No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Natural Products as Antifungal Agents against Invasive Fungi
Abstract
Background:
Invasive fungal infections (IFIs) are primarily caused by Candida spp., Cryptococcus neoformans, Aspergillus spp., Mucor spp., Sporothrix spp., and Pneumocystis spp., which attack human organs with a strong pathogenicity and exhibit drug resistance against commonly used chemical drugs. Therefore, the search for alternative drugs with high efficacy, low resistance rates, few side effects, and synergistic antifungal effects remains a major challenge. The characteristics of natural products with structural and bioactive diversity, lower drug resistance, and rich resources make them a major focus of the development of antifungal drugs.
Objective:
This review attempts to summarize the origin, structure, and antifungal activity of natural products and their derivatives with MIC ≤ 20 µg/mL or 100 µM, focusing on their MoA and SAR.
Methods:
All pertinent literature databases were searched. The search keywords were antifungal or antifungals, terpenoids, steroidal saponins, alkaloid, phenols, lignans, flavonoids, quinones, macrolide, peptide, tetramic acid glycoside, polyene, polyketide, bithiazole, natural product, and derivatives. All the related literature (covering the past 20 years, 2001-2022) was evaluated.
Results:
In total, 340 natural products and 34 synthesized derivatives with antifungal activity from 301 studies were included in this review. These compounds were derived from terrestrial plants, ocean life, and microorganisms and exhibited in vitro and in vivo potent antifungal activity alone or in combination. The MoA and SARs of reported compounds were summarized whenever applicable.
Conclusion:
In this review, we attempted to review the available literature on natural antifungal products and their derivatives. Most of the studied compounds showed potent activity against Candida species, Aspergillus species, or Cryptococcus species. Some of the studied compounds also demonstrated the ability to impair the cell membrane and cell wall, inhibit hypha and biofilms, and cause mitochondrial dysfunction. Although the MoAs of these compounds are not well understood yet, they can be used as lead components for the development of new, effective, and safe antifungal agents through their novel mechanisms.
... 98 Other antifungal mechanisms of NPs derived from plants have also been reported. 99 ...
Fungal resistance to the limited existing antifungal agents has become a considerable public health problem in the past few decades. However, natural products with multiple bioactivities are important sources for developing new therapeutic agents against fungi. Consequently, there is an urgent need to grasp the antifungal mechanism of natural products. In this review, the primary antifungal mechanisms of natural products derived from plants, including targeting the cell wall, cell membrane, mitochondria, biofilm and other mechanisms, are elaborated. This article critically describes the compounds with high antifungal activity, revealing opportunities for future research such as clinical trials or utilization as environmental disinfectants.
... Caspofungin is a safe rescue drug for patients with severe liver dysfunction (De Pauw et al., 2008), which has slight side effects on liver function compared with voriconazole (Wagner et al., 2006). In addition, TCM has been widely used in China and have been reported to contain potent antifungal potency and benefit for IFI patients (Dong et al., 2023;Xie et al., 2024). Therefore, most of the patients in our present study were treated by caspofungin and TCM. ...
Background and aim
The global burden of invasive fungal infections (IFIs) is emerging in immunologic deficiency status from various disease. Patients with acute-on-chronic hepatitis B liver failure (ACHBLF) are prone to IFI and their conditions are commonly exacerbated by IFI. However, little is known about the characteristics and risk factors for IFI in hospitalized ACHBLF patients.
Methods
A total of 243 hospitalized ACHBLF patients were retrospectively enrolled from January 2010 to July 2023. We performed restricted cubic spline analysis to determine the non-linear associations between independent variables and IFI. The risk factors for IFI were identified using logistic regression and the extreme gradient boosting (XGBoost) algorithm. The effect values of the risk factors were determined by the SHapley Additive exPlanations (SHAP) method.
Results
There were 24 ACHBLF patients (9.84%) who developed IFI on average 17.5 (13.50, 23.00) days after admission. The serum creatinine level showed a non-linear association with the possibility of IFI. Multiple logistic regression revealed that length of hospitalization (OR = 1.05, 95% CI: 1.02–1.08, P = 0.002) and neutrophilic granulocyte percentage (OR = 1.04, 95% CI: 1.00–1.09, P = 0.042) were independent risk factors for IFI. The XGBoost algorithm showed that the use of antibiotics (SHAP value = 0.446), length of hospitalization (SHAP value = 0.406) and log (qHBV DNA) (SHAP value = 0.206) were the top three independent risk factors for IFI. Furthermore, interaction analysis revealed no multiplicative effects between the use of antibiotics and the use of glucocorticoids (P = 0.990).
Conclusion
IFI is a rare complication that leads to high mortality in hospitalized ACHBLF patients, and a high neutrophilic granulocyte percentage and length of hospitalization are independent risk factors for the occurrence of IFI.
Fungal infections are a significant global public health challenge because of their widespread occurrence, morbidity, and profound social and economic consequences. Antifungal resistance is also an increasing concern, posing a substantial risk to public health. There is a growing interest in searching for new antifungal drugs isolated from natural sources. This study aimed to evaluate the antifungal activity of novel mollusk fractions against fungal strains resistant to nystatin and amphotericin B. In addition, the role of oxidative stress in the mechanism of damage was determined. The mucus from the garden snail Cornu aspersum (MCa/1-20) and the hemolymph fraction from the marine snail Rapana venosa (HLRv/3-100) were obtained and characterized via 12% sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometric -analyses. The results demonstrate that the spores and biomass of both mollusk fractions have a significant fungicidal effect against Penicillium griseofulvum, and Aspergillus niger. Compared to the control group, the release of intracellular proteins and reducing sugars was significantly increased in the treated groups. The data showed increased levels of oxidative stress biomarkers (lipid peroxidation and oxidatively damaged proteins) and a downregulated antioxidant enzyme defense, corresponding to increased antifungal activity. To our knowledge, this is the first study evaluating oxidative stress as a factor in mollusk fractions’ antifungal activity.
Covering: up to December 2023Decalin-containing tetramic acid derivatives, especially 3-decalinoyltetramic acids (3-DTAs), are commonly found as fungal secondary metabolites. Numerous biological activities of this class of compounds, such as antibiotic, antiviral, antifungal, antiplasmodial, and antiprotozoal properties, have been the subject of ongoing research. For this reason, these molecules have attracted a lot of interest from the scientific community and various efforts including semi-synthesis, co-culturing with bacteria and biosynthetic gene sequencing have been made to obtain more derivatives. In this review, 3-DTAs are classified into four major groups based on the absolute configuration of the bicyclic decalin ring. Their biosynthetic pathways, various biological activities, and structure-activity relationship are then introduced.
Vulvovaginal candidiasis, mostly caused by Candida albicans, remains a prevalent concern in women's health. Annona muricata L. (Annonaceae), a plant native from Brazil, is well-known for its therapeutic potential, including antitumor, anti-inflammatory, and antimicrobial properties. This study presents an innovative hydrogel formulation containing the ethanolic extract from A. muricata leaves designed to control C. albicans in an in vivo model of vulvovaginal candidiasis. Here, we report the development, thermal, physicochemical and rheological characterization of a Carbopol®-based hydrogel containing A. muricata extract. Furthermore, we evaluated its activity in a vulvovaginal candidiasis in vivo model. Thermal analyses indicated that the addition of the extract increased the polymer-polymer and polymer-solvent interactions. Rheological analysis showed a decrease in the viscosity and elasticity of the formulation as the A. muricata extract concentration increased, suggesting a liquid-like behavior. After treatment with the Carbopol®-based hydrogel with A. muricata, our in vivo results showed a significant reduction in vulvovaginal fungal burden and infection, as well as a reduction in mucosal inflammation. The current research opens up possibilities for the application of the Carbopol®-based hydrogel with A. muricata as a natural therapeutic option for the treatment of vulvovaginal candidiasis.
Six new β-resorcylic acid derivatives (1–5 and 7) were isolated from a halophyte-associated fungus, Colletotrichum gloeosporioides JS0419, together with four previously reported β-resorcylic acid lactones (RALs). The relative and absolute stereochemistry of 1 was completely established by a combination of spectroscopic data and chemical reactions. The structures of the isolated compounds were elucidated by analysis of HRMS and NMR data. Notably, compounds 1–3 had a β-resorcylic acid harboring a long unesterified aliphatic side chain, whereas the long aliphatic chains were esterified to form macrolactones in 4–9. Among the isolated compounds, monocillin I and radicicol showed potent antifungal activities against Cryptococcus neoformans, comparable to clinically available antifungal agents and radicicol showed weak antifungal activity against Candida albicans. These findings provide insight into the chemical diversity of fungal RAL-type compounds and their pharmacological potential.
