Antifungal activity of C3a and C3a-derived peptides against Candida

Antimicrobial peptides are generated during activation of the complement system [Nordahl et al. Proc. Natl. Acad. Sci. U. S. A. 2004, 101:16879-16884]. Here we show that the anaphylatoxin C3a exerts antimicrobial effects against the yeast Candida. Fluorescence microscopy and electron microscopy analysis demonstrated that C3a-derived peptides bound to the cell surface of Candida, and induced membrane perturbations and release of extracellular material. Various Candida isolates were found to induce complement degradation, leading to generation of C3a. Arginine residues were found to be critical for the antifungal and membrane breaking activity of a C3a-derived antimicrobial peptide, CNY21 (C3a; Cys⁵⁷-Arg⁷⁷). A CNY21 variant with increased positive net charge displayed enhanced antifungal activity. Thus, C3a-derived peptides can be utilized as templates in the development of peptide-based antifungal therapies.     

Antifungal activity of Syzygium samarangense leaf extracts against Candida

Candida species are opportunistic human fungal pathogens that cause acute and chronic infections against which only few antifungal agents are available. Here we have elucidated the antifungal effect of Syzygium samarangense leaf extracts (SSLE). Antifungal activity of SSLE was studied against Candida albicans, C. krusei, C. parapsilosis, C. glabrata, C. auris and C. tropicalis. Following experiments were performed: minimum fungicidal concentration (MFC) determination, agar well disc diffusion assays, fungal morphology analysis using scanning electron microscope (SEM), ex vivo fungal survival assays on porcine tongue and skin and in vivo fungal survival assays using Drosophila melanogaster fly model. Results demonstrated MFC of SSLE ranges between 100 and 125 mg ml⁻¹. SEM images showed cell wall degradation of C. albicans when treated with SSLE. Around 75% decrease in C. albicans viability was observed when infected porcine tongue and skin were treated using SSLE. The C. albicans infected D. melanogaster when fed with SSLE showed significant decrease (around 80%) of fungal count than the infected control. Furthermore, agar plate disc diffusion assays demonstrated that the antifungal activity of SSLE could be due to chalcone, which is one of the active constituents in SSLE. Our study demonstrated that SSLE could be used for the topical treatment of Candida infections.     

Antifungal activity of amniotic fluid against different Candida species

BackgroundAlthough Candida is a commensal of the urogenital tract, intrauterine fungal infections are extremely uncommon in clinical practice.AimsIn the present work we evaluated whether amniotic fluid (AF) possesses direct antifungal activity against clinical isolates of Candida albicans and other Candida species.MethodsA total of 23 AF samples from pregnant women with gestational age of 38–41 weeks were obtained under aseptic conditions by the aspiration of the amniotic sac during cesarean section. Different Candida species were inoculated in amniotic fluid and Sabouraud broth, used as control, and were incubated at 37 °C for 48 h. Quantitative cultures of test samples and controls were performed at 0, 4, 8, 12, 24, and 48 h.ResultsAF collected from 23 pregnant women had consistent and significant inhibitory activity against all Candida isolates tested. Nonetheless, a complete inhibition of growth by all 23 AF samples tested was observed only against Candida glabrata.ConclusionsIt is likely that the antifungal activity of the AF against C. albicans, C. glabrata and Candida parapsilosis observed in vitro also exists in vivo, contributing to protect against intrauterine fungal infections.     

Antifungal activity of a prenylated flavonoid from Dalea elegans against Candida albicans biofilms

