Influence of menthol on biofilm formation, ergosterol content, and cell surface hydrophobicity of Candida glabrata.

Resistance to synthetic antifungals has become one of the leading public health challenges around the world. Accordingly, novel antifungal products like naturally occurring molecules can be one of the potential ways to reach efficient curative approaches to control candidiasis. This work evaluated the effect of menthol on cell surface hydrophobicity (CSH), biofilm formation, growth, and ergosterol content of Candida glabrata, a yeast with a high resistance against antifungal agents. Disc diffusion method (susceptibility to synthetic antifungals), broth micro-dilution method (Susceptibility to menthol), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide reduction assay (biofilm formation), High-performance liquid chromatography (HPLC) technique (ergosterol content), and adherence to n-hexadecane (CSH) were employed to determine the influence of menthol against C. glabrata isolates. The minimum inhibitory concentration (MIC) range of menthol versus C. glabrata was 1250-5000 µg/mL (mean ± SD: 3375 ± 1375 µg/mL). The mean rate of C. glabrata biofilm formation was decreased up to 97.67%, 81.15%, 71.21%, 63.72%, 47.53%, 26.31%, and 0.051% at 625, 1250, 2500, 5000, 10 000, 20 000, and 40 000 µg/mL concentrations, respectively. The percentages of CSH were significant in groups treated with MIC/2 (17.51 ± 5.52%) and MIC/4 (26 ± 5.87%) concentrations of menthol. Also, the percentage changes in membrane ergosterol were 15.97%, 45.34%, and 73.40% at 0.125, 0.25, and 0.5 mg/mL concentrations of menthol, respectively, in comparison with untreated control. The results showed the menthol impact versus sessile and planktonic C. glabrata cells, and the interference with ergosterol content, CSH, and biofilm formation, which made it a potent natural antifungal.

Efficacy of Syzygium aromaticum essential oil on the growth and enzymatic activity of pathogenic Candida albicans strains.

Background and Purpose: Candida albicans (C. albicans) is the most common human pathogen owing to the most virulence factors. It seems that extracellular hydrolytic enzymes play a key role in C. albicans pathogenicity. The present study aimed to assess the susceptibility and enzymatic activity of pathogenic C. albicans isolates exposed to the Syzygium aromaticum (S. aromaticum) essential oil. Materials and Methods: S. aromaticum oil was characterized using gas chromatography-mass spectrometry (GC-MS). The broth microdilution technique (CLSI, M27-A3) was used to determine the minimum inhibitory concentration (MIC) of test compounds. Furthermore, before and after treatment with S. aromaticum essential oil, the yeasts were analyzed regarding the proteinase (Prz), hemolysin (Hz), and phospholipase (Phz) production/activity. Results: ß-caryophyllene (12.76%) was found to be the major constituent in the essential oil after eugenol (84.64%). Only one isolate of C. albicans showed the antifungal resistance to fluconazole. All isolates were susceptible to S. aromaticum essential oil with MIC of 625-1250 µg/ml. S. aromaticum oil represented the best antifungal effect against C. albicans at MIC 1000 µg/ml. The mean±SD enzyme activity of C. albicans not exposed to S. aromaticum essential oil was obtained at 0.55±0.03, 0.73±0.04, and 0.61±0.05 for proteinase, hemolysin, and phospholipase, respectively. The activities of these enzymes were reduced significantly (P<0.05) to 0.33±0.06, 0.40±0.04, and 0.16±0.03 for phospholipase, proteinase, and hemolysin, respectively, after the yeasts were subjected to S. aromaticum essential oil. Conclusion: The present study aimed to determine the ability of S. aromaticum essential oil to prevent the growth of C. albicans and decrease their enzymatic activity. As a natural antifungal agent, S. aromaticum can be utilized in pharmaceutical systems.

Molecular Identification and Antifungal Susceptibility Patterns of Dermatophytes Isolated from Companion Animals with Clinical Symptoms of Dermatophytosis.

