Characterization and antimicrobial activity of endophytic fungi from medicinal plant Agave americana.

Endophytic microorganisms associated with medicinal plants are of particular interest as they are a potential source of new bioactive chemicals effective against novel emerging and drug-resistant pathogens. Agave americana is a tropical medicinal plant with antibacterial, antifungal, and anticancer properties. We studied the biodiversity of fungal endophytes of A. americana and their antimicrobial production potential. Isolated endophytic fungi were classified into 32 morphotypes (15 from stem and 17 from leaf) based on their cultural and morphological characteristics. Among the fungal crude extracts tested, 82% of isolates from the leaves and 80% of the isolates from the stem showed antibacterial activity against the bacterial strains (Escherichia coli ATTC 25902, Staphylococcus aureus ATTC 14775, and Bacillus subtilis NRRL 5109) tested. Extracts from four fungal isolates from leaves showed antifungal activity against at least one of the fungal strains (Candida albicans ATTC 10231 and Aspergillus fumigatus NRRL 5109) tested. Crude extracts of seven fungal isolates showed a zone of inhibition of more than 11 mm at 10 mgml-1 against both Gram-positive and Gram-negative bacteria tested. Penicillium, Colletotrichum, Curvularia, Pleosporales, Dothideomycetes, and Pleurotus are the main endophytes responsible for bioactive potential. These results indicate that A. americana harbors endophytes capable of producing antimicrobial metabolites.

The antimicrobial property of JY-1, a complex mixture of Traditional Chinese Medicine, is linked to it abilities to suppress biofilm formation and disrupt membrane permeability.

The substantial increase of infections, caused by novel, sudden, and drug-resistant pathogens, poses a significant threat to human health. While numerous studies have demonstrated the antibacterial and antiviral effects of Traditional Chinese Medicine, the potential of a complex mixture of traditional Chinese Medicine with a broad-spectrum antimicrobial property remains underexplored. This study aimed to develop a complex mixture of Traditional Chinese Medicine (TCM), JY-1, and investigate its antimicrobial properties, along with its potential mechanism of action against pathogenic microorganisms. Antimicrobial activity was assessed using a zone of inhibition assay and the drop plate method. Hyphal induction of Candida albicans was conducted using RPMI1640 medium containing 10% FBS, followed by microscopic visualization. Quantitative real-time PCR (RT-qPCR) was employed to quantify the transcript levels of hyphal-specific genes such as HWP1 and ALS3. The impact of JY-1 on biofilm formation was evaluated using both the XTT reduction assay and scanning electron microscopy (SEM). Furthermore, the cell membrane integrity was assessed by protein and nucleic acid leakage assays. Our results clearly showed that JY-1 significantly inhibits the vegetative growth of Candida spp. and Cryptococcus spp. In addition, this complex mixture is effectively against a wide range of pathogenic bacteria, including Staphylococcus aureus, Vancomycin-resistant enterococci, Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae. More interestingly, JY-1 plays a direct anti-viral role against the mammalian viral pathogen vesicular stomatitis virus (VSV). Further mechanistic studies indicate that JY-1 acts to reduce the expression of hyphal specific genes HWP1 and ALS3, resulting in the suppression of the hyphal formation of C. albicans. The antimicrobial property of JY-1 could be attributed to its ability to reduce biofilm formation and disrupt the cell membrane permeability, a process resulting in microbial cell death and the release of cellular contents. Taken together, our work identified a potent broad-spectrum antimicrobial agent, a complex mixture of TCM which might be developed as a potential antimicrobial drug.

Uncovering the antimicrobial activity of G-type lysozyme 2 derived from Cyprinus carpio mucus against bacterial and fungal pathogens.

This study aimed to explore the antimicrobic potential of mucus samples collected from Cyprinus carpio and identify the specific antimicrobial peptides responsible for its activity. The crude extract was tested against various bacterial and fungal pathogens, and its protein content and profile were analyzed. Purification steps, including gel filtration chromatography, were employed to isolate the most active fraction (peak IV), which was further identified via liquid chromatography and mass spectroscopy. The results revealed varying degrees of antimicrobial activity of the crude extract against different bacterial and fungal strains, with Leclercia adecarboxylata, Candida glabrata, and Candida parapsilosis showing the highest susceptibility. SDS-PAGE analysis demonstrated the existence of multiple low molecular weight protein bands in the crude extract, while fraction IV obtained from gel filtration chromatography exhibited the strongest antimicrobial activity. Peak IV displayed a range of minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) values against the tested pathogens, spanning from 0.038 to 4.960 mg/mL. Further investigation identified the purified peptide derived from peak IV as G-type lysozyme 2, characterized by a molecular weight of 21 kDa. These findings shed light on the existence of a highly effective antimicrobial peptide, G-type lysozyme 2, within the mucus of Cyprinus carpio. This peptide demonstrates notable activity against diverse bacterial and fungal pathogens. The insights from this study enhance our understanding of the fish's antimicrobial defense mechanisms and hold promise for developing novel antimicrobial agents.

