar novas linhagens de fungos capazes de degradar com eficiência o herbicida onipresente na agricultura e no meio ambiente, o glifosato. Entre os 50 fungos isolados (usando meio de cultura suplementado com glifosato como única fonte de carbono), a maioria eram isolados do gênero Penicillium (60%) e os outros eram isolados de Aspergillus e Trichoderma (26 e 8%, respectivamente). Todos os 50 isolados de fungos foram capazes de usar glifosato como fonte de fósforo. Oito desses isolados cresceram melhor em meio suplementado com glifosato do que em meio Czapek Dox regular. LC-MS revelou que a degradação do glifosato por Penicillium 4A21 resultou nos metabólitos sarcosina e ácido aminometilfosfônico.

Amazonian soil fungi are efficient degraders of glyphosate herbicide; novel isolates of Penicillium, Aspergillus, and Trichoderma

Abstract Pesticide residues that contaminate the environment circulate within the hydrological cycle can accumulate within the food chain and cause problems to both environmental and human health. Microbes, however, are well known for their metabolic versatility and the ability to degrade chemically stable substances, including recalcitrant xenobiotics. The current study focused on bio-prospecting within Amazonian rainforest soils to find novel strains fungi capable of efficiently degrading the agriculturally and environmentally ubiquitous herbicide, glyphosate. Of 50 fungal strains isolated (using culture media supplemented with glyphosate as the sole carbon-substrate), the majority were Penicillium strains (60%) and the others were Aspergillus and Trichoderma strains (26 and 8%, respectively). All 50 fungal isolates could use glyphosate as a phosphorous source. Eight of these isolates grew better on glyphosate-supplemented media than on regular Czapek Dox medium. LC-MS revealed that glyphosate degradation by Penicillium 4A21 resulted in sarcosine and aminomethylphosphonic acid.

Resumo Resíduos de agrotóxicos que contaminam o meio ambiente circulam no ciclo hidrológico, podendo se acumular na cadeia alimentar e causar problemas tanto à saúde ambiental quanto humana. Por sua vez, microrganismos são bem conhecidos por sua versatilidade metabólica e capacidade de degradar substâncias quimicamente estáveis, incluindo xenobióticos recalcitrantes. O estudo atual se concentrou na bioprospecção nos solos da floresta amazônica para encontrar novas linhagens de fungos capazes de degradar com eficiência o herbicida onipresente na agricultura e no meio ambiente, o glifosato. Entre os 50 fungos isolados (usando meio de cultura suplementado com glifosato como única fonte de carbono), a maioria eram isolados do gênero Penicillium (60%) e os outros eram isolados de Aspergillus e Trichoderma (26 e 8%, respectivamente). Todos os 50 isolados de fungos foram capazes de usar glifosato como fonte de fósforo. Oito desses isolados cresceram melhor em meio suplementado com glifosato do que em meio Czapek Dox regular. LC-MS revelou que a degradação do glifosato por Penicillium 4A21 resultou nos metabólitos sarcosina e ácido aminometilfosfônico.

Amazonian soil fungi are efficient degraders of glyphosate herbicide; novel isolates of Penicillium, Aspergillus, and Trichoderma / Os fungos do solo amazônico são degradadores eficientes do herbicida glifosato; novos isolados de Penicillium, Aspergillus e Trichoderma

Abstract Pesticide residues that contaminate the environment circulate within the hydrological cycle can accumulate within the food chain and cause problems to both environmental and human health. Microbes, however, are well known for their metabolic versatility and the ability to degrade chemically stable substances, including recalcitrant xenobiotics. The current study focused on bio-prospecting within Amazonian rainforest soils to find novel strains fungi capable of efficiently degrading the agriculturally and environmentally ubiquitous herbicide, glyphosate. Of 50 fungal strains isolated (using culture media supplemented with glyphosate as the sole carbon-substrate), the majority were Penicillium strains (60%) and the others were Aspergillus and Trichoderma strains (26 and 8%, respectively). All 50 fungal isolates could use glyphosate as a phosphorous source. Eight of these isolates grew better on glyphosate-supplemented media than on regular Czapek Dox medium. LC-MS revealed that glyphosate degradation by Penicillium 4A21 resulted in sarcosine and aminomethylphosphonic acid.