Vulvovaginal candidiasis (VVC) is an inflammatory disease of the vagina mainly caused by Candida albicans (C. albicans), which affects around three-quarters of all women during their reproductive age. Although some antifungal drugs such as azoles have been applied clinically for many years, their therapeutic value is very limited due to the emergence of drug-resistant strains. Previous studies have shown that the adhesion of C. albicans to vaginal epithelial cells is essential for the pathogenesis of VVC. Therefore, preventing the adhesion of C. albicans to vaginal epithelial cells may be one of the most effective strategies for the treatment of VVC. Berberine (BBR) is a biologically active herbal alkaloid that was used to treat VVC. However, so far, its mechanism has remained unclear. This study shows BBR significantly inhibits the adhesion of C. albicans to vaginal epithelial cells by reducing the expressions of ICAM-1, mucin1, and mucin4 in vaginal epithelial cells, which play the most important role in modulating the adhesion of C. albicans to host cells, and balancing IL-2 and IL-4 expressions, which play a key effect on regulating the inflammatory response caused by C. albicans infection. Hence, our findings demonstrate that BBR may be a potential therapeutic agent for VVC by interfering with the adhesion of C. albicans to vaginal epithelial cells and represents a new pathway for developing antifungal therapies agents from natural herbs.
The incidence of fungal infections has significantly increased in recent years due to the emergence of antifungal resistance. Biofilm formation is considered to be a major contributor to both the infectious diseases and to antimicrobial resistance. Consequently, biofilm-associated infections are often problematic to treat with existing therapeutics. Adhesion of C. albicans to the host surface or implanted materials followed by hyphal invasion and biofilm formation enhances C. albicans pathogenicity and virulence. Thus, developing a therapeutic agent that inhibits candidal adherence, biofilm development and morphological switching could improve clinical management of infections. The present investigation studied two emerging and alternatives strategies, namely antibiofilm and combinatorial approach, to attenuate biofilm formation and the expression of Candida virulence factors. Piperine and thymol are major bioactive components of pepper and thyme, respectively. These phytochemicals are known to possess numerous biological activities, including recently reported antibiofilm effects against C. albicans. The minimum biofilm inhibitory concentration (MBIC) of both phytochemicals was determined to be 32 µg/ml. The phytochemical treatment of Candida biofilms using piperine and thymol revealed synergistic effects at four different combinations of concentrations, i.e. 8 and 8, 8 and 4, 8 and 2 and 4 and 8 µg/ml. These synergistic combinations resulted in the significant reduction in adherence of Candida, hyphal extension and morphological transformation. Moreover, limited exposure of synergistic combinations controlled the hyphal elongation. Results were validated through the gene expression analysis. Results from the present investigation suggest that piperine and thymol can be synergistically employed for the treatment of biofilm-associated C. albicans infection.
Vulvovaginal candidiasis (VVC) is the second most common vaginal infection that affects women of reproductive age. Its increased occurrence and associated treatment cost coupled to the rise in resistance of the causative pathogen to current antifungal therapies has necessitated the need for the discovery and development of novel effective antifungal agents for the treatment of the disease. We report in this study the anti-Candida albicans activity of Solanum torvum 70% ethanol fruit extract (STF), fractions and some isolated compounds against four (4) fluconazole-resistant strains of C. albicans. We further report on the effect of the isolated compounds on the antifungal activity of fluconazole and voriconazole in the resistant isolates as well as their inhibitory effect on C. albicans biofilm formation. STF was fractionated using n-hexane, chloroform (CHCl3) and ethyl acetate (EtOAc) to obtain four respective major fractions, which were then evaluated for anti-C. albicans activity using the microbroth dilution method. The whole extract and fractions recorded MICs that ranged from 0.25 to 16.00 mg/mL. From the most active fraction, STF- CHCl3 (MIC = 0.25–1.00 mg/mL), four (4) known compounds were isolated as Betulinic acid, 3-oxo-friedelan-20α-oic acid, Sitosterol-3-β-D-glucopyranoside and Oleanolic acid. The compounds demonstrated considerably higher antifungal activity (0.016 to 0.512 mg/mL) than the extract and fractions and caused a concentration-dependent anti-biofilm formation activity. They also increased the sensitivity of the C. albicans isolates to fluconazole. This is the first report of 3-oxo-friedelan-20α-oic acid in the plant as well as the first report of betulinic acid, sitosterol-3-β-D-glucopyranoside and oleanolic acid from the fruits of S. torvum. The present study has demonstrated the anti-C. albicans activity of the constituents of S. torvum ethanol fruit extract and also shown that the constituents possess anti-biofilm formation and resistance modulatory activities against fluconazole-resistant clinical C. albicans isolates.
Candida auris is an emerging healthcare-associated fungal pathogen that has become a serious global health threat. Current treatment options are limited due to drug resistance. New therapeutic strategies are required to target this organism and its pathogenicity. Plant polyphenols are structurally diverse compounds that present a vast range of biological properties. In the present study, plant-derived molecules ellagic acid (EA) and caffeic acid phenethyl ester (CAPE) were investigated for their antifungal and antivirulence activities against Candida auris. We also tested against C. albicans. The minimum inhibitory concentration (MIC) for EA ranged from 0.125 to 0.25 µg/mL and for CAPE ranged from 1 to 64 µg/mL against drug-resistant C. auris strains. Killing kinetics determined that after 4 h treatment with CAPE, there was a complete reduction of viable C. auris cells compared to fluconazole. Both compounds might act by modifying the fungal cell wall. CAPE significantly reduced the biomass and the metabolic activity of C. auris biofilm and impaired C. auris adhesion to cultured human epithelial cells. Furthermore, both compounds prolonged the survival rate of Galleria mellonella infected by C. auris (p = 0.0088 for EA at 32 mg/kg and p = 0.0028 for CAPE at 4 mg/kg). In addition, EA at 4 μg/mL prolonged the survival of C. albicans-infected Caenorhabditis elegans (p < 0.0001). CAPE was not able to prolong the survival of C. albicans-infected C. elegans. These findings highlight the antifungal and antivirulence effects of EA and CAPE against C. auris, and warrant further investigation as novel antifungal agents against drug-resistant infections.
Background
Current treatment of vulvovaginal candidiasis (VVC) is largely limited to azole therapy. Ibrexafungerp is a first-in-class triterpenoid antifungal with broad-spectrum anti-Candida fungicidal activity. The objective of this study was to evaluate the efficacy and safety of ibrexafungerp compared with placebo in patients with acute VVC.
Study Design
Patients were randomly assigned 2:1 to receive ibrexafungerp (300 mg twice for 1 day) or placebo. The primary endpoint was the percentage of patients with a clinical cure (complete resolution of vulvovaginal signs and symptoms [VSS]=0) at test-of-cure (day 11±3). Secondary endpoints included the percentage of patients with mycological eradication, overall success (clinical cure and mycological eradication), clinical improvement (VSS≤1) at test-of-cure, and symptom resolution at follow-up (day 25±4).
Results
Patients receiving ibrexafungerp had significantly higher rates of clinical cure (50.5% [95/188] vs 28.6% [28/98]; P=0.001), mycological eradication (49.5% [93/188] vs 19.4% [19/98]; P<0.001), and overall success (36.0% [64/178] vs 12.6% [12/95]; P<0.001) compared with placebo. Symptom resolution was sustained and further increased with ibrexafungerp compared with placebo (59.6% [112/188] vs 44.9% [44/98]; P=0.009) at follow-up. Post hoc analysis showed similar rates of clinical cure and clinical improvement at test-of-cure for African American patients (54.8% [40/73] and 63.4% [47/73], respectively) and patients with a body mass index >35 (54.5% [24/44] and 68.2% [30/44], respectively) compared with overall rates. Ibrexafungerp was well tolerated. Adverse events were primarily gastrointestinal and mild in severity.