Assessment of antimalarial activity of crude extract of Chan-Ta-Lee-La and Pra-Sa-Chan-Dang formulations and their plant ingredients for new drug candidates of malaria treatment: In vitro and in vivo experiments.

The emergence and spread of antimalarial drug resistance have become a significant problem worldwide. The search for natural products to develop novel antimalarial drugs is challenging. Therefore, this study aimed to assess the antimalarial and toxicological effects of Chan-Ta-Lee-La (CTLL) and Pra-Sa-Chan-Dang (PSCD) formulations and their plant ingredients. The crude extracts of CTLL and PSCD formulations and their plant ingredients were evaluated for in vitro antimalarial activity using Plasmodium lactate dehydrogenase enzyme and toxicity to Vero and HepG2 cells using the tetrazolium salt method. An extract from the CTLL and PSCD formulations exhibiting the highest selectivity index value was selected for further investigation using Peter's 4-day suppressive test, curative test, prophylactic test, and acute oral toxicity in mice. The phytochemical constituents were characterized using gas chromatography-mass spectrometry (GC-MS). Results showed that ethanolic extracts of CTLL and PSCD formulations possessed high antimalarial activity (half maximal inhibitory concentration = 4.88, and 4.19 g/mL, respectively) with low cytotoxicity. Ethanolic extracts of the CTLL and PSCD formulations demonstrated a significant dose-dependent decrease in parasitemia in mice. The ethanolic CTLL extract showed the greatest suppressive effect after 4 days of suppressive (89.80%) and curative (35.94%) testing at a dose of 600 mg/kg. Moreover, ethanolic PSCD extract showed the highest suppressive effect in the prophylactic test (65.82%) at a dose of 600 mg/kg. There was no acute toxicity in mice treated with ethanolic CTLL and PSCD extracts at 2,000 mg/kg bodyweight. GC-MS analysis revealed that the most abundant compounds in the ethanolic CTLL extract were linderol, isoborneol, eudesmol, linoleic acid, and oleic acid, whereas ethyl 4-methoxycinnamate was the most commonly found compound in the ethanolic PSCD extract, followed by 3-hydroxy-2-(4-hydroxy-3-methoxyphenyl)-4H-chromen-4-one, flamenol, oleic acid amide, linoleic acid, and oleic acid. In conclusions, ethanolic CTLL and PSCD extracts exhibited high antimalarial efficacy in vitro. The ethanolic CTLL extract at a dose of 600 mg/kg exhibited the highest antimalarial activity in the 4-day suppressive and curative tests, whereas the ethanolic PSCD extract at a dose of 600 mg/kg showed the highest antimalarial activity in the prophylactic test.

Control of pathogenic bacteria using marine actinobacterial extract with antiquorum sensing and antibiofilm activity.

OBJECTIVE: The objectives of this research were to screen the anti-quorum sensing and antibiofilm activity of marine actinobacteria, isolated from several aquatic environments in Indonesia against several pathogenic bacteria, such as Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Vibrio cholerae, Salmonella Typhimurium, and Pseudomonas aeruginosa. RESULTS: Ten out of 40 actinobacteria were found to have anti-quorum sensing activity against wild-type Chromobacterium violaceum (ATCC 12472); however, the validation assay showed that only eight of 10 significantly inhibited the quorum sensing system of Chromobacterium violaceum CV026. The crude actinobacteria extracts inhibited and disrupted biofilm formation produced by pathogens. The highest antibiofilm inhibition was discovered in isolates 11AC (90%), 1AC (90%), CW17 (84%), TB12 (94%), 20PM (85%), CW01 (93%) against Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Vibrio cholerae, Salmonella Typhimurium, and Pseudomonas aeruginosa, respectively. The highest biofilm destruction activity was observed for isolate 1AC (77%), 20PM (85%), 16PM (72%), CW01 (73%), 18PM (82%), 16PM (63%) against Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Vibrio cholerae, Salmonella Typhimurium, and Pseudomonas aeruginosa, respectively. Actinobacteria isolates demonstrated promising anti-quorum and/or antibiofilm activity, interfering with the biofilm formation of tested pathogens. Appropriate formulations of these extracts could be developed as effective disinfectants, eradicating biofilms in many industries.

Immunoregulatory effects of Huaier (Trametes robiniophila Murr) and relevant clinical applications.

