scholarly journals Effect of Allium cepa on LAC1 gene expression and physiological activities in Cryptococcus neoformans

2021 ◽  
Author(s):  
Seyed Afzal Musavinasab-Mobarakeh ◽  
Masoomeh Shams-Ghahfarokhi ◽  
Mehdi Razzaghi-Abyaneh
Keyword(s):  
Gene Expression ◽  
Cell Membrane ◽  
Cryptococcus Neoformans ◽  
Allium Cepa ◽  
Laccase Activity ◽  
Inhibitory Concentration ◽  
Biological Activities ◽  
Antifungal Drugs ◽  
Melanin Production ◽  
Ergosterol Content

Background and Purpose: This study aimed to investigate the effects of Allium cepa ethanolic extract (EAC) on Cryptococcus neoformans biological activities and LAC1 gene expression. Materials and Methods: The minimum inhibitory concentration (MIC) of EAC was determined based on the Clinical and Laboratory Standards Institute M27-A4 method at a concentration range of 125- 4000 µg/ml. The EAC synergism activity was determined in combination with fluconazole (FCZ) as an antifungal azole. Laccase activity, melanin production, and cell membrane ergosterol content of C. neoformans were assessed at the 0.5× MIC concentration of EAC (1000 μg/ml) and FCZ (64μg/ml) by approved methods. The expression of the LAC1 gene was studied in the fungus exposed to 0.5× MIC concentration of EAC and FCZ using the real-time polymerase chain reaction. Results: Based on obtained results, MIC of EAC and FCZ were 2000 and 128 μg/ml,respectively. A combinatory effect was reported for FCZ and EAC by a fractional inhibitory concentration index of 0.25. The cell membrane ergosterol content was inhibited in EAC- and FCZ-treated C. neoformans by 58.25% and 49.85%, respectively.The laccase activity and melanin production were reduced in EAC-treated C. neoformans by 45.37% and 51.57%, and in FCZ-treated fungus by 54.64% and 53.68%, respectively.The expression of fungal LAC1 at messenger RNA (mRNA) level was measured 0.46 and 0.58 folds and significantly decreased in both EAC- and FCZ-treated C. neoformans at the 0.5×MIC concentration, respectively (p <0.05). Conclusion: The findings revealed that EAC contains inhibitory compounds which interact with biological activities in C. neoformans and thereby, it could be considered as a potential source for the development of novel antifungal drugs.

scholarly journals The Organogermanium Compound THGP Suppresses Melanin Synthesis via Complex Formation with L-DOPA on Mushroom Tyrosinase and in B16 4A5 Melanoma Cells

2019 ◽  
Vol 20 (19) ◽  
pp. 4785
Author(s):  
Junya Azumi ◽  
Tomoya Takeda ◽  
Yasuhiro Shimada ◽  
Hisashi Aso ◽  
Takashi Nakamura
Keyword(s):  
Gene Expression ◽  
Kojic Acid ◽  
Biological Activities ◽  
Melanoma Cells ◽  
Tyrosinase Activity ◽  
Melanin Synthesis ◽  
Mushroom Tyrosinase ◽  
Melanin Production ◽  
Organogermanium Compound ◽  
Or Gene

The organogermanium compound 3-(trihydroxygermyl)propanoic acid (THGP) has various biological activities. We previously reported that THGP forms a complex with cis-diol structures. L-3,4-Dihydroxyphenylalanine (L-DOPA), a precursor of melanin, contains a cis-diol structure in its catechol skeleton, and excessive melanin production causes skin darkening and staining. Thus, the cosmetic field is investigating substances that suppress melanin production. In this study, we investigated whether THGP inhibits melanin synthesis via the formation of a complex with L-DOPA using mushroom tyrosinase and B16 4A5 melanoma cells. The ability of THGP to interact with L-DOPA was analyzed by 1H-NMR, and the influence of THGP and/or kojic acid on melanin synthesis was investigated. We also examined the effect of THGP on cytotoxicity, tyrosinase activity, and gene expression and found that THGP interacted with L-DOPA, a precursor of melanin with a cis-diol structure. The results also showed that THGP inhibited melanin synthesis, exerted a synergistic effect with kojic acid, and did not affect tyrosinase activity or gene expression. These results suggest that THGP is a useful substrate that functions as an inhibitor of melanogenesis and that its effect is enhanced by combination with kojic acid.

