Activity Of Cell Wall-degrading Enzymes Research Articles

Effects of ultraviolet irradiation on the in vitro antagonistic potential of Trichoderma spp. against soil-borne fungal pathogens

Development of new effective biocontrol agents is largely based on the antagonistic capacity of candidate agents against targeted pathogens in vitro. Different mechanisms contribute to such capacity, including the activity of cell wall-degrading enzymes, secretion of antimicrobial secondary metabolites, growth vigour and resistance to exogenous and endogenous toxins. In this study, a series of laboratory experiments were designed to improve the antagonistic activities of Trichoderma spp. against two plant fungal pathogens, Sclerotium rolfsii and Rhizoctonia solani. A simple but efficient mutagenesis programme was carried out using ultraviolet light to induce modifications in the genetic structure of two Trichoderma biocontrol agents, T. virens and T. asperellum. The obtained mutants were subjected to a) initial screening for media-permeable antifungal metabolites using the cellophane membrane-based method, and b) selected mutants were subjected to a series of antagonistic tests. Results revealed that the antagonistic potential of selected mutants was significantly improved against the two plant pathogens. Genetic stability test results indicated that the UV-derived mutant Tv3, maintained its elevated performance after 12 rounds of sub-culture. Gene expression analysis for five antagonism-associated genes were examined using real-Time PCR. Results revealed that the gene expression of two genes, chitinase 33, a cell wall degrading enzyme and, polyketide synthase, which is responsible for polyketide biosynthesis, a class of secondary metabolites with antimicrobial roles, were significantly upregulated in one of the mutated T. virens strains. Results of our in vitro antagonistic studies along with our molecular analysis indicate that the UV mutagenesis could be an effective strategy to improve Trichoderma antagonistic potential.

Innovative microscale workflow from fungi cultures to Cell Wall-Degrading Enzyme screening.

SummaryThis study aimed at developing a complete miniaturized high‐throughput screening workflow for the evaluation of the Cell Wall‐Degrading Enzyme (CWDE) activities produced by any fungal strain directly cultivated on raw feedstock in a submerged manner. In this study, wheat straw was selected as model substrate as it represents an important carbon source but yet poorly valorised to yield high added value products. Fungi were grown in a microbioreactor in a high‐throughput (HT) way to replace the fastidious shaking flask cultivations. Both approaches were compared in order to validate our new methodology. The range of CWDE activities produced from the cultures was assayed using AZO‐died and pNP‐linked substrates in an SBS plate format using a Biomek FXp pipetting platform. As highlighted in this study, it was shown that the CWDE activities gathered from the microbioreactor cultivations were similar or higher to those obtained from shake flasks cultures, with a lower standard deviation on the measured values, making this new method much faster than the traditional one and suitable for HT CWDE production thanks to its pipetting platform compatibility. Also, the results showed that the enzymatic activities measured were the same when doing the assay manually or using the automated method.

The MAP Kinase Kinase Gene AbSte7 Regulates Multiple Aspects of Alternaria brassicicola Pathogenesis.

Mitogen-activated protein kinase (MAPK) cascades in fungi are ubiquitously conserved signaling pathways that regulate stress responses, vegetative growth, pathogenicity, and many other developmental processes. Previously, we reported that the AbSte7 gene, which encodes a mitogen-activated protein kinase kinase (MAPKK) in Alternaria brassicicola, plays a central role in pathogenicity against host cabbage plants. In this research, we further characterized the role of AbSte7 in the pathogenicity of this fungus using ΔAbSte7 mutants. Disruption of the AbSte7 gene of A. brassicicola reduced accumulation of metabolites toxic to the host plant in liquid culture media. The ΔAbSte7 mutants could not efficiently detoxify cruciferous phytoalexin brassinin, possibly due to reduced expression of the brassinin hydrolase gene involved in detoxifying brassinin. Disruption of the AbSte7 gene also severely impaired fungal detoxification of reactive oxygen species. AbSte7 gene disruption reduced the enzymatic activity of cell wall-degrading enzymes, including cellulase, β-glucosidase, pectin methylesterase, polymethyl-galacturonase, and polygalacturonic acid transeliminase, during host plant infection. Altogether, the data strongly suggest the MAPKK gene AbSte7 plays a pivotal role in A. brassicicola during host infection by regulating multiple steps, and thus increasing pathogenicity and inhibiting host defenses.