Huaier (Trametes robiniophila Murr) is a medicinal fungus of traditional Chinese medicine with more than 1000 years of history of clinical application. Its remarkable anticancer activities has led to its application in treating diverse malignancies. In recent years, the immunomodulatory effects of Huaier have been uncovered and proved to be beneficial in a plethora of immune-related diseases including cancer, nephropathy, asthma, etc. In this review, we comprehensively summarized the active components of Huaier, its regulatory activities on multifaceted aspects of the immune system, its application in various clinical settings as well as toxicologic evidence. Based on currently available literature, Huaier possesses broad-spectrum regulatory activities on various components of the innate and adaptive immune system, including macrophages, dendritic cells, natural killer cells, T and B lymphocytes, etc. Versatile immunologic reactions are under the regulation of Huaier from expression of damage-associated molecular patterns, immune cell activation and maturation to cell proliferation, differentiation, antibody production, expression of cytokines and chemokines and terminal intracellular signal transduction. Moreover, some modulatory activities of Huaier might be context-dependent, typically promoting the restoration toward normal physiological status. With excellent efficacy and minimal side effects, we foresee more extensive application of Huaier for treating immune-related disorders.

Restoring Ampicillin Sensitivity in Multidrug-Resistant Escherichia coli Following Treatment in Combination with Coffee Pulp Extracts.

Escherichia coli, particularly multidrug-resistant (MDR) strains, is a serious cause of healthcare-associated infections. Development of novel antimicrobial agents or restoration of drug efficiency is required to treat MDR bacteria, and the use of natural products to solve this problem is promising. We investigated the antimicrobial activity of dried green coffee (DGC) beans, coffee pulp (CP), and arabica leaf (AL) crude extracts against 28 isolated MDR E. coli strains and restoration of ampicillin (AMP) efficiency with a combination test. DGC, CP, and AL extracts were effective against all 28 strains, with a minimum inhibitory concentration (MIC) of 12.5-50 mg/ml and minimum bactericidal concentration of 25-100 mg/ml. The CP-AMP combination was more effective than CP or AMP alone, with a fractional inhibitory concentration index value of 0.01. In the combination, the MIC of CP was 0.2 mg/ml (compared to 25 mg/ml of CP alone) and that of AMP was 0.1 mg/ml (compared to 50 mg/ml of AMP alone), or a 125-fold and 500-fold reduction, respectively, against 13-drug resistant MDR E. coli strains. Time-kill kinetics showed that the bactericidal effect of the CP-AMP combination occurred within 3 h through disruption of membrane permeability and biofilm eradication, as verified by scanning electron microscopy. This is the first report indicating that CP-AMP combination therapy could be employed to treat MDR E. coli by repurposing AMP.

In vitro antibacterial activity and synergetic effect of crude extract of the Wohlfahrtia nuba (Diptera: Sarcophagidae) flesh fly larvae.

Multidrug-resistant pathogens have become ubiquitous, and effective treatment alternatives are urgently required. Maggot therapy is a promising agent that is being studied to overcome antibiotic-resistant pathogens. This study evaluated the antibacterial activity of the larvae extract of the Wohlfahrtia nuba (wiedmann) (Diptera: Sarcophagidae) flesh fly on the growth of five pathogenic bacterial species (methicillin-sensitive Staphylococcus aureus [ATCC 29213], methicillin-resistant Staphylococcus aureus [ATCC BAA-1680], Pseudomonas aeruginosa [ATCC 27853], Escherichia coli [ATCC 25922], and Salmonella typhi [ATCC 19430]) in vitro by using different techniques. Resazurin-based turbidimetric assay demonstrated that the W. nuba maggot exosecretion (ES) was potent against all the bacterial species tested, and according to the determined minimum inhibitory concentration (MIC) for each bacterium, gram-negative bacteria were more sensitive than gram-positive bacteria. Additionally, colony-forming unit assay showed that maggot ES was able to inhibit bacterial growth rate for all bacterial species tested, where the highest bacterial reduction was observed with methicillin-sensitive S. aureus (MSSA) followed by S. typhi. Moreover, maggot ES was shown to be concentration-dependent, where 100 µL of ES at 200 mg/mL was bactericidal towards methicillin-resistant S. aureus (MRSA) and P. aeruginosa compared with 100 µL at the MIC of the ES. Moreover, based on the result of agar disc diffusion assay, maggot extract was more efficient against P. aeruginosa and E. coli than the remaining reference strains tested. Furthermore, the combination between regular antibiotics with maggot ES at different concentrations indicated that ES acts synergistically with the tested antibiotics against the five bacterial models.

[Effective substances and mechanism of Yishen Guluo Mixture in treatment of chronic glomerulonephritis based on metabolomics and serum pharmacochemistry].

