Aspergillus fumigatus conidia stimulate lung epithelial cells (TC-1 JHU-1) to produce IL-12, IFNγ, IL-13 and IL-17 cytokines: Modulatory effect of propolis extract.

Aspergillus fumigatus conidia are the most prevalent indoors fungal allergens. The interaction between Aspergillus antigens and lung epithelial cells (LECs) result in innate immune functions. The association between Aspergillus conidia and allergic reactions, like allergic bronchopulmonary aspergillosis (ABPA) and asthma have been repeatedly reported. Since conventional therapies for allergy and asthma are limited, finding new promising treatments are inevitable. This study was designed to evaluate the effect of A. fumigatus conidia on IL-12, IFNγ, IL-13 and IL-17 release from mouse LECs and to investigate the effect of propolis on cytokines modulation. Cells were divided to two groups, one was exposed to 3×104 conidia of Aspergillus fumigatus and another group was treated by propolis (25µg/mL) as well as exposed to A. fumigatus conidia. Cytokines IL-13, IL-12, IFNγ and IL-17 were measured at times 0, 6 and 12hours after exposure using ELISA assay. The results indicated that A. fumigatus could increase the release of the cytokines with IL-13 and IL-17 being the most affected ones whilst treatment with propolis decreased the effects of A. fumigatus on IL-13 and IL-17 production. The results showed that propolis has down regulatory effects on Th2 cytokine, IL-13, and IL-17 production, whereas it caused a significant induction of IL-12, as an important Th1 cytokines by LECs. With respect to the obtained results, propolis extract might be contributed to decrease Th2 responses in allergic asthma phenomenon. However more investigations must be done in future to fully understand its efficacy.

Chronic mucocutaneous candidiasis, a case study and literature review.

Chronic mucocutaneous candidiasis (CMC) is a clinically heterogeneous disease. Some immunologic and hormonal abnormalities have been associated with CMC. The factors that predispose host to CMC infection could be autosomal or acquisitive. The disease usually occurs in childhood. Here, we reviewed the published literature on chronic mucocutaneous candidiasis and a four years old girl is presented with CMC. She had a history of recurrent thrush and otomycosis since the age of one. Candida albicans was detected in skin scraping and biopsy samples. Serum iron was low. TSH hormone level was high and T4 level was low. Giardia cysts were found in stool sample. Mucocutaneous and nail manifestations of the disease were disappeared after a period of Itraconazole therapy.

Evaluation of murine lung epithelial cells (TC-1 JHU-1) line to develop Th2-promoting cytokines IL-25/IL-33/TSLP and genes Tlr2/Tlr4 in response to Aspergillus fumigatus.

OBJECTIVE: The aims of this study were to determine the role of live and heat-killed Aspergillus fumigatus conidia in releasing interleukin (IL)-25, IL-33 and thymic stromal lymphopoietin (TSLP) and to express Toll-like receptor (Tlr)2 and Tlr4 genes. MATERIALS AND METHODS: Murine lung epithelial cells were incubated with live and heat-killed A. fumigatus conidia at 37°C for 6, 24 and 48h. After treatments, ELISA was performed to measure the concentrations of IL-25, IL-33 and TSLP in the supernatants. Quantitative real-time PCR (qPCR) was performed to assess the expression levels of Tlr2 and Tlr4 genes. RESULTS: The concentrations of IL-25 and IL-33 significantly increased after exposure to live and heat-killed conidia for various times when compared with untreated control (P<0.05). The secretion of TSLP at different concentrations of heat-killed conidia was significantly higher than both live conidia and untreated control (P<0.05). qRT-PCR results indicated a up-regulation from 1.08 to 3.60-fold for Tlr2 gene expression and 1.20 to 1.80-fold for Tlr4 gene expression exposed to heat-killed conidia. CONCLUSION: A. fumigatus has a potential ability to stimulate murine lung epithelial cells to produce IL-25/IL-33/TSLP, as well as to express Tlr2/Tlr4 genes, indicating an important role of lung epithelial cells in innate immune responses to A. fumigatus interaction.

Immunomodulatory efficacy of ethanol extract of propolis on tumor-bearing mice with disseminated candidiasis.

