In Vitro Activity of Amphotericin B in Combination with Statins against Clinical and Environmental Rhizopus oryzae Strains.

BACKGROUND: Mucormycosis is an acute and invasive fungal infection with a high mortality rate. Mucorales are less sensitive than other types of fungi to most antifungal agents. Amphotericin B (AMB) is one treatment option for this infection, but in recent studies, the antifungal activity of statins against Mucorales was shown. Therefore, therapy that combines AMB with these agents may have better effects in management of patients with mucormycosis. We evaluated the in vitro activity of AMB alone and in combination with statins, against Mucorales. METHODS: Susceptibility profiles of AMB alone and in combination with two statins, atorvastatin (ATO) and lovastatin (LOV) determined against clinical (n: 15) and environmental (n: 5) Rhizopus oryzae isolates, obtained between Jan 2009 and Oct 2016 from patients with uncontrolled diabetes mellitus and cancer referred to the Department of Parasitology and Medical Mycology of Tehran University of Medical Sciences, Tehran, Iran. It was performed by microdilution method, based on the Clinical and Laboratory Standard Institute (CLSI) M38-A2 guideline. RESULTS: All clinical and environmental isolates were susceptible to AMB (MIC≤1 µg/mL). The results of the interactions between AMB and the two statins were positive. The AMB-ATO (GM: 0.13 µg/Ml) combination produced greater activity than the AMB-LOV (GM: 0.26 µg/mL) combination. AMB, in combination with ATO and LOV, reacts positively against clinical and environmental R. oryzae isolates. CONCLUSION: This combination strategy may lead to more effective treatment of mucormycosis and fewer side effects using low dose of AMB.

Increased Urine Interleukin-17 and Interleukin-22 Levels in Patients With Candidal Urinary Tract Infection.

INTRODUCTION: Candiduria is common in the hospitalized patients. This study aimed to quantify interleukin (IL)-17 and IL-22 levels in urine of candiduric patients. MATERIALS AND METHODS: A case-control study was conducted on inpatients at Hashemi Nejad Kidney Center. Thirty-four patients were identified with Candida species in their urine samples (> 103 colony-forming units per milliliter and presence of Candida species only). Urine samples with concomitant infections were excluded. Thirty-four patients with negative direct examination and culture were included as the control patients. Interleulin-17 and IL-22 levels were measured in the lyophilized and nonlyophilized urine. The relevant cytokine titers of the two groups were compared, and the association of cytokine elevation and candiduria was investigated. RESULTS: The majority of the candiduric patients were from the intensive care and urology units of women. Only 4 patients (11.7%) manifested fever and dysuria. Massive leukocyturia was observed in 4 patients. Candida glabrata was the most commonly isolated species (44%). Levels of the urine IL-17 and IL-22 were significantly elevated in the candiduric patients, when compared to the noncandiduric controls. While an increased IL-17 level was significantly associated with candiduria (odds ratio, 1.09; 95% confidence interval, 1.003 to 1.17; P = .04), an increased IL-22 level was not. The results showed that lyophilized urine samples maximized the detection power of urinary cytokines. CONCLUSIONS: Our results indicated that direct examination, fungal urine culture, and investigation of urine IL-17 and IL-22 levels are useful tools for diagnosis of Candida urinary tract infection.

The Utilization of RNA Silencing Technology to Mitigate the Voriconazole Resistance of Aspergillus Flavus; Lipofectamine-Based Delivery.

