Antioxidant, anticancer and antimicrobial activity of secondary metabolites from Fusarium species

The genus Lactobacillus has recently been the focal point of researchers due to their ability to produce secondary metabolites with antimicrobial effects. The aim of this study was to identify vaginal Lactobacillus sp. by analyzing 16S rRNA gene sequence, to investigate into antimicrobial activities of secondary metabolites produced by these isolates and to determine the quantities of lactic acid, acetic acid and hydrogen peroxide in secondary metabolites using High-Performance Liquid Chromatography. Antimicrobial activities of secondary metabolites were investigated against test microorganisms using Agar Well Diffusion and Agar Spot Methods. According to the results, 18 L. crispatus, 17 L. gasseri, 5 L. jensenii, 4 L. vaginalis, 3 L. fermentum, 2 L. coleohominis, 1 L. saerimneri, 1 L. reuteri, 1 L. johnsonii and 1 L. helveticus were identified. Isolates were frequently found to be effective against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Klebsiella pneumoniae RSKK 578 and Bacillus subtilis ATCC 6633. None of the isolates showed activity against Staphylococcus aureus ATCC 43300, S. epidermidis ATCC 12228, S. epidermidis ATCC 35984 and Candida albicans ATCC 10231. Secondary metabolites of all tested isolates contain hydrogen peroxide between 7.306 and 0.33 µg/µL range. It was found that the secondary metabolites of some isolates contained both acetic and lactic acid, while some of them contained either acetic or lactic acid.

Antimicrobial activity of secondary metabolites of fungi isolated from leaves of Bush mango against some selected microorganisms (Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus subtilis, Penicillium chrysogenum and Aspergillus fumigates) was carried out in this study. Agar well diffusion method was used to study the antimicrobial activity of the secondary metabolites and each of the secondary metabolites showed antagonist activity against the test organisms; two gram positive bacteria (Staphylococcus aureus and Bacillus subtilis), two gram negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and two fungi (Penicillium chrysogenum and Aspergillus fumigatus). Findings from the research indicates that the studies could also pave a way for new therapeutic agents which can be used as potential drugs against the selected microorganisms or its infections.

Background: The increasing interest in powerful biological activity of secondary metabolites outlined the necessity of determining their contents in medicinal plants. Aim: The present study intended to find out the total content of some important phytochemical constituents like phenols, flavonoids, tannins, non‑tannin phenols, condensed tannins and hydrolysable tannins in different parts of Svensonia hyderobadensis. Materials and Methods: By using Spectrophotometric methods. Results: The results showed that the plant is rich in phenols. Among the selected parts of the plant, the root showed maximum content of selected secondary metabolites, followed by leaf and stem. Roots are found to be rich in phenols [160.87 mg tannic acid equivalent (TAE)/g] and tannins (145.99 mg TAE/g); stem and leaf are rich in flavonoids (67.65 mg TAE/g and 133.95 mg TAE/g) followed by selected secondary metabolites. Conclusion: The findings of the present study will be helpful to phytochemists, pharmacologists and pharmaceutical industries. Key words: Quantitative analysis, secondary metabolites, Svensonia hyderobadensis

A PCR-denaturing gradient gel electrophoresis approach to assess Fusarium diversity in asparagus