Conclusion
Ibrexafungerp provides a promising safe and efficacious oral treatment that mechanistically differs from current azole treatment options for acute VVC.
Oral candidiasis is one of the most common types of fungal infection caused by Candida albicans (C. albicans). The present study aims to investigate the antifungal effects of phloretin (a dihydrochalcone flavonoid) against the C. albicans pathogenicity. In this work, we treated C. albicans SC5314 with 37.28, 74.55, or 149.10 μg/mL (equivalent to 0.5×, 1× or 2× MIC) phloretin in vitro. Besides, we established a mice model of oral candidiasis by a sublingual infection of C. albicans suspension (1 × 10⁷ colony-forming unit/mL), and mice were treated with phloretin (3.73 or 7.46 mg/mL, which were equivalent to 50× or 100× MIC) twice a day starting on day one post-infection. The results showed that the MIC of phloretin against C. albicans was 74.55 μg/mL. Phloretin exerted antifungal activity by inhibiting the biofilm formation and suppressing the yeast-to-hyphae transition upon the downregulation of hypha-associated genes including enhanced adherence to polystyrene 1, the extent of cell elongation gene 1, hyphal wall protein 1 gene, and agglutinin-like sequence gene 3. Next, phloretin repressed the secretion of proteases and phospholipases via reducing the expression of protease-encoding genes secreted aspartyl proteases (SAP)1 and SAP2, as well as phospholipase B1. Subsequently, the in vivo antifungal activity of phloretin was testified by the reverse of the enhanced lesion severity, inflammatory infiltration, and the increased colony-forming unit counts caused by C. albicans of tongue tissues in oral candidiasis mice. In conclusion, phloretin suppressed the pathogenicity and virulence factors against C. albicans both in vivo and in vitro.
Punica granatum L. (pomegranate) fruit is known to be an important source of bioactive phenolic compounds belonging to hydrolysable tannins. Pomegranate extracts have shown antifungal activity, but the compounds responsible for this activity and their mechanism/s of action have not been completely elucidated up to now. The aim of the present study was the investigation of the inhibition ability of a selection of pomegranate phenolic compounds (i.e., punicalagin, punicalin, ellagic acid, gallic acid) on both plant and human fungal pathogens. In addition, the biological target of punicalagin was identified here for the first time. The antifungal activity of pomegranate phenolics was evaluated by means of Agar Disk Diffusion Assay and minimum inhibitory concentration (MIC) evaluation. A chemoinformatic analysis predicted for the first time topoisomerases I and II as potential biological targets of punicalagin, and this prediction was confirmed by in vitro inhibition assays. Concerning phytopathogens, all the tested compounds were effective, often similarly to the fungicide imazalil at the label dose. Particularly, punicalagin showed the lowest MIC for Alternaria alternata and Botrytis cinerea, whereas punicalin was the most active compound in terms of growth control extent. As for human pathogens, punicalagin was the most active compound among the tested ones against Candida albicans reference strains, as well as against the clinically isolates. UHPLC coupled with HRMS indicated that C. albicans, similarly to the phytopathogen Coniella granati, is able to hydrolyze both punicalagin and punicalin as a response to the fungal attack. Punicalagin showed a strong inhibitory activity, with IC50 values of 9.0 and 4.6 µM against C. albicans topoisomerases I and II, respectively. Altogether, the results provide evidence that punicalagin is a valuable candidate to be further exploited as an antifungal agent in particular against human fungal infections.
Candida auris is an emerging pathogen that has rapidly spread to many countries on multiple continents. Invasive infections caused by this species are associated with significant mortality, and treatment options are limited due to antifungal resistance. Ibrexafungerp is the first-in-class member of the triterpenoids, which inhibit the production of (1, 3)-β-D-glucan and can be administered orally. We evaluated the in vitro activity and in vivo efficacy of ibrexafungerp against C. auris . Antifungal susceptibility was tested by broth microdilution against 54 C. auris isolates. Neutropenic mice were intravenously infected with a clinical isolate, and a 7-day treatment course began 24 hours post-inoculation with vehicle control, ibrexafungerp (20, 30, and 40 mg/kg orally twice daily), fluconazole (20 mg/kg orally once daily), or caspofungin (10 mg/kg intraperitoneally once daily). Fungal burden was assessed by colony counts in the kidneys on day 8, and on day 21 or as mice became moribund in the survival arm. Ibrexafungerp demonstrated consistent activity with MICs ranging between 0.25 to 2 μg/mL against all isolates. Marked improvements in survival were observed in mice treated with the higher doses of ibrexafungerp and caspofungin. Similarly, reductions in kidney fungal burden were also observed in these groups. No improvements in survival or reductions in fungal burden were observed with fluconazole, consistent with the in vitro resistance of the isolate used to establish infection to this azole. These results demonstrate that ibrexafungerp is effective in vivo against C. auris even when the start of therapy is delayed.
Cryptococcus neoformans is a serious human pathogen with limited options for treatment. We have interrogated extracts from fungal fermentations to find Cryptococcus-inhibiting natural products using assays for growth inhibition and differential thermosensitivity. Extracts from fermentations of four fungal strains from wild and domestic animal dung from Arkansas and West Virginia, USA were identified as Preussia typharum. The extracts exhibited two antifungal regions. Purification of one region yielded new 24-carbon macrolides incorporating both a phosphoethanolamine unit and a bridging tetrahydrofuran ring. The structures of these metabolites were established mainly by analysis of high-resolution mass spectrometry and 2D NMR data. Relative configurations were assigned using NOESY data, and the structure assignments were supported by NMR comparison with similar compounds. These new metabolites are designated preussolides A and B. The second active region was caused by the cytotoxin, leptosin C. Genome sequencing of the four strains revealed biosynthetic gene clusters consistent with those known to encode phosphoethanolamine-bearing polyketide macrolides and the biosynthesis of dimeric epipolythiodioxopiperazines. All three compounds showed moderate to potent and selective antifungal activity towards the pathogenic yeast C. neoformans.
Lack of novel antifungal agents and severe drug resistance has led to high incidence and associated mortality of invasive fungal infections. To tackle the challenges, novel antifungal agents with anti‐resistant potency are highly desirable. Thus, derivatives of curcumin were synthesized to restore the effectiveness of fluconazole (FLC) against FLC‐resistant Candida spp. and structure‐activity relationships were then discussed. Some novel derivatives showed promising features as novel antifungal lead compounds. Of them, compound 4 showed good alone or synergistic antifungal activity against FLC‐resistant Candida spp. Moreover, compound 4 was proven as a potent inhibitor of Candida albicans biofilm formation and yeast‐to‐hypha morphological transition whether used alone or in combination with FLC, which was further confirmed by the inhibitory effect on cellular surface hydrophobicity of C. albicans. Compound 4 also inhibits intracellular ATP production of C. albicans and disrupts membrane permeability of C. albicans when used in combination with FLC. The results highlighted the potential of curcumin derivatives to overcome fluconazole‐related and biofilm‐related drug resistance.
Four polyene macrolactams including the previously reported niizalactam C (4), and three new ones, streptolactams A–C (1–3) with a 26-membered monocyclic, [4,6,20]-fused tricyclic and 11,23-oxygen bridged [14,16]-bicyclic skeletons, respectively, were isolated from the fermentation broth of the deep-sea sediment-derived Streptomyces sp. OUCMDZ-3159. Their structures were determined based on spectroscopic analysis, X-ray diffraction analysis, and chemical methods. The abiotic formation of compounds 2 and 4 from compound 1 were confirmed by a series of chemical reactions under heat and light conditions. Compounds 1 and 3 showed a selective antifungal activity against Candida albicans ATCC 10231.
Candidiasis causes high morbidity and mortality among immunocompromised patients. Antifungal drug resistance and cytotoxicity highlight the need of effective antifungal therapeutics. In this study, we found that kalopanaxsaponin A (KPA), a triterpenoid saponin natural product, could inhibit the proliferation of various Candida species, and exerted a fungicidal effect against C. albicans. To further explore its antifungal action mode, spectrofluorophotometer, fluorescence microscopy and transmission electron microscopy were performed, showing that KPA treatment induced the accumulation of intracellular reactive oxygen species (ROS), resulting in mitochondrial dysfunction. Meanwhile, KPA treatment also broke down the membrane barrier of C. albicans causing the leakage of intracellular trehalose, the entrance of extracellular impermeable substance and the decrease of ergosterol content. Both ROS accumulation and membrane destruction contributed to the death of C. albicans cells. Our work preliminarily elucidated the potential mechanisms of KPA against C. albicans on a cellular level, and might provide a potential option for the treatment of clinical candidiasis.