Purpose: Introducing the effect of RNAi in fungi to downregulate essential genes has made it a powerful tool to investigate gene function, with potential strategies for novel disease treatments. Thus, this study is an endeavor to delve into the silencing potentials of siRNA on cyp51A and MDR1 in voriconazole-resistant Aspergillus flavus as the target genes. Methods: In this study, we designed three cyp51A-specific siRNAs and three MDR1-specific siRNAs and after the co-transfection of siRNA into Aspergillus flavus, using lipofectamine, we investigated the effect of different siRNA concentrations (5, 15, 25, 50nM) on cyp51A and MDR1 expressions by qRT-PCR. Finally, the Minimum Inhibitory Concentrations (MICs) of voriconazole for isolates were determined by broth dilution method. Results: Cyp51A siRNA induced 9, 22, 33, 40-fold reductions in cyp51A mRNA expres-sion in a voriconazole-resistant strain following the treatment of the cells with concentrations of 5, 15, 25, 50nM siRNA, respectively. Identically, the same procedure was applied to MDR1, even though it induced 2, 3, 4, 10-fold reductions. The results demonstrated a MIC for voriconazole in the untreated group (4µg per ml), when compared to the group treated with cyp51A-specific siRNA and MDR1-specific siRNA, both at concentrations of 25 and 50nM, yielding 2µg per ml and 1µg per ml when 25 nM was applied and 2µg per ml and 0.5µg per ml when the concentration doubled to 50 nM. Conclusion: In this study, we suggested that siRNA-mediated specific inhibition of cyp51A and MDR1 genes play roles in voriconazole-resistant A.flavus strain and these could be apt target genes for inactivation. The current study promises a bright prospect for the treatment of invasive aspergillosis through the effective deployment of RNAi and gene therapy.

Regulation of ERG3, ERG6, and ERG11 Genes in Antifungal-Resistant isolates of Candida parapsilosis

Background: Candida parapsilosis is one of the five common strains of yeasts involved in invasive candidiasis. The expression analysis of sterol biosynthesis pathway genes, which are associated with resistance, can assist the better understanding of antifungal resistance mechanisms. Method: The antifungal susceptibility of 120 clinical C. parapsilosis isolates was examined. The changes in the gene expression related to resistance were analyzed. Results: Eight strains were resistant to fluconazole (FLC), itraconazole (ITC), and amphotericin B (AMB). The regulation variations included increased mRNA levels of ERG3, ERG6, and ERG11 and decreased mRNA levels of ERG3 and ERG6 in response to FLC. ERG11 mRNA level increases in response to ITC and AMB. Conclusion: The mechanism of resistance to azoles in C. parapsilosis is very similar to C. Albicans. This feature may help to design new treatment strategy for candidiasis.

Expression of Efflux Pumps and Fatty Acid Activator One Genes in Azole Resistant Candida Glabrata Isolated From Immunocompromised Patients.

Acquired azole resistance in opportunistic fungi causes severe clinical problems in immunosuppressed individuals. This study investigated the molecular mechanisms of azole resistance in clinical isolates of Candida glabrata. Six unmatched strains were obtained from an epidemiological survey of candidiasis in immunocompromised hosts that included azole and amphotericin B susceptible and azole resistant clinical isolates. Candida glabrata CBS 138 was used as reference strain. Antifungal susceptibility testing of clinical isolates was evaluated using Clinical and Laboratory Standards Institute (CLSI) methods. Complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) technology, semi-quantitative RT-PCR, and sequencing were employed for identification of potential genes involved in azole resistance. Candida glabrata Candida drug resistance 1 (CgCDR1) and Candida glabrata Candida drug resistance 2 (CgCDR2) genes, which encode for multidrug transporters, were found to be upregulated in azole-resistant isolates (≥2-fold). Fatty acid activator 1 (FAA1) gene, belonging to Acyl-CoA synthetases, showed expression in resistant isolates ≥2-fold that of the susceptible isolates and the reference strain. This study revealed overexpression of the CgCDR1, CgCDR2, and FAA1 genes affecting biological pathways, small hydrophobic compounds transport, and lipid metabolism in the resistant clinical C.glabrata isolates.

Antifungal Susceptibility Analysis of Clinical Isolates of Candida parapsilosis in Iran.

