Bio-guided isolation of potential anti-inflammatory constituents of some endophytes isolated from the leaves of ground cherry (Physalis pruinosa L.) via ex-vivo and in-silico studies.

BACKGROUND: Due to the extensive potential of previously studied endophytes in addition to plants belonging to genus Physalis as a source of anti-inflammatory constituents, the present study aimed at isolation for the first time some endophytic fungi from the medicinal plant Physalis pruinosa. METHODS: The endophytic fungi were isolated from the fresh leaves of P. pruinosa then purified and identified by both morphological and molecular methods. Comparative evaluation of the cytotoxic and ex vivo anti-inflammatory activity in addition to gene expression of the three pro-inflammatory indicators (TNF-α, IL-1ß and INF-γ) was performed in WBCs treated with lipopolysaccharide (LPS) for the identified endophytes, isolated compounds and the standard anti-inflammatory drug (piroxicam). For prediction of the binding mode of the top-scoring constituents-targets complexes, the Schrödinger Maestro 11.8 package (LLC, New York, NY) was employed in the docking study. RESULTS: A total of 50 endophytic fungal isolates were separated from P. pruinosa leaves. Selection of six representative isolates was performed for further bioactivity screening based on their morphological characters, which were then identified as Stemphylium simmonsii MN401378, Stemphylium sp. MT084051, Alternaria infectoria MT573465, Alternaria alternata MZ066724, Alternaria alternata MN615420 and Fusarium equiseti MK968015. It could be observed that A. alternata MN615420 extract was the most potent anti-inflammatory candidate with a significant downregulation of TNF-α. Moreover, six secondary metabolites, alternariol monomethyl ether (1), 3'-hydroxyalternariol monomethyl ether (2), alternariol (3), α-acetylorcinol (4), tenuazonic acid (5) and allo-tenuazonic acid (6) were isolated from the most potent candidate (A. alternata MN615420). Among the tested isolated compounds, 3'-hydroxyalternariol monomethyl ether showed the highest anti-inflammatory potential with the most considerable reductions in the level of INF-γ and IL-1ß. Meanwhile, alternariol monomethyl ether was the most potent TNF-α inhibitor. The energy values for the protein (IL-1ß, TNF-α and INF-γ)-ligand interaction for the best conformation of the isolated compounds were estimated using molecular docking analysis. CONCLUSIONS: The results obtained suggested alternariol derivatives may serve as naturally occurring potent anti-inflammatory candidates. This study opens new avenues for the design and development of innovative anti-inflammatory drugs that specifically target INF-γ, IL-1ß and INF-γ.

Production, characterization and bio-emulsifying application of exopolysaccharides from Rhodotorula mucilaginosa YMM19.

Microbial exopolysaccharides (EPS) are high molecular weight polymers having different sugar residues. EPS have potential applications in different fields, such as medicine, food and environment. Therefore, there is a growing interest in production, characterization and application of EPS from different microorganisms. The present study designed to investigate the production and characterization of EPS from Rhodotorula mucilaginosa YMM19 isolated from Morus nigra L. fruits as well as to examine their potential emulsifying properties. Effect of NaCl concentration, incubation period and pH on the production of EPS was studied. The maximum EPS production by yeast was achieved at 10% NaCl (9741.84 mg/l). The best incubation time for production of EPS was 5 days. Production of EPS decreased under neutral condition and increased at acidic and alkaline condition. The structural feature of EPS was examined by FT-IR and NMR spectral analysis and confirmed the presence of glucose, glucopyranose and galactose. The isolated EPS showed higher emulsification capacity with emulsification activity of 71% and emulsifying index of 60%. The EPS gave strong emulsification for farnesol and was more effective than sodium dodecyl sulphate, a reference emulsifier, in enhancing the herbicidal activity of farnesol against Melilotus indicus under greenhouse condition. The results suggest that the EPS produced by YMM19 strain has a potential to be used as emulsifying agent in pesticide formulations.

Antibacterial and herbicidal properties of secondary metabolites from fungi.

Twenty-eight compounds were isolated from endophytic, soil and marine fungi and their structures were elucidated through spectroscopic methods. The isolated compounds were tested for their antibacterial and herbicidal activities against phytopathogenic bacteria and barnyard grass weed for the first time. Methyleurotinone (14) was the most potent compound against Pectobacterium carotovorum subsp. carotovorum, Pseudomonas syringae pv. syringae, Rhizobium radiobacter and Ralstonia solanacearum with minimum inhibitory concentration (MIC) values of 31.3, 125, 31.3 and 125 mg/L, respectively. Compounds 13-15 were highly effective in reducing the development of potato tuber soft rot disease caused by P. carotovorum subsp. carotovorum. Furthermore, twelve of the tested compounds induced a significant reduction in seed germination of Echinochloa crus-galli at 2 mM with compounds 8 and 26 causing complete inhibition of seed germination. Also, compounds 4, 22, 5, 8, 18 and 25-27 induced remarkable reduction of root and shoot growth of E. crus-galli at 2 mM.

Isolation and Identification of Plant Growth-Promoting Bacteria Highly Effective in Suppressing Root Rot in Fava Beans.