Insights into Salt Stress-Induced Biochemical, Molecular and Epigenetic Regulation of Spatial Responses in Pigeonpea (Cajanus cajan L.)

Pigeonpea (Cajanus cajan L.) is a rich source of nutritionally good quality proteins, carbohydrates, minerals, and vitamins. However, environmental stresses adversely affect its productivity. Only limited reports are available on biochemical/physiological responses of pigeonpea under salt stress. The objectives of the present study were to screen pigeonpea germplasm accessions for salt stress tolerance, followed by understanding their biochemical, epigenetic and molecular responses. Based on germination, growth, and vigor of seedlings under salt stress, the most contrasting pair of salt-responsive genotypes (ICP1071- most salt-sensitive, and ICP7- most salt-tolerant) were selected. Three-week-old seedlings subjected to 250 mM NaCl stress for 7 days showed a significant increase in proline and reducing sugar contents in the case of ICP7, whereas a considerable increase in cell wall-degrading enzyme activity and protein oxidation was observed in ICP1071. Superoxide dismutase, peroxidase, and glutathione reductase activity increased considerably in shoots of ICP7. We observed the CcCYP gene to be upregulated in root, whereas CcCDR was upregulated in shoots of the salt-tolerant genotype to provide protection against the stress. The extent of DNA hypomethylation in the contrasting pigeonpea genotypes under salt stress was correlated with their salt tolerance level. Bisulfite sequencing of CcCDR revealed that methylation of three cytosine residues in CHH context in shoots of the ICP7 genotype due to salt stress results in 2.6-fold upregulated expression of the gene. With a 6.8% increase in methylation of the coding region of CcCDR, its expression level increased by 22%. To the best of our knowledge, this is the first report on a comprehensive study of salt-induced biochemical, epigenetic and molecular responses of pigeonpea, which might be useful in the development of improved salt-tolerant variety.

Integrated transcriptomic-metabolomic analysis reveals cellular responses of harvested strawberry fruit subjected to short-term exposure to high levels of carbon dioxide

Harvested strawberry (Fragaria × ananassa) fruit has a short shelf life due to rapid postharvest metabolism, quick softening, mechanical damage, and fungal decay. To improve the storability of strawberry fruit, we subjected harvested fruit to short-term exposure to 30% carbon dioxide (CO2) and used transcriptomic and metabolomic analyses to identify the cellular responses induced by this treatment. Fruit was stored at 10 °C for 10 d after a 3-h exposure to 30% CO2 (treatment) or ambient air (control) at 25 °C, respectively. The CO2 treatment reduced fruit decay and softening compared to the control throughout the 10-d storage period. Transcriptomic analyses revealed that expression levels of genes encoding cell wall-degrading enzymes (expansin, pectinesterase, and β-xylosidase) decreased in response to the CO2 treatment. Within 1 d after the CO2 treatment, the expression levels of genes encoding heat-shock proteins significantly increased. Metabolite profiling revealed that glucose, quinic acid, and succinic acid increased in responses to the CO2 treatment at 1 d. Transmission electron microscopy showed that disintegration of the middle lamella in the cell wall was inhibited by the CO2 treatment. Polyuronide (insoluble pectin) content in cell walls was 30% higher, on average, in the treated fruit than those in the control fruit during the 10-d storage period. These results indicate that a short-term treatment with 30% CO2 reduces the degradation of pectin in the cell wall by reducing the activity of cell wall-degrading enzymes and induces abiotic stress-response genes in harvested strawberry fruit. Our results improve our understanding of the molecular mechanisms behind CO2-responsive genes in strawberry fruit and provide insight into ways to improve the postharvest quality of strawberries.

Application of sodium silicate retards apple softening by suppressing the activity of enzymes related to cell wall degradation.