scholarly journals Eugenol and Isoeugenol In Association with Antifungal Against Cryptococcus neoformans

2017 ◽  
Vol 9 (4) ◽  
Author(s):  
Pinheiro L. S. ◽  
Sousa J. P. ◽  
Sousa J. P. ◽  
Barreto N. A. ◽  
Dantas T B ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Inhibitory Concentration ◽  
Synergistic Effects ◽  
Antagonistic Effect ◽  
Antifungal Drugs ◽  
Antifungal Effect ◽  
Beneficial Effects ◽  
Combined Use ◽  
Antifungal Effects

The antifungal therapy combined is used in clinical practice of several mycoses as it may increase the efficacy of the treatment. The use of natural products (phytochemicals) in combination with conventional antifungal drugs has been related to beneficial effects, mainly synergistic effects. The aim of this study was to evaluate the effect of the combined use of eugenol / isoeugenol, compounds with recognized antimicrobial activity, in association with antifungal amphotericin B against strains of Cryptococcus neoformans. The combined antifungal effect were be determined from the Fraction Inhibitory Concentration index - checkerboard technique. The results obtained in this study showed that eugenol in combination with amphotericin B had antagonistic effect against the strains of C. neoformans, LM 615 and INCQS 40221 (FIC index 6.0 and 4.0), respectively. The combination of the isoeugenol and amphotericin B also showed antagonistic effects for both the LM 615 strain and INCQS 40221 (FIC index 6.0 and 5.0), respectively. This study contributed to the understanding of the antifungal effects of the association of phenylpropanoids (eugenol / isoeugenol) with amphotericin B. Further studies are needed to evaluate and compare the effects of the association of these phytochemicals with other conventional antifungal drugs used against C. neoformans.

scholarly journals Fungicidal Activity of Recombinant Javanicin against Cryptococcus neoformans Is Associated with Intracellular Target(s) Involved in Carbohydrate and Energy Metabolic Processes

Molecules ◽  
2021 ◽  
Vol 26 (22) ◽  
pp. 7011
Author(s):  
Santhasiri Orrapin ◽  
Sittiruk Roytrakul ◽  
Narumon Phaonakrop ◽  
Siriwan Thaisakun ◽  
Khajornsak Tragoolpua ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Laser Scanning ◽  
Antifungal Agents ◽  
Biological Activities ◽  
Microscopic Analysis ◽  
Inhibitory Effect ◽  
Antifungal Drugs ◽  
Confocal Laser ◽  
Label Free ◽  
Glycolysis Pathway

The occurrence of Cryptococcus neoformans, the human fungal pathogen that primarily infects immunocompromised individuals, has been progressing at an alarming rate. The increased incidence of infection of C. neoformans with antifungal drugs resistance has become a global concern. Potential antifungal agents with extremely low toxicity are urgently needed. Herein, the biological activities of recombinant javanicin (r-javanicin) against C. neoformans were evaluated. A time-killing assay was performed and both concentration- and time-dependent antifungal activity of r-javanicin were indicated. The inhibitory effect of the peptide was initially observed at 4 h post-treatment and ultimately eradicated within 36 to 48 h. Fungal outer surface alteration was characterized by the scanning electron microscope (SEM) whereas a negligible change with slight shrinkage of external morphology was observed in r-javanicin treated cells. Confocal laser scanning microscopic analysis implied that the target(s) of r-javanicin is conceivably resided in the cell thereby allowing the peptide to penetrate across the membrane and accumulate throughout the fungal body. Finally, cryptococcal cells coped with r-javanicin were preliminarily investigated using label-free mass spectrometry-based proteomics. Combined with microscopic and proteomics analysis, it was clearly elucidated the peptide localized in the intracellular compartment where carbohydrate metabolism and energy production associated with glycolysis pathway and mitochondrial respiration, respectively, were principally interfered. Overall, r-javanicin would be an alternative candidate for further development of antifungal agents.