Liupao tea is a postfermented dark tea with hypolipidemic activity. Research on the active substances in Liupao tea has primarily focused on those derived from the tea itself, overlooking the secondary metabolites produced by its predominant fungus, Euirotium cristatum. In this study, E. cristatum CPCC 401251, the predominant strain found in Liupao tea under investigation, was isolated and analyzed. A total of 19 representative metabolites, including prenylbenzaldehydes, diketopiperazines, and anthraquinones, were obtained from its culture. Subsequent analysis revealed the presence of multiple secondary metabolites of E. cristatum CPCC 401251 in Liupao tea. The 19 compounds significantly reduced the lipid content in free fatty acid (FFA)-stimulated hepatocyte AML-12 cells to varying degrees. Considering the content, chemical class, and biological activity of secondary metabolites from E. cristatum CPCC 401251, compounds 1, 7, and 13 were selected to detect their hypolipidemic activities and potential mechanisms in hyperlipidemia golden hamsters. Compound 1 exerted a hypolipidemic effect by activating the AMPK signaling pathway, decreasing Scd1, and improving intestinal flora. Compounds 7 and 13 played a role in the hypolipidemic activity by regulating the gene expression related to lipid synthesis and degradation, including upregulating the mRNA levels of Pparα, Hsl, and Atgl, and decreasing the mRNA level of Scd1. These findings help us understand the dominant fungus E. cristatum secondary metabolites presenting in Liupao tea and their potential hypolipidemic contributions. This work improves the understanding of the active substances in Liupao tea and highlights the health-promoting effects of microorganisms in the fermented tea.

Chemotherapy is one of the most effective approaches for cancer treatment. However, to improve efficacy, the therapeutic targets should be identified and characterised. Moreover, new drugs need to be discovered and developed to target different cancer pathways. Current therapeutics can eliminate most of the cancer cells. However, recurrence and metastasis still remain a major failure of cancer therapy. Emerging evidence demonstrates that multidrug resistance (MDR) and the existence of cancer stem cells (CSCs) are two major contributors for the failure of chemotherapy. MDR is a phenomenon in which cancer cells become resistant to structurally and functionally unrelated anticancer agents. CSCs are a small population of cells within cancer cells with capacity for self-renewal, tumor metastasis and differentiation. CSCs are also believed to be associated with chemoresistance. Thus, MDR and CSCs are the greatest challenges for cancer chemotherapy. Significant effort has been made to search for agents that specifically target MDR cells and CSCs. Consequently, some agents derived from nature have been developed to overcome MDR and CSCs. However, the developed chemotherapeutics cannot be used for all the cancers and some of them display severe cytotoxicity. Hence, there is an urgency to investigate the mechanism of drug resistance and to characterise cancer stem cells to identify potential new therapeutic targets. Natural products lie in the heart of the drug discovery. The developed chemotherapeutic compounds mainly originates from the secondary metabolites of microbes, terrestrial plants and marine organisms. In this study, MDR cancer cells were derived from tissue cultured cancer cells by the treatment the cells with fluorouracil (5-FU) and cisplatin (CDDP). CSCs were developed by treatment in serum-free medium with different factors. Fractions and compounds from Nature Bank (Griffith Institute for Drug Discovery, Griffith University), Compounds Australia (Griffith Institute for Drug Discovery, Griffith University) and Traditional Chinese Medicine (TCM) were screening by high through-put screening (HTS). As a result, one potential anticancer flavonoid was isolated from the Australian plant Cryptocarya (QID025519) which was identified by NMR spectroscopic data, in combination with LC-MS. Extracts, fractions and isolated pure compounds from Bruguiera gymnorrhiza andSchisandraviridis were identified as potential agents for the treatment of tongue cancer and breast cancer. The DCM and MeOH extracts and HPLC fractions of B. gymnorrhiza showed antiproliferation activity against cancer cells in a concentration-dependent manner. Further purification of the active fractions led to the isolation of five flavonoids namely rutin, myricetin 3-rutinoside, methoxyflavone, 5-Methoxyluteolin, and 7,3',4',5'-tetrahydroxy-5- gramrione. All five compounds showed antiproliferation activity against CAL27 and MCF7 and MDR cells in a concentration-dependent manner. Methoxyflavone demonstrated the strongest anticancer potential against CAL27 cells, MCF7 cells, CAL27 MDR cells while 7,3',4',5'-tetrahydroxy-5- gramrione illustrated the highest inhibitory effect on MCF7 MDR cells. Both aqueous and ethanol extracts showed activities against MCF7 and CAL27 cancer cells. Bioassay-guided fractionation and purification of the extracts from S.viridis resulted in six active principles, including five dibenzocyclooctene lignans namely gomisin H (1), schisandrin (2), angeloylgomisin H (3), (+)-gomisin M2 (4) and rubschisandrin (5), and one terpenoid, schisanol (6). Compounds 1-3 showed moderate anticancer activities with an IC50 value ranging from 100-200 μg/mL against MCF7 and CAL27 cell lines. Dioxane containing lignans 4-5 and triterpenoid 6 were 10 times more active with IC50 values of14.5, 13.4, 10.6 μg/mL against MCF7, and 21.2, 17.9, 11.7 μg/mL against CAL27, respectively. In addition, two compounds from Compounds Australia exhibited a potential application prospects for tongue cancer and breast cancer therapy. One compound SN00802961 exhibited significant inhibition on MCF7 cells, but low inhibitory effects on fibroblast cells. Meanwhile, it exhibited moderate inhibition on CAL27 MDR cells, CAL27 cells and CSCs. Compound SN00802961 has potently targeted the MAPK/ERK1/2 signaling pathway to induce cytotoxicity in MCF7 cells. Another agent SN00771077 for breast cancer cells in vitro was investigated. The effects of compound SN00771077 on cell viability in vitro were evaluated by treatment of MCF-7 and T47D cells. An in vitro viability assay demonstrated that compound SN00771077 inhibited the cell growth in a dose-dependent manner. The antiproliferative activity of compound SN00771077 is related to its activity on monomeric actin and the subsequent inhibition of polymerization of G-actin monomers. Exposure to compound SN00771077 induced the inhibition of Raf/MEK/ERK pathway in T47D cells. All the results indicated that compound SN00771077 had a strong cytotoxic effects on cancer cells, and shows potential in the treatment of breast cancer by causing the depolymerizing actin cytoskeleton.