INTRODUCTION: Dermatophytosis is a common skin disease in cats and dogs caused by Microsporum and Trichophyton fungi. Species identification and knowledge of their antifungal susceptibility are therapeutically and epidemiologically important. This study assessed the prevalence of feline and canine dermatophytosis in Iran, identified the aetiological agents molecularly and tested their antifungal susceptibility. MATERIAL AND METHODS: A total of 308 companion animals (134 dogs and 174 cats) with skin lesions were examined from March 2015 to March 2018. Hair and skin samples were examined by microscopy with 20% KOH and cultured on Sabouraud dextrose agar with cycloheximide and chloramphenicol. Fungal isolates were confirmed by sequencing of the internal transcribed spacer (ITS) r-DNA region. The antifungal susceptibility of dermatophytes was tested by broth microdilution assay using standard drugs. RESULTS: Dermatophytes were found in 130 (42.2%) samples, 62 of them feline and 68 canine. Based on sequencing of all strains, M. canis (78.5%, P<0.05), M. gypseum (10.7%), and T. mentagrophytes (10.7%) were the dermatophytes isolated. The non-dermatophyte species Nannizziopsis vriesii was also isolated from two feline dermatomycosis cases. Dogs and cats younger than one year (61.5%) showed a statistically significantly higher prevalence of infection (P<0.05). Caspofungin produced the lowest geometric mean MIC at 0.0018 µg/mL, followed by ketoconazole, terbinafine, itraconazole, miconazole, griseofulvin, clotrimazole and fluconazole, in a 0.038-1.53 µg/mL range. CONCLUSION: This is the first molecular study to identify the causes of pet dermatophytosis in north-western Iran. ITS-PCR was shown to be a useful and reliable method for the identification of closely related species of dermatophytes in clinical and epidemiological settings. The lowest MIC of caspofungin indicated that this drug was the most potent in vitro.

In vitro synergy of eugenol on the antifungal effects of voriconazole against Candida tropicalis and Candida krusei strains isolated from the genital tract of mares.

BACKGROUND: Due to the limited range of antifungals available to treat genital Candida infections and the emergence of resistant isolates, attention has focused on the antifungal potency of natural compounds with promising biological properties. OBJECTIVES: To examine whether eugenol synergises the in vitro efficacy of voriconazole against Candida strains isolated from the genital tract of mares. STUDY DESIGN: In vitro experiment. METHODS: The antifungal activity of eugenol and voriconazole was evaluated using the broth microdilution assay (CLSI- M27-A3). Synergism of eugenol and voriconazole against genital Candida isolates was evaluated by the microdilution checkerboard method. RESULTS: Minimum inhibitory concentration (MIC) values for eugenol and voriconazole ranged from 400 to 800 µg/mL and 1 to 8 µg/mL, respectively, for C. tropicalis isolates, and from 200 to 400 µg/mL for eugenol and 2 to 16 µg/mL for voriconazole against C. krusei isolates. Eugenol decreased the arithmetic mean MIC for voriconazole against C. tropicalis and C. krusei isolates from 2.66 to 0.46 µg/mL and 7.77 to 0.41 µg/mL respectively. The fractional inhibitory concentration index (FICI) values for the eugenol-voriconazole combination ranged from 0.25 to 0.88 and 0.19 to 0.63 for C. tropicalis and C. krusei isolates respectively. A synergistic effect of eugenol in combination with voriconazole was observed for 83.3% of C. tropicalis and 77.7% of C. krusei isolates. Antagonistic activity was not seen in any of the isolates tested. MAIN LIMITATIONS: Since in vitro antifungal susceptibility tests are not systematic analyses, any selection bias could influence the results. In addition, in vitro susceptibility does not uniformly predict clinical success in vivo. CONCLUSIONS: Eugenol showed fungistatic and fungicidal effects against genital Candida isolates and, in combination, synergised the antifungal effects of voriconazole. The eugenol-voriconazole combination can lay the foundation for a therapeutic approach against isolates in which azole resistance has increased over time.

Effects of Bunium persicum essential oil on the reduction of spore germination, growth, and expression of FUM1 and FUM14 genes in Fusarium verticillioides isolates.