In this study, 18 plant growth-promoting bacterial (PGPB) strains were isolated from the rhizosphere of both Red silk-cotton tree (Bombax malabaricum) and Chinese banyan (Ficus retusa). Culture morphology was observed and genotypic characterization was accomplished by sequencing partial 16S rRNA gene. Plant growth promotion traits and antagonistic activities of the strains against phytopathogenic fungi were evaluated. Among all strains, Bacillus thuringiensis (MN419208) exert the highest indole acetic acid (38 µg/ml), produced exopolysaccharides (587.2 µg/ml), and fixed nitrogen which in turn increased both fresh and dry weights of bean plants by 41.5% and 18.8%, respectively. In another greenhouse experiment studying the antifungal activities of seven strains and their co-culture against Rhizoctonia solani (LN735538), B. sonorensis MN419205, B. wiedmannii MN419207, B. subtilis MN419218, and the mixture of (MN419207) and (MN419208) reduced total damping off from 81.7% in control to 30%, 35%, 35%, and 38.3%, respectively, and reduced disease severity index from 33.3% to 20.5%, 22.5%, 14.2%, and 19.3% as well. Our data indicate that these strains are effective in promoting plant growth and in inhibiting R. solani infection nonetheless field experiments are needed to examine their effectiveness as a viable alternative to chemical fertilizers and conventional pesticides.

Bioherbicidal activity of terpenes and phenylpropenes against Echinochloa crus-galli.

This study was undertaken to evaluate the herbicidal activity of twelve natural compounds belonging to monoterpenes, phenylpropenes, and sesquiterpenes against Echinochloa crus-galli under laboratory and glasshouse conditions. Experiments were conducted to determine the impact of different concentrations (0.5, 1, 2, 4 and 8 mM) of these compounds on the seed germination and root and shoot growth of barnyard grass. trans-Cinnamaldehyde, eugenol, and thymol caused the highest impact on barnyard grass reducing its seed germination and shoot growth. p-Cymene (EC50 = 0.22 mM) and trans-cinnamaldehyde (EC50 = 0.34 mM) were the most potent compounds in limiting the root growth of the E. crus-galli. In a post-emergent experiment, thymol, trans-cinnamaldehyde, eugenol, farnesol, and nerolidol significantly reduced the shoot growth, fresh and dry weight of two-leaf stage barnyard grass after 2 days of the foliar treatment with the concentrations of 0.5, 1.0 and 2.0%. These compounds induced severe visible injury symptoms where trans-cinnamaldehyde, eugenol, farnesol and nerolidol showed a complete weed control at 1.0 and 2.0%. These compounds were successfully formulated as emulsifiable concentrates and showed higher herbicidal activities against barnyard grass. Altogether, our data showed that trans-cinnamaldehyde, eugenol, thymol, farnesol, and nerolidol can be developed as novel bioherbicides for managing E. crus-galli.

Insecticidal potential and repellent and biochemical effects of phenylpropenes and monoterpenes on the red flour beetle, Tribolium castaneum Herbst.

The main objectives of the present study are to introduce new, ecologically safe, and natural compounds for controlling red flour beetle, Tribolium castaneum, and to understand the possible mode of action of these compounds. Therefore, the insecticidal and repellent activities of two phenylpropenes and six monoterpenes have been evaluated against the adults of T. castaneum. The inhibitory effects of these compounds on the activity of adenosine triphosphatases (ATPases) and acetylcholinesterase (AChE) were also tested. In fumigant toxicity assay, (-)-terpinen-4-ol (LC50 = 20.47 µl/l air) and α-terpinene (LC50 = 23.70 µl/l air) exhibited the highest toxicity without significant differences between them. Moreover, (-)-menthone and p-cymene showed strong toxicity, while (-)-citronellal, trans-cinnamaldehde, and eugenol were not active. In contact toxicity assay, the two phenylpropenes, trans-cinnamaldehde and eugenol, had the highest toxicity with same LC50 value of 0.02 mg/cm2. The monoterpenes and phenylpropenes showed pronounced repellent effect on the adults of T. castaneum at 0.001 mg/cm2 with (-)-menthone, trans-cinnamaldehyde, and α-terpinene being the most effective after 2 h of exposure. Repellent activity depended on compound, exposure time, and concentration. On the other hand, the tested compounds exhibited strong inhibition of ATPases form the larvae of T. castaneum as their IC50 values ranged between 1.74 and 19.99 mM. In addition, (-)-citronellal (IC50 = 9.82 mM) and trans-cinnamaldehde (IC50 = 23.93 mM) caused the highest inhibitory effect on AChE, while α-pinene (IC50 = 53.86) and p-cymene (IC50 = 68.97 mM) showed the weakest inhibitory effect. The results indicated that the tested phenylpropenes and monoterpenes had the potential to be developed as natural insecticides and repellents for controlling T. castaneum.

Phragmalin limonoids from Chukrasia tabularis.

Six phragmalin limonoids, named tabulalin and tabulalides A-E, were isolated from the root bark of Chukrasia tabularis (Meliaceae). Their structures were determined by spectroscopic methods, and their antifeedant properties evaluated.