BACKGROUND: Candida parapsilosis is an emergent agent of invasive fungal infections. This yeast is one of the five most widespread yeasts concerned in invasive candidiasis. C. parapsilosis stands out as the second most common yeast species isolated from patients with bloodstream infections especially in neonates with catheter. Recently several reports suggested that its reduced susceptibility to azoles and polyene might become a cause for clinical concern, although C. parapsilosis is not believed to be intensely prone to the development of antifungal resistance. METHODS: In the present report, One hundred and twenty clinical isolates of C. parapsilosis complex were identified and differentiated by using PCR-RFLP analysis. The isolates were then analyzed to determine their susceptibility profile to fluconazole (FLU), itraconazole (ITC) and amphotericin B. The minimum inhibitory concentration (MIC) results were analyzed according to the standard CLSI guide. RESULTS: All of isolates were identified as C. parapsilosis. No C. metapsilosis and C. orthopsilosis strains were found. Evaluation of the antifungal susceptibility profile showed that only three (2.5%) C. parapsilosis were resistant to fluconazole, three (2.5%) C. parapsilosis were resistant to itraconazole and two (1.7%) C. parapsilosis were amphotericin B resistant. CONCLUSION: Profiles in clinical isolates of C. parapsilosis can provide important information for the control of antifungal resistance as well as distribution and susceptibility profiles in populations.

Sporothrix schenckii complex in Iran: Molecular identification and antifungal susceptibility.

Sporotrichosis is a global subcutaneous fungal infection caused by the Sporothrix schenckii complex. Sporotrichosis is an uncommon infection in Iran, and there have been no phenotypic, molecular typing or antifungal susceptibility studies of Sporothrix species. This study aimed to identify nine Iranian isolates of the S. schenckii complex to the species level using colony morphology, carbohydrate assimilation tests, and PCR-sequencing of the calmodulin gene. The antifungal susceptibilities of these Sporothrix isolates to five antifungal agents (amphotericin B (AMB), voriconazole (VRC), itraconazole (ITC), fluconazole (FLC), and terbinafine (TRB)) were also evaluated according to the M27-A3 and M38-A2 protocols of the Clinical and Laboratory Standards Institute for yeast and mycelial phases, respectively. Five of seven clinical isolates were identified as S. schenckii, and two clinical and two environmental isolates were identified as S. globosa. This is the first report of S. globosa in Iran. There was significant agreement (73%) between the results of the phenotypic and genotypic identification methods. TRB and ITC were the most effective antifungals against the Sporothrix isolates. The minimum inhibitory concentration (MIC) values of TRB for the yeast and mycelial phases of S. schenckii differed significantly. There was also a significant difference in the minimum fungicidal concentration (MFC) values of AMB and TRB for the two phases. Considering the low efficacy of VRC and FLC and the wide MIC ranges of AMB (1-16 µg/ml and 1-8 µg/ml for yeast and mycelial forms, respectively) observed in the present study, in vitro antifungal susceptibility testing should be performed to determine appropriate therapeutic regimens.

Identification of Azole Resistance Markers in Clinical Isolates of Candida tropicalis Using cDNA-AFLP Method.

BACKGROUND: Global reports have highlighted the increasing prevalence of Candida tropicalis infections as well as organism(') s drug resistance. This study aimed at identifying azole resistance markers in clinical isolates of C. tropicalis, which will be a great resource for developing new drugs. METHODS: Two susceptible and resistant isolates of C. tropicalis were recovered from an epidemiological investigation of candidiasis in immunocompromised patients. C. tropicalis ATCC 750 was used as reference strain. Antifungal susceptibility to fluconazole and itraconazole was determined using Clinical and Laboratory Standards Institute (CLSI) method. Complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) technology and real-time reverse-transcriptase (RT) PCR were used for identification of potential genes involved in azole resistance of C. tropicalis clinical isolates. RESULTS: Five genes encoding the following enzymes were identified as superoxide dismutase (SOD) implicated in antioxidant defense, ornithine aminotransferase (OAT), acetyl ornithine aminotransferase (ACOAT), adenosylmethionine-8-amino-7-oxononanoate aminotransferase (DAPA AT), and 4-aminobutyrate aminotransferase (ABAT)-belonging to pyridoxal phosphate (PLP) dependent enzymes and acting in an important physiological role in many fungal-cell cycles. Real-time RT-PCR confirmed mRNA level of the aforementioned genes. CONCLUSION: Our findings showed that factors such as PLP-dependent enzymes and SOD might be implicated in drug resistance in C. tropicalis clinical isolate. Therefore, further studies are required to explore the accurate biological functions of the mentioned genes that would be helpful for effective drug development.

