Prevalence, Molecular Identification, and Genotyping of Candida Species Recovered from Oral Cavity among Patients with Diabetes Mellitus from Tehran, Iran.

Background: Oral candidiasis (OC) has been noticed as a common mucous membrane infection in immunocompromised patients such as that diabetes. This study, focused on the genotyping of Candida albicans and enzymatic activities of Candida species recovered from oral mucosa among diabetes patients and healthy individuals. Materials and Methods: Specimens were obtained from oral mucosa of One-hundred and sixty patients with type 2 diabetic and 108 healthy individuals. All isolates were definitely identified by ribosomal DNA (rDNA) gene sequencinghHydrophobicity, hemolytic activities of Candida species and genotypes of C. albicans were determined through polymerase chain reaction (CA-INT). Results: , Eighty eight (55%) samples out of 160, were positive for Candida species in diabetic patients. Moreover, 79.5% (70/88) and 20.5% (18/88) isolates belonged to the C. albicans and non-albicans Candida species respectively. Three genotypes of C. albicans have recovered in diabetic patients: genotype A (71.42%), B (21.42%), and C (7.14%). In healthy individuals, 42.6% (46/102) Candida species recovered from oral cavity, with the highest prevalence of genotype A (76.6% of C. albicans). Additionally, hydrophobicity and hemolytic activities from Candida species were significantly greater in diabetes patients than healthy nondiabetic subjects. Conclusion: Collectively, C. albicans was the most causative agent isolated from diabetes patients and non-diabetes healthy individuals. Genotype A, as the most remarkable genotype, should be mentioned in both groups. Higher potential hydrophobicity and hemolytic activities of Candida species in diabetic patients compared to healthy cases suggest these features triggering pathogenicity of OC in diabetes patients.

Biofilm formation in clinically relevant filamentous fungi: a therapeutic challenge.

Biofilms are highly-organized microbial communities attached to a biotic or an abiotic surface, surrounded by an extracellular matrix secreted by the biofilm-forming cells. The majority of fungal pathogens contribute to biofilm formation within tissues or biomedical devices, leading to serious and persistent infections. The clinical significance of biofilms relies on the increased resistance to conventional antifungal therapies and suppression of the host immune system, which leads to invasive and recurrent fungal infections. While different features of yeast biofilms are well-described in the literature, the structural and molecular basis of biofilm formation of clinically related filamentous fungi has not been fully addressed. This review aimed to address biofilm formation in clinically relevant filamentous fungi.

Differential genes expression analysis of invasive aspergillosis: a bioinformatics study based on mRNA/microRNA.

Invasive aspergillosis is a severe opportunistic infection with high mortality in immunocompromised patients. Recently, the roles of microRNAs have been taken into consideration in the immune system and inflammatory responses. Using bioinformatics approaches, we aimed to study the microRNAs related to invasive aspergillosis to understand the molecular pathways involved in the disease pathogenesis. Data were extracted from the gene expression omnibus (GEO) database. We proposed 3 differentially expressed genes; S100B, TDRD9 and TMTC1 related to pathogenesis of invasive aspergillosis. Using miRWalk 2.0 predictive tool, microRNAs that targeted the selected genes were identified. The roles of microRNAs were investigated by microRNA target prediction and molecular pathways analysis. The significance of combined expression changes in selected genes was analyzed by ROC curves study. Thirty-three microRNAs were identified as the common regulator of S100B, TDRD9 and TMTC1 genes. Several of them were previously reported in the pathogenesis of fungal infections including miR-132. Predicted microRNAs were involved in innate immune response as well as toll-like receptor signaling. Most of the microRNAs were also linked to platelet activation. The ROC chart in the combination mode of S100B/TMTC1, showed the sensitivity of 95.65 percent and the specificity of 69.23 percent. New approaches are needed for rapid and accurate detection of invasive aspergillosis. Given the pivotal signaling pathways involved, predicted microRNAs can be considered as the potential candidates of the disease diagnosis. Further investigation of the microRNAs expression changes and related pathways would lead to identifying the effective biomarkers for IA detection.

The effect of Candida cell wall beta-glucan on treatment-resistant LL/2 cancer cell line: in vitro evaluation.