During the storage of apples, apple softening is one of the main problems. Sodium silicate has been used to enhance disease resistance and maintain quality of fruits. In the present study, apple fruit (cv. Golden delicious) were treated with 100 mmol L-1 sodium silicate for 10 min and stored at 20 °C to investigate its effects on weight loss, flesh firmness, and the activity of cell wall-degrading enzymes. The results indicated that 100 mmol L-1 of sodium silicate treatment delayed the increase of weight loss and decrease of the flesh firmness in apples. Sodium silicate treatment also suppressed the activity of polygalacturonic acid transeliminase and pectin methyltranseliminase, pectin methylgalacturonase, polygalacturonase, cellulase and β-galactosidase in the fruit. Delaying apple softening by sodium silicate treatment is closely related to the inhibition of the activity of cell wall-degrading enzymes and weight loss. © 2018 Society of Chemical Industry.

Population structure and virulence analysis of Alternaria carthami isolates of India using ISSR and SSR markers.

Alternaria leaf spot caused by Alternaria carthami is one of the most devastating diseases of safflower. Diversity among 95 isolates of A. carthami was determined using virulence assays, enzyme assays, dominant (ISSR) and co-dominant (SSR) markers. Collections and isolations were made from three major safflower producing states of India. The virulence assays categorised the population into four groups based on level of virulence. Estimation of activities of cell wall degrading enzymes (CWDE) yielded concurrent results to virulence assays with maximum CWDE activities in most virulent group. Eighteen ISSR primers were used and 23 polymorphic microsatellite markers were developed to assess the genetic diversity and determine the population structure of A. carthami. Analysis of ISSR profiles revealed high genetic diversity (Nei's Genetic diversity index; h = 0.36). Microsatellite markers produced a total of 56 alleles with an average of 2.43 alleles per microsatellite marker and Nei's genetic diversity index as h = 0.43. Unweighted Neighbor-joining and population structure analysis using both the marker systems differently arranged the isolates into three clusters. Distance analysis of the marker profiles provided no evidence for geographical clustering of isolates, indicating that isolates are randomly spread across India, signifying high potential of the fungus to adapt to diverse regions. Microsatellite markers clustered the isolates in consonance to the virulence groups in the dendrogram. This implies that the fungus has a high potential to adapt to resistant cultivars or fungicides. The information can aid in the breeding and deployment of A. carthami resistant varieties, and in early blight disease management in all safflower growing regions of the world.

Potential biocontrol efficacy of Trichoderma atroviride with cellulase expression regulator ace1 gene knock-out.

The biocontrol function of the repressor of cellulase expression I (ACE1) in Trichoderma atroviride was verified through constructing Δace1 mutant strain by Agrobacterium tumefaciens-mediated transformation. The activities of cell wall-degrading enzymes (cellulase, xylanase, chitinase, β-1,3-glucanase, and protease) in the supernatant of Δace1 mutant strain were distinctly higher than those of control strain, followed with the elevation of related genes transcript levels. Besides, the Δace1 mutant resulted in an elevating transcript level of xyr1, but no obvious change in the expression of cre1, which suggested that ACE1 was negative regulator of the xyr1 transcription, but not involved in cre1 transcription. On core polyketide synthases of four biosynthesis gene clusters for antibiotic secondary metabolites, only the transcription levels of encoding genes Try83179/TryH and Aza79482/AzaJ in Δace1 mutant strain were significantly higher than that in wild-type during antagonizing with pathogenic fungi Fusarium oxysporum and Rhizoctonia solani (with the inhibition rate of 30.7 and 19.8%, respectively). The biocontrol function of Δace1 mutant strain was remarkably enhanced. The results indicated that ACE1, indeed, acted as a repressor for cell wall-degrading enzymes and PKSs expression in T. atroviride, and the Δace1 mutant strain effectively made related enzymes activities improved with potential enhancement of biocontrol potency.