Glandular trichomes (GTs) are one of the epidermal tissues of medicinal plants which function in the synthesis, storage, and secretion of secondary metabolites. The active ingredients of Chinese medicinal materials are mostly secondary metabolites of plants. Accordingly, it is of great research value to explore the quality of medicinal materials using the GTs of medicinal plants as the starting point. However, most of the current studies on GTs of medicinal plants are still at the simple morphological identification stage, and there are few studies on the compounds secreted by GTs and secondary metabolic processes. Here, we reviewed the literature, summarized the morphological types of medicinal plant GTs, separation and purification technology, analysis technology, and biological activities of secondary metabolites, and established a research approach to medicinal plant GTs. We hope to provide a reference for future research on GT inclusions and secondary metabolism.

The nutritional requirements and environmental conditions for a submerged culture of Streptomyces sp. 8812 were determined. Batch and fed-batch Streptomyces sp. 8812 fermentations were conducted to obtain high activity of secondary metabolites. In the study several factors were examined for their influence on the biosynthesis of the active metabolites-7-hydroxy-6-oxo-2,3,4,6-tetrahydroisoquinoline-3-carboxy acid (C10H9NO4) and N-acetyl-3,4-dihydroxy-L-phenylalanine (C11H13NO5): changes in medium composition, pH of production medium, various growth phases of seed culture, amino acid supplementation and addition of anion exchange resin to the submerged culture. Biological activities of secondary metabolites were examined with the use of DD-carboxypeptidase 64-575 and horseradish peroxidase. Streptomyces sp. 8812 mycelium was evaluated under fluorescent microscopy and respiratory activity of the strain was analyzed. Moreover, the enzymatic profiles of the strain with the use of Api ZYM test were analyzed and genetic analysis made. Phylogenetic analysis of Streptomyces sp. 8812 revealed that its closest relative is Streptomyces capoamus JCM 4734 (98%), whereas sequence analysis for 16S rRNA gene using NCBI BLAST algorithm showed 100% homology between these two strains. Biosynthetic processes, mycelium growth and enzyme inhibitory activities of these two strains were also compared.