OBJECTIVE: This study was aimed at investigating the effect of propolis on immunosurveillance by measuring the levels of serum interleukin (IL)-4, IL-10, IL-17, tumor necrosis factor (TNF)-α and interferon (IFN)-γ in tumor-bearing mice with disseminated candidiasis. METHODS: The ethanol extract of propolis was selected for this study. Balb/C female mice were infected with Candida albicans (C. albicans) and inoculated with spontaneous mouse mammary tumor (SMMT). The serum levels of tissue inhibitor of metalloproteinase-1(TIMP-1) were assessed by enzyme- linked immunosorbent assay (ELISA). Mice were treated daily with propolis solution (100mg/kg, 0.1 mL, orally) for 3 days before IV challenge with C. albicans and SC challenge with SMMT and continued for 10 days. The rates of survival and tumor growth of understudy mice were investigated as well. The levels of TNF-α, IFN-γ, IL-4, IL-10 and IL-17 cytokines in culture supernatants were determined by ELISA. RESULTS: The mean tumor size was significantly increased in tumor-bearing mice infected with C. albicans (16.98 ± 0.49 mm(2)) as compared to other mice groups (P<0.05). The results showed a significant decline of IL-4 and IL-10 levels after propolis administration to tumor-bearing mice infected with C. albicans (53.41 pg/mL, 156.81 pg/mL and 63.45 pg/mL) (P < 0.05). The increment of TNF-α (433.85 pg/mL) and IFN-γ (120.43 pg/mL) levels were also observed. CONCLUSION: Data revealed that propolis has remarkable immunomodulatory effect, which provides a scientific validation for the popular use of this natural substance, and further investigation will help to understand propolis usefulness during immunosuppressive conditions.

Potential effects of Trachyspermum copticum essential oil and propolis alcoholic extract on Mep3 gene expression of Microsporum canis isolates.

OBJECTIVE: The purpose of this study was to evaluate the effects of Trachyspermum copticum (T. copticum) essential oil and propolis alcoholic extract on growth and transcription of Mep3 gene of Microsporum canis (M. canis) strains. METHODS: The antifungal activity was assayed by broth macrodilution method. Fungal isolates were grown in soy peptone liquid medium and treated with T. copticum oil and propolis extract. Total RNAs of M. canis were subjected to reverse transcription-polymerase chain reaction (RT-PCR). Specific primers of Actin and Mep3 genes were used. RESULTS: The results revealed that MIC values of T. copticum oil against M. canis strains were ranged from 0.2-30.5 µg/mL, with 42.3% of the strains inhibited at 0.9 µg/mL. In addition, MIC values of propolis extract against M. canis strains were ranged from 0.2-488.2 µg/mL, with 34.6% of the strains inhibited at 0.9 µg/mL. RT-PCR analysis of Mep3 and Actin expression showed DNA fragments of 661 and 690 bp amplified in all isolates before treatments with T. copticum essential oil and propolis extract. Both T. copticum and propolis completely inhibited the expression of Mep3 gene. CONCLUSION: We reported for the first time that T. copticum and propolis inhibits the expression of Mep3 gene in M. canis strains in relation to a remarkable inhibition in protease production by the fungus.

The potential effects of Zataria multiflora Boiss essential oil on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes of toxigenic Aspergillus parasiticus.

This study aims at evaluating the effects of Zataria multiflora (Z. multiflora) essential oil (EO) on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes. Total RNAs of Aspergillus parasiticus (A.parasiticus) ATCC56775 grown in yeast extract sucrose (YES) broth medium treated with Z. multiflora EO were subjected to reverse transcription- polymerase chain reaction (RT-PCR). Specific primers of nor-1, ver-1, omt-A and aflR genes were used. In parallel mycelial dry weight of samples were measured and all the media were assayed by high-pressure liquid chromatography (HPLC) for aflatoxinB1 (AFB1), aflatoxinB2 (AFB2), aflatoxinG1 (AFG1), aflatoxinG2 (AFG2) and aflatoxin total (AFTotal) production. The results showed that mycelial dry weight and aflatoxin production reduce in the presence of Z. multiflora EO (100 ppm) on day 5 of growth. It was found that the expression of nor-1, ver-1, omt-A and aflR genes was correlated with the ability of fungus to produce aflatoxins on day 5 in YES medium. RT-PCR showed that in the presence of Z.multiflora EO (100 ppm) nor-1, ver-1 and omtA genes expression was reduced. It seems that toxin production inhibitory effects of Z. multiflora EO on day 5 may be at the transcription level and this herb may cause reduction in aflatoxin biosynthesis pathway genes activity.