Detection of Fungal Elements in Atherosclerotic Plaques Using Mycological, Pathological and Molecular Methods.

BACKGROUND: The aim of this study was to detect fungi in atherosclerotic plaques and investigate their possible role in atherosclerosis. METHODS: Coronary atherosclerotic plaques specimen were obtained from patients with atherosclerosis. Direct examination, culture, histopathology study, PCR and sequencing were performed to detect/identify the mycotic elements in the plaques. Age, sex, smoking, obesity, hypertension, hyperlipidemia, family history of heart diseases and diabetes were considered and data were analyzed using Chi Square test by SPSS version 15. RESULTS: A total of 41 specimens were analyzed. Direct examination for fungal elements was negative in all cases but in culture only one specimen grew as a mold colony. The presence of fungal elements were confirmed in 6 and 2 tissue sections stained by Gomori methenamine silver and Hematoxylin and Eosin methods, respectively. Using PCR, 11 cases were positive for fungi. The DNA sequence analysis of six positive specimens which were randomly selected revealed fungi as Candida albicans (n=3), Candida guilliermondii (n=2) and Monilia sp. (n=1). CONCLUSION: A significant association between the presence of fungi in atherosclerotic plaques and severity of atherogenesis and atherosclerotic disease was not found. This could be due to limited numbers of patients included in our study. However, the presence of fungal elements in 26.8% of our specimens is considerable and the results does not exclude the correlation between the presence of fungi with atherosclerosis and coronary artery disease.

In Vitro Activity of Caspofungin Against Fluconazole-Resistant Candida Species Isolated From Clinical Samples in Iran.

BACKGROUND: Candida spp. is the most common organisms involved in fungal infections in the high risk patients. It causes the greatest number of invasive candidiasis. Fluconazole is effective in treating mucosal candidiasis. However, resistance to fluconazole and other azoles antifungal drugs is an important clinical problem to treat candidiasis. Caspofungin is more effective against Candida species such as some azoles-resistant isolates. OBJECTIVES: The current study aimed to investigate the susceptibilities of clinical fluconazole-resistant and fluconazole - susceptible dose- dependent Candida species to caspofungin. MATERIALS AND METHODS: In the Minimum Inhibitory Concentration (MIC) test, 207 Candida species and other yeasts isolated from Iranian patients (each isolated from a high-risk patient) were evaluated. The yeasts were differentiated by standard mycological methods, CHROM agar Candida, and verified by API20C.AUX. In vitro susceptibilities were determined using Broth Micro Dilution (BMD) method described in the Clinical Laboratory Standards Institute M27-A3. MICs were noted after 24 and 48 hours of incubation. RESULTS: The most frequently isolated species were Candida albicans (52.2%), C. glabrata (24.6%), followed by C. tropicalis (7.7%) and C. krusei (3.4%). MICs of caspofungin against 87% of C. albicans and 90% of C. glabrata and C. tropicalis isolates were 2 µg/mL and for C. krusei were 4 µg/mL, respectively. The results revealed that only 20 out of 207 isolates (9.7%) were non-sensitive to caspofungin. Caspofungin non-susceptible isolates were isolated from the patients with cancer, diabetes and AIDS; and not in the species isolated from patients with other underlying diseases. CONCLUSIONS: Caspofungin appears more effective in vitro against Iranian fluconazole-resistant Candida isolates and some other yeasts.