Search for new species of medicinal plant raw materials, promising for use as a source of biologically active compounds is one of the priority directions of scientific research in the development of effective and safe drugs. Among the representatives of the domestic flora the leafy tree – Phellodendron amurense Rupr. is of interest for research. This plant, along with Phellodendron chinense C.K. Schneid, is the subject of research of scientists from different countries of the world, aimed at identifying phellodendron-specific secondary metabolites and assessing the prospects of their use in therapeutic and prophylactic purposes. In this connection, the purpose of the present work was to summarise the data of scientific literature on systematics, composition of biologically active substances (BAS) accumulated in different parts of plants of the genus Phellodendron and their pharmacological properties. The analysis allowed to generalise and systematise the data available today on the chemical composition of Phellodendron amurense and Phellodendron chinense. Particular attention in the article is paid to the issue of establishing the structure of the dominant component of Phellodendron amurense leaves – prenyl flavonoid phellamurin. Along with this, the data on biological activity of secondary metabolites of plants of the genus Phellodendron are given. The questions of standardisation of raw materials are covered. On the basis of the conducted information analysis it is shown that Phellodendron amurense is a source of obtaining BAS. Phellamurin and other flavonoids with high biological activity are of the greatest interest for study.

Miliusa velutina Hook. f. and Thomson var. deviyarina (Annonaceae) is a lesser-known medicinal tree traditionally used by tribal and local communities in Maharashtra, India. However, its pharmacological profile has not been systematically characterized. This study aimed to provide a comprehensive in vitro evaluation of the antioxidant, α-amylase inhibition, antimicrobial, antibiofilm and anticancer activities of different extracts of this variety. Bark, leaf, fruit and flower powders were successively extracted with water, methanol, ethanol, ethyl acetate and alcohol–water mixtures. Antioxidant activity was assessed by DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging, α-amylase inhibition by α-amylase inhibition, antimicrobial and antibiofilm activities by agar well diffusion and biofilm inhibition assays, and anticancer activity by sulforhodamine B (SRB) assays on MCF-7 and Vero cell lines. Methanol, ethanol and ethyl-acetate bark and leaf extracts showed strong DPPH scavenging with low IC₅₀ values comparable to quercetin and vitamin C. Green and ripe fruit extracts produced up to 89.57% α-amylase inhibition, while bark extracts showed moderate inhibition (37.50–61.25%). Bark extracts exhibited pronounced antimicrobial and antibiofilm effects, with maximum biofilm inhibition of 95.45% and zones of inhibition comparable to penicillin and streptomycin at 50 µg/mL. The ethyl-acetate bark extract displayed potent cytotoxicity toward MCF-7 cells (GI₅₀ < 10 µg/mL) with relatively lower effects on Vero cells, approaching the activity of Adriamycin. These in vitro findings highlight M. velutina var. deviyarina as a promising source of bioactive extracts and provide a multi-assay dataset to guide future phytochemical characterization and in vivo evaluation of this understudied medicinal plant.

The incidence of <i>Fusarium</i> spp. in stem bases of selected crops of winter wheat in the Midlands, UK

