Silver colloidal nanoparticles: antifungal effect against adhered cells and biofilms of Candida albicans and Candida glabrata

The aim of this study was to evaluate the effect of silver nanoparticles (SN) against Candida albicans and Candida glabrata adhered cells and biofilms. SN (average diameter 5 nm) were synthesized by silver nitrate reduction with sodium citrate and stabilized with ammonia. Minimal inhibitory concentration (MIC) tests were performed for C. albicans (n = 2) and C. glabrata (n = 2) grown in suspension following the Clinical Laboratory Standards Institute microbroth dilution method. SN were applied to adhered cells (2 h) or biofilms (48 h) and after 24 h of contact their effect was assessed by enumeration of colony forming units (CFUs) and quantification of total biomass (by crystal violet staining). The MIC results showed that SN were fungicidal against all strains tested at very low concentrations (0.4–3.3 μg ml^?1). Furthermore, SN were more effective in reducing biofilm biomass when applied to adhered cells (2 h) than to pre-formed biofilms (48 h), with the exception of C. glabrata ATCC, which in both cases showed a reduction ～90%. Regarding cell viability, SN were highly effective on adhered C. glabrata and respective biofilms. On C. albicans the effect was not so evident but there was also a reduction in the number of viable biofilm cells. In summary, SN may have the potential to be an effective alternative to conventional antifungal agents for future therapies in Candida-associated denture stomatitis.     

Plasma membrane damage to Candida albicans caused by chlorine dioxide (ClO2)

Aim:  To investigate the plasma membrane damage of chlorine dioxide (ClO₂) to Candida albicans ATCC10231 at or below the minimal fungicidal concentration (MFC).
Methods and Results:  ClO₂ at MFC or below was adopted to treat the cell suspensions of C. albicans ATCC10231. Using transmission electron microscopy, no visible physiological alteration of cell shape and plasma membrane occurred. Potassium (K⁺) leakages were significant; likewise, it showed time- and dose-dependent increases. However, adenosine triphosphate (ATP) leakages were very slight. Research shows that when 99% of the cells were inactivated, the leakage was measured at 0·04% of total ATP. Compared with the mortality-specific fluorescent dye of DiBAC₄(3), majority of the inactivated cells were poorly stained by propidium iodide, another mortality-specific fluorescent dye which can be traced by flow cytometry.
Conclusion:  At or below MFC, ClO₂ damages the plasma membranes of C. albicans mainly by permeabilization, rather than by the disruption of their integrity. K⁺ leakage and the concomitant depolarization of the cell membrane are some of the critical events.
Significance and Impact of the Study:  These insights into membrane damages are helpful in understanding the action mode of ClO₂.     

Silver nanoparticles: influence of stabilizing agent and diameter on antifungal activity against Candida albicans and Candida glabrata biofilms

Aim: The purpose of this work was to evaluate the size‐dependent antifungal activity of different silver nanoparticles (SN) colloidal suspensions against Candida albicans and Candida glabrata mature biofilms. Methods and Results: The research presented herein used SN of three different average sizes (5, 10 and 60 nm), which were synthesized by the reduction of silver nitrate through sodium citrate and which were stabilized with ammonia or polyvinylpyrrolidone. Minimal inhibitory concentration (MIC) assays were performed using the microdilution methodology. The antibiofilm activity of SN was determined by total biomass quantification (by crystal violet staining) and colony forming units enumeration. MIC results showed that all SN colloidal suspensions were fungicidal against the tested strains at very low concentrations (0·4–3·3 μg ml^?1). With regard to biomass quantification, SN colloidal suspensions were very effective only against C. glabrata biofilms, achieving biomass reductions around 90% at a silver concentration of 108 μg ml^?1. In general, all SN suspensions promoted significant log₁₀ reduction of the mean number of cultivable biofilm cells after exposure to silver concentrations at or higher than 108 μg ml^?1. Moreover, the results showed that the particle size and the type of stabilizing agent used did not interfere in the antifungal activity of SN against Candida biofilms. Conclusions: This study suggests that SN have antifungal therapeutic potential, but further studies are still required namely regarding formulation and delivery means. Significance and Impact of the Study: SN may contribute to the development of new strategies for the improvement of oral health and quality of life particularly of the complete denture wearers.     