Ergosterol, an essential constituent of membrane lipids of yeast, is distributed in both the cell membrane and intracellular endomembrane components such as vacuoles. Honokiol, a major polyphenol isolated from Magnolia officinalis, has been shown to inhibit the growth of Candida albicans. Here, we assessed the effect of honokiol on ergosterol biosynthesis and vacuole function in C. albicans. Honokiol could decrease the ergosterol content and upregulate the expression of genes related with the ergosterol biosynthesis pathway. The exogenous supply of ergosterol attenuated the toxicity of honokiol against C. albicans. Honokiol treatment could induce cytosolic acidification by blocking the activity of the plasma membrane Pma1p H+-ATPase. Furthermore, honokiol caused abnormalities in vacuole morphology and function. Concomitant ergosterol feeding to some extent restored the vacuolar morphology and the function of acidification in cells treated by honokiol. Honokiol also disrupted the intracellular calcium homeostasis. Amiodarone attenuated the antifungal effects of honokiol against C. albicans, probably due to the activation of the calcineurin signaling pathway which is involved in honokiol tolerance. In conclusion, this study demonstrated that honokiol could inhibit ergosterol biosynthesis and decrease Pma 1p H+-ATPase activity, which resulted in the abnormal pH in vacuole and cytosol. Keywords: Candida albicans, honokiol, ergosterol biosynthesis.
Background:
Urinary tract infections (UTIs) caused by opportunistic pathogens are among the leading health challenges globally. Most available treatment options are failing as a result of antibiotic resistance and adverse effects. Natural sources such as plants may serve as promising alternatives.
Methods:
Compounds were isolated from the South African weed Chromolaena odorata through column chromatography. Purified compounds were tested for antimicrobial activity using the p-iodonitrotetrazolium chloride (INT) colorimetric method, against uropathogenic Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Aspergillus fumigatus and Cryptococcus neoformans. Anti-biofilm, anti-adhesion and metabolic inhibition activities were investigated against selected strains. Safety of the compounds was determined against Vero monkey kidney, C3A human liver and colon (Caco2) cells.
Results:
Four compounds identified as pectolinaringenin (1), (±)-4',5,7-trimethoxy flavanone (2), 5-hydroxy-3,7,4'-trimethoxyflavone (3) and 3,5,7-trihydroxy-4'-methoxyflavone) (4) were isolated. Minimum inhibitory concentration (MIC) varied between 0.016 and 0.25 mg/mL. Compounds 2 and 3 showed promising antimicrobial activity against E. coli, S. aureus, K. pneumoniae, A. fumigatus and C. neoformans with MIC between 0.016 and 0.125 mg/mL, comparable to gentamicin, ciprofloxacin and amphotericin B used as positive controls. Compounds 2 and 3 showed good anti-biofilm and metabolic inhibition activities against E. coli and S. aureus but weak anti-adhesion activity against the organisms. Low toxicity with selectivity indexes between 1 and 12.625 were recorded with the compounds, indicating that the compounds were rather toxic to the microbial strains and not to the human and animal cells.
Conclusion:
Pharmacological activities displayed by compounds 2 and 3 isolated from C. odorata and low toxicity recorded credits it as a potential lead for the development of useful prophylactic treatments and anti-infective drugs against UTIs. Although known compounds, this is the first time these compounds have been isolated from the South African weed C. odorata and tested for antimicrobial, anti-biofilm, metabolic inhibition and anti-adhesion activities.
Candida albicans is the primary etiological agent associated with the pathogenesis of candidiasis. Unrestricted growth of C. albicans in the oral cavity may lead to oral candidiasis, which can progress to systemic infections in worst scenarios. Biofilm of C. albicans encompasses yeast and hyphal forms, where hyphal formation and yeast to hyphal morphological transitions are contemplated as the key virulence elements. Current clinical repercussions necessitate the identification of therapeutic agent that can limit the biofilm formation and escalating the susceptibility of C. albicans to immune system and conventional antifungals. In the present study, a plant-derived alkaloid molecule, piperine, was investigated for the antibiofilm and antihyphal activities against C. albicans. Piperine demonstrated a concentration-dependent antibiofilm activity without exerting negative impact on growth and metabolic activity. Inhibition in the hyphal development was witnessed through confocal laser-scanning microscopy and scanning electron microscopy. Interestingly, piperine displayed a tremendous potential to inhibit the virulence-associated colony morphologies, such as filamentation and wrinkling. Furthermore, piperine regulated morphological transitions between yeast and hyphal forms by inhibiting hyphal extension and swapping hyphal phase to yeast forms yet under filamentation-inducing circumstances. Remarkably, piperine-challenged C. albicans exhibited low potential for spontaneous antibiofilm resistance development. In addition, piperine effectively reduced in vivo colonization and prolonged survival of C. albicans-infected Caenorhabditis elegans, thereby expounding the distinct antivirulent potential. Transcriptomic analysis revealed piperine significantly downregulating the expression of several biofilm related and hyphal-specific genes (ALS3, HWP1, EFG1, CPH1, etc.). Furthermore, no acute toxicity was observed in the HBECs and nematodes exposed to piperine. Altogether, results from this study reveals the potential of piperine to inhibit biofilm and hyphal morphogenesis, and its in vivo efficacy and innocuous nature to HBECs suggests that piperine may be considered as a potential candidate for the treatment of biofilm-associated C. albicans infection, especially for oral candidiasis.
Purpose:
We characterized the effects of Honokiol (HNK) on Aspergillus fumigatus-caused keratomycosis and the underlying mechanisms. HNK is known to have anti-inflammatory and antifungal properties, but the influence on fungal keratitis (FK) remains unknown.
Methods:
In ex vivo, minimum inhibitory concentration and Cell Count Kit-8 assay were carried out spectrophotometrically to provide preferred concentration applied in vivo. Time kill assay pointed that HNK was fungicidal and fungistatic chronologically. Adherence assay, crystal violet staining, and membrane permeability assay tested HNK effects on different fungal stages. In vivo, clinical scores reflected the improvement degree of keratitis outcome. Myeloperoxidase (MPO) assay, flow cytometry (FCM), and immunohistofluorescence staining (IFS) were done to evaluate neutrophil infiltration. Plate count detected HNK fungicidal potentiality. RT-PCR, Western blot, and enzyme-linked immunosorbent assay (ELISA) verified the anti-inflammatory activity of HNK collaboratively.
Results:
In vitro, MIC90 HNK was 8 µg/mL (no cytotoxicity), and Minimal Fungicidal Concentration (MFC) was 12 µg/mL for A. fumigatus. HNK played the fungistatic and fungicidal roles at 6 and 24 hours, respectively, inhibiting adherence at the beginning, diminishing biofilms formation, and increasing membrane permeability all the time. In vivo, HNK improved C57BL/6 mice outcome by reducing disease severity (clinical scores), neutrophil infiltration (MPO, FCM, and IFS), and fungal loading (plate count). RT-PCR, Western blot, and ELISA revealed that HNK downregulated mRNA and protein expression levels of Toll-like receptor-2 (TLR-2), high mobility group box 1 (HMGB1), IL-1β, and TNF-α.
Conclusions:
Our study suggested HNK played antifungal and anti-inflammatory roles on keratomycosis by reducing survival of fungi, infiltration of leucocytes, and expression of HMGB1, TLR-2, and proinflammatory cytokines, providing a potential treatment for FK.
Notonesomycin A is a 32-membered bioactive glycosylated macrolactone known to be produced by Streptomyces aminophilus subsp. notonesogenes 647-AV1 and S. aminophilus DSM 40186. In a high throughput antifungal screening campaign, we identified an alternative notonesomycin A producing strain, Streptomyces sp. A793, and its biosynthetic gene cluster. From this strain, we further characterized a new more potent antifungal non-sulfated analogue, named notonesomycin B. Through CRISPR–Cas9 engineering of the biosynthetic gene cluster, we were able to increase the production yield of notonesomycin B by up to 18-fold as well as generate a strain that exclusively produces this analogue.