Corn or Maize product is regarded as one of the essential products in the world and stands third product after the rice and wheat crops. Different fungal pathogens attack corn plants; one of them is ear rot, brought on by Fusarium species and whose occurrence is primarily influenced by environmental factors. In order to isolate and identify Fusarium species from corn plants and their prevalence, 50 samples of corn were collected during September, October, and November of 2021 from 30 corn fields in 14 regions of different places in the Kurdistan region of Iraq. From all samples, 39 isolates of Fusarium were detected and based on morphological characteristics, six other species of Fusarium were identified, namely F. verticillioides (33.34%), F. proliferatum (25.64%), F. oxysporum (12.82%), F. incarnatum and F. equiseti (10.25% each), then F. fujikuroi (7.7%). The most prevalent species was F. verticillioides which was isolated from seven corn fields and significantly higher than all other isolated species. All Fusarium isolates were also molecularly identified depending on amplifying the internal transcribed spacer (ITS) universal region using forward ITS1 and reverse ITS4 primers and indicated DNA fragments ranged from 550 to 570 bp. The PCR fragments of the amplified ITS region were sequenced, aligned and registered in NCBI GeneBank with specified accession numbers. The phylogenetic tree and all analyzes were performed using the MEGA program version 11.0.13. The current study concluded that the corn fields in the Kurdistan region are infected with different Fusarium species, and the most common species is F. verticillioides. As well as the Fusarium species in the Kurdistan region have close evolutionary history to the same species in other countries. Thus, the study recommends more research to investigate the occurrence of toxigenic Fusarium species associated with cereal grains in the region.

The accurate identification of Fusarium species can take 2-3 weeks. Preliminary exoantigen studies indicate that a mature culture suspected of being a Fusarium species may be immunologically identified 48 h after receipt. Exoantigen extracts of 10-day-old slant cultures of Fusarium chlamydosporum, Fusarium moniliforme (= Fusarium verticilloides), Fusarium oxysporum, Fusarium proliferatum and Fusarium solani and partially purified reference homologous and heterologous shake culture extracts (6-week-old) were reacted against rabbit anti-F. chlamydosporum, F. moniliforme, F. oxysporum, F. proliferatum and F. solani sera, in a micro-immunodiffusion procedure. The results indicated that all the strains belonging to a given species produced 1-3 bands of identity only when tested against its homologous antiserum and reference antigen. No cross-reactions were observed with the heterologous antisera. Furthermore, extracts from isolates of Fusarium dimerum, Fusarium equiseti, Fusarium roseum complex, Acremonium species, Cylindrocarpon, Fonsecaea pedrosoi and Trichoderma species did not react with any of the prepared Fusarium species' antisera. Our data suggest that the exoantigen procedure is a rapid and reliable tool for the accurate immuno-identification of the medically important Fusarium species studied.

Pre-B-Cell Leukemia Transcription Factor 1 Regulates Expression of Valosin-Containing Protein, a Gene Involved in Cancer Growth

Sixty‐four isolates of Fusarium species isolated from 44 sorghum samples collected during 2004–2005 were subjected to polymerase chain reaction (PCR) analysis. PCR detection was performed on all Fusarium species using two different sets of primers, namely, internal transcribed spacer (ITS) and FUM1. The developed protocol is rapid for small‐scale extraction of DNA from fungal mycelia for the detection of Fusarium species using diagnostic PCR. Sixty‐four Fusarium isolates, namely, Fusarium verticillioides (45), Fusarium proliferatum (4), Fusarium anthophilum (4), Fusarium sporotrichioides (3), Fusarium pallidoroseum (6) and Fusarium oxysporum (2), were analyzed by PCR using the ITS and FUM1 set of primers. All Fusarium species scored positive with the ITS set of primers. Among the 64 isolates, 53 have scored positive with the FUM1 set of primers. These 53 isolates represent F. verticillioides (45), F. proliferatum (4) and F. anthophilum (4), respectively. The results of the study revealed that PCR‐based technique could be used to identify a group of potential fumonisin‐producing Fusarium species.PRACTICAL APPLICATIONSPolymerase chain reaction (PCR) method provides the basis for a simple, accurate, rapid and precise detection of potential fumonisin producing Fusarium species, which are of considerable risk to animal and human health. The detection of Fusarium species is therefore crucial for the prevention of toxins entering the food chain. The protocols developed in this study are helpful for a rapid and small‐scale extraction of nucleic acid from fungal mycelia, occurring on contaminated sorghum samples. A large number of samples can be screened in relatively less time using this PCR protocol when compared with the conventional methods.