ABSTRACT
In recent decades, beta-glucans have been used as important complementary and alternative medicines for numerous immunocompromised individuals and those with advanced cancer. The most active form of beta-glucans is beta[1,3]D-glucan and its most common source is cell wall of Candida albicans. Recently it has been introduced as a nano particle design to be used as a carrier for drug delivery. The current study researches a rapid method for the extraction of beta[1,3]D-glucans. The present study was conducted at Tarbiat Modares Medical University in 2012. Candida solubilized beta-glucans were obtained by oxidation of the cell wall with sodium hypochlorite and sodium hydroxide. The particle part could be solubilized by treatment with dimethylsulfoxide [DMSO] and zymolyase digestion to extract beta[1,3]Dglucan. The soluble fractions were lyophilized. We performed the Callose test to verify the presence of beta[1,3]D-glucans. Solubilized fractions were dissolved in D2O and 1HNMR spectra were measured. The soluble beta[1,3]D-glucan fraction which was derived from 1 g of dried Candida albicans germ tube weighed 190 mg. beta[1,3]D-glucan was verified by the Callose test and [1]H-NMR test compared with Curdlan [standard]. [1]H-NMR spectra verified the existence of beta[1,3]D-glucan in the final product. In the present study, extraction of beta[1,3]D-glucan by oxidation of the cell wall using sodium hypochlorite yielded more pure beta[1,3]D-glucans in comparison with other extraction methods. Thus it might represent a rapid method of extraction.
Subject(s)
Candida albicans , Cell Wall , Oxidation-Reduction , Sodium Hypochlorite , Sodium HydroxideABSTRACT
Dermatophytosis is one of the most common pandemic fungal infections that is a major health problem in cities and villages. This study aims to evaluate PCR sensitivity and accuracy in the detection of nail dermatophytosis compared to conventional direct and culture detection methods, and performs an assessment of Trichophyton rubrum in patients suspected of having nail dermatophytosis. This experiment was a descriptive-experimental study carried out on 71 nail samples obtained from patients with suspected nail dermatophytosis. All clinical samples of nails or chips were divided into three sections and each section underwent direct examination, culture and molecular tests. In the molecular test, we used fungal rRNA universal primers [ITS1 and ITS4] and Trichophyton rubrum-specific primers. In this study, for the first time in Iran and based on a modified protocol, DNA was directly extracted from tissues of infected nails in less than five hours. Additionally a comparison of the results obtained from routine laboratory methods such as direct examination and culture with PCR verified the high sensitivity and accuracy of PCR compared to the other studied methods. PCR, as a rapid, accurate method, can be a good replacement for conventional culture and direct examination
ABSTRACT
Candida albicans is one of the most important opportunistic pathogens that suppress immunologic mechanisms of the host. It is speculated that structural and secretory proteins of C. albicans have immunomodulatory effects in cancer. To evaluate the effects of C. albicans structural and secreted proteins on intratumoral CD4/CD8 ratio as well as the survival rate in BALB/c tumor model. Structural and secretory proteins from C. albicans were isolated and examined for their effects on tumor growth and survival of adenocarcinoma bearing mice. The results indicated that in mice treated with C. albicans structural protein, the survival rate significantly decreased compared with the control groups. Also, mice treated with secretory proteins showed a decrease in survival rate but it was not statistically significant [p>0.05]. Investigating the frequency of tumor infiltrated CD4+ and CD8+ T lymphocytes indicated that the percentages of tumor infiltrated CD4+ T lymphocytes in response to structural and secreted proteins were higher compared to the control groups. Our study suggests that C. albicans structural and secreted proteins modulate intratumor T lymphocyte infiltration
ABSTRACT
The secreted aspartic proteinases [Sap2] of Candida albicans has prominent role on Candida adherence, invasion, and pathogenicity. The aim of this study was cloning, expression and characterizing of Sap2 enzyme. Also in this study for the first time, the expression system P. pasturis was used for expressing the recombinant protein. C. albicans Sap2 gene was amplified by PCR with sticky ends, EcoR1 and SacII, and it was subcloned into the T/A vector. The sequencing of this gene was done with universal primers and then the Sap2 gene was cloned into pGAPZ?A expression vector. The construct was transformed into P. pasturis yeast; the Sap2 gene integration into the yeast genome was accomplished by the homologous recombination. The expressed protein was confirmed by western blotting using monoclonal antibody against Sap2 protein. Finally, the recombinant protein was purified by Ni-NTA chromatography column, and the activity of the enzyme was confirmed. In this study, we successfully amplified C. albicans Sap2 gene and subsequently integrated into the yeast pichia pasturis genome by homologous recombination. Moreover, we were able to identify a yeast clone secreting the recombinant protein. The optimum over expression of sap2 protein was obtained after 96 h, at 30 C. Expression of Sap2 gene in P. pasturis, in comparison to bacterial expression system, leads to a high-level expression, and also need for post translation modifications, that might be required for the activity of enzyme, is obviated in the yeast system. Based on our results, the purified acid aspartyl proteinase purified from P. pasturis was capable of degrading BSA as a substrate in-vitro. The recombinant Sap2 protein had maximum activity in an acidic pH