Resumo Resíduos de agrotóxicos que contaminam o meio ambiente circulam no ciclo hidrológico, podendo se acumular na cadeia alimentar e causar problemas tanto à saúde ambiental quanto humana. Por sua vez, microrganismos são bem conhecidos por sua versatilidade metabólica e capacidade de degradar substâncias quimicamente estáveis, incluindo xenobióticos recalcitrantes. O estudo atual se concentrou na bioprospecção nos solos da floresta amazônica para encontrar novas linhagens de fungos capazes de degradar com eficiência o herbicida onipresente na agricultura e no meio ambiente, o glifosato. Entre os 50 fungos isolados (usando meio de cultura suplementado com glifosato como única fonte de carbono), a maioria eram isolados do gênero Penicillium (60%) e os outros eram isolados de Aspergillus e Trichoderma (26 e 8%, respectivamente). Todos os 50 isolados de fungos foram capazes de usar glifosato como fonte de fósforo. Oito desses isolados cresceram melhor em meio suplementado com glifosato do que em meio Czapek Dox regular. LC-MS revelou que a degradação do glifosato por Penicillium 4A21 resultou nos metabólitos sarcosina e ácido aminometilfosfônico.

Advances in research and application of Trichoderma for inducing resistance against root rot diseases in root and rhizome of Chinese medicinal materials / 中国中药杂志

Root rot is a microbial disease that is difficult to control and can result in serious losses in the planting of most Chinese medicinal materials. As high as 87.6% of roots or rhizomes of Chinese medicinal materials are susceptible to root rot, which seriously affects the cultivation development of Chinese medicinal materials. Trichoderma fungi, possessing biological control functions, can induce plants to improve their resistance to microbial diseases, promote plant growth, and effectively reduce the losses caused by various microbial diseases on cultivation. At present, Trichoderma is rarely used in the cultivation of Chinese medicinal materials, so it has great application potential for the prevention and control of root rot diseases in farmed Chinese medicinal materials. Based on the above situation, after comparison and discussion, it is believed that compared with chemical control and physical control, biological control of root rot diseases of Chinese medicinal materials is more efficient and meets the development needs of Chinese medicinal materials ecological planting in China. This paper reviewed the progress in the research and application of Trichoderma in the control of root rot diseases in the root and rhizome of farmed Chinese medicinal materials in the past 10 years and found that most of the current research on the biological control of root rot diseases in Chinese medicinal materials was mostly limited to the verification of the inhibitory effect of Trichoderma strains on the growth of the pathogenic microbes. Studies on the induction effect of Trichoderma on Chinese medicinal materials are not in depth. Studies on the responding mechanisms of most Chinese medicinal materials to Trichoderma are highly absent. Moreover, there are few reports on field experiments, which indicates that there is a long way to go before Trichoderma is widely applied in the farming practice of Chinese medicinal materials. To sum up, this paper aimed to link the present and the future and advocated further relevant research and more experiments on the application of Trichoderma in the farming of Chinese medicinal materials.

Influence of microbial consortia on the incidence of grey mold (Botrytis cinerea) in strawberry (Monterey variety)

Botrytis cinerea, the causal agent of grey mold disease, is one of the most destructive pathogens of strawberry crops, both in vegetative development and postharvest. The control of this pathogen is complex due to its aggressiveness and ability to attack and infect various plant tissues and is mainly based on chemical control; however, the incorrect use of pesticides, mainly due to overdosing, causes the presence of traces of these agrochemicals in the fruits, as well as the selection of pathogen resistance to fungicides, making it a risk to human health and the environment. The objective of the study was to use biological regulation strategies, with the application of microbial consortia made up of mycorrhizal fungi, antagonistic bacteria and Trichoderma harzianum, as an alternative for the management of grey mold in strawberry crops (Monterey variety) under field conditions. Treatments T4 (mycorrhizal fungi), T8 (mycorrhizal fungi, antagonistic bacteria and T. harzianum) and T2 (T. harzianum) presented the lowest incidence of the pathogen with 2.6, 3.1 and 3.6 %, respectively, compared to control plants with 16.6%. The influence of all biological treatments on the regulation of B. cinerea was greater than the control.