胶体纳米银对白假丝酵母菌的抑制作用

目的 探讨胶体纳米银对白假丝酵母菌的抑制作用。方法 以白假丝酵母菌为研究对象,采用倍比稀释及涂布法,检测胶体纳米银对白假丝酵母菌的抑制作用及影响因素。结果 化学纳米银及物理纳米银对白假丝酵母菌的最小抑菌浓度及最小杀菌浓度相同,分别为3.13 μg/mL和6.25 μg/mL。12.50 μg/mL的化学纳米银及物理纳米银可在5 min内完全杀死白假丝酵母菌。胶体纳米银的抑菌作用受到杀菌环境的影响,特别氯离子对胶体纳米银的杀菌作用有明显的影响。结论 胶体纳米银对白假丝酵母菌具有明显的抑菌作用,其抑制作用易受到氯离子及蛋白质的的影响。     

Efficacy of surface-generated nitric oxide against Candida albicans adhesion and biofilm formation

This report details the efficacy of nitric oxide (NO)-releasing xerogel surfaces composed of N-(6-aminohexyl)aminopropyl trimethoxysilane (AHAP3) and isobutyltrimethoxysilane (BTMOS) against Candida albicans adhesion, viability, and biofilm formation. A parallel plate flow cell assay was used to examine the effect of NO on planktonic fungal cells. Nitric oxide fluxes as low as 14 pmol cm^?2 s^?1 were sufficient to reduce fungal adhesion by ～49% over the controls after 90 min. By utilizing a fluorescence live/dead assay and replicate plating, NO flux was determined to reduce fungal viability in a dose-dependent manner. The formation of C. albicans biofilms on NO-releasing xerogel-coated silicon rubber (SiR) coupons was impeded when compared to control (non-NO-releasing) and bare SiR surfaces. The synergistic efficacy of NO and silver sulfadiazine against adhered fungal cells and biofilms is reported with increased killing and biofilm inhibition over NO alone.     

Evaluation of Candida albicans adhesion and biofilm formation on a denture base acrylic resin containing silver nanoparticles

Aim: This study firstly evaluated the activity of a silver nanoparticle (AgNPs) solution against Candida albicans and then the effect of incorporation of AgNPs into a denture base acrylic resin on the material’s hydrophobicity, C. albicans adhesion and biofilm formation. Methods and Results: The AgNPs solution was synthesized by chemical reduction and characterized. Minimum inhibitory (MIC) and minimum fungicidal (MFC) concentrations for planktonic cells and sessile cells (MFCs) of the AgNPs solution against C. albicans were determined. Specimens (n = 360) of silver‐incorporated acrylic resin at concentrations of 1000, 750, 500, 250 and 30 ppm were also prepared and stored in PBS for 0, 7, 90 and 180 days. Control was acrylic resin without AgNPs (0 ppm). After the storage periods, contact angles were measured and the specimens were used for C. albicans adherence (37°C; 90 min; n = 9) and biofilm formation (37°C; 48 h; n = 9) by XTT reduction assay. MIC, MFC and MFCs values were 3·98, 15·63 and 1000 ppm, respectively. Incorporation of AgNPs reduced the hydrophobicity of the resin. No effect on adherence and biofilm formation was observed. At 90 and 180 days of storage, there was significant increase in adherence and biofilm formation. Conclusions: Although the AgNPs solution had antifungal activity, no effect on C. albicans adherence and biofilm formation was observed after its incorporation into a denture base resin. Significance and Impact of the Study: The synthesized AgNPs solution is a promising antifungal agent, warranting investigations of more efficient methods of incorporation into denture base resins.     