Action of selenium against Sclerotinia sclerotiorum: Damaging membrane system and interfering with metabolism

Selenium (Se) in soil is beneficial for environmental stress tolerance of plants, and it has widespread toxic effects on pathogens. Based on the fact that Se significantly inhibited the growth of Sclerotinia sclerotiorum, we set experiments with different concentrations of Se to investigate the action of Se against S. sclerotiorum in this study. The results showed that Se (>0.5 mg L−1) changed the morphology of S. sclerotiorum mycelia, and higher Se concentrations severely damaged mycelial structures. Fourier transform infrared spectroscopy (FTIR) analysis indicated that Se treatment induced the chemical composition of mycelia with much abundance of functional groups such as alcohols, ketones, ammonium and esters, and 0.5 mg L−1 Se maximized their concentrations. Under Se treatments, the electrical conductivity of mycelia increased in a time-dependent manner, and osmolyte concentrations of mycelia increased as well. Se supplementation significantly reduced polymethylgalacturonase (PMG) and carboxymethylcellulase (Cx) activities, which protecting plants from infection, and increased the energy expenditure in S. sclerotiorum. Combined action of Se damage on membrane system, osmoregulation, reduction of cell wall degrading enzymes activities and improvement of energy expenditure resulted in the inhibition of S. sclerotiorum growth. Findings in this study provided evidences for using Se as a potential fungicide to control S. sclerotiorum.

Phomopsis longanae Chi-Induced Changes in Activities of Cell Wall-Degrading Enzymes and Contents of Cell Wall Components in Pericarp of Harvested Longan Fruit and Its Relation to Disease Development.

The main goal of this study was to investigate the influences of Phomopsis longanae Chi infection on activities of cell wall-degrading enzymes (CWDEs), and contents of cell wall components in pericarp of harvested “Fuyan” longan (Dimocarpus longan Lour. cv. Fuyan) fruit and its relation to disease development. The results showed that, compared with the control samples, P. longanae-inoculated longans showed higher fruit disease index, lower content of pericarp cell wall materials (CWMs), as well as lower contents of pericarp cell wall components (chelate-soluble pectin (CSP), sodium carbonate-soluble pectin, hemicelluloses, and cellulose), but higher content of pericarp water-soluble pectin (WSP). In addition, the inoculation treatment with P. longanae significantly promoted the activities of CWDEs including pectinesterase, polygalacturonase, β-galactosidase, and cellulase. The results suggested that the P. longanae stimulated-disease development of harvested longans was due to increase in activities of pericarp CWDEs, which might accelerate the disassembly of pericarp cell wall components. In turn, resulting in the degradation of pericarp cell wall, reduction of pericarp mechanical strength, and subsequently leading to the breakdown of longan pericarp tissues. Eventually resulting in development of disease development and fruit decay in harvested longans during storage at 28°C.

The Kynurenine 3-Monooxygenase Encoding Gene, BcKMO, Is Involved in the Growth, Development, and Pathogenicity of Botrytis cinerea.

A pathogenic mutant, BCG183, was obtained by screening the T-DNA insertion library of Botrytis cinerea. A novel pathogenicity-related gene BcKMO, which encodes kynurenine 3-monooxygenase (KMO), was isolated and identified via thermal asymmetric interlaced PCR, bioinformatics analyses, and KMO activity measurement. The mutant BCG183 grew slowly, did not produce conidia and sclerotia, had slender hyphae, and presented enhanced pathogenicity. The phenotype and pathogenicity of the BcKMO-complementing mutant (BCG183/BcKMO) were similar to those of the wild-type (WT) strain. The activities of polymethylgalacturonase, polygalacturonase, and toxins were significantly higher, whereas acid production was significantly decreased in the mutant BCG183, when compared with those in the WT and BCG183/BcKMO. Moreover, the sensitivity of mutant BCG183 to NaCl and KCl was remarkably increased, whereas that to fluconazole, Congo Red, menadione, H2O2, and SQ22536 and U0126 [cAMP-dependent protein kinase (cAMP) and mitogen-activated protein kinase (MAPK) signaling pathways inhibitors, respectively] were significantly decreased compared with the other strains. Furthermore, the key genes involved in the cAMP and MAPK signaling pathways, Pka1, Pka2, PkaR, Bcg2, Bcg3, bmp1, and bmp3, were significantly upregulated or downregulated in the mutant BCG183. BcKMO expression levels were also upregulated or downregulated in the RNAi mutants of the key genes involved in the cAMP and MAPK signaling pathways. These findings indicated that BcKMO positively regulates growth and development, but negatively regulates pathogenicity of B. cinerea. Furthermore, BcKMO was found to be involved in controlling cell wall degrading enzymes activity, toxins activity, acid production, and cell wall integrity, and participate in cAMP and MAPK signaling pathways of B. cinerea.