This study aimed to investigate the effective substances and mechanism of Yishen Guluo Mixture in the treatment of chronic glomerulonephritis(CGN) based on metabolomics and serum pharmacochemistry. The rat model of CGN was induced by cationic bovine serum albumin(C-BSA). After intragastric administration of Yishen Guluo Mixture, the biochemical indexes related to renal function(24-hour urinary protein, serum urea nitrogen, and creatinine) were determined, and the efficacy evaluations such as histopathological observation were carried out. The serum biomarkers of Yishen Guluo Mixture in the treatment of CGN were screened out by ultra-performance liquid chromatography-quadrupole time-of-flight/mass spectrometry(UPLC-Q-TOF-MS) combined with multivariate statistical analysis, and the metabolic pathways were analyzed. According to the mass spectrum ion fragment information and metabolic pathway, the components absorbed into the blood(prototypes and metabolites) from Yishen Guluo Mixture were identified and analyzed by using PeakView 1.2 and MetabolitePilot 2.0.4. By integrating metabolomics and serum pharmacochemistry data, a mathematical model of correlation analysis between serum biomarkers and components absorbed into blood was constructed to screen out the potential effective substances of Yishen Guluo Mixture in the treatment of CGN. Yishen Guluo mixture significantly decreased the levels of 24-hour urinary protein, serum urea nitrogen, and creatinine in rats with CGN, and improved the pathological damage of the kidney tissue. Twenty serum biomarkers of Yishen Guluo Mixture in the treatment of CGN, such as arachidonic acid and lysophosphatidylcholine, were screened out, involving arachidonic acid metabolism, glycerol phosphatide metabolism, and other pathways. Based on the serum pharmacochemistry, 8 prototype components and 20 metabolites in the serum-containing Yishen Guluo Mixture were identified. According to the metabolomics and correlation analysis of serum pharmacochemistry, 12 compounds such as genistein absorbed into the blood from Yishen Guluo Mixture were selected as the potential effective substances for the treatment of CGN. Based on metabolomics and serum pharmacochemistry, the effective substances and mechanism of Yishen Guluo Mixture in the treatment of CGN are analyzed and explained in this study, which provides a new idea for the development of innovative traditional Chinese medicine for the treatment of CGN.

The crude extract from the flowers of Trollius chinensis Bunge exerts anti-influenza virus effects through modulation of the TLR3 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The flowers of Trollius chinensis Bunge (Ranunculaceae) is a traditional Chinese medicine used to treat various inflammatory diseases, including upper respiratory infections, chronic tonsillitis, and pharyngitis. Recently, there has been growing research on the antiviral role of the flowers of T. chinensis Bunge. However, little is known about its anti-influenza virus effects and the underlying mechanisms. AIM OF THE STUDY: This study aims to evaluate the therapeutic effects of the crude extract from the flowers of T. chinensis Bunge (CEFTC) on mice infected with influenza virus. We further explored its mechanism by detecting the expression of vital proteins (TLR3, TBK1, TAK1, IKKα, IRF3, and IFN-ß) related to TLR3 signaling pathway. MATERIALS AND METHODS: Mice were infected with influenza A virus (H1N1) through the nasal cavity and were intragastrically administered CEFTC at the dose of 0.2 mg/g once daily. The therapeutic effects of CEFTC were evaluated by blood cell count, lung index, spleen index, alveolar lavage fluid testing, and HE staining. Network pharmacology analysis predicted the potential signaling pathway between the flowers of T. chinensis Bunge and pneumonia. The expression of TLR3, TBK1, TAK1, IKKα, IRF3, and IFN-ß in lung tissues were examined by Western blot assay. In addition, the immunofluorescence assay was applied to assess the effect of CEFTC on the distribution of IRF3 and IFN-ß between nuclei and cytoplasm. RESULTS: Compared with the infected group, the lung index was markedly reduced, and the pathological damage of the lungs was also attenuated in the CEFTC treatment group. The network pharmacology analysis indicated that the NF-κB pathway was a potential signaling pathway in the flowers of T. chinensis Bunge for the treatment of pneumonia, TLR3, IRF3, and TBK1 were crucial targets associated with pneumonia. Western blot assay demonstrated that in the high-dose virus infected group, CEFTC reduced the expression of TLR3, TAK1, TBK1, and IRF3. Furthermore, CEFTC could increase the nuclear distribution of IRF3 in alveolar epithelial cells after virus infection. CONCLUSIONS: These results suggested that different doses of influenza virus could cause varying infection symptoms in mice. Moreover, CEFTC could exert anti-influenza virus effects by regulating the expression of TLR3, IRF3, IFN-ß, TAK1, and TBK1 in the TLR3 signaling pathway.

Methanolic Extract of Aerva javanica Leaves Prevents LPS-Induced Depressive Like Behavior in Experimental Mice.