Evaluation of mRNA Expression Levels of cyp51A and mdr1, Candidate Genes for Voriconazole Resistance in Aspergillus flavus.

BACKGROUND: Voriconazole Resistance (VRC-R) in Aspergillus flavus isolates impacts the management of aspergillosis, since azoles are the first choice for prophylaxis and therapy. However, to the best of our knowledge, the mechanisms underlying voriconazole resistance are poorly understood. OBJECTIVES: The present study was designed to evaluate mRNA expression levels of cyp51A and mdr1 genes in voriconazole resistant A. flavus by a Real-Time Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) technique. MATERIALS AND METHODS: Five A. flavus isolates with resistance to VRC were examined by a RT-PCR approach. RESULTS: Four out of five isolates revealed cyp51A and mdr1 mRNA overexpression. Interestingly, the isolate, which was negative for cyp51A and mdr1 mRNA expression showed a high voriconazole Minimum Inhibitory Concentration (MIC). Furthermore, a computational-based analysis predicted that voriconazole resistance could be mediated through cooperation with a network protein interaction. CONCLUSIONS: Our experimental and in silico findings may provide new insight in the complex molecular pathways of drug resistance and also could assist design an efficient therapeutic strategy for aspergillosis treatment.

Identification and Sequencing of Candida krusei Aconitate Hydratase Gene Using Rapid Amplification of cDNA Ends Method and Phylogenetic Analysis.

BACKGROUND: The production and development of an effective fungicidal drug requires the identification of an essential fungal protein as a drug target. Aconitase (ACO) is a mitochondrial protein that plays a vital role in tricarboxylic acid (TCA) cycle and thus production of energy within the cell. OBJECTIVES: The current study aimed to sequence Candida krusei ACO gene and determine any amino acid residue differences between human and fungal aconitases to obtain selective inhibition. MATERIALS AND METHODS: Candida krusei (ATCC: 6258) aconitase gene was determined by 5'Rapid Amplification of cDNA Ends (RACE) method and degenerate Polymerase Chain Reaction (PCR) and analyzed using bioinformatics softwares. RESULTS: One thousand-four hundred-nineteen nucleotide of C. krusei aconitase gene were clarified and submitted in Genbank as a partial sequence and then taxonomic location of C. krusei was determined by nucleotide and amino acid sequences of this gene. The comparison of nucleotide and amino acid sequences of Candida species ACO genes showed that C. krusei possessed characteristic sequences. No significant differences were observed between C. krusei and human aconitases within the active site amino acid residues. CONCLUSIONS: Results of the current study indicated that aconitase was not a suitable target to design new anti-fungal drugs that selectively block this enzyme.

Multilocus variable-number tandem-repeat analysis of clinical isolates of Aspergillus flavus from Iran reveals the first cases of Aspergillus minisclerotigenes associated with human infection.