Edible mushrooms are an important source of nutraceuticals and for the discovery of bioactive metabolites as pharmaceuticals. In this work, the OSMAC (One Strain, Many Active Compounds) approach was used to isolate two new compounds (1 and 2) along with seven known compounds (3–9) from a mycelial culture of a unique North American edible mushroom Hericium sp. The fruiting body was collected in Marine on St. Croix, Minnesota (USA), and mycelial cultures were grown on four different solid and liquid media. Extracts from the mycelial cultures were screened for antimicrobial activity and only the extract from the Cheerios substrate culture exhibited antifungal activity. Bioassay guided fractionation and HPLC analysis were used to isolate nine pure compounds and the structures of the known compounds were established by analysis of the NMR and mass spectrometry data and comparison to published reports. Compound 1 is a new erinacerin alkaloid and 2 is an aldehyde derivative of 4-hydroxy chroman. Four chlorinated orcinol derivatives (3–6), a pyran (7), erinaceolactone (8), and erinacine (9) were identified. Compound 4 showed antifungal activity against C. albicans and C. neoformans (MIC = 31.3–62.5 μg/mL, respectively). Compound 4 also inhibited biofilm formation of C. albicans and C. neoformans at 7.8 μg/mL. These results suggest that mycelial cultures of edible fungi may provide useful, bioactive compounds.
Background
The pathogenicity of Candida albicans is attributed to various virulence factors including adhesion to the surface of epithelial cells or mucosa, germ tube formation, hyphal morphogenesis, development of drug resistant biofilms, and so on. The objective of this study was to investigate the effects of Kalopanaxsaponin A (KPA) on the virulence of C. albicans.
Methods
The effect of KPA on the virulence of C. albicans was characterized by an XTT reduction assay and fluorescent microscopic observation. The action mechanism was further explored using GC/MS system and BioTek Synergy2 spectrofluorophotometry. The cytotoxicity and therapeutic effect of KPA were evaluated by the Caenorhabditis elegans-C. albicans infection model in vivo.
Results
The minimum inhibitory concentration (MIC) of KPA was 8∼16 μg/mL for various genotypes of C. albicans. The compound was identified as having remarkable effect on the adhesion, morphological transition and biofilm formation of C. albicans. The results of fluorescent microscopy and GC/MS system suggested that KPA could promote the secretion of farnesol by regulating the expression of Dpp3 and decrease the intracellular cAMP level, which together inhibited morphological transition and biofilm formation. Notably, KPA showed low toxicity in vivo and a low possibility of developing resistance.
Conclusion
Our results demonstrated that KPA had remarkable efficacy against C. albicans pathogenicity, suggesting that it could be a potential option for the clinical treatment of candidiasis.
FK506, also known as tacrolimus, is a clinically important immunosuppressant drug and has promising therapeutic potentials owing to its antifungal, neuroprotective, and neuroregenerative activities. To generate various FK506 derivatives, the structure of FK506 has been modified by chemical methods or biosynthetic pathway engineering. Herein, we describe the mode of the antifungal action of FK506 and the structure-activity relationship of FK506 derivatives in the context of immunosuppressive and antifungal activities. In addition, we discuss the neurotrophic mechanism of FK506 known to date, along with the neurotrophic FK506 derivatives with significantly reduced immunosuppressive activity. This review suggests the possibility to generate novel FK506 derivatives as antifungal as well as neuroregenerative/neuroprotective agents.
The antifungal activity of magnolol and honokiol, two naturally occurring hydroxylated biphenyls, and of their synthetic derivatives was evaluated on a collection of representative isolates of Fusarium oxysporum, F. solani and F. verticillioides of clinical and ecological concern. The tested compounds were proposed as a ‘natural’ alternative to conventional fungicides, even though a larger range of concentrations (5–400 μg/ml) was applied. The activity of magnolol and honokiol was compared with that of terbinafine (0.1–10 μg/ml), and fluconazole (1–50 μg/ml), two fungicides widely used in treating fungal infections on humans. Magnolol showed similar fungicidal activity compared to fluconazole, whereas honokiol was more effective in inhibiting mycelium growth compared to this fungicide on all tested clinical Fusarium spp. isolates. Compared to terbinafine, honokiol showed similar antifungal activity when tested on clinical F. solani isolates, whereas magnolol was less effective at all selected concentrations (5–400 μg/ml). The different position of the phenol-OH group, as well as its protection, explain different in vitro activities between magnolol, honokiol, and their derivatives. Furthermore, magnolol showed mycelium dry weight reduction at a concentration of 0.5 mM when tested on a set of agricultural isolates of Fusaria, leading to complete inhibition of some of them. Magnolol and honokiol are proposed as efficient and safe candidates for treating clinically relevant Fusaria.
Fungal drug resistance is a major health threat, and reports of clinical resistance worldwide are becoming increasingly common. In a research program to discover new molecules to help overcome this problem, 14 new lanostane-type triterpenoids, gibbosicolids A-G (2-8) and gibbosic acids I-O (9-15), were isolated from the fruiting bodies of Ganoderma gibbosum, along with seven known triterpenoid derivatives. These compounds featured high levels of oxidation, epimerization, and γ-lactonization. Structures were elucidated by comprehensive spectroscopic analyses and HRMS data. Absolute configurations were assigned based on quantum chemical calculations, including calculated chemical shift with DP4+ analysis, coupling constants, and electronic circular dichroism (ECD) methods. Results show that the calculated NMR with DP4+ analysis could not reliably establish the overall spatial configuration of molecules possessing independent and free-rotational stereoclusters. All these compounds significantly increased the sensitivity of fluconazole (FLC)-resistant C. albicans to FLC. Compounds 2, 5, 9, 12, 16, 17, and 21 exhibited strong antifungal activity against FLC-resistant C. albicans when combined with FLC, with MIC50 values ranging from 3.8 to 8.8 μg/mL.
Four new triterpenoids, 3β,12β,16β,21β,22-pentahydroxyhopane (1), 12β,16β,21β,22-tetrahydroxyhopan-3-one (2), 3-oxo-olean-12-ene-28,30-dioic acid (3), and 3β-hydroxyoleana-11,13(18)-diene-28,30-dioic acid 30-methyl ester (4); 21 new triterpenoid saponins, glinusopposides A–U (5–25); and 12 known compounds (26–37) were isolated from the whole plants of Glinus oppositifolius. The structures of the new compounds were elucidated based on the analysis of one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR) and mass spectrometry (MS) data. All compounds from the plants were measured for antifungal activities against Microsporum gypseum and Trichophyton rubrum. Glinusopposide B (6), glinusopposide Q (21), glinusopposide T (24), and glinusopposide U (25) showed strong inhibitory activities against M. gypseum (MIC50 7.1, 6.7, 6.8, and 11.1 μM, respectively) and T. rubrum (MIC50 14.3, 13.4, 11.9, and 13.0 μM, respectively). For those active compounds with an oleanane skeleton, glycosylation (21–26) or oxidation (3) of 3-OH was helpful in increasing the activity; replacement of the 30-methyl group (29) by a carboxymethyl group (26) enhanced the activity; the presence of 11,13(18) double bonds (20) decreased the activity.
Background: Resistance of bacteria and fungi to antibiotics is one of the biggest problems that faces public health. The present work was designated to evaluate the antimicrobial activities of saponins from Melanthera elliptica and their synergistic effects with standard antibiotics against pathogenic phenotypes. The plant extract was prepared by maceration in methanol. The methanol extract was partitioned into ethyl acetate and n-butanol extracts. Column chromatography of the n-butanol extract followed by purification of different fractions led to the isolation of four saponins. Their structures were elucidated on the basis of spectra analysis, and by comparison with those from the literature. The antimicrobial activities of the extracts/compounds alone and their combinations with tetracycline and fluconazole were evaluated using the broth microdilution method through the determination of minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC).