Aim: Depression is a chronic recurrent neuropsychiatric disorder associated with inflammation. This study explored the pharmacological activities of Aerva javanica leaves crude extract (Aj.Cr) on lipopolysaccharide (LPS)-induced depressive-like behavior in experimental mice. Methods: Aj.Cr was evaluated for its phenolic and flavonoid contents, bioactive potential, amino acid profiling and enzyme inhibition assays using different analytical techniques followed by in-silico molecular docking was performed. In addition, three ligands identified in HPLC analysis and standard galantamine were docked to acetyl cholinesterase (AchE) enzyme to assess the ligand interaction along with their binding affinities. In in-vivo analysis, mice were given normal saline (10 mL/kg), imipramine (10 mg/kg) and Aj.Cr (100, 300, and 500 mg/kg) orally for 14-consecutive days. On the 14th day, respective treatment was given 30-minutes before intra-peritoneal administration of (0.83 mg/kg) LPS. Open field, forced swim and tail suspension tests were performed 24-hours after LPS injection, followed by a sucrose preference test 48-hours later. Serum corticosterone levels, as well as levels of nitric oxide (NO), malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), tumor necrosis factor-alpha (TNF-), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), brain-derived neurotrophic factor (BDNF) and catecholamines were determined in brain tissues. Results: In-vitro results revealed that crude extract of Aj.Cr possesses anti-depressant agents with solid antioxidant potential. In-vivo analysis showed that LPS significantly increased depressive-like behavior followed by alteration in serum and tissue biomarkers as compared to normal control (p < 0.001). While imipramine and Aj.Cr (100, 300, and 500 mg/kg) treated groups significantly (p<0.05) improved the depressive-like behavior and biomarkers when compared to the LPS group. Conclusion: The mitigation of LPS-induced depressive-like behavior by Aj.Cr may be linked to the modulation of oxidative stress, neuro-inflammation and catecholamines due to the presence of potent bioactive compounds exerting anti-depressant effects.

Antibacterial and Anticancer Properties of Microbispora sp., AL22: An Endophyte of Alpinia galanga (L.) Willd.

<b>Background and Objective:</b> The AL22 strain was isolated from the rhizosphere soil of <i>Alpinia galanga</i> (L.) Willd (Zingiberaceae) and identified as <i>Microbispora</i> sp., by analysing its morphology, chemotaxonomy and 16S rDNA sequence. Previous studies demonstrated the bactericidal effects of its crude extract against <i>Bacillus cereus</i>, <i>Bacillus subtilis</i>, <i>Staphylococcus aureus</i> and methicillin-resistant <i>Staphylococcus aureus</i>. The present study aimed to isolate the major compounds and evaluate their biological properties. <b>Materials and Methods:</b> Silica gel column chromatography and thin-layer chromatography were used for the purification and identification of 3,4-dihydro-lactucin (compound <b>1</b>) and umbelliferone (compound <b>2</b>) by NMR and mass spectrometry, respectively. Antibacterial and anticancer activities were carried out. <b>Results:</b> The bioassay studies illustrated that compound <b>1</b> had antibacterial activity against gram-positive bacteria, with its minimum inhibitory concentration and minimum bactericidal concentration of 16-32 and 64-128 µg mL¹, respectively. The crude extract and purified compounds showed weak cytotoxic activity on the L929 and Vero cells with IC₅₀ values >512.00 µg mL¹. The cytotoxicity of compound <b>1</b> was observed in the MDA-MB-231 and HeLa cells with IC₅₀ values of 37.62 and 75.34 µg mL¹, respectively, while its IC₅₀ value against the HepG2 cells was 456.67 µg mL¹. <b>Conclusion:</b> These findings showed that compound <b>1</b> of <i>Microbispora</i> sp., AL22 exhibited antibacterial and anticancer activities. Extensive studies on 3,4-dihydro-lactucin could lead to the development of beneficial approaches for managing bacterial infections and cancer.

Effect of occupational exposure to welding fumes and noise on heart rate variability: An exposed-unexposed study on welders and airport workers' population.

Introduction: Welding fumes (WF) are a complex mixture of gas and particles. Action of occupational exposure to WF on cardiovascular system has been recently studied as for noise. Research question: The main objectives of our study are therefore to evaluate the impact of exposure to WF, noise, and combined WF and noise on autonomic nervous system as assessed by heart rate variability (HRV). Methods: The study groups were 16 welders and eight airport workers (as a control group). All the participants underwent ambulatory electrocardiogram, personal WF, and noise exposure monitoring, respectively, with dust track and calibrated noise dosimeter during workday. Atmospheric environmental assessments at workplaces have been also performed. HRV parameters were summarized for all the workday and hourly. Correlation tests were used to examine relation between HRV parameters and levels of noise exposure in the two population. Analysis of covariance (ANCOVA) was used for mean of each HRV parameters. Results: For HRV parameters, we found significant higher levels for mean range of high frequency (HF), standard deviation of normal-to-normal R-R interval (SDNN), and root mean square of successive heartbeat interval difference (RMSSD) in welders which suggested an imbalance between sympathetic and parasympathetic nervous system in this population. For relation between noise and HRV parameters, we noted that levels of low frequency (LF), HF, and SDNN were significantly correlated with mean noise levels for welders (respectively, r = 0.62, r = 0.357, r = 0.48), not in control group. Using ANCOVA, we found that working as a welder significantly increases mean of HF (p = 0.01) and RMSSD (p = 0.02) and decreases in LF/HF (p = 0.008). Indeed, the interaction between exposure to WF and mean noise levels for HF (p = 0.005), LF/HF (p = 0.01), and RMSSD (p = 0.007) was significant. Conclusion: This study shows an impact of WF and noise on ANS balance. One hypothesis is WF exposure could increase sensibility to noise exposure on autonomic nervous system or there is a synergic effect.