The potential effects of Zataria multiflora Boiss essential oil on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes of toxigenic Aspergillus parasiticus

This study aims at evaluating the effects of Zataria multiflora (Z. multiflora) essential oil (EO) on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes. Total RNAs of Aspergillus parasiticus (A.parasiticus) ATCC56775 grown in yeast extract sucrose (YES) broth medium treated with Z. multiflora EO were subjected to reverse transcription-polymerase chain reaction (RT-PCR). Specific primers of nor-1, ver-1, omt-A and aflR genes were used. In parallel mycelial dry weight of samples were measured and all the media were assayed by high-pressure liquid chromatography (HPLC) for aflatoxinB1 (AFB1), aflatoxinB2 (AFB2), aflatoxinG1 (AFG1), aflatoxinG2 (AFG2) and aflatoxin total (AFTotal) production. The results showed that mycelial dry weight and aflatoxin production reduce in the presence of Z. multiflora EO (100 ppm) on day 5 of growth. It was found that the expression of nor-1, ver-1, omt-A and aflR genes was correlated with the ability of fungus to produce aflatoxins on day 5 in YES medium. RT-PCR showed that in the presence of Z.multiflora EO (100 ppm) nor-1, ver-1 and omtA genes expression was reduced. It seems that toxin production inhibitory effects of Z. multiflora EO on day 5 may be at the transcription level and this herb may cause reduction in aflatoxin biosynthesis pathway genes activity.

Severe tinea corporis due to Trichophyton verrucosum mimicking discoid lupus erythematosus.

OBJECTIVE: This study was carried out to show possible similarities between discoid lupus erythematosus clinical signs and histopathological findings and dermatophytosis. PATIENT AND METHODS: A 43-year-old woman with diffused erythematous scaling lesions on her neck was studied. Skin biopsy was performed using H&E and PAS staining. Direct microscopic examination and mycological culture of specimens were done as well. RESULTS: Histopathological examination suggested she was suffering from discoid lupus erythematosus; however mycological findings confirmed Trichophyton verrucosum as the etiologic agent of lesions. She was successfully treated with oral terbinafine and clotrimazole cream. CONCLUSION: Regarding to the results of this study, there are some similarities between dermatophytosis and skin infections such as discoid lupus erythematosus.

Trichophyton Schoenleinii-induced widespread tinea corporis mimicking parapsoriasis.

We report a case of extensive tinea corporis in an 80-year-old woman on her forearms, thighs, legs, buttocks and trunk, mimicking parapsoriasis due to Trichophyton schoenleinii, without scalp involvement. Diagnosis of Trichophyton schoenleinii was confirmed by microscopy and mycological culture specimens.

Propolis efficacy on TNF-α, IFN-γ and IL2 cytokines production in old mice with and without systemic candidiasis.

OBJECTIVE: The purpose of this study was to investigate the efficacy of propolis on cytokine levels in old healthy mice and old mice with systemic candidiasis. METHODS: Fifty 8-month-old Balb/C mice were divided into five groups. Group 1 received orally the ethanolic extract of propolis (100mg/kg per day) for 7 days, group 2 received intravenously Candida albicans (C. albicans) (2 × 10(5) cell) and orally the ethanolic extract of propolis, and group 3 received intravenously C. albicans. After 18 days of experiments, all mice were euthanized. Then, blood samples were collected, and the spleens were excised. Splenocytes were isolated immediately and cultured in RPMI-1640 medium without stimulation and/or stimulated in the presence of Concanavalin A (Con A) for 48 h. The supernatant of splenocyte cultures and the sera of mice were tested for cytokines such as TNF-α, IFN-γ and IL2 by Enzyme-linked immunosorbent assay (Elisa). RESULTS: Orally-administered propolis treatments showed that it alone suppressed all cytokines understudy (TNF-α, IFN-γ and IL2) in the sera of mice when compared to controls, whereas the cytokine production was strongly stimulated in old mice receiving propolis altogether with C. albicans. CONCLUSION: Our results indicated that propolis was not able to stimulate cytokine production in old mice, but it may have a beneficial effect on pathogenesis of systemic candidiasis by modulating levels of cytokines such as TNF-α, IFN-γ and IL2.