Results: Four compounds: 3-O-β-D-glucuronopyranosyl-oleanolic acid (1), 3-O-β-D-glucuronopyranosyloleanolic acid 28-O-β-D-glucopyranosyl ester (2), 3-O-β-D-glucopyranosyl(1→2)-β-D-glucuronopyranosyl oleanolic acid (3) and 3-O-β-D-glucopyranosyl(1→2)-β-D-glucuronopyranosyl oleanolic acid 28-O-β-D-glucopyranosyl ester (4) were isolated. Compounds 1, 2 and 3 showed the largest antibacterial activities (MIC = 8 – 128 μg/ml) whereas compound 4 displayed the highest antifungal activities (MIC = 8 – 16 μg/ml). The antibacterial activities of compounds 1 and 2 (MIC = 16 – 32 μg/ml) against multi-drug-resistant Escherichia coli S2 (1) and Shigella flexneri SDINT are equal to those of vancomycin (MIC = 16 – 32 μg/ml) used as reference antibiotic.
Conclusions: The present study showed significant antimicrobial activity of compounds 1, 2, 3 and 4 against the tested microorganisms. The saponins act in synergy with the tested standard antibiotics. This synergy could lead to new options for the treatment of infectious diseases and emerging drug resistance.
Purpose
To investigate the effect of Glabridin (GLD) in Aspergillus fumigatus keratitis and its associated mechanisms.
Methods
Aspergillus fumigatus (A. fumigatus) conidia was inoculated in 96-well plate, and minimal inhibitory concentration (MIC) and biofilm formation ability were evaluated after GLD treatment. Spore adhesion ability was evaluated in conidia infected human corneal epithelial cells (HCECs). Keratitis mouse model was created by corneal intrastromal injection with A. fumigatus conidia, and GLD treatment started at the day after infection. The number of fungal colonies was calculated by plate count, and degree of corneal inflammation was assessed by clinical score. Flow cytometry, myeloperoxidase (MPO), and immunofluorescence staining (IFS) experiments were used to assess neutrophil infiltrations. PCR, ELISA and Western blot were conducted to determine levels of TLR4, Dectin-1 as well as downstream inflammatory factors.
Results
GLD treatment suppressed the proliferation, biofilm formation abilities and adhesive capability of A. fumigatus. In mice upon A. fumigatus infection, treatment of GLD showed significantly decreased severity of corneal inflammation, reduced number of A. fumigatus in cornea, and suppressed neutrophil infiltration in cornea. GLD treatment obviously inhibited mRNA and protein levels of Dectin-1, TLR4 and proinflammatory mediators such as IL-1β, HMGB1, and TNF-α in mice corneas compared to the control group.
Conclusion
GLD has antifungal and anti-inflammatory effects in fungal keratitis through suppressing A. fumigatus proliferation and alleviating neutrophil infiltration, and repressing the expression of TLR4, Dectin-1 and proinflammatory mediators.
Ibrexafungerp (BREXAFEMME®) is an orally active triterpenoid antifungal drug being developed by SCYNEXIS, Inc. for the treatment of fungal infections. The inhibition of β-1,3-d glucan synthetase by ibrexafungerp compromises the integrity of fungal cell walls. Ibrexafungerp has been recently approved for the treatment of vulvovaginal candidiasis (VVC), and it is the first novel antifungal drug class to be approved in more than 20 years. Development for the treatment of recurrent VVC and invasive fungal infections is ongoing. This article summarizes the milestones in the development of ibrexafungerp leading to this first approval for the treatment of VVC in adult and postmenarchal paediatric females.
Background:
Voriconazole (VRC) is widely recommended as the first-line therapy for invasive aspergillosis. However, surveillance studies have demonstrated that there is an increase in the frequency of azole resistance among Aspergillus fumigates isolates. In recent years, more studies on effective synergisms between natural agents and antifungal drugs have been published.
Aims:
To evaluate the synergistic antifungal effect of glabridin (Gla) and VRC against A. fumigatus isolates.
Methods:
Potential interactions between Gla and VRC were studied by using a microdilution checkerboard method based on the CLSI reference technique. To assess the interaction of drugs the fractional inhibitory concentration index (FICI) was calculated based on the Loewe Additivity model.
Results:
The minimum inhibitory concentrations (MIC) obtained with Gla alone were relatively high (MIC50 16μg/ml). However, our results showed synergistic interaction between Gla and VRC against A. fumigatus strains, with FICI range values between 0.15 and 0.5.
Conclusions:
Synergistic activity of Gla and VRC against both VRC-sensitive and -resistant A. fumigatus isolates may lead to design new antifungal agents, especially for inhibiting those azole-resistant strains.
Seven new polyketides including a phenol (1), two diphenyl ethers (2 and 3), two depsidones (4 and 5), and two phthalides (6 and 7) were isolated from the fungus Aspergillus unguis PSU-MF16 along with 27 known compounds. Their structures were determined by extensive spectroscopic analysis. The absolute configurations of 1 and 4-7 were established using comparative analyses of calculated and experimental ECD spectra. Among the new metabolites, 2 exhibited the best antimicrobial activity against Staphylococcus aureus, methicillin-resistant S. aureus, and Microsporum gypseum with equal MIC values of 16 μg/mL. In addition, known emeguisin A displayed potent antimicrobial activity against S. aureus, methicillin-resistant S. aureus, and Cryptococcus neoformans with equal MIC values of 0.5 μg/mL, compared with the standard drugs, vancomycin and amphotericin B. The structure-activity relationship study of the isolated compounds for antimicrobial activity is discussed.
Bioactivity-driven LC/MS-based phytochemical analysis of the root bark extract of Ulmus davidiana var. japonica led to the isolation of 10 compounds including a new coumarin glycoside derivative, ulmusakidian (1). The structure of the new compound was elucidated using extensive spectroscopic analyses via 1D and 2D NMR spectroscopic data interpretations, HR-ESIMS, and chemical transformation. The isolated compounds 1–10 were tested for their antifungal activity against human fungal pathogens Cryptococcus neoformans and Candida albicans. Compounds 9 and 10 showed antifungal activity against C. neoformans, with the lowest minimal inhibitory concentration (MIC) of 12.5–25.0 µg/mL, whereas none of the compounds showed antifungal activity against C. albicans.
In our screening program for new biologically active compounds, a new polyene macrolide, lavencidin (1), along with known compound RKGS-A2215A (2), was isolated from the fermentation broth of Streptomyces lavendulae FRI-5 by changing the composition of liquid medium normally used for the strain. Their structures were elucidated by spectral methods (high-resolution fast-atom bombardment mass spectrometry (HRFABMS) and nuclear magnetic resonance (NMR)). Compound 1 includes a conjugated pentaene moiety together with six hydroxy groups and a carboxylic acid as a side chain. Lavencidin (1) showed moderate growth-inhibitory activity against yeast and was cytotoxic against human cancer cell lines with low-micromolar IC50 values.
Objective:
In order to prevent and control the infection of Candida albicans, the antifungal activity, possible mechanism of myriocin against C. albicans and its biofilm were studied.
Methods:
The antifungal activity of myriocin was investigated by microdilution method. The effect of myriocin on fungal cell wall or membrane was evaluated by adding sorbitol, ergosterol or phytosphingosine (PHS). The damage to the cell membrane was investigated with propidium iodide (PI) staining and visualized by Scanning electron microscope (SEM). The effects on biofilms and extracellular polysaccharides (EPS) were observed by crystal violet staining method and phenol-sulfuric acid method respectively. The adhesion of C. albicans cells to hydrocarbons was tested to evaluate cell surface hydrophobic (CSH). The combined effects of myriocin and antifungal drugs commonly used in clinical practice were investigated by using the checkerboard microdilution method.
Results:
MICs were found to be 0.125~4 µg/ml. Myriocin was found to affect both cell wall and cell membrane. After exposure to myriocin, biofilm and EPS were found to be inhibited and removed, and the CSH was decreased. The combined fungistasis of myriocin and voriconazole (VCZ) or amphotericin B (AMB) were additive.
Conclusion:
Myriocin had significant antifungal activity against C. albicans, and the antifungal mechanisms might be cell wall and membrane damage. Myriocin effectively inhibited and eliminated biofilms, and its mechanism may be related to the inhibition of EPS and CSH.
We previously reported medicinal chemistry efforts that identified MK-5204, an orally efficacious β-1,3-glucan synthesis inhibitor derived from the natural product enfumafungin. Further extensive optimization of the C2 triazole substituent identified 4-pyridyl as the preferred replacement for the carboxamide of MK-5204, leading to improvements in antifungal activity in the presence of serum, and increased oral exposure. Reoptimizing the aminoether at C3 in the presence of this newly discovered C2 substituent, confirmed that the (R) t-butyl, methyl aminoether of MK-5204 provided the best balance of these two key parameters, culminating in the discovery of ibrexafungerp, which is currently in phase III clinical trials. Ibrexafungerp displayed significantly improved oral efficacy in murine infection models, making it a superior candidate for clinical development as an oral treatment for Candida and Aspergillus infections.