Botrytis cinerea, el agente causal de la enfermedad del moho gris, es uno de los patógenos más destructivos del cultivo de fresa, tanto en el desarrollo vegetativo como en poscosecha. El control de este patógeno es complejo, debido a su agresividad y capacidad de atacar e infectar diversos tejidos de la planta y se basa, principalmente, en el control químico; sin embargo, el uso incorrecto de plaguicidas, principalmente por sobredosificación, provoca la presencia de trazas de estos agroquímicos en los frutos, así como la selección de resistencia del patógeno a los fungicidas, convirtiéndolo en un riesgo para la salud humana y el ambiente. El objetivo del estudio fue utilizar estrategias de regulación biológica, con la aplicación de consorcios microbianos, conformados por hongos micorrícicos, bacterias antagonistas y Trichoderma harzianum, como alternativa para el manejo del moho gris, en cultivos de fresa (variedad Monterey), en condiciones de campo. Los tratamientos T4 (hongos micorrízicos), T8 (hongos micorrízicos, bacterias antagonistas y T. harzianum) y T2 (T. harzianum) presentaron la menor incidencia del patógeno, con 2,6, 3,1 y 3,6 %, respectivamente, en comparación con las plantas control, con 16,6 %. La influencia de todos los tratamientos biológicos en la regulación de B. cinerea fue mayor respecto al control.

Growth of Schizolobium parahyba var. amazonicum seedlings by strains of Trichoderma spp. under boron rates

Brazil has many important native species, which includes Schizolobium parahyba var. amazonicum (Huber ex Ducke) Barneby, however, the growth of this species is inhibited in soils with low boron contents, or excess boron, which causes phytotoxicity. Applications of strains of Trichoderma spp. increase the plants' tolerance to abiotic stresses, including tolerance to low or excess levels of nutrients such as boron (B). Thus, the objective of this work was to evaluate plant development and the effect of strains of Trichoderma spp. in Schizolobium parahyba var. amazonicum seedlings grown under different boron rates. A randomized block experimental design was used, with a 4×5 factorial scheme, consisting of 4 strains of Trichoderma spp. and 5 B rates (0, 0.5, 1.0, 1.5, and 2.0 mg dm-3), with seven replications, in greenhouse. Plant height, stem diameter, fresh and dry mass, leaf, stem and total were evaluated at 120 days after emergence. The stains of Trichoderma spp. and B rates presented significant interaction for all evaluated variables, decreasing phytotoxicity. The strain IBLF006WP (T. harzianum) showed a higher capacity of increasing the plants' tolerance to boron, followed by URM5911 (T. asperellum). However, the beneficial effect of increasing this tolerance with the application of these strains is only feasible for soils with high contents of this micronutrient in the soil.

Methods of plant growth-promoting fungi application to enhance the growth and yield of wheat var. Ibaa99

Aims@#The main aim of the study was to evaluate some methods of application of Aspergillus niger AD1 and Trichoderma hamatum T-113 for enhancing the growth and yield of wheat var. Ibaa99 in pots and field conditions.@*Methodology and results@#Plant growth-promoting fungi (PGPF) loaded with peat moss were used at a rate of 100, 150 and 200 mL pot-1 or m-2 in filed soil; seed treatment (coating) with fungi suspension 19 × 107, soil treatment and combination of all the three methods was employed in the study. Wheat seeds were sown in pots and field plots during 2018-2019, and data regarding various growth and yield attributes were recorded. In both pot and field trials, the results revealed that the best treatments for the desired plant growth and yield attributes were peat moss 150 mL alone or in combination with soil and seed treatments. The soil physicochemical parameters were also improved after inoculation with selected fungal isolates in different application methods compared with un-inoculated control treatment in both pot and field conditions.@*Conclusion, significance and impact of study@#The PGPF play a vital role represented phytoremediation, phytostimulation and bio-fertilization. The isolates of PGPF, which were applied with peat moss at 150 mL to the pot and in the field alone or combined with seed treatment and soil application, were significantly the best effective method for improving wheat attributes.