Evaluation of protective efficacy of Spirulina platensis in Balb/C mice with candidiasis.

OBJECTIVE: This study was aimed at evaluating the immunostimulatory effect of Spirulina platensis in prophylaxis of Balb/C mice with systemic candidiasis. MATERIALS AND METHODS: In first experiment, 40 mice were divided into four groups, ten mice per each group, for cytokines assay. Animals received a dose of 800mg/kg of S. platensis for 4days and then were intravenously inoculated with 1×10(6) Candida albicans. Control groups received 0.2mL and 0.1mL normal saline for prophylaxis and inoculation, respectively. Five mice from each group were euthanized after 24hours and 72hours and the serum levels of IFN-γ and TNF-α were measured by Enzyme-linked immunosorbent assay (ELISA) method. In second experiment, two mice groups with systemic candidiasis, 11 mice per each group, were included to evaluate the survival rate. Animals were monitored for 30days and the kidneys, liver, lungs and spleen were analyzed for fungal invasion. RESULTS: The results indicated that the Spirulina-treated mice produced more IFN-g and TNF-α level than their control groups. This infected group showed that the mean survival time (28.86±2.7) was significantly (P<0.05) higher than control group (13.9±3.34). They also exhibited that fungal clearance in selected organs at death time represents significant differences between spleen and liver (P<0.05). CONCLUSION: Prophylaxis with S. platensis had synergistic effect through producing cytokines such as TNF-α and IFN-γ. Our results provide important information for the potential application of S. platensis in the treatment and resistance of Balb/C mice with systemic candidiasis.

The potential inhibitory effect of cuminum cyminum, ziziphora clinopodioides and nigella sativa essential oils on the growth of Aspergillus fumigatus and Aspergillus flavus

The goals of this study were to evaluate the effectiveness of Cuminum cyminum, Ziziphora clinopodioides and Nigella sativa essential oils to inhibit the growth of Aspergillus fumigatus and A. flavus and to evoke ultrastructural changes. The fungi were cultured into RPMI 1640 media in the presence of oils at concentrations of 8, 6, 5, 4, 3, 2, 1.5, 1.25, 1, 0.75 and 0.5 mg/ml in broth microdilution and 2, 1.5, 1 and 0.5 mg/ml in broth macrodilution methods with shaking for 48 h at 28ºC. Conidial and mycelial samples exposed to 0.25, 0.5, 1, 1.5 and 2 mg essential oils/ml for 5 days in 2 percent yeast extract granulated plus 15 percent Saccharose media were processed for transmission electron microscopy (TEM). Based on broth dilution methods, C. cyminum and to a lesser extent Z. clinopodioides oils exhibited the strongest activity against A. fumigatus and A. flavus with MIC90 ranging from 0.25 to 1.5 mg/ml, while the oil from N. sativa exhibited relatively moderate activity against two above fungi with MIC90 ranging from 1.5 to 2 mg/ml. The main changes observed by TEM were in the cell wall, plasma membrane and membranous organelles; in particular, in the nuclei and mitochondria. These modifications in fungal structure were associated with the interference of the essential oils with the enzymes responsible for cell wall synthesis, which disturbed normal growth. Moreover, the essential oils caused high vacuolation of the cytoplasm, detachment of fibrillar layer of cell wall, plasma membrane disruption and disorganization of the nuclear and mitochondrial structures. Aspergillus fumigatus and A. flavus growth inhibition induced by these oils were found to be well-correlated with subsequent morphological changes of the fungi exposed to different fungistatic concentrations of the oils. Our results show the anti-Aspergillus activities of C. cyminum, Z. clinopodioides and N. sativa essential oils, which strengthens the potential use of these substances as anti-mould in the future.