Effects of paper containing 1-MCP postharvest treatment on the disassembly of cell wall polysaccharides and softening in Younai plum fruit during storage

Disassembly of cell wall polysaccharides accompanied with softening is very common in harvested fruits. To develop a facile postharvest approach, which can be used at ambient temperature, for suppressing softening and maintaining higher nutritive cell wall polysaccharides of Younai plums, influences of paper containing 1-methylcyclopropene (1-MCP) on firmness, activities of cell wall-degrading enzymes, and contents of cell wall polysaccharide in Younai plums during storage at 25 ± 1 °C were investigated. As compared to the control plums, 1.2 μL·L−1 1-MCP-treated plums exhibited higher firmness, lower activities of cell wall-degrading enzymes (pectinesterase, polygalacturonase, cellulase and β-galactosidase), higher contents of cell wall polysaccharides (sodium carbonate-soluble pectin, chelate-soluble pectin, cellulose, and hemicelluloses), and lower content of water-soluble pectin. The results suggested that paper containing 1-MCP, which was convenient to apply under ambient temperature, could significantly inhibit activities of cell wall degrading-enzymes and decrease disassembly of cell wall polysaccharides, and subsequently retard softening in Younai plums.

Impact of chitosan/PVA with salicylic acid, cell wall degrading enzyme activities and berries shattering of ‘Thompson seedless’ grape vines during shelf life

Abstract The viability of chitosan/polyvinyl alcohol (CS/PVA), mixed with salicylic acid (SA) at various doses (0, 1, and 2 mM) of ‘Thompson seedless’ grapes coating to delay fruit clusters aging during shelf life. Bunches were covered with various CS/PVA-SA treatments and placed at room temperature (26 ± 1 °C and relative humidity 46 ± 1% RH) for five days and fruit samples were picked each day. Physical and chemical of quality attributes parameters were determined to assess the impact CS/PVA-SA coating on berry quality during shelf life period (days). The physical parameters, for example, water loss, rachis browning index record, berry shattering rate, berry color (hue angle), berry separation force and firmness. Be that as it may, the chemical characteristics, for example, total soluble solids content (SSC%), titratable acidy (TA%), such phenolic compounds (add up to phenol and flavonoid content), polyphenol oxidase (PPO) beside the cell wall degrading enzyme activities: cellulase (CEL, EC:3.2.1.4), polygalacturonase (PG, EC:3.2.1.15), xylanase (XYL, EC:3.2.1.8) and cell plasma membrane permeability rate were evaluated. CS/PVA-SA 2 mM treatment reduce remarkable water loss and berry shattering and postponed the subjective change in berry color hue angle and controlled on SSC%, TA%, and SSC/TA ratio. Additionally, the progressions in TP, FL substance, and PPO activity are maintained and reduced during shelf life. Likewise, the cell wall enzyme activities (CEL, PG, and XYL) were inhibited compared with different treatments. Relationship investigation confirmed that numerous quality attributes are independent. Postharvest of CS/PVA-SA 2 mM postponed fruit clusters ripening, kept berry quality, and expanded the shelf life of 'Thompson seedless' grapes.

Improving fruit cluster quality attributes of ‘Flame Seedless’ grapes using preharvest application of ascorbic and salicylic acid

Abstract The effectiveness of ascorbic acid (AA) and salicylic acid (SA) at different concentrations (AA, 3 and 6 mM; and SA, 2 and 4 mM) of ‘Flame Seedless’ grapes spraying to improve fruit cluster quality or delay cluster deterioration during shelf life. Vines were received different AA and SA concentration tow time at veraison stages and 14 days before harvesting time. Fruit clusters were harvested when total soluble solid content reaches to 16%, then fruit clusters were stored at ambient air for four-day shelf life (28 ± 1 °C and air humidity 48 ± 2 during shelf life period). Samples were picked every day to measure physical and chemical attribute quality properties such as water loss, soluble solid content (SSC%), titrable acidity (TA%), berry firmness and separation force, rachis browning index, and berry color hue angle (h°). Riches browning parameters such as total phenol content (TP), flavonoids (TF), and poly phenoloxidase (PPO) activity. While cell wall degrading enzyme activities like polygalacturonase (PG; EC: 3.2.1.15), xylanase (XYL; EC:3.2.1.8), and cellulase (CEL; EC: 3.2.1.4). The AA 6 mM + SA 4 mM treatment was delayed fruit cluster deterioration during shelf life by minimizing cell wall degradation enzyme activities, maintaining the phenolic compounds. So, less rachis browning symptoms, and preserving the chemical and physical of a fruit cluster quality attribute during shelf life.