BackgroundThe continuing emergence of infections with antifungal resistant Candida strains requires a constant search for new antifungal drugs, with the plant kingdom being an important source of chemical structures.PurposeThe present study investigated the antifungal effect of 2′,4′-dihydroxy-5′-(1′′′,1′′′-dimethylallyl)-8-prenylpinocembrin (8PP, formerly 6PP), a natural prenylflavonoid, on Candida albicans biofilms, and compared this with an azole antifungal (fluconazole) by studying the cellular stress and antioxidant response.Study design/methodsThe fluconazole sensitive (SCa) and azole-resistant (RCa) C. albicans strains were used, with biofilm formation being studied using crystal violet (CV) and confocal scanning laser microscopy (CSLM). The minimal inhibitory concentration for sessile cells (SMIC) was defined as the concentration of antifungal that caused a 50% (SMIC 50) and 80% (SMIC 80) reduction of treated biofilms. The reactive oxygen species (ROS) were detected by the reduction of nitro blue tetrazolium (NBT), and reactive nitrogen intermediates (RNI) were determined by the Griess assay. The activities of the superoxide dismutase (SOD) and catalase (CAT) antioxidant enzymes and the total antioxidant capacity of the biofilms were measured by spectrophotometric methods. ROS accumulation was also detected inside biofilms by using the fluorogenic dye 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA), which was visualized by CSLM.ResultsThe SCa and RCa biofilms were strongly inhibited by 8PP at 100 µM (SMIC 80). We observed that cellular stress affected biofilms growth, resulting in an increase of ROS and also of reactive nitrogen intermediates (RNI), with SOD and CAT being increased significantly in the presence of 8PP. The basal level of the biofilm total antioxidant capacity was higher in RCa than SCa. Moreover, in SCa, the total antioxidant capacity rose considerably in the presence of both 8PP and fluconazole.ConclusionOur data suggest that 8PP may be useful for the treatment of biofilm-related Candida infections, through an accumulation of endogenous ROS and RNI that can induce an adaptive response based on a coordinated increase in antioxidant defenses. 8PP may also have a therapeutic potential in C. albicans infections.     

Antifungal activity of Latarcin 1 derived cell-penetrating peptides against Fusarium solani

Cell-penetrating peptides and antimicrobial peptides share physicochemical characteristics and mechanisms of interaction with biological membranes, hence, termed as membrane active peptides. The present study aims at evaluating AMP activity of CPPs. LDP-NLS and LDP are Latarcin 1 derived cell-penetrating peptides and in the current study we have evaluated antifungal and cell-penetrating properties of these CPPs in Fusarium solani. We observed that LDP-NLS and LDP exhibited excellent antifungal activity against the fungus. Cellular uptake experiments with LDP-NLS and LDP showed that LDP-NLS acted as a CPP but LDP uptake into fungal spores and hyphae was negligible. CPP and AMP activity of mutated version of LDP-NLS was also evaluated and it was observed that both the activities of the peptide were compromised, signifying the importance of arginines and lysines present in LDP-NLS for initial interaction of membrane active peptides with biological membranes. Dextrans and Propidium Iodide uptake studies revealed that the mode of entry of LDP-NLS into fungal hyphae is through pore formation. Also, both LDP-NLS and LDP showed no cytotoxicity when infiltered into leaf tissues. Overall, our results suggest that LDP-NLS and LDP are selectively cytotoxic to F. solani and can be a potent peptide based antifungal agents.     

白念珠菌SSK1突变株对氟康唑敏感性的初步研究

目的探讨氟康唑作用于白念珠菌双组分信号传导途径SSK1突变株SSK21后药物敏感性的变化。方法采用微量液体稀释法和固醇测定法测定野生株(CAF2-1)和突变株(SSK21)的最小抑菌浓度(MIC);并应用RT-PCR观察氟康唑作用前后,SSK1的表达变化。结果氟康唑对CAF2-1的MIC为16μg/mL,对SSK21为0.032μg/mL。加入氟康唑后,CAF2-1的SSK1表达明显增加,60min时达到最多。结论 SSK21对氟康唑高度敏感,SSK1基因及其相关的重要基因与药物敏感性的关系值得进一步研究,从而为新的抗真菌药物和治疗途径的研发提供参考。     

Antifungal activity of Juniperus essential oils against dermatophyte, Aspergillus and Candida strains