BACKGROUND AND PURPOSE: Black Cumin of Kerman (Bunium persicum) is an Iranian plant that is commonly used as an antispasmodic, carminative, and antimicrobial substance. The present study aimed to assess different components of the essence of B. persicum and its effect on antifungal activity, spore germination inhibition, and expressions of FUM1 and FUM14 genes in Fusarium verticillioides strains. MATERIALS AND METHODS: The essence was extracted by hydrodistillation and analyzed through gas chromatography-mass spectroscopy. A broth microdilution method was used for the determination of the minimum inhibitory concentration (MIC). In addition, the expression of FUM1 and FUM14 genes of toxigenic F. verticillioides was assessed by using the real-time polymerase chain reaction (RT-PCR) technique. RESULTS: Based on the findings, most of the essence consisted of γ-terpinene (15.56%), propanal, and 2-methyl-3-phenyl (14.18%). The oil showed a good antifungal activity (mean MIC value: 2556.8 µg/ml) as well as the inhibition of spore germination and mycelial growth (P<0.05). The RT-PCR demonstrated that the expression levels of FUM1 and FUM14 of B. persicum-treated F. verticillioides were 0.43 and 0.53 folds lower than the control samples, respectively. CONCLUSION: These findings revealed that the essential oil of B. persicum has different components responsible for the inhibition of mycelial growth and spore germination of F. verticillioides as well as reduction of expressions of FUM1 and FUM14 genes involving fumonisin production.

Isolation and characterization of mycoflora of chicken hatcheries in Mazandaran province, north of Iran.

Fungal infections cause significant economic losses in the poultry industry either due to their direct infectious nature or due to mycotoxins production. Hatchery contamination with fungi can threaten chicken health. In this regard, geographical and seasonal distributions of airborne fungal contamination of 25 hatcheries in Mazandaran province, northern Iran, were investigated using an open plate method. The results of this study showed that hatcheries have various fungal contaminations, among which the most common were respectively Cladosporium (31.07%), Penicillium (24.00%), Aspergillus (20.63%), sterile hyphae (14.70%) and Alternaria (6.20%) from different regions. The results revealed that the highest level of fungal isolation was in spring and autumn. This study also showed that the concentration of fungal air spora in forest and seaside locations was significantly greater than mountainous ones. In spite of the regular disinfection in commercial hatcheries, fungal contamination was found in different parts.

Synergistic anticandidal activity of menthol in combination with itraconazole and nystatin against clinical Candida glabrata and Candida krusei isolates.

BACKGROUND: Candida glabrata (C. glabrata) and C. krusei are now emerging as serious hospital acquired infections in immunocompromised patients. Menthol, a terpenic compound, has been reported to have antifungal activity. OBJECTIVES: The aim of this study was to investigate the effect of menthol in combination with itraconazole or nystatin against C. glabrata and C. krusei isolates. METHODS: The effects of menthol along with itraconazole and nystatin, were evaluated by the Clinical Laboratory Standards Institute (CLSI) M44-A and CLSI M27-A3 methods. The fractional inhibitory concentration index (FICI) was determined for menthol plus itraconazole and nystatin combinations using the checkerboard method. RESULTS: The mean of minimum inhibitory concentration (MIC) values of menthol, nystatin and itraconazole were 53.2, 2.30 and 1.50 µg/ml for C. glabrata isolates and 121, 1.08 and 0.38 µg/ml for C. krusei isolates, respectively. Menthol in combination with itraconazole or nystatin exhibited the synergistic effects against all species of Candida tested. FICI values for menthol plus itraconazole and nystatin combinations ranged from 0.250 to 0.561 and 0.139 to 0.623 for C. glabrata isolates, and 0.182 to 0.750 and 0.188 to 0.760 for C. krusei, respectively. CONCLUSIONS: These results support the potential use of menthol as an anticandidal agent, and it can be used complementarily with other conventional antifungal agents.

In vitro lymphoproliferative response and cytokine production in mice with experimental disseminated candidiasis.

OBJECTIVES: Systemic candidiasis is an infection of Candida albicans (C. albicans) causing disseminated disease and sepsis, invariably when host defenses are compromised. We investigated the histopathological changes as well as the lymphoproliferative responses and cytokine production of splenic cells after stimulation with Concanavalin A (Con A) and Pokeweed mitogen (PWM) in mice with disseminated candidiasis. MATERIALS AND METHODS: Lymphoproliferative responses were stimulated in vitro with Con A (1 µg/ml) and PWM (1 µg/ml) mitogens in Roswell Park Memorial Institute (RPMI) 1640 media, and the production of interferon (IFN)-γ and interleukin-4 (IL-4) in the supernatants was measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: The results revealed that C. albicans organisms multiplied to a greater extent in the kidneys than in the liver and spleen of infected mice. The most predominant forms of C. albicans in different parts of the kidneys were yeast mixed with hyphal forms. Infected mice had a significantly increased proliferative response when splenocytes were stimulated with PWM (2.0±0.16) and Con A (1.9±0.19) (P<0.05). PWM and Con A-stimulated production of IFN-γ significantly tended to be higher in infected mice (PWM: 68.4±14.0 pg/ml; Con A: 53.7±17.3 pg/ml) when compared to controls (P<0.05). Stimulation with PWM and Con A showed no differences in IL-4 production between infected mice and controls. CONCLUSION: These findings demonstrated a significant increase in both cell proliferation and IFN-γ secretion in supernatants of PWM and Con A- stimulated splenocyte cultures obtained from mice with disseminated candidiasis.

