Patrícia de Souza Bonfim-Mendonça

Propolis Is an Efficient Fungicide and Inhibitor of Biofilm Production by Vaginal Candida albicans

Vulvovaginal candidiasis (VVC) is one of the most common genital infections in women. The therapeutic arsenal remains restricted, and some alternatives to VVC treatment are being studied. The present study evaluated the influence of a propolis extractive solution (PES) on biofilm production by Candida albicans isolated from patients with VVC. Susceptibility testing was used to verify the minimum inhibitory concentration (MIC) of PES, with fluconazole and nystatin as controls. The biofilm formation of 29 vaginal isolates of C. albicans and a reference strain that were exposed to PES was evaluated using crystal violet staining. Colony-forming units were evaluated, proteins and carbohydrates of the matrix biofilm were quantified, and scanning electron microscopy was performed. The MIC of PES ranged from 68.35 to 546.87 μg/mL of total phenol content in gallic acid. A concentration of 546.87 μg/mL was able to cause the death of 75.8% of the isolates. PES inhibited biofilm formation by C. albicans from VVC. Besides antifungal activity, PES appears to present important antibiofilm activity on abiotic surfaces, indicating that it may have an additional beneficial effect in the treatment of VVC.

β-Glucan Induces Reactive Oxygen Species Production in Human Neutrophils to Improve the Killing of Candida albicans and Candida glabrata Isolates from Vulvovaginal Candidiasis

Vulvovaginal candidiasis (VVC) is among the most prevalent vaginal diseases. Candida albicans is still the most prevalent species associated with this pathology, however, the prevalence of other Candida species, such as C. glabrata, is increasing. The pathogenesis of these infections has been intensely studied, nevertheless, no consensus has been reached on the pathogenicity of VVC. In addition, inappropriate treatment or the presence of resistant strains can lead to RVVC (vulvovaginal candidiasis recurrent). Immunomodulation therapy studies have become increasingly promising, including with the β-glucans. Thus, in the present study, we evaluated microbicidal activity, phagocytosis, intracellular oxidant species production, oxygen consumption, myeloperoxidase (MPO) activity, and the release of tumor necrosis factor α (TNF-α), interleukin-8 (IL-8), IL-1β, and IL-1Ra in neutrophils previously treated or not with β-glucan. In all of the assays, human neutrophils were challenged with C. albicans and C. glabrata isolated from vulvovaginal candidiasis. β-glucan significantly increased oxidant species production, suggesting that β-glucan may be an efficient immunomodulator that triggers an increase in the microbicidal response of neutrophils for both of the species isolated from vulvovaginal candidiasis. The effects of β-glucan appeared to be mainly related to the activation of reactive oxygen species and modulation of cytokine release.

Colonization of the oral cavity by yeasts in patients with chronic renal failure undergoing hemodialysis

OBJECTIVES To determine the frequency of yeast in the oral cavity of patients with chronic renal failure, undergoing hemodialysis (PCRFH); identification and antifungal susceptibility profile of yeast and demographic profile of patients. METHODS We performed mouthwash in 146 PCRFH; the rinse fluid was collected and cultured, yeasts grown were identified by phenotypic and molecular methods. The antifungal susceptibility profile was determined against nystatin, amphotericin B, fluconazole, voriconazole, and caspofungin based in Clinical and Laboratory Standards Institute (document M27-A3). RESULTS Positive culture was observed in 39% of patients, of whom 53% were women; the median of dialysis time was 2.9 years. The age of the colonized patients varied between 26 and 84 years, with a median of 52.5 years. PCRFH over 45 years were significantly more colonized (P = 0.0108) as well as denture wearers (84.0%). We isolated 81 yeasts, predominantly Candida albicans (63%) followed by Candida glabrata. In general, yeasts were sensitive to the evaluated antifungal agents, but there was significant variation in the minimum inhibitory concentration, especially among non-C. albicans Candida (NCAC) compared to fluconazole, caspofungin, and amphotericin B. NCAC required significantly higher concentrations of fluconazole (P < 0.01). CONCLUSION The rate of colonization by yeasts in PCRFH was high, and there was variability in species distribution and antifungal susceptibility profile. These results are little known in this group of patients and are important for controlling the risk of developing invasive fungal infections.