AIMS: The increasing resistance to antifungal compounds and the reduced number of available drugs led us to search therapeutic alternatives among aromatic plants and their essential oils, empirically used by antifungal proprieties. In this work the authors report on the antifungal activity of Juniperus essential oils (Juniperus communis ssp. alpina, J. oxycedrus ssp. oxycedrus and J. turbinata). METHODS AND RESULTS: Antifungal activity was evaluated by determination of MIC and MLC values, using a macrodilution method (NCCLS protocols), on clinical and type strains of Candida, Aspergillus and dermatophytes. The composition of the oils was ascertained by GC and GC/MS analysis. All essential oils inhibited test dermatophyte strains. The oil from leaves of J. oxycedrus ssp. oxycedrus is the most active, with MIC and MLC values ranging from 0.08-0.16 microl ml(-1) to 0.08-0.32 microl ml(-1), respectively. This oil is mainly composed of alpha-pinene (65.5%) and delta-3-carene (5.7%). CONCLUSIONS: J. oxycedrus ssp. oxycedrus leaf oil proved to be an emergent alternative as antifungal agent against dermatophyte strains. delta-3-Carene, was shown to be a fundamental compound for this activity. SIGNIFICANCE AND IMPACT OF THE STUDY: Results support that essential oils or some of their constituents may be useful in the clinical management of fungal infections, justifying future clinical trials to validate their use as therapeutic alternatives for dermatophytosis.     

Antifungal activity of novel synthetic peptides by accumulation of reactive oxygen species (ROS) and disruption of cell wall against Candida albicans

In the present work, we investigated the antifungal activity of two de novo designed, antimicrobial peptides VS2 and VS3, incorporating unnatural amino acid α,β-dehydrophenylalanine (ΔPhe). We observed that the low-hemolytic peptides could irreversibly inhibit the growth of various Candida species and multidrug resistance strains at MIC₈₀ values ranging from 15.62 μM to 250 μM. Synergy experiments showed that MIC₈₀ of the peptides was drastically reduced in combination with an antifungal drug fluconazole. The dye PI uptake assay was used to demonstrate peptide induced cell membrane permeabilization. Intracellular localization of the FITC-labeled peptides in Candida albicans was studied by confocal microscopy and FACS. Killing kinetics, PI uptake assay, and the intracellular presence of FITC-peptides suggested that growth inhibition is not solely a consequence of increased membrane permeabilization. We showed that entry of the peptide in Candida cells resulted in accumulation of reactive oxygen species (ROS) leading to cell necrosis. Morphological alteration in Candida cells caused by the peptides was visualized by electron microscopy. We propose that de novo designed VS2 and VS3 peptides have multiple detrimental effects on target fungi, which ultimately result in cell wall disruption and killing. Therefore, these peptides represent a good template for further design and development as antifungal agents.     

Antifungal activity of ovotransferrin towards genus Candida

The inhibiting activity of ovotransferrin was tested towards different species belonging to genus Candida.Of one hundred strains tested, only C. krusei showed a noticeable resistance, while the other species appeared to be more sensitive than bacteria to the action of ovotransferrin. The influence of anions, such as bicarbonate and citrate, on the inhibiting activity of ovotransferrin was also investigated. Moreover it was observed that iron saturated ovotransferrin retained its activity, thus suggesting an interaction between the protein and Candida cells.     