Down-regulatory effect of Thymus vulgaris L. on growth and Tri4 gene expression in Fusarium oxysporum strains.

The aims of this study were to evaluate the efficacy of Thymus vulgaris (T. vulgaris) essential oil on the fungal growth and Tri4 gene expression in Fusarium oxysporum (F. oxysporum) strains. The oil was obtained by water-distillation using a Clevenger-type system. The chemical composition of the essential oil was obtained by gas chromatography- mass spectroscopy (GC-MS) and by retention indices. The antifungal activity was evaluated by broth microdilution assay. A quantitative real time RT-PCR (qRT-PCR) assay was also developed specific for F. oxysporum on the basis of trichothecene biosynthetic gene, Tri4, which allowed discrimination from F. oxysporum. Results showed thymol (32.67%) and p-cymene (16.68%) as the main components of T. vulgaris. Minimum inhibitory concentration (MIC) values varied from 5 to 20 µg/ml with T. vulgaris (mean: 10.50 µg/ml), while minimum fungicidal concentration (MFC) values ranged from 8 to 30 µg/ml with mean value of 16.20 µg/ml qRT-PCR results revealed a downregulation from 4.04 to 6.27 fold of Tri4 gene expression of the fungi exposed to T. vulgaris essential oil. The results suggest that T. vulgaris oil can be considered potential alternative natural fungicide to the synthetic chemicals that are currently used to prevent and control seed-borne diseases.

Evaluation of APR1 Gene Expression in Candida albicans Strains Isolated From Patients With Multiple Sclerosis.

BACKGROUND: Intracellular aspartic proteinase A enzyme is expressed by the APR1 gene and is one of the important factors in the development of systemic candidiasis caused by Candida albicans. OBJECTIVES: The aim of this study was to evaluate the expression of the APR1 gene in C. albicans isolates obtained from patients with multiple sclerosis (MS) and from controls. PATIENTS AND METHODS: The samples were obtained from 135 MS patients with candidiasis and 100 matched controls of healthy individuals during 2010 - 2011. The clinical and control isolates of C. albicans obtained from individuals were cultured onto sabouraud dextrose agar (SDA). The evaluation of APR1 gene expression was performed using the reverse transcriptase-polymerase chain reaction (RT-PCR) method. RESULTS: There was a statistically significant difference in APR1 gene expression of C. albicans strains between MS patients (mean ± SD: 0.5208 ± 0.11518) and the control group (mean ± SD: 0.7603 ± 0.11405) (P = 0.000). Significant correlations were found between the APR1 gene expression of C. albicans strains from MS patients with regard to age and the expanded disability status scale (EDSS) (P = 0.000). The mean values of EDSS were 1.6074 ± 0.1081 after antifungal treatment and 2.2519 ± 0.1323 before antifungal treatment (P = 0.000). No significant correlation was observed between the APR1 gene expression with regard to sex and MS subtypes. CONCLUSIONS: The results suggested that APR1 gene expression in C. albicans strains isolated from MS patients may be an important factor for invasive C. albicans strains in the progression of MS disease.

Differential Gene Expression of Heat Shock Protein 90 (Hsp90) of Candida albicans obtained from Malaysian and Iranian Patients.

BACKGROUND: Candida albicans (C. albicans) has several virulence factors, in particular heat shock protein 90 (Hsp90), which is expressed by Hsp90 gene. The purposes of this study were to assess the expression of Hsp90 gene in clinical and control isolates of C. albicans obtained from different geographical regions (Malaysia and Iran), different temperatures (25°C, 37°C and 42°C) and mice with candidiasis. METHODS: C. albicans isolates were cultured onto sabouraud dextrose agar (SDA). The assessment of the expression of Hsp90 gene was performed using real time-polymerase chain reaction (RT-PCR). RESULTS: The results showed a significant increase in the expression of C. albicans Hsp90 gene under high thermal shock (42°C) when compared to other temperatures tested (P-value = 0.001). The mean differences in the expression of Hsp90 gene at 37°C were 0.20 (95% confidence interval (CI) 0.13-0.29) between Malaysian and Iranian controls (P-value = 0.040) and 0.47 (95% CI 0.27-0.60) between Malaysian and Iranian patients (P-value = 0.040). CONCLUSION: The results demonstrated that the expression of C. albicans Hsp90 gene varied between Malaysian and Iranian subjects, representing the efficacy of geographical and thermal conditions on virulence gene expression.