Six new terpenoids (1-6) and two new phloroglucinol glucosides (7 and 8) were isolated from the extract of Syzygium szemaoense leaves. Among the isolates, compounds 1 and 2 (named syzygiumursanolides A and B) were unusual 28-norursane type triterpenoids with 19(18→17)-abeo spirocyclic skeleton. And syzygiumone B (8) was rare ascorbyl-modified phloroglucinol glucoside. Their structures were elucidated on the basis of comprehensive 1D and 2D NMR, HRESIMS spectroscopic data analysis, as well as experimental and calculated ECD spectra and acid hydrolysis. Antimicrobial bioassay revealed that syzygiumursanolide D (4) displayed the most potent antifungal activities with MIC values ranging from 6.25 to 25 μg/mL against a panel of fungi.
Ethnopharmacological relevance
Boesenbergia rotunda (L.) Mansf. (Zingiberaceae) is an indigenous plant of Southeast Asia. Based on ethnopharmacological use, the rhizome is recommended in the treatment of stomachache, leukoplakia, abscesses, and leukorrhea in Thailand primary health care system. Candida albicans often causes leukorrhea, and infection of many mucosal sites. Its infection leads to serious illness.
Aim of the study
: This study aimed to investigate the effects of the ethanolic extract of the B. rotunda rhizome on C. albicans ATCC10231 in the stages of planktonic and biofilm formation and to explore the underlying mechanisms.
Materials and methods
The chemical composition of the extract was determined using ultra-performance liquid chromatography (UPLC). The planktonic growth of C. albicans was evaluated by the microdilution method, following EUCAST guidelines. For each stage of biofilm formation, the biofilm was assessed by the MTT assay. The biofilm structure was examined under a light microscope. The degree of cell surface hydrophobicity was measured. The mRNA levels of ALS1, ALS3, and ACT1 were determined by RT-qPCR.
Results
The extract of B. rotunda consisted of 25% (w/w) pinostrobin and 12% (w/w) pinocembrin. All stages of C. albicans biofilm formation were significantly inhibited by the extract, whereas the planktonic growth did not change. Biofilm development greatly decreased due to the extract in a concentration-dependent manner, with an IC50 value of 17.7 μg/mL. Pinostrobin and pinocembrin demonstrated inhibitory effects during this stage. These results were in accordance with the microscopic evaluation. The filamentous form decreased with pinocembrin rather than pinostrobin. Moreover, the cell surface hydrophobicity was significantly decreased by 6.25 and 12.5 μg/mL of the extract and 100 μM of pinocembrin. The ALS3 mRNA level was noticeably decreased by 12.5 μg/mL of the extract, 100 μM of pinostrobin, and 100 μM of pinocembrin. The ACT1 mRNA level decreased significantly with pinocembrin. However, the ALS1 mRNA level was not altered following all treatments.
Conclusion
The ethanolic extract of B. rotunda could inhibit biofilm formation of C. albicans, especially during the biofilm development stage, by means of reducing the cell surface hydrophobicity and suppressing the ALS3 mRNA expression. Pinocembrin had a stronger effect on ALS3 mRNA expression than pinostrobin. Only pinocembrin significantly decreased the ACT1 mRNA level.
Candida albicans is a member of pathogens with potential drug resistance threat that needs novel chemotherapeutic strategies. Considering the multifarious biological activities including bioenhancer activity, anti-Candida potential of piperine was evaluated against planktonic/biofilm and hyphal growth of C. albicans alone or in combination as a synergistic agent with fluconazole. Piperine inhibits planktonic growth at or less than 15 μg/ml, hyphae induction at 5 μg/ml concentration, and exhibits stage-dependent activity against biofilm growth of a fluconazole-resistant strain of C. albicans (ATCC10231). Though piperine couldn't kill inoculum completely at minimum inhibitory concentration (MIC), it is fungicidal at higher concentrations, as shown in apoptosis assay. FIC index values indicate that piperine exhibits excellent synergistic activity with fluconazole against planktonic (0.123) and biofilm (0.215) growth of an FLC resistant strain. Mode of anti-Candida activity was studied by identifying piperine responsive proteins wherein the abundance of 25 proteins involved in stress response, signal transduction and cell cycle were modulated (22 up and 3 down-regulated) significantly in response to piperine (MIC50). Modulation of the proteins involved suggests that piperine affects membrane integrity leading to oxidative stress followed by cell cycle arrest and apoptosis in C. albicans. Flow cytometry-based mitochondrial membrane potential (MMP), cell cycle and apoptosis assay, as well as real-time quantitative polymerase chain reaction analysis of selected genes, confirms piperine induced oxidative stress (TRR1), cell cycle arrest and apoptosis (CaMCA1). Based on our results, we conclude that piperine inhibits planktonic and difficult-to treat-biofilm growth of C. albicans by affecting membrane integrity thereby inducing oxidative stress and apoptosis.
Lay abstract:
Piperine inhibit Candida albicans growth (planktonic and biofilm) significantly in our study. Piperine exhibits excellent synergistic potential with fluconazole The proteome analysis suggests that piperine induced membrane damage leads to oxidative stress followed by cell cycle arrest and apoptosis.
Woodfordia uniflora is a flowering shrub unique to the Dhofar region of Oman and is used locally as a sedative and remedy for skin infection. However, no study to date has examined the pharmacological properties of this plant, and studies regarding phytochemicals present in W. uniflora are limited. Herein, phytochemical screening of the extract of W. uniflora was performed using LC/MS. Three new phenolic compounds, (±)-woodfordiamycin (1), woodfordic acid (2), and rhamnetin 3-O-(6″-galloyl)-β-d-glucopyranoside (3), together with 16 known compounds 4-19, were isolated from the antifungal fraction of the extract. The structures of the new compounds were established by NMR and HR-MS data, and their absolute configurations were established using chemical transformations, including Mosher's method, comparison of experimental and calculated electronic circular dichroism data, and gauge-including atomic orbital NMR chemical shift calculations, followed by DP4+ analysis. The isolated compounds (1-19) were tested for antifungal activity against human fungal pathogens Cryptococcus neoformans and Candida albicans. Compounds (±)-1 and 8 showed antifungal activity against C. neoformans, with the lowest minimum inhibitory concentrations of 1.8-1.9 μM, which was ∼10-fold lower than that of the currently available antifungal drug fluconazole, while (±)-1, 8, and 19 showed antifungal activity against C. albicans.
Two new trichothecene sesquiterpenes, trichobreols D (1) and E (2), were isolated from the culture broth of marine-derived Trichoderma cf. brevicompactum together with trichobreol A (3). The structures of 1 and 2 were assigned on the basis of their spectroscopic data. Compound 1 inhibited the growth of two yeast-like fungi, Candida albicans and Cryptococcus neoformans, with equivalent MIC values (6.3 μg/mL), while 2 gave MIC values of 12.5 and 25 μg/mL, respectively. The antifungal activities of five semisynthetic derivatives (4–8) prepared from 3 were evaluated and compared to investigate the preliminary structure-activity relationship.
Fourteen new terpenoids plagicosins A-N (1-14), including seven sesquiterpenoids (1-7) consisting of six ent-bicyclogermacrenes and one ent-2,3-seco-aromadendrane, as well as seven diterpenoids (8-14) comprising five fusicoccanes, a eunicellane, and a rare gersemiane, were isolated from the Chinese liverwort Plagiochila fruticosa Mitt. The structures of these terpenoids were determined on the basis of comprehensive analysis of MS and NMR spectroscopic data coupled with electronic circular dichroism (ECD) and coupling constant calculations. Plagicosin F (6) displayed potent antivirulence activity through inhibiting the hyphal morphogenesis, adhesion, and biofilm formation of Candida albicans. The genes related to hyphal formation were regulated by 6.