Antifungal activities of origanum oil against Candida albicans

The antimicrobial properties of volatile aromatic oils from medicinal as well as other edible plants has been recognized since antiquity. Origanum oil, which is used as a food flavoring agent, possesses a broad spectrum of in vitro antimicrobial activities attributed to the high content of phenolic derivatives such as carvacrol and thymol. In the present study, antifungal properties of origanum oil were examined both in vitro and in vivo. Using Candida albicans in broth cultures and a micro dilution method, comparative efficacy of origanum oil, carvacrol, nystatin and amphotericin B were examined in vitro. Origanum oil at 0.25 mg/ml was found to completely inhibit the growth of C. albicans in culture. Growth inhibitions of 75% and >50% were observed at 0.125 mg/ml and 0.0625 mg/ml level, respectively. In addition, both the germination and the mycelial growth of C. albicans were found to be inhibited by origanum oil and carvacrol in a dose-dependent manner. Furthermore, the therapeutic efficacy of origanum oil was examined in an experimental murine systemic candidiasis model. Groups of mice (n = 6) infected with C. albicans (5 × LD₅₀) were fed varying amounts of origanum oil in a final vol. of 0.1 ml of olive oil (vehicle). The daily administration of 8.6 mg of origanum oil in 100 l of olive oil/kg body weight for 30 days resulted in 80% survivability, with no renal burden of C. albicans as opposed to the group of mice fed olive oil alone, who died within 10 days. Similar results were obtained with carvacrol. However, mice fed origanum oil exhibited cosmetically better clinical appearance compared to those cured with carvacrol. The results from our study encourage examination of the efficacy of origanum oil in other forms of systemic and superficial fungal infections and exploration of its broad spectrum effect against other pathogenic manifestations including malignancy.     

Antifungal activity of streptavidin C1 and C2 against pathogens causing Fusarium wilt

Fusarium wilt is caused by the soil-inhabiting fungus Fusarium oxysporum ff. spp. and is one of the most devastating plant diseases, resulting in losses and decreasing the quality and safety of agricultural crops. We recently reported the structures and biochemical properties of two biotin-binding proteins, streptavidin C1 and C2 (isolated from Streptomyces cinnamonensis strain KPP02129). In the present study, the potential of the biotin-binding proteins as antifungal agent for Fusarium wilt pathogens was investigated using recombinant streptavidin C1 and C2. The minimum inhibitory concentration of streptavidin C2 was found to be 16 µg ml^–1 for inhibiting the mycelial growth of F. oxysporum f.sp. cucumerinum and F. oxysporum f.sp. lycopersici, while that of streptavidin C1 was found to be 64 µg ml^–1. Compared with the nontreated control soil, the population density of F. oxysporum f.sp. lycopersici in the soil was reduced to 49·5% and 39·6% on treatment with streptavidin C1 (500 µg ml^–1) and C2 (500 µg ml^–1), respectively. A greenhouse experiment revealed that Fusarium wilt of tomato plants was completely inhibited on soil drenching using a 50-ml culture filtrate of the streptavidin-producing strain KPP02129.     

Antifungal activity of hypocrellin compounds and their synergistic effects with antimicrobial agents against Candida albicans

Candida albicans is a common human fungal pathogen. The previous study revealed that quinone compounds showed antimicrobial activity against C. albicans by inhibiting cell growth. However, it was unclear whether quinones have other antifungal effects against C. albicans in addition to fungicidal effects. In this study, we assessed the inhibitory activity of a total of 25 quinone compounds against C. albicans morphological transition, which is essential for the pathogenicity of C. albicans. Several quinones exhibited strong inhibition of mycelium formation by C. albicans SC5314. Three leading compounds, namely hypocrellins A, B and C, also exhibited marked attenuation of C. albicans SC5314 virulence in both human cell lines and mouse infection models. These three compounds significantly suppressed the proliferation of C. albicans SC5314 cells in a mouse mucosal infection model. Intriguingly, hypocrellins not only attenuated the cytotoxicity of a nystatin-resistant C. albicans strain but also showed excellent synergistic effects with antifungal agents against both wild-type C. albicans SC5314 and the drug-resistant mutant strains. In addition, hypocrellins A, B and C interfered with the biological functions and virulence of various clinical Candida species, suggesting the promising potential of these compounds for development as new therapeutic agents against infections caused by Candida pathogens.     

Antifungal activity of nanoemulsion from Cleome viscosa essential oil against food-borne pathogenic Candida albicans

Pathogenic and spoilage fungi cause enormous challenges to food related fatal infections. Plant essential oil based classical emulsions can functions as antifungal agents. To investigate the antifungal spectrum, that is the scope of the nanoemulsion composed of Cleome viscosa essential oil and Triton-x-100 fabricated by ultrasonication method. Minimum inhibitory and fungicidal concentration of essential oil nanoemulsion (EONE) was tested against food borne pathogenic C. albicans. The MIC and MFC values ranged from 16.5 to 33 µl/ml with significant reduction on biofilm of C. albicans isolates. The alteration of molecular fingerprints was confirmed by Fourier transformed infrared spectroscopy and subsequent reduction of chitin levels in cell walls was noted by spectroscopic analysis. The EONE and their bioactive compounds cause collateral damage on C. albicans cells.     