MOLECULAR TYPING OF Candida albicans ISOLATES FROM HOSPITALIZED PATIENTS

SUMMARY Introduction: The majority of nosocomial fungal infections are caused by Candida spp. where C. albicans is the species most commonly identified. Molecular methods are important tools for assessing the origin of the yeasts isolated in hospitals. Methods: This is a study on the genetic profifiles of 39 nosocomial clinical isolates of C. albicans using two typing methods: random amplifified polymorphic DNA (RAPD) and microsatellite, two different primers for each technique were used. Results: RAPD provided 10 and 11 different profiles with values for SAB of 0.84 ± 0.126 and 0.88 ± 0.08 for primers M2 and P4, respectively. Microsatellite using two markers, CDC3 and HIS3, allowed the observation of six and seven different alleles, respectively, with combined discriminatory power of 0.91. Conclusions: Although genetic variability is clear, it was possible to identify high similarity, suggesting a common origin for at least a part of isolates. It is important to emphasize that common origin was proven from yeasts isolated from colonization (urine, catheter or endotracheal secretions) and blood culture from the same patient, indicating that the candidemia must have started from a site of colonization. The combination of RAPD and microsatellite provides a quick and efficient analysis for investigation of similarity among nosocomial isolates of C. albicans.

Oxidative Stress Triggered by Apigenin Induces Apoptosis in a Comprehensive Panel of Human Cervical Cancer-Derived Cell Lines

Recently, the cytotoxic effects of apigenin (4′,5,7-trihydroxyflavone), particularly its marked inhibition of cancer cell viability both in vitro and in vivo, have attracted the attention of the anticancer drug discovery field. Despite this, there are few studies of apigenin in cervical cancer, and these studies have mostly been conducted using HeLa cells. To evaluate the possibility of apigenin as a new therapeutic candidate for cervical cancer, we evaluated its cytotoxic effects in a comprehensive panel of human cervical cancer-derived cell lines including HeLa (human papillomavirus/HPV 18-positive), SiHa (HPV 16-positive), CaSki (HPV 16 and HPV 18-positive), and C33A (HPV-negative) cells in comparison to a nontumorigenic spontaneously immortalized human epithelial cell line (HaCaT). Our results demonstrated that apigenin had a selective cytotoxic effect and could induce apoptosis in all cervical cancer cell lines which were positively marked with Annexin V, but not in HaCaT (control cells). Additionally, apigenin was able to induce mitochondrial redox impairment, once it increased ROS levels and H2O2, decreased the Δψm, and increased LPO. Still, apigenin was able to inhibit migration and invasion of cancer cells. Thus, apigenin appears to be a promising new candidate as an anticancer drug for cervical cancer induced by different HPV genotypes.

Nanoparticles of waste material of propolis and gelatin as a novel system for delivery of L-ascorbic acid

BACKGROUND The waste material from the preparation of propolis extracts is a potential natural compound for application in pharmaceutical and medicine nanostructured products. Ascorbic acid is an excellent antioxidant and an important cofactor of several physiological and biochemical processes. OBJECTIVE The aim of this study was to develop and characterize nanoparticles containing L-ascorbic acid prepared with propolis byproduct. METHOD Nanoparticles physicochemical characteristics (surface morphology, particle size, zeta potential, and entrapment efficiency), antioxidant activity, in vitro release profile, and in vitro cytotoxicity were evaluated. RESULTS Nanoparticles showed to be spherical, with agglomeration, mean diameter between 110.93 and 480.59 nm, zeta potential near zero and good entrapment efficiency. Antioxidant activity of L-ascorbic acid increased when nanoencapsulated and the drug release was prolonged, controlled mainly by the phenomenon of relaxation of polymer chains and dependent of propolis residue concentration. The application of technology provided a reduction in the level of cytotoxicity of L-ascorbic acid, and the nanoparticles showed a protective effect on macrophages.

Propolis: a potential natural product to fight Candida species infections.

AIM To evaluate the effect of propolis against Candida species planktonic cells and its counterparts biofilms. MATERIALS & METHODS The MIC values, time-kill curves and filamentation form inhibition were determined in Candida planktonic cells. The effect of propolis on Candida biofilms was assessed through quantification of CFUs. RESULTS MIC values, ranging from 220 to 880 µg/ml, demonstrated higher efficiency on C. albicans and C. parapsilosis than on C. tropicalis cells. In addition, propolis was able to prevent Candida species biofilms formation and eradicate their mature biofilms, coupled with a significant reduction on C. tropicalis and C. albicans filamentation. CONCLUSION Propolis is an inhibitor of Candida virulence factors and represents an innovative alternative to fight candidiasis.