Candida albicans (C. albicans) cell wall beta-glucan has been considered as a potential agent in the treatment of cancers due to its anti-tumor properties. Therefore, in the present study, we investigated the anti-cancer effects of Candida cell wall beta-glucan on Lewis lung carcinoma cell line (LL/2) cells. Beta-glucan of C. albicans cell wall was extracted. LL/2 cell line was cultured, then sphere cells and parental cells were exposed to the different concentrations of beta-glucan extracted from C. albicans (10-6000 µg/ml), for 24, 48 and 72 h. Cytotoxicity of beta-glucan was assayed by MTT test, then RNA extracted from cells population (treated and untreated cells), cDNA synthetized and expression level of Sox2, Oct4, C-myc, Nanog genes were also investigated using Real-time methods. At optimal concentrations of 800 and 1000 µg/ml, the extracted beta-glucan showed a significant cytotoxic effect on both parental and sphere cell populations (p < 0.05). Real-time PCR analysis revealed a decreased expression of Oct4 and Sox2 genes in treatment of cells with beta-glucan compared with control group. Since the extracted beta-glucan showed an inhibitory effect on the expression of Oct4 and Sox2 genes involved in LL/2 metastasis, therefore, beta-glucan can be considered as an anti-tumor agent because of its anti-metastatic properties, however, more in vitro and in vivo studies are needed to provide further evidence on this topic in the future.

Candida albicans Beta-Glucan Induce Anti- Cancer Activity of Mesenchymal Stem Cells against Lung Cancer Cell Line: An In-Vitro Experimental Study.

OBJECTIVE: ß-glucan, glucopyranosyl polymers of fungi cell wall, represent an immune stimulating effects with potential anti-cancer activity. Mesenchymal stem cells (MSC) have immunomodulating properties in cancer microenvironment. The aim of this study was to investigate the anti-cancer effect of Candida albicans (C. albicans) beta-glucan on MSCs supernatant for apoptosis assay of lung cancer cells in vitro. METHODS: Beta-glucan was extracted from cell wall of C.albicans. MSC isolated from adipose tissue of patients and confirmed using specific surface markers expression which examined by flow cytometry. MSCs treated with various concentrations of ß-glucans for 48 hours. Cytotoxic effect of ß-glucans was evaluated using MTT assay. MSC and lung cancer line cocultured and treated with ß-glucans and apoptosis assay was done by flow cytometry. RESULTS: Cytotoxicity findings showed a significant decrease in MSC viability during 48h, however it was dose-dependent (P<0.05). According to the obtained findings, supernatant of mesenchymal stem cells treated with ß-glucans increased cancer cells apoptosis (P<0.05). CONCLUSION: Beta glucan may highlight a potential and novel promising candidate in future strategies to cause apoptosis of cancer cells and consider as therapeutic  agent against tumor growth as well. Definitely, more in vitro and in vivo studies are required to understand its functions.

A new formulation of graphene oxide/fluconazole compound as a promising agent against Candida albicans.

Candida albicans (C. albicans) belongs to the opportunistic fungal pathogens, which cause a wide spectrum of infections in immune-compromised patients. Graphene oxide (GO), a biocompatibility agent, has been reported to exhibit effective antimicrobial activity. In the present study, a graphene oxide/fluconazole (GO/Flu) compound was synthesized and characterized using Fourier transform infrared spectroscopy (FTIR) and Raman spectroscopy. The antifungal activity of GO/Flu was examined against fluconazole-resistant C. albicans (ATCC 10231) compared to GO and Flu using the broth microdilution method, according to CLSI protocol. DNA fragmentation was assessed through the antifungal mechanism of GO/Flu. The release of Fluin PBS medium was measured. Moreover, the cytotoxicity effect of GO/Flu on SW480 cell line was evaluated. Indeed, adhesion ability of C. albicans-treated GO/Flu against SW480 cell line was assessed. The minimum inhibitory concentration (MIC) of GO, Flu, and GO/Flu was determined at 800 µg/mL, 16 µg/mL, and 400-9 µg/mL, respectively. Notably, GO/Flu exhibited an intense antifungal activity compared to GO and Flu. In addition, GO/Flu showed much less cell toxicity against SW480 cell line than GO and Flu (P < 0.05). The release determination of Flu in PBS (pH 7.4) medium was 72.42%. GO/Flu reduced the adhesion ability of C. albicans to SW480 cell line significantly. DNA fragmentation assay indicated that GO/Flu potentially degraded the DNA of C. albicans and caused a fungicidal influence. According to the findings, GO/Flu could enhance the antifungal activity against C.albicans through DNA fragmentation with low cytotoxicity effect.

