Isolation and characterization of Trichoderma harzianum L-methioninase with promising a powerful anticancer.

Bioactive components derived from medicinal herbs have recently acquired popularity due to their efficacy in treating various ailments, including cancer and infectious diseases. In this study, the anticancer enzyme, L-methioninase isolated from medicinal plants endophytic fungi, then evaluated its promising therapeutic agents against different types of human cancers. L methionine was purified using column chromatography with the stationary phase of Sephadex G-200 with 6.6-fold purification, which increased the specific activity of 71.3 U/mg of protein with a recovery rate of 48.2 %. On the SDS-PAGE chromatogram, the apparent molecular mass of the isolated enzyme was 48 kDa, and its highest activity was observed at pH 8 and 35 °C. The enzyme was catalytically stable within the pH range of 6.0-9.0 and below 40 °C. This study demonstrates that isolated L-methioninase is particularly efficient against tumour cell lines in vitro. The crude and purified L-methioninase inhibited 60 and 80 % of the growth of the breast cancer cell line (MCF-7), respectively, with an estimated IC50 = 12.6 µg/ml (crude) and IC50 = 5.0 µg/ml for purified L-methioninase from isolate 8 with accession no MZ675362. Because of this, pure L-methioninase has better catalytic characteristics and significant thermal stability, which could be used as a cancer-fighting substance than the enzyme purified from other sources.

Biocontrol potential of the endophytic Epicoccum nigrum HE20 against stripe rust of wheat.

Biological control using endophytic microorganisms represents an eco-friendly and effective alternative to the health-hazardous chemical fungicides used to control devastating plant diseases such as stripe rust in wheat. In this study, the inhibitory potential of the endophytic Epicoccum nigrum HE20, isolated from a healthy wheat plant, was screened against uredospores germination in vitro. A high suppression (96%) in the germination of the uredospores was recorded. GC-MS analysis of the culture filtrate of E. nigrum HE20 showed a production of various secondary metabolites with an antifungal background such as butyric acid, α-linolenic acid, hexanoic acid, lactic acid, 10,12-Tricosadiynoic acid, and pentadecanoic acid. Results from the greenhouse experiment revealed that the application of E. nigrum HE20 suspension led to a reduction in the disease severity by 87.5%, compared with the untreated-infected plants. Real-time PCR results exhibited an overexpression in three defensive genes (JERF3, GLU, and PR1) in the infected wheat plants, in response to the application of E. nigrum HE20, recorded 8-, 15.8-, and 3.5-fold, respectively. In addition, an increment in the phenolic content, activity of POD, PPO, and CAT, and a reduction in the lipid peroxidation were recorded due to the endophyte application. Transmission electron microscopic observations indicated mitigation of the pathogen in wheat cells after the treatment with E. nigrum HE20 metabolite. Furthermore, a growth-promoting effect was also observed due to E. nigrum HE20 application, as well as an increment in the total photosynthetic pigments in wheat leaves. Based on these results, it can be concluded that E. nigrum HE20 is a probable efficient bioagent against stripe rust in wheat. However, its field evaluation is highly necessary in the future studies.

Mycorrhizal colonization and Streptomyces viridosporus HH1 synergistically up-regulate the polyphenol biosynthesis genes in wheat against stripe rust.

BACKGROUND: Stripe rust is considered one of the most devastating diseases of wheat all over the world, resulting in a high loss in its production. In this study, time-course changes in expression of the polyphenol biosynthesis pathways genes in wheat against stripe rust were investigated. The defense mechanisms triggered by mycorrhizal colonization and/or spraying with Streptomyces viridosporus HH1 against this disease were also investigated. RESULTS: Results obtained revealed that C3H, which is considered the key gene in lignin biosynthesis, was the most expressed gene. Furthermore, most of the chlorogenic acid and flavonoid biosynthesis genes were also overexpressed. Volcano plots of the studied genes reveal that the dual treatment led to a high significant overexpression of 10 out of the 13 studied genes. Heatmap of these genes showed that the most frequent expressed gene in response to all applied treatments along the study period was DFR, the key gene in the biosynthesis of anthocyanidins. Gene co-expression network of the studied genes showed that HQT was the most central gene with respect to the other genes, followed by AN2 and DFR, respectively. Accumulation of different flavonoids and phenolic acids were detected in response to the dual treatment, in particular, cinnamic acid, coumarin, and esculetin, which recorded the highest elevation level recording 1000, 488.23, and 329.5% respectively. Furthermore, results from the greenhouse experiment showed that application of the dual treatment led to an 82.8% reduction in the disease severity, compared with the control treatment. CONCLUSIONS: We can conclude that the biosynthesis of lignin, chlorogenic acid, and flavonoids contributed to the synergistic triggering effect of the dual treatment on wheat resistance to stripe rust.

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

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

Synergism between Streptomyces viridosporus HH1 and Rhizophagus irregularis Effectively Induces Defense Responses to Fusarium Wilt of Pea and Improves Plant Growth and Yield.