In vitro evaluation of antifungal susceptibility and keratinase, elastase, lipase and DNase activities of different dermatophyte species isolated from clinical specimens in Iran.

The aims of this study were to evaluate the enzymatic activity of various dermatophyte species and their antifungal susceptibility profiles. A total of 60 dermatophyte isolates, including Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis and Microsporum gypseum, were examined. Fungal isolates were analysed for the production of keratinase, lipase, elastase and deoxyribonuclease (DNase). A broth microdilution method was performed on the basis of M38-A2 Clinical and Laboratory Standards Institute (CLSI) guidelines. T. mentagrophytes, M. canis and M. gypseum isolates were capable of producing keratinase, lipase, elastase and DNase, while T. rubrum isolates were elastase negative. The highest mean diameter of the clear zone around the colonies (PZ) was associated with keratinase (PZ: 4.56 ± 1.29 mm), followed by lipase (PZ: 1.53 ± 0.90 mm), DNase (PZ: 0.65 ± 0.54 mm) and elastase (PZ: 0.22 ± 0.27 mm) (P < 0.05). The mean minimum inhibitory concentration 90 (MIC90 ) of all strains were as follows: itraconazole (MIC90 : 0.28 ± 0.31 µg ml-1 ), ketoconazole (MIC90 : 0.48 ± 0.51 µg ml-1 ), griseofulvin (MIC90 : 0.86 ± 1.00 µg ml-1 ) and fluconazole (MIC90 : 18.57 ± 20.10 µg ml-1 ). Dermatophyte isolates had higher keratinolytic activity than other enzymes. Itraconazole was the most effective antifungal drug and fluconazole had the poorest activity.

A review on the inhibitory potential of Nigella sativa against pathogenic and toxigenic fungi.

Nigella sativa (N. sativa) grows in various parts of the world, particularly in Iran. It has been traditionally used as a folk remedy to treat a number of diseases. The seeds of this plant contain moisture, proteins, carbohydrates, crude fiber, alkaloids, saponins, ash, fixed oils and essential oil. The major components of the essential oil are thymoquinone, p-cymene, trans-anethole, 2-methyl-5(1-methyl ethyl)-Bicyclo[3.1.0]hex-2-en and γ-terpinene. So far, several pharmacological effects such as anti-oxidant, anti-inflammatory, anti-cancer and anti-microbial have been reported for N. sativa or its active compounds. Thymoquinone, thymohydroquinone and thymol are the most active constituents which have different beneficial properties. The oil, extracts and some of N. sativa active components possessed moderate in vitro and in vivo inhibitory activity against pathogenic yeasts, dermatophytes, non-dermatophytic filamentous fungi and aflatoxin-producing fungi. The main morphological changes of pathogenic and toxigenic fungi treated with N. sativa oil were observed in the cell wall, plasma membrane and membranous organelles, particularly in the nuclei and mitochondria. Although this review represents first step in the search for a new anti-fungal drug, the full potential of N. sativa as a fungitoxic agent has not been exploited and necessitates further investigations.

Antifungal activity of essential oils from Iranian plants against fluconazole-resistant and fluconazole-susceptible Candida albicans.

OBJECTIVE: The purpose of this study was to assay the antifungal activity of selected essential oils obtained from plants against both fluconazole (FLU)-resistant and FLU-susceptible C. albicans strains isolated from HIV positive patients with oropharyngeal candidiasis (OPC). MATERIALS AND METHODS: The essential oils were obtained by hydrodistillation method from Myrtus communis (My. communis), Zingiber officinale roscoe (Z. officinale roscoe), Matricaria chamomilla (Ma. chamomilla), Trachyspermum ammi (T. ammi) and Origanum vulgare (O. vulgare). The susceptibility test was based on the M27-A2 methodology. The chemical compositions of the essential oils were obtained by gas chromatography- mass spectroscopy (GC-MS). RESULTS: In GC-MS analysis, thymol (63.40%), linalool (42%), α-pinene (27.87%), α-pinene (22.10%), and zingiberene (31.79%) were found to be the major components of T. ammi, O. vulgare, My. communis, Ma. chamomilla and Z. officinale roscoe, respectively. The results showed that essential oils have different levels of antifungal activity. O. vulgare and T. ammi essential oils were found to be the most efficient (P<0.05). The main finding was that the susceptibilities of FLU-resistant C. albicans to essential oils were higher than those of the FLU-susceptible yeasts. CONCLUSION: Results of this study indicated that the oils from medicinal plants could be used as potential anti FLU-resistant C. albicans agents.