Lithium-mediated suppression of morphogenesis and growth in Candida albicans

Hyphal development in Candida albicans contributes to virulence, and inhibition of filamentation is a target for the development of antifungal agents. Lithium is known to impair Saccharomyces cerevisiae growth in galactose-containing media by inhibition of phosphoglucomutase, which is essential for galactose metabolism. Lithium-mediated phosphoglucomutase inhibition is reverted by Mg(2+). In this study we have assessed the effect of lithium upon C. albicans and found that growth is inhibited preferentially in galactose-containing media. No accumulation of glucose-1-phosphate or galactose-1-phosphate was detected when yeasts were grown in the presence of galactose and 15 mM LiCl, though we observed that in vitro lithium-mediated phosphoglucomutase inhibition takes place with an IC(50) of 2 mM. Furthermore, growth inhibition by lithium was not reverted by Mg(2+). These results show that lithium-mediated inhibition of growth in a galactose-containing medium is not due to inhibition of galactose conversion to glucose-6-phosphate but is probably due to inhibition of a signaling pathway. Deletion of the Ser-Thr protein phosphatase SIT4 and treatment with rapamycin have been shown to inhibit filamentous differentiation. We observed that C. albicans filamentation was inhibited by lithium in solid medium containing either galactose as the sole carbon source or 10% fetal bovine serum. These results suggest that suppression of hyphal outgrowth by lithium could be related to inhibition of the target of rapamycin (TOR) pathway.     

Effect of sub-inhibitory concentration of chlorhexidine on lipid and sterol composition of shape Candida albicans

The effect of a sub-inhibitory concentration of chlorhexidine on lipid and sterol composition of Candida albicans was investigated. The total lipid content of this yeast grown in the presence of chlorhexidine was reduced whilst the total sterol content was increased compared with control-grown cells. Lipids and sterol analyses of this yeast grown in the presence and absence of chlorhexidine are presented. Chlorhexidine-grown yeast had a higher level of phosphatidylethanolamine, phosphatidylcholine and monogalactosyldiacylglycerol. Lower proportions of phosphatidylinositol plus phosphatidylserine, phosphatidic acid and cardiolipin were found in C. albicans grown in the presence of the drug when compared with control-grown yeast. The major fatty acids in control-grown cells were C16 and C18. Drug grown-cells had higher proportions of palmitic acid (16 : 0) and stearic acid (18 : 0), but lower proportions of palmitoleic acid (16 : 1) and oleic acid (18 : 1). Chlorhexidine also decreased the unsaturated-to-saturated fatty acid ratio, while the C16/C18 ratios increased compared to control-grown cells. Differences in the fatty acid composition of major phospholipids and neutral lipids between drug and control-grown yeast were also detected. Sterol analysis of control-grown cells showed that the major sterol present was ergosterol (55.4% wt). A significant increase in ergosterol and obtusifoliol was observed in chlorhexidine-treated cells and a significant decrease in squalene and lanosterol. Our results suggested that chlorhexidine affected the lipid and sterol composition of C. albicans. This revised version was published online in August 2006 with corrections to the Cover Date.     

间接免疫荧光法快速鉴别白念珠菌和都柏林念珠菌的初步研究

目的 用一种新制备的单克隆抗体MAb03．2Cl-C2鉴别生物学形态相近的白念珠菌和都柏林念珠菌。方法 用小鼠体内诱导法制备抗白念珠菌芽管胞壁外膜单克隆抗体MAb03．2Cl-C2。用不完全RPMI1640培养液、L—DMEM、H—DMEM、完全1640液、小牛血清诱导白念珠菌和都柏林念珠菌芽管及菌丝形成,间接免疫荧光（IIF）方法检测都柏林念珠菌芽管或菌丝表面有无可与该单抗相结合的成分。收集临床口腔念珠菌病标本涂片,直接做IIF试验。结果 用不完全RP-MI1640培养液37℃,6h可同时最高效率地诱导白念珠菌和都柏林念珠菌芽管或菌丝形成。单抗MAb03．2Cl-C2仅与白念珠菌芽管或菌丝特异性地结合,与都柏林念珠菌的孢子和菌丝不能结合。结论 单抗MAh03．2Cl-C2可用于白念珠菌和都柏林念珠菌实验室的速鉴别。     