Curcumin-loaded nanoliposomes linked to homing peptides for integrin targeting and neuropilin-1-mediated internalization.

CONTEXT: Curcumin, a naturally occurring polyphenol, has been extensively studied for its broad-spectrum anticancer effects. The potential benefits are, however, limited due to its poor water solubility and rapid degradation which result in low bioavailability on administration. OBJECTIVES: This study encapsulates curcumin in nanoliposomes including an integrin-homing peptide combined with a C end R neuropilin-1 targeting motif for targeted delivery and receptor-mediated internalization, respectively. MATERIALS AND METHODS: The linear GHHNGR (Glycine-Histidine-Histidine-Asparagine-Glycine-Arginine) was synthesized through F-moc chemistry on 2-chlorotrityl chloride resin and conjugated to oleic acid. The lipoyl-peptide units were then co-assembled with lecithin and 0-75 mole % Tween-80 into liposomes. Curcumin was passively entrapped using a film hydration technique and its degradation profile was examined within seven consecutive days. The cytotoxic effects of the curcumin-loaded liposomes were studied on MCF-7 and MDA-MB-468, during 24 h exposure in MTT assay. RESULTS: The maximum curcumin entrapment (15.5% W/W) and minimum degradation (< 23%) were obtained in a pH switch loading method from 5.7 to 8, in nanoliposomes (< 50 nm) containing oleyl-peptide, lecithin and Tween-80 (1:1:0.75 mole ratio). The oleyl-peptide did not prove any haemolytic activity (< 1.5%) up to 10-fold of its experimental concentration. The curcumin-loaded liposomes displayed significant reduction in the viabilities of MCF-7 (IC50 3.8 µM) and MDA-MB-468 (IC50 5.4 µM). DISCUSSION AND CONCLUSION: This study indicated potential advantages of the peptide-conjugated liposomes in drug transport to the cancer cells. This feature might be an outcome of probable interactions between the targeted nanoliposomes with the integrin and neuropilin-1 receptors.

Effect of long-term exposure to mobile phone radiation on alpha-Int1 gene sequence of Candida albicans.

Over the last decade, communication industries have witnessed a tremendous expansion, while, the biological effects of electromagnetic waves have not been fully elucidated. Current study aimed at evaluating the mutagenic effect of long-term exposure to 900-MHz radiation on alpha-Int1 gene sequences of Candida albicans. A standard 900 MHz radiation generator was used for radiation. 10 ml volumes from a stock suspension of C. albicans were transferred into 10 polystyrene tubes. Five tubes were exposed at 4 °C to a fixed magnitude of radiation with different time periods of 10, 70, 210, 350 and 490 h. The other 5 tubes were kept far enough from radiation. The samples underwent genomic DNA extraction. PCR amplification of alpha-Int1 gene sequence was done using one set of primers. PCR products were resolved using agarose gel electrophoresis and the nucleotide sequences were determined. All samples showed a clear electrophoretic band around 441 bp and further sequencing revealed the amplified DNA segments are related to alpha-Int1 gene of the yeast. No mutations in the gene were seen in radiation exposed samples. Long-term exposure of the yeast to mobile phone radiation under the above mentioned conditions had no mutagenic effect on alpha-Int1 gene sequence.

Fluconazole Resistance Candida albicans in Females With Recurrent Vaginitis and Pir1 Overexpression.

BACKGROUND: Some genes may be associated with Candida albicans resistance to azoles. Pir1 gene is described as responsible to induce resistance in C. albicans. OBJECTIVES: The current study aimed to find the relationship between fluconazole resistance and Pir1 protein (Pir1p) overexpression in the females with recurrent C. albicans vaginitis requiring longer fluconazole therapy. PATIENTS AND METHODS: A total of 52l vaginal samples were obtained from the females with C. albicans vaginitis. The azole susceptibility phenotype was determined according to the Clinical Laboratory for Standards Institute (CLSI) protocol for disk diffusion method and inhibition zone for fluconazole. Expression of pir1 gene and fluconazole -resistance were evaluated using polymerase chain reaction (PCR) in C. albicans. RESULTS: In the 52 isolates, 49 (94%) were resistant to fluconazole. Overexpression of Pir1 gene was detected in 47 (96%) fluconazole-resistant C. albicans isolates. CONCLUSIONS: The findings show fluconazole -resistance in C. albicans isolates with overexpression of Pir1p.