Performance of a selected Trichoderma strain as plant pathogen inhibitor and biofertilizer

Aims@#The application of beneficial microbes is a suitable alternative to synthetic pesticides and fertilizers for agriculture. This study was aimed to evaluate the potential of a selected Trichoderma strain as a biocontrol agent against Rhizoctonia sp. and as a biofertilizer to improve paddy growth.@*Methodology and results@#Four Bipolaris strains were identified via DNA barcoding as the cause of brown spot disease, whereas two Rhizoctonia strains were similarly identified as the cause of sheath blight disease in Brunei Darussalam. Eight Trichoderma strains were initially screened using confrontation assay and were found to substantially inhibit the growth of Rhizoctonia sp. Hybrid rice named BDR5 was treated with Trichoderma sp. UBDFM01 and/or Rhizoctonia sp. It was found that the selected strain showed the potential as a biofertilizer by significantly increasing the vigour index I, chlorophyll a, chlorophyll b, total chlorophyll and dry shoot weight of the rice plants. The pathogen negatively affected the plants by significantly reducing the vigour index II, chlorophyll a, chlorophyll a/b ratio, total chlorophyll, and total weight of grains. Trichoderma strain showed the potential as a biocontrol agent by significantly diminishing the negative effects of the pathogen on the chlorophyll a, chlorophyll a/b ratio and total chlorophyll.@*Conclusion, significance and impact of study@#This study highlights the potential of Trichoderma sp. UBDFM01 as a biocontrol agent against Rhizoctonia sp. and also as a biofertilizer for rice plants. In addition, this study is the first to provide DNA-based evidence of Bipolaris sp. and Rhizoctonia sp. as the fungi that caused rice diseases in Brunei Darussalam.

Suppression and management of Meloidogyne incognita in soil using Trichoderma harzianum NFCF160 and Trichoderma virens Isf-77

Aims@#Meloidogyne incognita adversely affects numerous crop plants worldwide. Therefore, the modern world has been moving towards biocontrol methods to prevent nematode attacks. This study was aimed to (i) investigate the potential use of Trichoderma harzianum NFCF160 and T. virens Isf-77 in managing M. incognita in soil and (ii) identify trapping mechanisms employed by both Trichoderma strains to suppress M. incognita.@*Methodology and results@#Three weeks old, Basella alba L. plants were subjected to five different treatments. The above and below ground growth parameters and the galling indices of these plants were measured every four weeks for three sampling times. Trapping mechanisms employed by Trichoderma strains were examined following plate assays. Plants treated with T. harzianum NFCF160 and T. virens Isf-77 had significantly higher values for the total number of leaves (34 ± 2.84) and (27 ± 2.61), fresh weight of the shoot (81 ± 9.51 g) and (91 ± 9.70 g), dry weight of the shoot (71 ± 5.24 g) and (62 ± 5.81 g), respectively eight weeks after inoculation of M. incognita. Significantly low galling indices (2 and 2) were recorded in B. alba treated with Trichoderma strains. Both Trichoderma strains exhibited various nematode-trapping mechanisms, such as non-constricting rings and adhesive spores.@*Conclusion, significance and impact of study@#This investigation highlighted the potential of both Trichoderma strains as biocontrol agents to control M. incognita effect in sustainable agriculture.