Effects of a nanoparticulate silica substrate on cell attachment of Candida albicans

AIMS: To investigate the influence of silica nanoparticles on the attachment and growth of Candida albicans cells. METHODS AND RESULTS: Spherical silica nanoparticles with diameters of 4, 7, 14 or 21 nm were attached to tissue culture polystyrene by a polycationic binding layer using a simple deposition procedure. The modified surfaces were shown to reduce the attachment and growth of C. albicans cells by a range of different measurements including microscopy, staining cells and measuring the amount of dye taken up and total cell activity measured using a dye reduction assay. For those cells that did attach and grow, the nanoparticle-coated surface inhibited the yeast to hyphal transition that is induced in the presence of serum. The greatest effect was observed for 7 and 14 nm diameter silica particles and we propose that the mechanism for these effects are related to either the topography of the surface or the slow dissolution of the bound silica. CONCLUSIONS: The attachment and growth of C. albicans is reduced by surface modification with silica nanoparticles. SIGNIFICANCE AND IMPACT OF THE STUDY: The modification of surfaces by nanoparticulate coatings is a simple process that may have applications in reducing the prevalence of Candida sp. cells on medical devices thus, limiting the incidence of this pathogenic yeast in clinical environments.     

The effect of denture adhesives on Candida albicans growth in vitro

doi: 10.1111/j.1741‐2358.2011.00478.x
The effect of denture adhesives on Candida albicans growth in vitro Objective: Denture‐wearing favours the growth of Candida. In view of the fact that many denture wearers regularly use adhesives to enhance denture retention, stability and function, the aim of this work was to study the effect of denture adhesives on Candida albicans growth in vitro. Materials and methods: The denture adhesives tested were Corega^® cream, Kukident^® cream, Novafix^® cream, Polident^® cream, Protefix^® cream, Steradent^® cream, Aderyn^® powder, Corega^® ultra powder, Protefix^® powder and Corega^® strip. C. albicans growth curves were obtained in the presence or absence of a 1% solution of the denture adhesive diluted in Sabouraud broth. Macro‐ and microscopic morphological changes in C. albicans were analysed, as was microbial contamination of the denture adhesive. Results: Most of the denture adhesives studied induced morphological changes in C. albicans cells and colonies, but only two had any significant inhibitory effect on yeast growth. Kukident^® cream markedly inhibited C. albicans growth in a concentration‐dependent way, reducing the growth rate by 95%, whereas Corega^® cream also inhibited C. albicans growth but in a non‐concentration‐dependent way, reducing the growth rate by 37%. In addition, denture adhesives available as powders had detectable microbial contamination. Conclusion: Some commercially available denture adhesives showed microbial contamination and some had significant inhibitory effect on C. albicans growth.     

Purification and germination of Candida albicans and Candida dubliniensis chlamydospores cultured in liquid media

Candida albicans and Candida dubliniensis are the only Candida sp. that have been observed to produce chlamydospores. The function of these large, thick-walled cells is currently unknown. In this report, we describe the production and purification of chlamydospores from these species in defined liquid media. Staining with the fluorescent dye FUN-1 indicated that chlamydospores are metabolically active cells, but that metabolic activity is undetectable in chlamydospores that are >30 days old. However, 5–15-day-old chlamydospores could be induced to produce daughter chlamydospores, blastospores, pseudohyphae and true hyphae depending on the incubation conditions used. Chlamydospores that were preinduced to germinate were also observed to escape from murine macrophages following phagocytosis, suggesting that these structures may be viable in vivo . Mycelium-attached and purified chlamydospores rapidly lost their viability in water and when subjected to dry stress, suggesting that they are unlikely to act as long-term storage structures. Instead, our data suggest that chlamydospores represent an alternative specialized form of growth by C. albicans and C. dubliniensis .     