The role of Candida albicans in the severity of multiple sclerosis.

The purpose of this study was to compare the specific activity of proteinase A in Candida albicans (C. albicans) between multiple sclerosis (MS) patients and controls. A total of 135 and 100 C. albicans strains were isolated from superficial surfaces of MS patients and healthy controls. Analytical models (regression and neural network) were applied to predict the severity of MS considering specific enzyme activity (SEA) and other factors which affect the expanded disability status scale (EDSS). The SEA of C. albicans in MS patients (3466.95 ± 277.25 µmol min-1 mg-1 ) was significantly more than that of healthy controls (1108.98 ± 294.51 µmol min-1 mg-1 ) that was confirmed by regression model (P < 0.001). The SEA had a positive correlation with the severity of MS (P < 0.001, r = 0.65). Analytical models showed that SEA played the most important role (among all included factors that affect on EDSS) in the severity of MS. The SEA of C. albicans in MS patients was significantly more than the healthy controls. The results suggest that the level of SEA of proteinase A and probably the capacity of C. albicans isolates to invade the host tissue is associated with the severity of MS.

Factors in Etiology and Predisposition of Adult Tinea Capitis and Review of Published Literature.

BACKGROUND: Tinea capitis is a common fungal infection in children but is less frequently encountered in adults, especially in immunocompromised individuals. OBJECTIVES: To determine the incidence of tinea capitis in adults, the predisposing factors and causative species. METHODS: A retrospective study was conducted over a period of 5 years, from 2010 to 2015, on cases of tinea capitis diagnosed in the Department of Dermatology and Mycology Research Center in Tehran, Iran. The information was collected from the patients including age, gender, location of the lesions, results of direct examination and culture, cause of immunosuppression and the prescribed treatment. RESULTS: Twenty-five (20.6 %) patients (10 men and 15 women) with a mean age of 45.28 years were affected by tinea capitis among a total number of 121 positive cases. Most of these adults (80 %) had a grade of immunodeficiency due to the underlying syndromes or diseases, and the rest were immunocompetent. Trichophyton species were isolated from 84 % of these adult patients, indicating Trichophyton violaceum (T. violaceum) as the most common fungal agent. Treatment with oral terbinafine or itraconazole was successful in all these cases. CONCLUSIONS: The results showed that most cases affecting the adult population were caused by species of the genus Trichophyton. T. violaceum was the most common dermatophyte of adult patients. Thus, it is important to consider tinea capitis as a differential diagnosis in immunocompromised adults, even though it is considered to be rare in adults.

Occurrence and distribution of Malassezia species on skin and external ear canal of horses.

The aim of this study was to investigate the prevalence of Malassezia species from the body skin and external ear canal of healthy horses. The samples were obtained by scraping the skin surface from the nose, groin and dorsum and swabbing from the external ear canal of 163 animals, and then incubated on sabouraud dextrose agar and modified Dixon agar. Malassezia species were isolated from 34.9% of horses. The percentages of Malassezia species were 64.3% for Arab, 35.7% for Persian, 35.4% for Thoroughbred and 27.1% for Turkmen breeds. The greatest abundance of Malassezia species was found in the external ear canal (47.7%, representing significant difference with other sites), followed by nose (26.3%), groin (15.8%) and dorsum (10.5%) (P < 0.05). A total of 57 strains from six Malassezia species were detected with a frequency rate as follows: M. pachydermatis (33.3%), M. globosa (26.3%), M. sympodialis (14.1%), M. restricta (10.5%), M. obtusa (8.8%) and M. furfur (7%). The most common age-group affected was 1-3 years (59.4%). This study confirmed that cutaneous Malassezia microbiota in healthy horses varies by body site and age but not by breed and gender, representing M. pachydermatis as the most prevalent species on horse skin.