Socioeconomic disparities and smoking habits in metabolic syndrome: evidence from isfahan healthy heart program.

BACKGROUND: The metabolic syndrome (Mets) consists of major clustering of cardiovascular disease (CVD) risk factors. This study determines the association of socioeconomic determinants and smoking behavior in a population-based sample of Iranians with Mets. METHODS: This cross-sectional survey comprised 12600 randomly selected men and women aged ≥ 19 years living in three counties in central part of Iran. They participated in the baseline survey of a community-based program for CVD prevention entitled" Isfahan Healthy Heart Program" in 2000-2001. Subjects with Mets were selected based on NCEP- ATPIII criteria. Demographic data, medical history, lifestyle, smoking habits, physical examination, blood pressure, obesity indices and serum lipids were determined. RESULTS: The mean age of subjects with Mets was significantly higher. The mean age of smokers in both groups was higher than non-smokers but with lower WC and WHR. Marital status, age and residency were not significantly different in smokers with Mets and non-smokers with Mets. Smoking was more common in the middle educational group in the income category of Quartile 1-3. Mets was significantly related to age, sex and education. Middle-aged and elderly smokers were at approximately 4-5 times higher risk among Mets subjects. Low education decreased the risk of Mets by 0.48; similarly in non-smokers, 6-12 years of education decreased the risk of Mets by 0.72. CONCLUSION: More educated persons had a better awareness and behavior related to their health and role of smoking. In the lower social strata of the Iranian population, more efforts are needed against smoking habits.

Comparative efficacies of Zataria multiflora essential oil and itraconazole against disseminated Candida albicans infection in BALB/c mice / Eficiência comparada do óleo essencial de Zataria multiflora e itraconazol contra infecção disseminada de Candida albicans em camundongos BALB/c

Disseminated candidiasis is a serious problem in public health that results from the invasion of Candida species, in particular Candida albicans. The aim of this study was to compare the efficacies of Zataria multiflora essential oil and itraconazole in clearing C. albicans from the visceral organs of BALB/c mice suffered from disseminated candidiasis. Zataria multiflora essential oil was extracted using Clevenger-type apparatus and analyzed by gas chromatography mass spectrometry (GC-MS). For clearance experiment, mice (20-25 g, N=8 per group) received essential oil at doses of 30, 48 and 64 mg/kg and itraconazole at dose of 200 mg/kg intraperitoneally (IP) 2 days before and after intravenous inoculation of 0.5×10(6) C. albicans blastospores. The treated animals were sacrificed on day 20, and 0.1 g of the tissue homogenates was plated onto specific media. In GC-Mass, the main components of the essential oil were carvacrol (61.29 percent) and thymol (25.18 percent). The results demonstrated that IP administration of 64 mg/kg of the essential oil had the highest efficacy in reducing C. albicans and produced 39.5, 21.8, 141.5, 174 and 501-fold reductions in mean CFUs per 0.1 gram in Candida infections of the liver, spleen, lungs, brain and kidneys, respectively, compared to positive control. Itraconazole showed significantly more responsiveness than the essential oil at dose of 30 mg/kg in clearing C. albicans from the kidneys (P<0.02), brain (P<0.02) and spleen (P<0.04), and less responsiveness than that of 64 mg/kg in clearing the organism from the brain (P<0.01), lungs (P<0.0005) and kidneys (P<0.0005), whereas no significant difference was observed between this drug and Z. multiflora at dose of 48 mg/kg. These data explain the increased rate of yeast clearance and reduced dissemination to the viscera of Z. multiflora treated mice.