Chimerization of lactoferricin and lactoferrampin peptides strongly potentiates the killing activity against Candida albicans

Bovine lactoferrin harbors 2 antimicrobial sequences (LFcin and LFampin), situated in close proximity in the N1-domain. To mimic their semi parallel configuration we have synthesized a chimeric peptide (LFchimera) in which these sequences are linked in a head-to-head fashion to the α- and ε-amino group, respectively, of a single lysine. In line with previously described bactericidal effects, this peptide was also a stronger candidacidal agent than the antimicrobial peptides LFcin17-30 and LFampin265-284, or a combination of these 2. Conditions that strongly reduced the candidacidal activities of LFcin17-30 and LFampin265-284, such as high ionic strength and energy depletion, had little influence on the activity of LFchimera. Freeze-fracture electron microscopy showed that LFchimera severely affected the membrane morphology, resulting in disintegration of the membrane bilayer and in an efflux of small and high molecular weight molecules such as ATP and proteins. The differential effects displayed by the chimeric peptide and a mixture of its constituent peptides clearly demonstrate the synergistic effect of linking these peptides in a fashion that allows a similar spatial arrangement as in the parent protein, suggesting that in bovine lactoferrrin the corresponding fragments act in concert in its candidacidal activity.     

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.     

Antifungal activity of Rubus chingii extract combined with fluconazole against fluconazole‐resistant Candida albicans

This study aimed to investigate the antifungal activity of Rubus chingii extract in combination with fluconazole (FLC) against FLC‐resistant Candida albicans 100 in vitro. A R. chingii extract and FLC‐resistant C. albicans fungus suspension were prepared. The minimum inhibitory concentration and fractional inhibitory concentration index of R. chingii extract combined with FLC against C. albicans were determined, after which growth curves for C. albicans treated with R. chingii extract, FLC alone and a combination of these preparations were constructed. Additionally, the mechanisms of drug combination against C. albicans were explored by flow cytometry, gas chromatographic mass spectrometry and drug efflux pump function detection. R. chingii extract combined with FLC showed significant synergy. Flow cytometry suggested that C. albicans cells mainly arrest in G1 and S phases when they have been treated with the drug combination. The drug combination resulted in a marked decrease in the ergosterol content of the cell membrane. Additionally, efflux of Rhodamine 6G decreased with increasing concentrations of R. chingii extract. R. chingii extract combined with FLC has antifungal activity against FLC‐resistant C. albicans.     

Antifungal and antitumor activities of a monoclonal antibody directed against a stress mannoprotein of Candida albicans

Immunization of mice with a stress mannoprotein of >200 kDa from the cell wall of Candida albicans led to the production of monoclonal antibody (Mab) C7. The immunogen is a major target of secretory IgA and its expression is regulated by different environmental conditions including temperature, pH, glucose concentration and ammonium sulphate in the culture medium. Mab C7 reacted with a peptide epitope present in the >200 kDa antigen as well as in a number of antigens from the blastoconidium and germ tube cell wall, including enolase. In addition to its reactivity with C. albicans, Mab C7 also reacted with antigens present in C. krusei, C, tropicalis, C. glabrata, C. dubliniensis and C. lusitaniae, as well as in Cryptococcus neoformans, Scedosporium prolificans and Aspergillus fumigatus. Mab C7 exhibited four important biological activities, namely inhibition of adhesion of C. albicans to a variety of surfaces, inhibition of germination of C. albicans, direct candidacidal activity and direct tumoricidal activity. In tumor cells, Mab C7 reacted with nucleoporin Nup88, a reactivity that can be utilized for diagnostic and prognostic purposes.