scholarly journals Evaluation of laccases and melanization in clinical and environmental Cryptococcus neoformans samples by non-denaturing PAGE

2009 ◽  
Vol 58 (5) ◽  
pp. 563-566 ◽  
Author(s):  
Cristiane B. Pereira ◽  
Frank L. Bueno ◽  
Amanda L. T. Dias ◽  
Maísa R. P. L. Brigagão ◽  
Claudete R. Paula ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Enzyme Production ◽  
Laccase Activity ◽  
Opportunistic Pathogen ◽  
Clinical Strain ◽  
Melanin Production ◽  
Densitometry Analysis ◽  
Serotype B ◽  
Colour Quantification ◽  
Laccase Isoforms

The increased incidence of infections caused by the opportunistic pathogen Cryptococcus neoformans, which mainly affects immunocompromised patients but can also infect immunocompetent individuals, has needed additional studies on this micro-organism's pathogenicity and factors related to virulence, such as enzyme production, for a better understanding of the aetiology of cryptococcosis. The aim of this study was to verify the applicability of non-denaturing PAGE for analysis of laccases by quantification of the amount of melanin pigment produced by clinical and environmental strains of C. neoformans. After incubation of the gel with the substrate l-dopa, strains produced melanin spots of a bright brown to black colour. Quantification of these spots was performed by densitometry analysis and the amount of melanin produced was calculated and compared among the strains. All strains showed laccase activity. Serotype B strains showed a higher melanin intensity than serotype A strains. Over half of the clinical strains (56.2 %) showed the lowest melanin intensities, suggesting that melanin production may not be the main virulence factor against host defence. The clinical strain ICB 88 revealed two melanin spots on the gel, indicating the presence of two laccase isoforms. The environmental strains showed the highest values of melanin intensity, which may be related to previous exposure to environmental stress conditions.

scholarly journals Laccase Affects the Rate of Cryptococcus neoformans Nonlytic Exocytosis from Macrophages

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Stefânia de Oliveira Frazão ◽  
Herdson Renney de Sousa ◽  
Lenise Gonçalves da Silva ◽  
Jéssica dos Santos Folha ◽  
Kaio César de Melo Gorgonha ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cryptococcus Neoformans ◽  
Laccase Activity ◽  
Severe Disease ◽  
Cell Types ◽  
Great Variability ◽  
Melanin Production ◽  
Content Type ◽  
Fungal Melanin ◽  
Laccase Enzyme

ABSTRACT Nonlytic exocytosis is a process in which previously ingested microbes are expelled from host phagocytes with the concomitant survival of both cell types. This process has been observed in the interaction of Cryptococcus spp. and other fungal cells with phagocytes as distant as mammalian, bird, and fish macrophages and ameboid predators. Despite a great amount of research dedicated to unraveling this process, there are still many questions about its regulation and its final benefits for host or fungal cells. During a study to characterize the virulence attributes of Brazilian clinical isolates of C. neoformans, we observed great variability in their rates of nonlytic exocytosis and noted a correlation between this process and fungal melanin production/laccase activity. Flow cytometry experiments using melanized cells, nonmelanized cells, and lac1Δ mutants revealed that laccase has a role in the process of nonlytic exocytosis that seems to be independent of melanin production. These results identify a role for laccase in virulence, independent of its role in pigment production, that represents a new variable in the regulation of nonlytic exocytosis. IMPORTANCE Cryptococcus neoformans is a yeast that causes severe disease, primarily in immunosuppressed people. It has many attributes that allow it to survive and cause disease, such as a polysaccharide capsule and the dark pigment melanin produced by the laccase enzyme. Upon infection, the yeast is ingested by cells called macrophages, whose function is to kill them. Instead, these fungal cells can exit from macrophages in a process called nonlytic exocytosis. We know that this process is controlled by both host and fungal factors, only some of which are known. As part of an ongoing study, we observed that C. neoformans isolates that produce melanin faster are more-frequent targets of nonlytic exocytosis. Further experiments showed that this is probably due to higher production of laccase, because fungi lacking this enzyme are nonlytically exocytosed less often. This shows that laccase is an important signal/regulator of nonlytic exocytosis of C. neoformans from macrophages.