A candidíase disseminada é um problema sério de saúde publica decorrente da invasão por espécies de Candida, e Candida albicans em particular. O objetivo deste estudo foi comparar a eficiência do óleo essencial de Zataria multiflora e itraconazol na remoção de C. albicans das vísceras de camundongos BALB/c com candidíase disseminada. O óleo essencial de Zataria multiflora foi extraído empregando um aparelho do tipo Clevenger e analisado por cromatografia a gás e espectrometria de massa (GC-MS). Para os experimentos de remoção, camundongos (20-25g, n=8 por grupo) receberam óleo essencial nas doses de 30, 48 e 64 mg/kg e itraconazol na dose de 200 mg/kg via intraperitoneal (IP) por dois dias antes e após a inoculação intravenosa de 0,5 x 10(6) blastósporos de C. albicans. Os animais tratados foram sacrificados no vigésimo dia e 0,1g dos tecidos homogeneizados foram semeados em meios específicos. De acordo com o GC-MS, os principais componentes do óleo essencial foram carvacrol (61,29 por cento) e timol (25,28 por cento). Os resultados mostraram que a administração IP de 64 mg/kg de óleo essencial apresentou a eficiência mais alta na redução de C. albicans e resultou na redução de 39,5, 21,8, 141,5, 174 e 501 vezes na contagem média de C. albicans por 0,1g do fígado, baço, pulmões, cérebro e rins, respectivamente, quando comparado ao controle positivo. O itraconazol apresentou redução de C. albicans maior do que o óleo essencial na dose de 30mg/kg nos rins (P<0,02), cérebro (P<0,02) e baço (P<0,04) e menor no cérebro (P<0,01), pulmões (P<0,0005) e rins (P<0,0005) na dose de 64 mg/kg, enquanto não houve diferença entre esse droga e óleo essencial na dose de 48mg/kg. Estes resultados explicam a remoção aumentada de leveduras e a disseminação reduzida para as vísceras de camundongos tratados com Z. multiflora.

Comparative efficacies of Zataria multiflora essential oil and itraconazole against disseminated Candida albicans infection in BALB/c mice.

Disseminated candidiasis is a serious problem in public health that results from the invasion of Candida species, in particular Candida albicans. The aim of this study was to compare the efficacies of Zataria multiflora essential oil and itraconazole in clearing C. albicans from the visceral organs of BALB/c mice suffered from disseminated candidiasis. Zataria multiflora essential oil was extracted using Clevenger-type apparatus and analyzed by gas chromatography mass spectrometry (GC-MS). For clearance experiment, mice (20-25 g, N=8 per group) received essential oil at doses of 30, 48 and 64 mg/kg and itraconazole at dose of 200 mg/kg intraperitoneally (IP) 2 days before and after intravenous inoculation of 0.5 × 10(6) C. albicans blastospores. The treated animals were sacrificed on day 20, and 0.1 g of the tissue homogenates was plated onto specific media. In GC-Mass, the main components of the essential oil were carvacrol (61.29%) and thymol (25.18%). The results demonstrated that IP administration of 64 mg/kg of the essential oil had the highest efficacy in reducing C. albicans and produced 39.5, 21.8, 141.5, 174 and 501-fold reductions in mean CFUs per 0.1 gram in Candida infections of the liver, spleen, lungs, brain and kidneys, respectively, compared to positive control. Itraconazole showed significantly more responsiveness than the essential oil at dose of 30 mg/kg in clearing C. albicans from the kidneys (P<0.02), brain (P<0.02) and spleen (P<0.04), and less responsiveness than that of 64 mg/kg in clearing the organism from the brain (P<0.01), lungs (P<0.0005) and kidneys (P<0.0005), whereas no significant difference was observed between this drug and Z. multiflora at dose of 48 mg/kg. These data explain the increased rate of yeast clearance and reduced dissemination to the viscera of Z. multiflora treated mice.

Outbreak of severe disseminated aspergillosis in a flock of ostrich (Struthio camelus).

This study was undertaken to describe clinical, mycological and histopathological findings in black neck ostriches affected with severe aspergillosis in a flock including 80 birds, near Tehran, Iran. The signs included anorexia, depression, notable weight loss, diarrhoea, severe respiratory distress and death. Grossly, the lungs showed numerous white to yellow caseous nodules and the walls of the thoracic and abdominal air sacs were thickened with inflammatory exudates containing cellular debris, necrotic masses and green mold colonies. Multiple nodules were observed in the liver, spleen and gastrointestinal tract as well. Histopathologically, there were conidial heads and fungal hyphae in the air sacs and multifocal necrotic and granulomatous lesions with septated and dichotomously branched hyphae in various tissues, which were stained with haematoxylin and eosin and Grocott's methenamine silver nitrate. Aspergillus fumigatus was isolated in various tissues taken from affected ostriches.