Gomisin A enhances the antitumor effect of paclitaxel by suppressing oxidative stress in ovarian cancer.

Gomisin A (GA) is an effective component of Schisandra. The crude extracts of Schisandra chinensis and its active ingredients have been shown to inhibit multidrug resistance in tumour cells. Reactive oxygen species (ROS) have different roles in cancer and may contribute to therapy resistance. The human ovarian cancer (OC) cell lines SKOV3 and A2780, and a mouse model of OC, were used in the present study. MTT assay, colony formation assay, flow cytometry, western blot analysis, and haematoxylin and eosin (H&E) staining were performed to determine the antitumor effect of GA and paclitaxel (PTX) in vitro and in vivo. The ROS inhibitor N­acetyl cysteine (NAC) was used to assess the mechanism underlying the chemosensitizing effects of GA. Notably, the proliferation of OC cells was inhibited by PTX, which could be enhanced by the ROS inhibitor NAC or GA. Treatment with NAC + PTX or GA + PTX enhanced the cell cycle arrest, but not apoptosis, induced by PTX. Moreover, the molecular mechanism underlying this effect may be that GA decreases the levels of ROS in ovarian cancer cells and inhibits cell cycle progression by downregulating the expression of the cell cycle proteins cyclin­dependent kinase 4 and cyclin B1. In conclusion, the combination of PTX and the ROS inhibitor GA may be a novel strategy in OC chemotherapy.

First Discovery of Beta-Sitosterol as a Novel Antiviral Agent against White Spot Syndrome Virus.

The outbreak of white spot syndrome (WSS) is a looming challenge, due to dramatic losses to the crustacean aquaculture industry. However, at present, there are no prophylactic or therapeutic means to control this infectious viral disease. Here, we screened fifteen medicinal plants for their inhibitory activity on the white spot syndrome virus (WSSV), using red swamp crayfish (Procambarus clarkii) as a model species. The results showed that the crude extracts of Pinellia ternata (Thunb.) Breit. had the highest inhibitory effect (91.59%, 100 mg/kg) on WSSV proliferation, and its main component, beta-sitosterol, showed a much higher activity (95.79%, 50 mg/kg). Further, beta-sitosterol potently reduced (p < 0.01) viral loads and viral gene transcription levels in a concentration-dependent fashion, and significantly promoted the survival rate of WSSV-challenged crayfish (57.14%, 50 mg/kg). The co-incubation assay indicated that beta-sitosterol did not influence the infectivity of WSSV particles. Both pre- and post-treatment of beta-sitosterol exerted a significant inhibitory effect (p < 0.01) on the viral load in vivo. Mechanistically, beta-sitosterol not only interfered with the expression of viral genes (immediate early gene 1, ie1; DNA polymerase, DNApol) that are important in initiating WSSV transcription, but it also attenuated the hijacking of innate immune signaling pathways (Toll, IMD, and JAK/STAT pathways) by viral genes to block WSSV replication. Moreover, the expression of several antiviral immune, antioxidant, pro-inflammatory, and apoptosis-related genes changed significantly in beta-sitosterol-treated crayfish. Beta-sitosterol is a potent WSSV inhibitor and has the potential to be developed as an effective anti-WSSV agent against a WSS outbreak in crustacean aquaculture.

Characterization of a new Bacillus velezensis as a powerful biocontrol agent against tomato gray mold.

Biocontrol microbes are environment-friendly and safe for humans and animals. To seek biocontrol microbes effective in suppressing tomato gray mold is important for tomato production. Therefore, serial experiments were conducted to characterize the antagonism of Bacillus velezensis HY19, a novel self-isolated biocontrol bacterium, against Botrytis cinerea in vitro and the control on tomato gray mold in greenhouse. This bacterium produced extracellular phosphatase, protease, cellulose and siderophores, and considerably inhibited the growth of B. cinerea. A liquid chromatography-mass spectrometry (LC-MS) detected salicylic acid and numerous antifungal substances present in B. velezensis HY19 fermentation liquid (BVFL). When B. cinerea was grown on potato glucose agar, BVFL crude extract remarkably suppressed the fungal growth and reduced protein content and the activities of catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD). Transcriptome studies showed that BVFL crude extract significantly induced different expression of numerous genes in B. cinerea, most of which were down-regulated. Theses differently expressed genes were involved in the biological process, cell compartment, molecular functions, and metabolisms of glycine, serine, threonine, and sulfur in pathogen hyphae. Thus, this biocontrol bacterium antagonized B. cinerea in multiple ways due to the production of numerous antifungal substances that acted on multiple targets in the cells. BVFL significantly increased antioxidant enzyme activities in tomato leaves and decreased the incidence of tomato gray mold, with the control efficacies of 73.12-76.51%. Taken together, B. velezensis HY19 showed a promising use potential as a powerful bioagent against tomato gray mold.