Morphological and molecular characterization of Trichoderma species isolated from rhizosphere soils in Malaysia

Aims@#Knowledge of the Trichoderma taxa is important for both control efficiency and environmental conservation. Therefore, the objective of this study is to isolate and identify Trichoderma species from various rhizosphere soil samples using phenotypic and molecular characterization.@*Methodology and results@#Native Trichoderma spp. were isolated from agricultural fields in 17 sites from seven states of Malaysia. These isolates were characterized via morphological observation and molecular phylogenetic analysis based on the translation elongation factor-1α (tef1-α) gene. About 42 isolates were classified into eight Trichoderma species, which are Trichoderma asperellum, T. hamatum, T. harzianum, T. koningiopsis, T. rodmanii, T. spirale, T. viride and T. virens. Comparison of DNA sequences of tef1-α showed that the isolates were 98-100% similar to respective Trichoderma species from GenBank, thus confirming the fungal identity. Phylogenetic trees of maximum likelihood (ML) dataset of tef1-α inferred that the isolates were clustered according to species.@*Conclusion, significance and impact of study@#Findings in the present study will be beneficial for the purposes of biodiversity conservation and plant disease management using biocontrol agents.

Evaluation on the effectiveness of combination of biocontrol agents in managing Ganoderma boninense of oil palm

Aims@#The development of an effective biocontrol formulation for inhibition of Ganoderma boninense, a well-known destructive pathogen in oil palm plantation is important to prolong the palm’s lifespan and reduce the losses due to this disease. In this paper, we present some new bioformulations with combination of different types of biocontrol agents in managing basal stem rot (BSR) disease. @*Methodology@#The effectiveness of the treatments designed as T1 (Trichoderma harzianum + Lecanicillium spp. + Streptomyces sundarbansensis + Pseudomonas aeruginosa), T2 (Penicillium simplicissimum + Lecanicillium sp. + S. sundarbansensis + P. aeruginosa), T3 (P. simplicissimum + P. aeruginosa) and T4 (LEStani®) was evaluated through treatment on the oil palm seedlings artificial infected by G. boninense and the results were expressed in disease severity index (DSI), bole severity index (BSI) and ergosterol content.@*Conclusion, significance and impact of study@#All tested treatments (T1-T4) managed to control the severity of the Ganoderma infection from continuously increasing when the treatments were applied either one month before or after artificial inoculation. The disease severity of infected seedlings without treatments had increased for almost 2-fold at the end of the trial. Moreover, T1 had the greatest inhibition of G. boninense with the lowest ergosterol content (a bioindicator of Ganoderma colonization) detected (676.67 g/mL), which is about 1.9-fold lower than control (1273.33 ug/mL) without treatments and a BSI score of 1. Based on the effectiveness among the four tested biocontrol formulations, T1 is the most promising formulation to be further evaluated in the field for control of BSR disease. However, more research is needed in the future to estimate the effective amount for application in open environment.

Antagonistic microorganisms and nitrogen fertilization in control of tomato southern blight

The present study assessed the efficacy of formulated biocontrol agents and nitrogen fertilization on southern blight control. Antagonism test in vitro was performed to assess the inhibitory activity of Bacillus methylotrophicus and Trichoderma asperellum against the growth of Sclerotium rolfsii. Tomato seedlings were transplanted into the substrate added with ammonium nitrate doses and inoculated with the formulated biocontrol agents Ônix (B. methylotrophicus) or Quality (T. asperellum). Subsequently, seedlings were inoculated with S. rolfsii. Plant mortality, shoot and root weight were assessed 11 days after the last inoculation. Agents had effective inhibitory activity against S. rolfsii; thus, they could reduce southern blight severity when combined with ammonium nitrate. However, plant mortality was not reduced by them.(AU)

Selection and characterization of novel zinc-tolerant Trichoderma strains obtained by protoplast fusion