Can intrauterine contraceptive devices lead to VulvoVaginal Candidiasis (VVC) and Anemia in Iranian new users?

OBJECTIVES: To assess the prevalence of anemia and VulvoVaginal Candidiasis (VVC) in women before and 3 months after copper Intra Uterine Device (IUD) insertion. METHODS: Longitudinal prospective study was performed with 101 women aged 15-45 year who wanted to use the IUD at six health centers in Tehran, Iran from November 2011 to August 2012. The pattern of bleeding, Hemoglobin and Hematocirt levels, and Candida colony count/cultures in the women were assessed before and after 3 months of IUD insertion. Data analysis was performed by descriptive and analytical statistics using the SPSS software for Windows. RESULTS: At the end of 3 months, a significant increase in menstrual blood loss and a significant decrease of Hb and HCT (P=0.047 and 0.001, respectively) were reported. Moreover, no difference in the prevalence of anemia before and after IUD insertion was observed. The mean±SD Candida colony counts significantly increased (P=0.001), but positive Candida cultures were not significantly different before and 3 months after IUD insertion. Also, no clinical VVC was reported 3 months after IUD insertion. While BMI≥29 had a positive relationship with Candida colony counts, the results remained unchanged after adjusting for potential risk factors. CONCLUSIONS: Despite an increase in bleeding and Candida colony counts in copper IUD users in this study, clinical VVC or anemia cases were not increased, which indicates relative safety of this contraception method. The study findings can be helpful to healthcare professionals and midwives to counsel women who want to start using IUD and also current users who are contemplating IUD removal due to its complications.

Changes in copper and zinc serum levels in women wearing a copper TCu-380A intrauterine device.

OBJECTIVE To assess the effects of the copper intrauterine device (IUD) TCu-380A, on copper and zinc serum levels. MATERIAL AND METHODS This longitudinal study enrolled 121 women attending Health Centres in Tehran between November 2011 and August 2012. A blood sample was obtained before use and three months after insertion of a TCu-380A IUD. Serum levels of copper and zinc were measured for the 101 women who had completed three months with the device in situ. Analyses of change included paired t-tests, McNemar tests and linear regression. RESULTS Significant elevations in mean serum levels were found for both copper (170.22 µg/dl at three months vs.160.40 µg/dl at baseline, p = 0.034) and zinc (107.67 µg/dl at three months vs. 94.61 µg/dl at baseline, p < 0.001) three months after IUD insertion. CONCLUSIONS A slight, but significant increase in copper serum levels, not reaching toxic levels, was observed three months after TCu-380A IUD insertion. Zinc levels too had risen significantly, which was quite unexpected, and warrants further investigation.

The effects of Candida albicans cell wall protein fraction on dendritic cell maturation.

BACKGROUND: Candida albicans is a member of the normal human microflora. C. albicans cell wall is composed of several protein and carbohydrate components which have been shown to play a crucial role in C. albicans interaction with the host immune system. Major components of C. albican cell wall are carbohydrates such as mannans, beta glucans and chitins, and proteins that partially modulate the host immune responses. Dendritic cells (DC), as the most important antigen-presenting cells of the immune system, play a critical role in inducing immune responses against different pathogens. OBJECTIVE: We investigated the effect of the cell wall protein fraction (CPF) of C. albicans on DC maturation. METHODS: The CPF of C. albicans cells was extracted by a lysis buffer containing sodium dodecyl sulphate, 2-mercaptoethanol and phosphate-buffered saline. The extract was dialyzed and its protein pattern was evaluated by electrophoresis. Dendritic cells were purified from Balb/c mice spleens through a three-step method including mononuclear cell separation, as well as 2-h and overnight cultures. The purified CPF was added at different concentrations to DC. The purity and maturation status of DC were determined by flow cytometry using monoclonal antibodies against CD11c, MHC-II, CD40 and CD86. RESULTS: Treatment of DC with 10 microg/ml of CPF increased the expression of maturation markers including MHC-II, CD86 and CD40 on DC compared to the control group. CONCLUSION: In this study we used C. albicans CPF with the molecular weight of 40-45 kDa for pulsing and maturation of dendritic cells. Since according to our results CPF significantly increased the expression of maturation markers on DC, we suggest that CPF may act as an efficient immunomodulator, or may be used as a potential adjuvant to boost the host immune system against infections.