Glutathione levels during thermal induction of the yeast-to-mycelial transition in Candida albicans

Glutathione (GSH) levels were directly monitored by reverse phase HPLC during the thermal yeast-to-mycelial induction of Candida albicans. The GSH levels decreased approximately 100-fold within 120 min which corresponded to the time of maximal yeast-to-mold conversion. The yeast to mold conversion was inhibited by 1-p-chlorophenyl-4,4-dimethyl-5-diethylamino-1-penten-3-one (CDDP), a thiol-specific alkylator, which prevented the decline in GSH levels. These results are discussed with respect to the potential involvement of intracellular GSH levels in regulation of the yeast-to-mold dimorphism in Candida albicans.     

Nitric oxide-dependent killing of Candida albicans by murine peritoneal cells during an experimental infection

Abstract The phagocytic and candidacidal activities of the peritoneal cells of Candida albicans -infected mice were studied 20 days following experimental infection. Both activities were enhanced during infection. The production of nitric oxide (NO) by the peritoneal cells of infected mice was determined, and an increase in the nitrite concentration in supernatants of peritoneal cell cultures was detected. The period of NO production by the peritoneal cells coincided partially with the period of enhanced C. albicans killing. The inhibition of NO synthesis by N -monomethyl- l -arginine was concomitant with inhibition of candidacidal activity. We conclude that NO systhesis is the primary candidacidal mechanism of the murine peritoneal cells activated by C. albicans infection.     

Identification of salivary components that induce transition of hyphae to yeast in Candida albicans

Candida albicans , the major human fungal pathogen, undergoes a reversible morphological transition from single yeast cells to pseudohyphae and hyphae filaments. The hyphae form is considered the most invasive form of the fungus. The purpose of this study is to investigate the effect of saliva on hyphae growth of C. albicans. Candida albicans hyphae were inoculated in Roswell Park Memorial Institute medium with whole saliva, parotid saliva or buffer mimicking the saliva ion composition, and cultured for 18 h at 37 °C under aerobic conditions with 5% CO₂. Whole saliva and parotid saliva induced transition to yeast growth, whereas the culture with buffer remained in the hyphae form. Parotid saliva was fractionated on a reverse-phase C8 column and each fraction was tested for inducing transition to yeast growth. By immunoblotting, the salivary component in the active fraction was identified as statherin, a phosphoprotein of 43 amino acids that has been implicated in remineralization of the teeth. Synthetically made statherin induced transition of hyphae to yeast. By deletion of five amino acids at the negatively charged N-terminal site (DpSpSEE), yeast-inducing activity and binding to C. albicans were increased. In conclusion, statherin induces transition to yeast of C. albicans hyphae and may thus contribute to the oral defense against candidiasis.     

白念珠菌能量代谢与致病性

白念珠菌是人类正常菌群,也是致死性真菌感染最重要的病原体之一。目前,白念珠菌致病的决定性机制仍未明确,其对宿主的致病性主要取决于菌体的毒力因子、菌体与宿主相互作用两方面。能量代谢是白念珠菌生长繁殖的基础,也是影响其致病性的重要因素。深入研究白念珠菌能量代谢特征,探索其在致病过程中的作用,或可为发现新的药物靶点奠定基础。白念珠菌是人类正常菌群,也是致死性真菌感染最重要的病原体之一。目前,白念珠菌致病的决定性机制仍未明确,其对宿主的致病性主要取决于菌体的毒力因子、菌体与宿主相互作用两方面。能量代谢是白念珠菌生长繁殖的基础,也是影响其致病性的重要因素。深入研究白念珠菌能量代谢特征,探索其在致病过程中的作用,或可为发现新的药物靶点奠定基础。