scholarly journals Voriconazole Inhibits Melanization in Cryptococcus neoformans

2007 ◽  
Vol 51 (12) ◽  
pp. 4396-4400 ◽  
Author(s):  
Luis R. Martinez ◽  
Patricia Ntiamoah ◽  
Attila Gácser ◽  
Arturo Casadevall ◽  
Joshua D. Nosanchuk
Keyword(s):  
Gene Expression ◽  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Antifungal Drug ◽  
Melanin Production ◽  
Melanin Pigmentation ◽  
Ergosterol Synthesis ◽  
Subinhibitory Concentrations

ABSTRACT Voriconazole is a triazole antifungal drug that inhibits ergosterol synthesis and has broad activity against yeast and molds. While studying the interaction of voriconazole and Cryptococcus neoformans, we noted that cells grown in the presence of subinhibitory concentrations of voriconazole reduced melanin pigmentation. We investigated this effect systematically by assessing melanin production in the presence of voriconazole, amphotericin B, caspofungin, itraconazole, and fluconazole. Only voriconazole impeded the formation of melanin at subinhibitory concentrations. Voriconazole did not affect the autopolymerization of l-dopa, and 0.5 MIC of voriconazole did affect the gene expression of C. neoformans. However, voriconazole inhibited the capacity of laccase to catalyze the formation of melanin. Hence, voriconazole affects melanization in C. neoformans by interacting directly with laccase, which may increase the efficacy of this potent antifungal against certain pigmented fungi.

scholarly journals Antifungal activity of eugenol on Cryptococcus neoformans biological activity and Cxt1p gene expression

2020 ◽  
Author(s):  
Parviz Hassanpour ◽  
Masoomeh Shams-Ghahfarokhi ◽  
Mehdi Razzaghi-Abyaneh
Keyword(s):  
Gene Expression ◽  
Biological Activity ◽  
Growth Inhibition ◽  
Cryptococcus Neoformans ◽  
Urease Activity ◽  
Inhibition Zone ◽  
Antifungal Drugs ◽  
Treated Culture ◽  
Polymerase Chain ◽  
A3 Method

Background and Purpose: The present study was targeted toward investigating the effects of eugenol on Cryptococcus neoformans biological activity and Cxt1p gene expression. Materials and Methods: For the purpose of the study, the growth, urease, synergism activity, and disk diffusion of C. neoformans were assessed in eugenol-treated culture. The minimum inhibitory concentration (MIC) was determined by the Clinical and Laboratory Standards Institute M27-A3 method at a concentration range of 0.062-2 mg/mL. Subsequently, the expression of Cxt1p genes was studied at the MIC50 concentration of eugenol using real-time polymerase chain reaction. Results: The obtained results showed that eugenol at the concentrations of 125 and 500 μg/mL resulted in 50% and 100% growth inhibition in C. neoformans, respectively. In terms of urease activity, the results showed that the addition of MIC50 of eugenol and fluconazole to urea medium reduced urease activity in C. neoformans. In the culture treated with eugenol, the inhibition zone of antifungal drugs, namely amphotericin B, itraconazole, and fluconazole, was increased to 36±0.002, 22±0.001, and 12±0.002 mm, respectively. The expression levels of Cxt1p in the eugenol-treated, fluconazole-treated, and non-treated samples were estimated at 46%, 58%, and 100%, respectively. Conclusion: The findings of the current study revealed that eugenol could cause C. neoformans growth inhibition and reduce Cxt1p expression in this species. As the results indicated, the susceptibility of C. neoformans to fluconazole was increased when combined with eugenol.

scholarly journals A Novel Antimicrobial Peptide Sparamosin26–54 From the Mud Crab Scylla paramamosain Showing Potent Antifungal Activity Against Cryptococcus neoformans

2021 ◽  
Vol 12 ◽  
Author(s):  
Yan-Chao Chen ◽  
Ying Yang ◽  
Chang Zhang ◽  
Hui-Yun Chen ◽  
Fangyi Chen ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Amino Acids ◽  
Cell Wall ◽  
Cell Membrane ◽  
Antimicrobial Peptide ◽  
Cryptococcus Neoformans ◽  
Antifungal Drugs ◽  
Scylla Paramamosain ◽  
Mud Crab ◽  
Mature Peptide