Antibiofilm and Anti-Quorum Sensing Potential of Cycloartane-Type Triterpene Acids from Cameroonian Grassland Propolis: Phenolic Profile and Antioxidant Activity of Crude Extract.

Propolis is very popular for its beneficial health properties, such as antimicrobial activity and antioxidant effects. It is one of the most long-serving traditional medicines to mankind due to its interesting chemical diversity and therapeutic properties. The detailed chemical information of propolis samples is very necessary to guarantee its safety and for it to be accepted into health care systems. The phenolic profile of the hydroethanolic extract was determined using HPLC-DAD, and the antioxidant was evaluated using five complementary methods. Triterpenoids were isolated using column chromatography and characterized using 1H NMR and 13C NMR. The effects of the extract and the isolated compounds on quorum sensing mediated processes and biofilm formation in bacteria were evaluated. Protocatechic acid (40.76 ± 0.82 µg/g), 4-hydroxybenzoic acid (24.04 ± 0.21 µg/g), vanillic acid (29.90 ± 1.05 µg/g), quercetin (43.53 ± 1.10 µg/g), and luteolin (4.44 ± 0.48 µg/g) were identified and quantified. The extract showed good antioxidant activity in the DPPH•, ABTS•+, CUPRAC, and metal chelating assays, and this antioxidant effect was confirmed by cyclic voltammetry. 27-Hydroxymangiferonic acid (1), Ambolic acid (2), and Mangiferonic acid (3) were isolated from anti-quorum sensing activity at MIC, and it was indicated that the most active sample was the extract with inhibition diameter zone of 18.0 ± 1.0 mm, while compounds 1, 2, and 3 had inhibition zones of 12.0 ± 0.5 mm, 9.0 ± 1.0 mm, and 12.3 ± 1.0 mm, respectively. The samples inhibited the P. aeruginosa PA01 swarming motility at the three tested concentrations (50, 75, and 100 µg/mL) in a dose-dependent manner. The propolis extract was able to inhibit biofilm formation by S. aureus, E. coli, P. aeruginosa, C. albicans, and C. tropicalis at MIC concentration. Compound 1 proved biofilm inhibition on S. aureus, L. monocytogenes, E. faecalis, E. coli, and C. tropicalis at MIC and MIC/2; compound 2 inhibited the formation of biofilm at MIC on S. aureus, E. faecalis, E. coli, S. typhi, C. albicans, and C. tropicalis; and compound 3 inhibited biofilm formation on E. faecalis, E. coli, C. albicans, and C. tropicalis and further biofilm inhibition on E. coli at MIC/4 and MIC/8. The studied propolis sample showed important amounts of cycloartane-type triterpene acids, and this indicates that there can be significant intra-regional variation probably due to specific flora within the vicinity. The results indicate that propolis and its compounds can reduce virulence factors of pathogenic bacteria.

Antibacterial effect of Asphodelus fistulosus aqueous and ethanolic crude extracts on gram positive and gram negative bacteria.

Asphodelus fistulosus (A. fistulosus) is a wild plant grows in Jordan. Traditionally, it is used to treat different medical conditions and diseases such as respiratory ailments, against burns and dermatomucosal infections.This study aims to find out the effects of A. fistulosus aqueous and ethanolic crude extracts on Staphylococcus aureus(S. aureus) as gram positive bacteria and Escherichia coli (E. coli) as gram negative bacteria and to investigate which one will be affected either by aqueous and/or ethanolic crude extracts of A. fistulosus shooting parts that were collected from Jerash in the north of Jordan. Agar well diffusion method was used to evaluate the antibacterial activity of the crude extracts. In addition, MIC (minimum inhibitory concentration) as well as MBC (minimum bactericidal concentration) were determined against both types of bacteria. The results showed that flower aqueous extract of A. fistulosus was very effective against E. coli (20.0 ± 0.50) mm and caused a (14.0 ± 0.50) mm inhibition to S. aureus. The ethanolic extract of stem was very effective cauesed a (19.0 ± 0.50) mm inhibition in both bacterial species. Respectively, both S. aureus and E. coli were inhibited by ethanolic and aqueous extracts (mixture1 and mixture2) (15.0 ± 0.00 mm and 10.5 ± 0.50 mm). The highest antimbacterial activity was observed for the leaves aqueous extract against E.coli (0.06120 mg/mL). The obtained MIC values from A. fistulosus parts extracts demonstrated antibacterial activity ranged between 7.606 and 0.06120 mg/mL. The highest antimicrobial activity was recorded in the leaves aqueous extract against E. coli.The MBC value of stem aqueous extract was 5.00 mg/mL against both S. aureus and E. coli. On the other hand, ethanolic and aqueous extracts of the leaves gave MBC values 5.00 mg/mL, and 0.156 mg/mL, respectively, against E. coli.Based on the results of this study, it can be concluded that there is good inhibitory effect of aqueous and ethanolic of A. fistulosus shooting parts extracts on growth of E. coli and S. aureus. Adding to that, stem ethanolic extract has the most effective against S. aureus while aqueous extract of flower has the most effective against E. coli.So, it is recommended to have further future studies on the A. fistulosus shooting parts crude extract bioactive components and the mechanism of how these constituents affect these types of bacteria.