Aim: To develop new Trichoderma strains, capable of removing toxic heavy metal ions from polluted environments, via protoplast fusion.Methodology: Trichoderma parental strains (T. viride and T. koningii) along with their ten fusants (Tk+Tv 1, Tk+Tv 2, Tk+Tv 3, Tk+Tv 4, Tk+Tv 5, Tk+Tv 6, Tk+Tv 7, Tk+Tv 8, Tk+Tv 9 and Tk+Tv 10) were obtained from the Department of Plant Pathology, Junagadh Agricultural University, Junagadh. The strains obtained by protoplast fusion were examined for their ability to remove toxic heavy metal ions, especially zinc ion. Fourier-transform infrared spectroscopy (FTIR) was conducted to detect the zinc uptake mechanism of Trichoderma parental and their fusant strains. Results: FTIR results demonstrated the Zn ion uptake capacity of fusant strains was found to be higher than that of the parental strains (12.8 to 10.7 mg g-1 on a dry weight basis at 1300 ppm). The highest Zn ion mobility observed was 62.1 mg. kg-1 and the highest Zn ion mobility observed per strain was 12.4% in Tk+Tv 3, followed by 11.86 % in Tk + Tv 7, 11.84% in Tk + Tv 9 and 11.28% in Tk + Tv 10. Parental and fusant strains Tk + Tv 3, Tk + Tv 8 and Tk + Tv 10 confirmed the involvement of different functional groups for different concentrations of zinc during adsorption by the fungus. Interpretation: FTIR results identified greater metal removal capacity in the fusant strains, particularly for soil Zn ion. Zinc tolerance was higher in the fusant strains than in the parental strains. Thus, protoplast fusion is an effective and feasible method for constructing new strains that can be used for bioremediation of contaminated environments.

Factors affecting the chitinase activity of Trichoderma asperellum isolated from agriculture field soils

In the present study, 20 fungal strains were isolated from tomato rhizosphere of Senegal. Of 20 strains, five showedthe chitinolytic activity on chitin agar medium. Of the five strains, NG4 showed the maximum solubilization zone.This strain was identified by preliminary biochemical and 18S rRNA sequencing analysis. Enzyme productionstarted after 3 days of incubation and maximum was observed after 5 days of incubation. Culture filtrate amendedwith 0.1% colloidal chitin was used in the production medium. The optimum conditions for maximum chitinaseactivity are – 6 days of growth and temperature of 30°C at pH 6.0. The chitinase activity was also influenced by theaddition of carbon and nitrogen sources in the production medium.

A simple and accurate method for specific quantification of biomass in mixed cultures of filamentous fungi by quantitative PCR / Un método simple y preciso para la cuantificación específica de biomasa en cultivos mixtos de hongos filamentosos por PCR cuantitativa

Abstract Production of lignocellulolytic enzymes by filamentous fungi have a great potential at industrial level due to their widespread applications. Mixed fungal cultures and particularly mixed fungal biofilms constitute a promising fermentation system for an enhanced enzyme production. However, it has not been addressed how much of this enhancement depends on the mixed biomass proportion. In this sense, the aim of this study was to develop a method to specifically and accurately quantify mixed fungal biomass. For this purpose, mixed biofilm cultures composed of Aspergillus niger and Trichoderma reesei, two filamentous fungi used industrially for cellulase production, were collected from 48 to 120 h of growth; mycelia were pulverized, and DNA was extracted for qPCR assays with specific primers for each fungus. Primers were designed from non-conserved regions of sequences of actin and β-tubulin genes of both A. niger and T. reesei. Specificity of these primers was tested in silico and experimentally. A statistically significant correlation was obtained between qPCR-calculated biomass and dry weight biomass data. By this method, it was possible to detect changes on mycelia proportions in biofilms over time, suggesting a competitive interaction between these two fungi. In conclusion, this method allows a specific and accurate quantification of mixed fungal biomass and could be also applied to different mixed culture systems for studying microbial interactions.