The effect of ethylene absorbent treatment on the softening of blueberry fruit

The potential of ethylene absorbent (EA) to delay softening of ‘Lanfeng’ blueberry (Vaccinium spp.) fruit in conjunction with cold storage was evaluated. The fruit quality was evaluated after 60 days of storage at 0 °C again kept at 20 °C, with or without EA. Changes in quality attributes and ethylene biosynthesis and fruit softening indicators were assessed. The results indicated that EA treatment inhibited fruit softening, reduced weight loss and decay, and prevented the loss of total phenolic content. It also decreased the fruit ethylene production by inhibiting 1-aminocyclopropane-1-carboxylic acid oxidase and 1-aminocyclopropane-1- carboxylic acid synthase activities, whilst maintaining firmness by hampering cell wall-degrading enzyme activities, especially after more than 30 days of cold storage. In conclusion, EA treatment can inhibit the softening of harvested blueberry fruit during storage at 0 °C and shelf life after cold storage. After being refrigerated for more than 30 days at 0 °C, the EA has a good effect on blueberries storage.

High temperature enhances the ability of Trichoderma asperellum to infect Pleurotus ostreatus mycelia.

Trichoderma asperellum is one of the species which can be isolated from contaminated Pleurotus ostreatus cultivation substrate with green mold disease. This study focused on the relationship between high temperature and infectivity of T. asperellum to P. ostreatus. Antagonism experiments between T. asperellum and P. ostreatus mycelia revealed that high temperature-treated P. ostreatus mycelia were more easily infected by T. asperellum and covered by conidia. Microscopic observation also showed that P. ostreatus mycelia treated with high temperature could adsorb more T. asperellum conidia. Furthermore, conidia obtained from T. asperellum mycelia grown at 36°C featured higher germination rate compared with that incubated at 28°C. High temperature-treated T. asperellum mycelia can produce conidia in shorter periods, and T. asperellum mycelia were less sensitive to high temperature than P. ostreatus. Deactivated P. ostreatus mycelia can induce T. asperellum cell wall-degrading enzymes (CWDEs) and P. ostreatus mycelia subjected to high temperature showed induced CWDEs more effective than those incubated at 28°C. Moreover, T. asperellum showed higher CWDEs activity at high temperature. In dual cultures, hydrogen peroxide (H2O2) increased after 36°C, and high concentration of H2O2 could significantly inhibit the growth of P. ostreatus mycelia. In summary, our findings indicated for the first time that high temperature can induce a series of mechanisms to enhance infection abilities of T. asperellum to P. ostreatus mycelia and to cause Pleurotus green mold disease.

Cell wall degrading enzymes activity is altered by high carbon dioxide treatment in postharvest ‘Mihong’ peach fruit