Antibacterial Effects of Artemisia afra Leaf Crude Extract Against Some Selected Multi-Antibiotic Resistant Clinical Pathogens.

Background: Due to improper use of antibiotics, some pathogenic bacteria that cause serious and deadly infections have become resistant to commonly used broad spectrum antibiotics. This antibiotic resistance has become major global healthcare problem. Therefore, there is an urgent need to develop novel antibacterial agents; hence, much attention has been made on medicinal plants such as Artemisia afra. Thus, the current study was aimed to evaluate the antibacterial activity of ethanolic, methanolic and n-hexane extracts of this plant leaf against four multi-antibiotic resistant clinical pathogens. Methods: Crude extracts from A.afra leaf were prepared using ethanol, methanol and n-hexane and the antibacterial effect of each extract was tested against Escherichia coli, Streptococcus pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus. In addition, minimum inhibitory concentrations (MIC) and minimum bactericidal concentrations (MBC) were evaluated. Results: Among the crude extracts, the highest zone of inhibition (25.33±0.58mm) was recorded against E. coli when methanolic extract was applied. On the other hand, the lowest inhibition was exhibited when n-hexane extract was applied against S.aureus (5.67±1.56 mm). Concerning MIC values of the different extracts, varied results were obtained. MIC value of 6.25mg/mL was recorded when methanolic extract was applied against all clinical pathogens. Moreover, both methanolic and ethanolic extracts showed MBC value of 12.5mg/mL against the four clinical pathogens. However, the methanolic extract gave MBC value of 6.25mg/mL against E. coli. Conclusion: From this study, it can be concluded that it is possible to develop and formulate of new, efficacious, less toxic and inexpensive herbal medicine from A.afra leaf extract that act against multi-antibiotic resistant clinical pathogens.

Community composition, bacterial symbionts, antibacterial and antioxidant activities of honeybee-associated fungi.

BACKGROUND: Fungi associated with insects represent one potentially rich source for the discovery of novel metabolites. However, a comprehensive understanding of the fungal communities of Apis mellifera ligustica remains elusive. RESULTS: Here, we investigated the phylogenetic diversity and community composition of honeybee-associated fungi using combination of culture-dependent and culture-independent approaches. A total of forty-five fungi were isolated and purified from the Apis mellifera ligustica, royal jelly, and honeycomb, which belonged to four classes and eleven different genera. Furthermore, 28 bacterial 16S rRNA gene sequences were obtained by PCR from the fungal metagenome. High-throughput sequencing analyses revealed that the fungal communities were more diverse, a total of 62 fungal genera were detected in the honeybee gut by culture-independent method, whereas only 4 genera were isolated by culture-dependent method. Similarly, 247 fungal genera were detected in the honeycomb, whereas only 4 genera were isolated. In addition, we assessed the antibacterial and antioxidant activities of fungal isolates. Most fungal crude extracts obtained from the cultivation supernatant exhibited antioxidant activities. Only two fungal crude extracts displayed moderate activity against Escherichia coli and Staphylococcus aureus. Chemical analysis of Chaetomium subaffine MFFC22 led to the discovery of three known compounds, including cochliodinol (1), emodin (2), chrysophanol (3). Among them, cochliodinol (1) showed intense DPPH radical scavenging activity with the 50% inhibitory concentration (IC50) of 3.06 µg/mL, which was comparable to that of the positive ascorbic acid (IC50 = 2.25 µg/mL). Compound 2 displayed weak inhibitory activities against Micrococcus tetragenus and S. aureus. CONCLUSIONS: This research provided a fundamental clue for the complex interactions among honeybees, fungi, bacterial symbionts, and the effects on the honeybee. Furthermore, the diversity of honeybee-associated fungi had great potential in finding the resource of new species and antioxidants.