ABSTRACT
Fungal nail infection [onychomycosis] is a common disease in all communities consisting about 50 percent of nail disorders. Yeasts are one of the important causative agents of onychomycosis. Identification of the yeast species is important in the epidemiological and therapeutical point of views. The aim of the present study is the precise species identification of the pathogenic yeast isolated from fungal nail infections, using the DNA-based methods. The isolates were preliminary studied according to study of morphological characteristics. For species identification, the genomic DNA of each sample was extracted by boiling method and the ITS region of ribosomal DNA was amplified by polymerase chain reaction [PCR]. The amplified DNA was digested by the restriction enzyme MspI and each isolate was identified according to the electrophoretic patterns. A new enzymatic profile was used for final differentiation of Candida albicans and C. dubliniensis. A few of yeast isolates were identified by using ITS-sequensin. C. albicans with the prevalence of 45.6% was the most common isolate, followed by C. parapsilosis with 22.5% and C. tropicalis with 21.8%. The less common species were C. glabrata, C. krusei, C. kefyr, C. lusitaniae, C. guilliermondii and C. pulcherrima that consisted 2.72%, 2%, 1.36%, 0.68% and 0.68% of the isolates, respectively. No C. dubliniensis was found among C. albicans isolates. Two isolates [1.36%] were identified as Trichosporon spp. The most common group of the patients was in the age range of 40-70 years old and the majority [83.2%] of the patients were women with finger nail infections. Although C. albicans is still the most prevalent isolates of nail candidiasis, the increasing number of non-albicans species is notable. The study showed that for identification of some rare species, the routine phenotypical approaches are not efficient and application of the ITS-PCR-RFLP can improve the level of differentiation up to 98%. The remaining isolates can be identified by more expensive methods such as sequencing
ABSTRACT
In this study the susceptibility of Candida albicans to inhibitory effect of polyphenols under varying time [24 and 48 hours] conditions were evaluated. Green tea leaf polyphenols were extracted and analyzed by chromatography.Among polyphenols, Catechin showed stronger antifungal activity against C. albicans PTCC-5027. Catechin1 s M1C90 [The concentration of Catechin causing 90% growth inhibition of tested strain of C. albicans] and MFC [The minimum antifungal susceptibility of Catechin] were determined by Macro dilution test and calculation after 24 and 48 hours.; The antifungal activity of Catechin was time dependent. Catechin's MIC for 0.5 xl0[3], Ixl0[3] and 2x10[3] cells/ml was 12.5, 25 and 100 mg/ml after 24h respectively. The results after 48h for 0.5xl0[3], Ixl0[3] and 2x10[3] cells/ml were 6.25, 12.5 and 25 mg/ml respectively. Fluconazol was tested on C. albicans PTCC-5027 and the results indicated that this strain of Candida is fluconazol resistant. Data shown are from three separate experiments and were analysed statistically. C. albicans PTCC-5027 is fluconazol resistant, however green tea leaf polyphenols especially Catechin could inhibit the growth of this yeast at MIC and MFC concentrations. On the basis of the obtained results the green tea leaf contains effective antifungal components. Since the common and generic antifungal drugs possess some side effects and also there is an increasing drug resistance, it is hoped that consumption of herbal drugs may help to cure fungal diseases and to avoid the side effects of antimycotics as a good replacement
ABSTRACT
Ajowan is an annual herbaceous essential oil of Carum copticom. The main components of the oil are Tymol, beta-pinene, gamma- terpinene and Sabinene. The fruit oil of Carum copticum has been reported to have several therapeutic effects including anti fungal, anti bacterial and anti viral, Candida albicans is an opportunistic fungus and transforms into pathogenic form in favorable conditions, causing fungal diseases. In this study essential oil and alcoholic extract of Carum copticum were gained and Microdilution Broth method were used for detection of minimum inhibition concentration [MIC] and minimum fungicide concentration [MFC] of 11 clinical isolates of Candida albicans and Standard strain [PTCC50-27]. Results show that MIC for essential oil is 0.43 micro g/ml, 0.87 micro g/ml and for alcoholic extract is 3.51 micro g/ml, 7.03 micro g/ml, 1/75 micro g/ml. Thus, it seems that Carum copticum could inhibit Candida albicans growth by a similar mechanism which occurs by Fluconazole [FLZ]. In general, the results obtained in this study indicate that Carum copticum has potential values for growth inhibition of Candida albicans in vitro. In recent years, systemic fungal infections due to Candida species have been received major consideration about inducing mortality in nosocomial patients because of increasing in immunocompromised disorders such as AIDS and hematological disorders as well as long term use of broad spectrum antibiotics and corticosteroids. The present study was done with the aim of evaluating antifungal effects of essential oil and alcoholic extract from Carum copticum against Fluconazole [FLZ] susceptible and Fluconazole resistance Candida albicans strains isolated from different types of Candidiasis. Standard drug susceptibility tests with broth dilution technique were used to measure the in vitro antifungal activity of essential oil and alcoholic extract from Carum copticum. According to our results, it seems that Carum copticum could inhibit Candida albicans growth by a similar mechanism which occurs by FLZ and could be used as a potential antifungal agent especially with FLZ