In vitro effect of Paullinia cupana (guaraná) on hydrophobicity, biofilm formation, and adhesion of Candida albicans’ to polystyrene, composites, and buccal epithelial cells

OBJECTIVE In vitro evaluation of the effect of guaraná (GUAR) on cell surface hydrophobicity (CSH), on biofilm formation, and on adhesion of C. albicans to polystyrene, to composite resins, and to buccal epithelial cells (BEC). MATERIALS AND METHODS Lyophilised aqueous extract of GUAR was tested on C. albicans ATCC (90028). The effect of GUAR was evaluated by examining the CSH of C. albicans, as determined by microbial adhesion to hydrocarbons test, by assessing biofilm production and through adhesion assays (microplates of polystyrene, BEC and composites). One nanoparticle (Z350(®)) and two microhybrid (LLis(®), Opallis(®)) composites were tested. Scanning electron microscopy (SEM) was used to analyse adhesion of C. albicans composites. Assays were performed in triplicate and the results analysed by Chi-square test, Kruskal-Wallis test and Dunns Multiple Comparison post hoc test at 5% significance level. RESULTS GUAR did not inhibit growth of C. albicans at any concentration, but it reduced adhesion to polystyrene surface (p < 0.001). Exposure to GUAR did not change CSH and biofilm formation, but it increased adhesion of C. albicans to the nanoparticle composite (p = 0.042) and reduced its adhesion to BEC (p < 0.001). SEM confirmed an aggregatory pattern of adhesion of C. albicans to composites. CONCLUSION GUAR increased the adhesion of C. albicans to the surface of the nanoparticle composite. However, it reduced the adhesion of C. albicans to BEC and to polystyrene, which reveals its potential use in prevention of oral diseases.

Effect of highly active antiretroviral therapy on vaginal Candida spp. isolation in HIV-infected compared to HIV-uninfected women.

Vulvovaginal candidiasis (VVC) in HIV-infected women contributed to the impairment of their quality of life. The aim of this study was to evaluate the effect of highly active antiretroviral therapy (HAART) use on the vaginal Candida spp. isolation in HIV-infected compared to HIV-uninfected women. This cross-sectional study included 178 HIV-infected (HIV group) and 200 HIV-uninfected women (control) that were studied at the Specialized Assistance Service (SAE) for sexually transmitted diseases (STD)/AIDS of the city of Maringá, Brazil, from April 1 to October 30, 2011. The yeasts were isolated and identified by phenotypic and molecular methods. The in vitro antifungal susceptibility to fluconazole, itraconazole, nystatin and amphotericin B was tested by the reference microdilution method. Higher frequencies of total vaginal Candida spp. isolation were found in the HIV-infected group than in the control group. However, both groups showed a similar frequency of colonization and VVC. Although C. albicans was the most frequent and sensitive to azolics and polyenes in both HIV-infected and uninfected women, the emerging resistance of C. glabrata to amphotericin B in the HIV-infected women was observed. Although higher frequency of vaginal Candida spp. isolation had been observed in the HIV-infected than in HIV-uninfected women, colonization and VVC showed similar frequency in both groups, indicating that HAART appears to protect against vaginal colonization and VVC.

Highlights Regarding Host Predisposing Factors to Recurrent Vulvovaginal Candidiasis: Chronic Stress and Reduced Antioxidant Capacity.

We studied host factors that could predispose women to develop recurrent vulvovaginal candidiasis (RVVC), including glycemia, insulin resistance, chronic stress, antioxidant capacity, overall immune status, local inflammation and vaginal microbiota. The presence of yeasts in vaginal culture was screened in 277 women, with or without signs and symptoms of VVC and RVVC. The presence of an inflammatory process and microbiota were analyzed through vaginal bacterioscopy and cervical-vaginal cytology, respectively. Fasting-blood samples were collected by standard venipuncture for biochemical analyses. Flow cytometry was employed to obtain the T helper/T cytotoxic lymphocyte ratio, and insulin resistance was assessed by the HOMA index (HI). Yeasts were isolated from 71 (26%) women: 23 (32.4%) with a positive culture but without symptoms (COL), 22 (31%) in an acute episode (VVC), and 26 (36.6%) with RVVC. C. albicans was the main yeast isolated in all clinical profiles. The control group (negative culture) comprised 206 women. Diabetes mellitus and insulin resistance were more associated with the positive-culture groups (COL, VVC and RVVC) than with negative ones. The RVVC group showed lower mean levels of cortisol than the control group and lower antioxidant capacity than all other groups. The T Helper/T cytotoxic lymphocyte ratio was similar in all groups. The RVVC group showed a similar level of vaginal inflammation to the control group, and lower than in the COL and VVC groups. Only the CVV group showed a reduction in vaginal lactobacillus microbiota. Our data suggest that both chronic stress (decreased early-morning cortisol levels) and reduced antioxidant capacity can be host predisposing factors to RVVC.