Evaluation of virulence factors and antifungal susceptibility patterns of different Candida species isolated from the female camel (Camelus dromedarius) genital tract.

The purposes of this study were to investigate the enzymatic activity of different Candida species and their antifungal susceptibility patterns. The study involved a total of 83 isolates of Candida from the genital tract of the female Camelus dromedarius. After species identification, the isolates were analysed for the production/activity of phospholipase, proteinase and haemolysin. In addition, the agar disc diffusion method was performed on the basis of CLSI guidelines M44-A2 protocol for antifungal susceptibility testing. All the isolates were able to produce phospholipase, proteinase and haemolysin. A total of 35.48%, 87.09% and 64.51% of C. albicans isolates exhibited very high phospholipase, proteinase and haemolytic activities, respectively, whereas very high phospholipase, proteinase and haemolytic activities were determined in 5.76%, 23.07% and 45.16% of non-C. albicans isolates respectively. Overall, 61 (73.5%) of Candida isolates were susceptible to fluconazole, 70 (84.3%) susceptible to clotrimazole, 82 (98.8%) susceptible to voriconazole, 76 (91.6%) susceptible to itraconazole, 75 (90.4%) susceptible to ketoconazole, 83 (100%) susceptible to amphotericin B, 81 (97.6%) susceptible to nystatin and 36 (43.4%) susceptible to flucytosine. Candida isolates showed higher haemolytic activity than that of other secreted hydrolases among vaginal Candida species. In addition, amphotericin B was the most in vitro effective antifungal drug and flucytosine had the poorest activity under such conditions.

Efficacy of Cuminum cyminum essential oil on FUM1 gene expression of fumonisin-producing Fusarium verticillioides strains.

OBJECTIVES: The purpose of this study was to evaluate the effect of Cuminum cyminum (C. cyminum) essential oil on the growth and FUM1 gene expression of fumonisin-producing Fusarium verticillioides (F. verticillioides) strains isolated from maize. MATERIALS AND METHODS: All fungal strains were cultured on potato dextrose agar (PDA) slopes at 30°C for 7 days. The antifungal activity was evaluated by broth microdilution assay. One set of primers was F. verticillioides species specific, which selectively amplified the intergenic space region of rDNA. The other set of primers was specific to FUM1 gene region of fumonisin-producing F. verticillioides. FUM1 transcript levels were quantified using a reverse transcription-polymerase chain reaction (RT-PCR) protocol. RESULTS: Minimum inhibitory concentration (MIC) values of C. cyminum oil against F. verticillioides strains varied from 0.195 to 0.781 µl.ml(-1) (mean MIC value: 0.461 µl.ml(-1)) indicating 54.5% of the fungal strains inhibited at 0.390 µl.ml(-1). PCR analysis of FUM1 gene expression revealed that DNA fragment of 183 bp was amplified in all the isolates of F. verticillioides before treatment with C. cyminum essential oil. Based on RT-PCR analyses, reduction in the expression of fumonisin biosynthetic genes was significant only for FUM1 gene (p<0.05), while no effect was observed on ITS gene. CONCLUSIONS: This study showed that all F. verticillioides isolates were susceptible to C. cyminum essential oil, indicating a significant reduction in the growth of fungal isolates. In addition, this oil completely inhibited the expression of FUM1 gene in concentrations dose-dependently.

Repetitive sequences based on genotyping of Candida albicans isolates obtained from Iranian patients with human immunodeficiency virus.

OBJECTIVES: Candidiasis infection caused by Candida albicans has been known as a major problem in patients with immune disorders. The objective of this study was to genotype the C. albicans isolates obtained from oral cavity of patients with positive human immunodeficiency virus (HIV(+)) with or/and without oropharyngeal candidiasis (OPC). MATERIALS AND METHODS: A total of 100 C. albicans isolates from Iranian HIV(+)patients were genotyped using specific PCR primers of the 25S rDNA and RPS genes. RESULTS: The frequencies of genotypes A, B and C which were achieved using 25S rDNA , were 66, 24 and 10 percent, respectively. In addition, genotypes D and E were not found in this study. Each C. albicans genotype was further classified into four subtypes (types 2, 3, 2/3 and 3/4) by PCR amplification targeting RPS sequence. CONCLUSION: In general, genotype A3 constituted the majority of understudy clinical isolates obtained from oral cavity of Iranian HIV(+) patients.