Resumen La producción de enzimas lignocelulolíticas por hongos filamentosos tiene un gran potencial a nivel industrial debido a sus diversas aplicaciones. Los cultivos fúngicos mixtos y particularmente las biopelículas fúngicas mixtas constituyen un sistema de fermentación prometedor para una mayor producción enzimática. Sin embargo, no se ha abordado cuánto de esta mejora depende de la proporción de biomasa mixta. En este sentido, el objetivo de este estudio fue desarrollar un método para cuantificar de forma específica y precisa la biomasa fúngica mixta. Para este propósito, se recolectaron cultivos mixtos de biopelículas de 48 a 120 h de crecimiento compuestos por Aspergillus niger y Trichoderma reesei, dos hongos filamentosos utilizados industrialmente para la producción de celulasas; el micelio se pulverizó y el ADN se extrajo para ensayos de qPCR con cebadores específicos para cada hongo. Los cebadores se diseñaron a partir de regiones no conservadas de las secuencias de los genes de actina y β-tubulina de A. niger y T. reesei. La especificidad de estos cebadores se probó in silico y experimentalmente. Se obtuvo una correlación estadísticamente significativa entre la biomasa calculada mediante qPCR y los datos de biomasa en peso seco. Mediante este método, fue posible detectar cambios en las proporciones de los micelios en las biopelículas a lo largo del tiempo, lo que sugiere una interacción competitiva entre estos dos hongos. En conclusión, este método permite una cuantificación específica y precisa de la biomasa fúngica mixta y también podría aplicarse a diferentes sistemas de cultivo mixto para estudiar interacciones microbianas.

Screening of endophytic fungi against southern blight disease pathogen-Sclerotium delphinii in Dendrobium catenatum / 中国中药杂志

In order to screen the endophytic fungi that can enhance the host(Dendrobium catenatum) resistance to Sclerotium delphinii, the antagonism between each of the 43 endophytic fungi and the pathogen S. delphinii were tested. The results showed that 6 endophytic fungi(DCR2, DCR5, DCR21, DCR22, DCR42, DCR43) have strong activities against the pathogen, the inhibition rates were 49.2%, 49.2%, 47.2%, 56.2%, 53.2%, 48.0%, respectively. Then D. catenatum plantlets were inoculated with both S. delphinii and each of these six endophytic fungi. As a result, the incidence rates of leaves and stems of the D. catenatum plantlets inoculated with DCR2 and the pathogen were both significantly lower than those with other treatments, and the plantlet death rate was 0. It showed that DCR2 Trichoderma polysporum could effectively inhibit the southern blight disease of D. catenatum. Through the endophytic fungal re-isolation test, it was found that DCR2 can colonize in the roots, stems, and leaves of D. catenatum. The research will provide new ideas for the prevention and treatment of the southern blight disease of D. catenatum. It is also significant for reducing pesticide use, ensuring food safety, and promoting the sustainable development of D. catenatum industry. Furthermore, it will provide a basis for the disease control in other crops.

Enhance of Cellulase Production and Biomass Degradation by Transformation of the Trichoderma reesei RUT-C30∆ zface1 Strain

Abstract The second-generation bioethanol employs lignocellulosic materials degraded by microbial cellulases in their production. The fungus Trichoderma reesei is one of the main microorganisms producing cellulases, and its genetic modification can lead to the optimization in obtaining hydrolytic enzymes. This work carried out the deletion of the sequence that encodes the zinc finger motif of the transcription factor ACE1 (cellulase expression repressor I) of the fungus T. reesei RUT-C30. The transformation of the RUT-C30 lineage was confirmed by amplification of the 989 bp fragment relative to the selection marker, and by the absence of the zinc finger region amplification in mutants, named T. reesei RUT-C30Δzface1. The production of cellulases by mutants was compared to RUT-C30 and measured with substrates carboxymethylcellulose (CMC), microcrystalline cellulose (Avicel®) and Whatman filter paper (PF). The results demonstrated that RUT-C30Δzface1 has cellulolytic activity increased 3.2-fold in Avicel and 2.1-fold in CMC and PF. The mutants presented 1.4-fold higher sugar released in the hydrolysis of the biomass assays. These results suggest that the partial deletion of ace1 gene is an important strategy in achieving bioethanol production on an industrial scale at a competitive price in the fuel market.