Due to the increasing prevalence of drug-resistant fungi and the limitations of current treatment strategies to fungal infections, exploration and development of new antifungal drugs or substituents are necessary. In the study, a novel antimicrobial peptide, named Sparamosin, was identified in the mud crab Scylla paramamosain, which contains a signal peptide of 22 amino acids and a mature peptide of 54 amino acids. The antimicrobial activity of its synthetic mature peptide and two truncated peptides (Sparamosin1–25 and Sparamosin26–54) were determined. The results showed that Sparamosin26–54 had the strongest activity against a variety of Gram-negative bacteria, Gram-positive bacteria and fungi, in particular had rapid fungicidal kinetics (killed 99% Cryptococcus neoformans within 10 min) and had potent anti-biofilm activity against C. neoformans, but had no cytotoxic effect on mammalian cells. The RNA-seq results showed that after Sparamosin26–54 treatment, the expression of genes involved in cell wall component biosynthesis, cell wall integrity signaling pathway, anti-oxidative stress, apoptosis and DNA repair were significantly up-regulated, indicating that Sparamosin26–54 might disrupt the cell wall of C. neoformans, causing oxidative stress, DNA damage and cell apoptosis. The underlying mechanism was further confirmed. Sparamosin26–54 could bind to several phospholipids in the cell membrane and effectively killed C. neoformans through disrupting the integrity of the cell wall and cell membrane observed by electron microscope and staining assay. In addition, it was found that the accumulation of reactive oxygen species (ROS) increased, the mitochondrial membrane potential (MMP) was disrupted, and DNA fragmentation was induced after Sparamosin26–54 treatment, which are all hallmarks of apoptosis. Taken together, Sparamosin26–54 has a good application prospect as an effective antimicrobial agent, especially for C. neoformans infections.

Anti-inflammatory and analgesic effects of methanol extract of red cultivar Allium cepa bulbs in rats and mice

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Adeoye Joshua Oyewusi ◽  
Olayinka Ayotunde Oridupa ◽  
Adebowale Bernard Saba ◽  
Ibironke Kofoworola Oyewusi ◽  
Jonny Olufemi Olukunle
Keyword(s):  
Allium Cepa ◽  
Methanol Extract ◽  
Biological Activities ◽  
Hot Water ◽  
Control Group ◽  
Tail Flick ◽  
Control Groups ◽  
Inflammatory Models ◽  
Paw Oedema ◽  
Anti Inflammatory

Abstract Objectives Several cultivars of Allium cepa L. have been studied for anti-inflammatory and analgesic activities but there is inadequate information on such biological activities of the concentrated extracts of the Nigerian grown red cultivar A. cepa bulb. Methods The anti-inflammatory models used in this study were Carrageenan-induced paw oedema and formalin-induced paw lick in rats, while acetic acid-induced abdominal writhing, hot plate reaction, hot water tail flick tests in mice were the analgesic models. Results At 30 min post-induction (pi), the inhibition of paw oedema (62.50%) by 200 mg/kg of methanol extract of red cultivar A. cepa bulb (MERCACB) was significantly (p<0.001) higher than that of indomethacin (15.63%) at 10 mg/kg. The paw oedema inhibition at 60 min pi by MERCACB (76.92%) was significantly higher than that of indomethacin (41.03%). At the early phase of formalin paw-lick test, the pain reaction time (PRT) of rat treated with MERCACB (400 mg/kg) was significantly lower than that of indomethacin and the control groups. The hotplate test revealed that PRT of mice treated with 800 mg/kg of MERCACB were significantly (p<0.01) longer in comparism to indomethacin and control groups. The PRT of mice subjected to thermal pain due to hot water and treated with 800 mg/kg of MERCACB was significantly (p<0.05) longer than that of the control group. Conclusions These findings indicate that MERCACB possesses potent anti-inflammatory and analgesic properties which confirm the traditional use of the plant for the treatment of inflammatory diseases and may be useful as a future therapeutic agent.

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