Peach fruits are ripen rapidly after harvest, especially at room temperature. It is difficult to handle because they are scars easily and easy to decay. It is generally acknowledged that modified atmospheres with elevated CO2 inhibits fruit ripening by reduces ethylene biosynthesis. This study was to investigate the affect of high concentration CO2 treatment on ethylene biosynthesis, ACO and softening-related enzymatic activity. Peaches were treated with high CO2 (90 pKa) at the time of sealing in the plastic chambers and then the chambers were ventilated after 3 and 12h. Fruits treated with 90kPa CO2 for 3 or 12h exhibited a significant delay in firmness when compared to control fruits. In addition to fruit softening inhibition, the decay rate was also inhibited by CO2 treatment. Four days after storage at room temperature, CO2 untreated peaches had decay rate of 14%, whereas the decay rate was 6% and 3.5% in fruits treated with 90kPa CO2 for 3 and 12h. The ACO was inhibited by CO2 and the inhibition was enhanced with higher CO2 exposure time. Also High CO2 treatment inhibited the activities of softening-related enzyme such as PG, β-Galactosidase, PEL promoted by ethylene biosynthesis in peaches. These results indicate that increasing levels of CO2 inhibit ethylene biosynthesis by reducing the ACO activity of peach fruits. Results suggest that the effect of CO2 on ethylene production in ‘Mihong’ peach fruit depended on exposure time and CO2 level. Also a short-term treatment with a high concentration of CO2 may be used to reduce ethylene biosynthesis, which may be induced after postharvest of fruit.

Enhanced virulence of Fusarium species associated with spear rot of oil palm following recovery from osmotic stress

ABSTRACTFusarium spp., which are common inhabitants of oil palm leaves, are weak pathogens of common spear rot (CSR). We investigated the influence of osmotic stress on the growth, virulence, and activity of cell wall-degrading enzymes of CSR fungi, using potato dextrose agar (PDA) supplemented with KCl or sucrose (hyperosmotic medium). Hyperosmotic stress significantly inhibited mycelial growth, but growth rapidly recovered when mycelia were transferred to control medium. When inoculated into oil palm spear leaflets, Fusarium sp., and F. incarnatum precultured on 1.0 and 1.5 M KCl-hyperosmotic medium induced lesions that were two to four times larger than those in non-stressed cultures, suggesting enhanced virulence of the weak pathogens. Lesion size was not greatly affected in hyperosmotic cultures of moderately virulent F. sacchari. No activity of pectin lyase was detected in liquid cultures of the Fusarium isolates. All isolates except F. incarnatum BT48 secreted polygalacturonase (PG), which was active in both liquid cultures and inoculated leaves. Significantly increased PG activity (5–32-fold) was observed on leaves inoculated with hyperosmotic cultures of Fusarium sp. and F. sacchari. These findings suggest that Fusarium sp., F. incarnatum, and F. sacchari exhibit an adaptive physiological plasticity to hyperosmotic stress that results in enhanced virulence.

The gene task1 is involved in morphological development, mycoparasitism and antibiosis of Trichoderma asperellum

ABSTRACTCompetitive activity, mycoparasitism and antibiosis of Trichoderma asperellum are considered essential mechanisms in its suppressive activity against soil-borne plant pathogens. The role of the mitogen-activated protein kinase encoding gene task1 on morphological development, mycoparasitic interaction and the production of cell wall degrading enzymes and secondary metabolites were examined in T. asperellum. The Δtask1 mutant had altered growth morphology, lost its ability to parasitise plant pathogens and showed increased expression of several cell wall degrading enzymes during confrontation with Rhizoctonia solani. T. asperellum task1 expression was negatively correlated with cell wall degrading enzyme activities during inducing experiments using pathogen cell wall compounds. In antibiosis assays, task1 deletion caused increased output of 6-pentyl-α-pyrone and inhibition of pathogen growth.

Molecular Identification, Enzyme Assay, and Metabolic Profiling of Trichoderma spp.

The goal of this study was to identify and characterize selected Trichoderma isolates by metabolic profiling and enzyme assay for evaluation of their potential as biocontrol agents against plant pathogens. Trichoderma isolates were obtained from the Rural Development Administration Genebank Information Center (Wanju, Republic of Korea). Eleven Trichoderma isolates were re-identified using ribosomal DNA internal transcribed spacer (ITS) regions. ITS sequence results showed new identification of Trichoderma isolates. In addition, metabolic profiling of the ethyl acetate extracts of the liquid cultures of five Trichoderma isolates that showed the best anti-Phytophthora activities was conducted using gas chromatography-mass spectrometry. Metabolic profiling revealed that Trichoderma isolates shared common metabolites with well-known antifungal activities. Enzyme assays indicated strong cell walldegrading enzyme activities of Trichoderma isolates. Overall, our results indicated that the selected Trichoderma isolates have great potential for use as biocontrol agents against plant pathogens.