Fusarium wilt is a detrimental disease of pea crop, resulting in severe damage and a reduction in its yield. Developing synergistically enhanced bioagents for disease management and growth promotion is pivotal for food safety, security, and sustainability. In this study, biocontrol potential of treating pea plants with Streptomycesviridosporus HH1 and/or their colonization with Rhizophagusirregularis against infection with Fusarium wilt was investigated. Impacts on the expression profiles of defense-related genes, biochemical, and ultrastructural levels, as well as the growth and yield of pea plants in response to these treatments, were also investigated. Data obtained indicated the antifungal activity of S. viridosporus HH1 against F. oxysporum f.sp. pisi in vitro. Furthermore, the GC-MS analysis revealed production of different bioactive compounds by S. viridosporus HH1, including 2,3-butanediol, thioglycolic acid, and phthalic acid. The results from the greenhouse experiment exhibited a synergistic biocontrol activity, resulting in a 77% reduction in disease severity in pea plants treated with S. viridosporus HH1 and colonized with R. irregularis. In this regard, this dual treatment overexpressed the responsive factor JERF3 (5.6-fold) and the defense-related genes ß-1,3-glucanase (8.2-fold) and the pathogenesis-related protein 1 (14.5-fold), enhanced the total phenolic content (99.5%), induced the antioxidant activity of peroxidase (64.3%) and polyphenol oxidase (31.6%) enzymes in pea plants, reduced the antioxidant stress, and improved their hypersensitivity at the ultrastructural level in response to the Fusarium wilt pathogen. Moreover, a synergistic growth-promoting effect was also recorded in pea plants in response to this dual treatment. In this regard, due to this dual treatment, elevated levels of photosynthetic pigments and improved growth parameters were observed in pea leaves, leading to an increment in the yield (113%). In addition, application of S. viridosporus enhanced the colonization levels with R. irregularis in pea roots. Based on the obtained data, we can conclude that treating pea plants with S. viridosporus HH1 and colonization with R. irregularis have synergistic biocontrol activity and growth-promoting effects on pea plants against Fusarium wilt. Despite its eco-safety and effectiveness, a field evaluation of this treatment before a use recommendation is quite necessary.

Ascophyllum nodosum Extract and Mycorrhizal Colonization Synergistically Trigger Immune Responses in Pea Plants against Rhizoctonia Root Rot, and Enhance Plant Growth and Productivity.

Rhizoctonia root rot is one of the most destructive diseases affecting pea crops, resulting in up to 75% loss. In this study, the biocontrol activity of seaweed (Ascophyllum nodosum) extract at 1, 2, and 3% and/or mycorrhization of pea roots was investigated against Rhizoctonia root rot under greenhouse conditions. In addition, their effects on the transcriptional, physiological, ultrastructural, and growth status of pea plants were also studied. The results showed that the mycorrhizal colonization of pea roots and the application of the seaweed extract at 3% synergistically overexpressed the responsive factor (JERF3) recording 18.2-fold, and the defense-related genes peroxidase (23.2-fold) and chitinase II (31.8-fold). In addition, this treatment improved the activity of the antioxidant enzymes POD and PPO, increased the phenolic content in pea roots, and triggered multiple hypersensitivity reactions at the ultrastructural level of the cell, leading to a 73.1% reduction in disease severity. Moreover, a synergistic growth-promoting effect on pea plants was also observed. The photosynthetic pigments in pea leaves were enhanced in response to this dual treatment, which significantly improved their yield (24 g/plant). The inducing effect of mycorrhizal colonization on plant resistance and growth has been extensively studied. However, developing improved and synergistically acting biological agents for plant disease control and growth promotion as alternatives to the chemical fungicides is crucial for safety and food security. Based on these results, it can be concluded that the mycorrhizal colonization of pea roots and soaking their seeds in the A. nodosum extract at 3% have a promising and improved biocontrol activity against R. solani, and a growth-promoting effect on pea plants. However, field applications should be evaluated prior to any use recommendations.

Effects of Mycorrhizal Colonization on Transcriptional Expression of the Responsive Factor JERF3 and Stress-Responsive Genes in Banana Plantlets in Response to Combined Biotic and Abiotic Stresses.

Banana plants (Musa acuminata L.) are exposed to various biotic and abiotic stresses that affect their production worldwide. Banana plants respond to these stresses, but their responses to combined stresses are unique and differ from those to various individual stresses. This study reported the effects of the mycorrhizal colonization of banana roots and/or infection with root rot on the transcriptional expression of the responsive factor JERF3 and stress-responsive genes (POD, PR1, CHI, and GLU) under different salinity levels. Different transcriptional levels were recorded in response to the individual, dual, or triple treatments. All the applied biotic and abiotic stresses triggered the transcriptional expression of the tested genes when individually applied, but they showed different influences varying from synergistic to antagonistic when applied in combinations. The salinity stress had the strongest effect when applied in combination with the biotic stress and/or mycorrhizal colonization, especially at high concentrations. Moreover, the salinity level differentially affects the banana responses under combined stresses and/or mycorrhizal colonization in addition, the mycorrhizal colonization of banana plantlets improved their growth, photosynthesis, and nutrient uptake, as well as greatly alleviated the detrimental effects of salt and infection stresses. In general, the obtained results indicated that the responses of banana plantlets under the combined stresses are more complicated and differed from those under the individual stresses depending on the crosstalks between the signaling pathways.