Iseolides A–C (1–3), three new glycosylated macrolides, were identified from the culture extract of Streptomyces sp. DC4-5 isolated from a stony coral Dendrophyllia. Extensive analysis of one- and two-dimensional NMR data, coupled with MS/MS analytical data, revealed that iseolides are new congeners of 36-membered macrolides, PM100117 and PM100118, previously reported from a marine-derived Streptomyces. Iseolides showed potent antifungal activity against a plant pathogen Glomerella cingulata and human pathogens Candida albicans and Trichophyton rubrum with MIC in the range of 0.19–6.25 μg/mL.
Genome-guided discovery of a novel linear polyene-polyol that induces ergosterol aggregation and inhibits hyphal growth.
Latterly, the upsurge in use of antifungal drugs has brought about the emergence of several drug-resistance strains, making it skeptical to continue relying on current therapeutic regime. In the necessity of resistance-free antifungal agent, flavonoids presented possibilities of replacing existing drugs, displaying antifungal activity against pathogenic fungi. Among them, quercetin, one of the most representative flavonoids, exhibited antifungal activity against Candida albicans. To inspect the further understanding regarding quercetin, the antifungal mode of action of quercetin was investigated. In the initial step, the apoptosis was monitored after quercetin treatment. Moreover, intracellular levels of Mg2+ was assessed and was determined that Mg2+ increase occurred under the influence of quercetin. In addition, several features of mitochondrial dysfunction were monitored. Mitochondrial dysfunction triggers decrease in mitochondrial redox levels and leads to disruption in mitochondrial antioxidant system. Increased intracellular ROS and decreased intracellular redox levels were also displayed, indicating the occurrence of overall disruption in antioxidant systems. Sequentially, DNA fragmentation was observed and this DNA damage in turn induces apoptosis. In analyses, hexaamminecobalt(III) chloride (Cohex) was applied to inhibit Mg2+ transport between cytosol and mitochondria. Cohex attenuated the effects induced by quercetin, which demonstrates that the presence of Mg2+ is essential in quercetin-induced apoptosis.
The resistance of pathogenic fungi and failure of drug therapy increased dramatically. Numerous studies have reported the individual or synergistic antifungal potency of natural and synthesized flavonoids, especially against drug-resistant fungi. This brief review summarizes the structure and individual or synergistic antifungal activity of natural and synthesized flavonoids (literatures mainly cover the past 10 years 2009-2019), with a special focus on the antifungal spectra, structure-activity relationship and mechanisms of actions. These may contribute to a better understanding of flavonoids as multi-target agents in the treatment of mycoses and provide some ideas on the development of novel flavonoids-based antifungals.
A reduction in the strong immunosuppressive activity of FK506 (1) is essential for developing this compound as an antifungal agent. Seven new FK506 analogues modified at both the FK506-binding protein 12- and the calcineurin-binding regions were biosynthesized. 9-DeoxoFK520 (7) exhibited a >900-fold reduction in the in vitro immunosuppressive activity but maintained significant antifungal activity, indicating that the C-9 and C-21 positions are critical for separation of immunosuppressive and antifungal activities. 7 exhibited robust synergistic antifungal activity with fluconazole. FK506 (1) is a 23-membered macrolide produced by several Streptomyces species and is used as an immunosuppressive drug to prevent the rejection of transplanted organs. FK506 has also exhibited antifungal, neuroprotective, and neuroregenerative activities. In humans, FK506 binds to FK506-binding protein (FKBP) 12, and the resulting FKBP12–FK506 complex interacts with a Ca²⁺-calmodulin-dependent phosphatase, calcineurin (CaN). Inactivation of CaN by forming the FKBP12–FK506–CaN ternary complex prevents the activation of nuclear factor of activated T cells (NF-AT), inhibiting the production of interleukin-2 and subsequent T-cell proliferation. This CaN signaling pathway also plays a critical role in the growth and pathogenesis of major fungal pathogens such as Cryptococcus neoformans, Candida albicans, and Aspergillus fumigatus. Therefore, the synthesis of FK506 analogues that can discriminate human FKBP12/CaN from its fungal counterparts may separate antifungal activity from the immunosuppressive activity, thereby allowing the development of a novel antifungal agent.
Cladosporine A (1), a new indole diterpenoid alkaloid, was isolated from the extract of a fungal strain Cladosporium sp. JNU17DTH12-9-01. Its structure was elucidated by extensive spectroscopic analysis, and the absolute configurations were determined by electronic circular dichroism (ECD) experiments. This is the first report of the presence of indole diterpenoid alkaloid in the genus Cladosporium. The antimicrobial activities against Staphylococcus aureus 209P, Escherichia coli ATCC0111, Aspergillus niger R330, and Candida albicans FIM709 were evaluated. Compound 1 showed MICs of 4 μg/mL and 16 μg/mL against S. aureus 209P and C. albicans FIM709, respectively.
Three natural glycosylated macrolide compounds, known irumamycin 1 and X-14952B 2, as well as new isoirumamycin 3, were isolated from ethyl acetate mycelium extract of Streptomyces sp. INA-Ac-5812. Structures of the compounds were elucidated using 1D and 2D NMR. Isoirumamycin 3 was found to be an isomer of irumamycin with an 18-membered macrolactone ring instead of 20-membered macrolide in irumamycin. A previously unknown stereo configuration of irumamycin epoxide (C23,C24) and hemiketal (C3,C7) fragments was deduced from NMR data (ROESY/NOESY and HSQMBC). Cytotoxic, antifungal and antibacterial activities were studied for all isolated compounds. Comparison of the collected data showed crucial importance of 20-membered macrolactone ring for antimicrobial properties of this antibiotic family.
Three new sesquiterpenes (1-3) and one new monoterpene (4) were isolated from the soil-derived fungus Trichoderma reesei PSU-SPSF013 along with five known compounds including hydroheptelidic acid (5), 5-hydroxy-3-hydroxymethyl-2-methyl-7-methoxychromone (6), adenosine (7), uridine (8) and tyrosol (9). Their structures were elucidated by spectroscopic analyses. The absolute configuration of the secondary alcohol moiety in 2 was assigned using Mosher’s method whereas that of 1, 3 and 4 was established by comparison of their specific rotations with those of structurally related compounds. Compounds 3-5 displayed interesting antifungal activity against Cryptococcus neoformans ATCC90113 with the respective MIC values of 8, 8 and 4 μg/mL. Moreover, only 3 was active against C. neoformans ATCC90112 with the MIC value of 32 μg/mL. Interestingly, 4 and 5 were noncytotoxic towards noncancerous Vero cells.
Lippia species share various pharmacological activities and are used in traditional cooking and medicine worldwide. Combined chromatographic techniques such as column chromatography, high-performance liquid chromatography, and countercurrent chromatography led to the purification of two new antifungal phenylpropanoid glycosides, lippiarubelloside A (1) and lippiarubelloside B (2), by bioactivity-directed fractionation of an ethanol-soluble extract from Lippia rubella, in addition to the known active related compounds forsythoside A (3), verbascoside (4), isoverbascoside (5), and poliumoside (6). The structures of compounds 1 and 2 were determined by comparison of their NMR spectroscopic data with the prototype active compound 4. Cryptococcus neoformans, which causes opportunistic lung infections, was sensitive to compounds 1-6 in the concentration range of 15-125 μg/mL. A synergistic effect (FICindex = 0.5) between 3 and amphotericin B was demonstrated. The glycosylated flavonoids pectolinarin (7), linarin (8), and siparunoside (9) were also isolated.
Three new depsides, aspergisides A-C, and one new depsidone, aspergisidone, were isolated from the soil-derived fungus Aspergillus unguis PSU-RSPG204 together with one phthalide derivative and 11 depsidones including emeguisin A, aspersidone and folipastatin. The structures were determined by spectroscopic evidence. Aspersidone and emeguisin A showed remarkably antibacterial activity against Staphylococcus aureus and methicillin-resistance S. aureus with equal MIC values of 0.5 μg/mL. Emeguisin A, which displayed the highest activity with 87.06% inhibition of human colon carcinoma (HCT-116) cell viability, decreased numbers of live cells/numbers of dead cells in a dose-dependent manner with the IC50 values of 34.8–84.7 μM in a 3D culture model depending on durations of incubation. In addition, folipastatin dose-dependently inhibited forskolin-stimulated chloride secretion in human intestinal epithelial (T84) cells with an IC50 value of 0.5 μM. These depsidones were considered to be inactive against noncancerous Vero Cells with the IC50 values of >10 μM.