BACKGROUND: Aspergillus flavus is intensively studied for its role in infecting crop plants and contaminating produce with aflatoxin, but its role as a human pathogen is less well understood. In parts of the Middle East and India, A. flavus surpasses A. fumigatus as a cause of invasive aspergillosis and is a significant cause of cutaneous, sinus, nasal and nail infections. METHODS: A collection of 45 clinical and 10 environmental A. flavus isolates from Iran were analysed using Variable-Number Tandem-Repeat (VNTR) markers with MICROSAT and goeBURST to determine their genetic diversity and their relatedness to clinical and environmental A. flavus isolates from Australia. Phylogeny was assessed using partial ß-tubulin and calmodulin gene sequencing, and mating type was determined by PCR. Antifungal susceptibility testing was performed on selected isolates using a reference microbroth dilution method. RESULTS: There was considerable diversity in the A. flavus collection, with no segregation on goeBURST networks according to source or geographic location. Three Iranian isolates, two from sinus infections and one from a paranasal infection grouped with Aspergillus minisclerotigenes, and all produced B and G aflatoxin. Phylogenic analysis using partial ß-tubulin and calmodulin sequencing confirmed two of these as A. minisclerotigenes, while the third could not be differentiated from A. flavus and related species within Aspergillus section flavi. Based on epidemiological cut-off values, the A. minisclerotigens and A. flavus isolates tested were susceptible to commonly used antifungal drugs. CONCLUSIONS: This is the first report of human infection due to A. minisclerotigenes, and it raises the possiblity that other species within Aspergillus section flavi may also cause clinical disease. Clinical isolates of A. flavus from Iran are not distinct from Australian isolates, indicating local environmental, climatic or host features, rather than fungal features, govern the high incidence of A. flavus infection in this region. The results of this study have important implications for biological control strategies that aim to reduce aflatoxin by the introduction of non-toxigenic strains, as potentially any strain of A. flavus, and closely related species like A. minisclerotigenes, might be capable of human infection.

Overexpression of aldo-keto-reductase in azole-resistant clinical isolates of Candida glabrata determined by cDNA-AFLP.

BACKGROUND: Candida glabrata causes significant medical problems in immunocompromised patients. Many strains of this yeast are intrinsically resistant to azole antifungal agents, and treatment is problematic, leading to high morbidity and mortality rates in immunosuppressed individuals. The primary goal of this study was to investigate the genes involved in the drug resistance of clinical isolates of C. glabrata. METHODS: The clinical isolates of C. glabrata were collected in an epidemiological survey of candidal infection in immunocompromised patients and consisted of four fluconazole and itraconazole resistant isolates, two fluconazole and itraconazole sensitive isolates, and C. glabrata CBS 138 as reference strain. Antifungal susceptibility patterns of the organisms were determined beforehand by the Clinical and Laboratory Standards Institute (CLSI). The potential gene(s) implicated in antifungal resistance were investigated using complementary DNA- Amplified Fragment Length Polymorphism (cDNA-AFLP). Semi-quantitative RT-PCR was carried out to evaluate the expression of gene(s) in resistant isolates as compared to sensitive and reference strains. RESULTS AND CONCLUSIONS: The aldo-keto-reductase superfamily (AKR gene) was upregulated in the resistant clinical isolates as assessed by cDNA-AFLP. Semi-quantitative RT-PCR revealed AKR mRNA expression approximately twice that seen in the sensitive isolates. Overexpression of the AKR gene was associated with increased fluconazole and itraconazole resistance in C. glabrata. The data suggest that upregulation of the AKR gene might give a new insight into the mechanism of azole resistance.

Molecular epidemiology of dermatophytosis in Tehran, Iran, a clinical and microbial survey.

In the framework of a survey on dermatophytoses, 14,619 clinical specimens taken from outpatients with symptoms suggestive of tinea and referred to a Medical Mycology laboratory in Tehran, Iran, were analyzed by direct microscopy and culture. In total, 777 dermatophyte strains recovered in culture were randomly identified by a formerly established RFLP analysis method based on the rDNA ITS regions. For confirmation of species identification, 160 isolates representing the likely entire species spectrum were subjected to ITS-sequencing. Infection was confirmed in 5,175 collected samples (35.4%) by direct microscopy and/or culture. Tinea pedis was the most prevalent type of infection (43.4%), followed by tinea unguium (21.3%), tinea cruris (20.7%), tinea corporis (9.4%), tinea manuum (4.2%), tinea capitis (0.8%) and tinea faciei (0.2%). Trichophyton interdigitale was the most common isolate (40.5%) followed by T. rubrum (34.75%), Epidermophyton floccosum (15.6%), Microsporum canis (3.9%), T. tonsurans (3.5 %) and M. gypseum (0.5%). Other species included M. ferrugineum, T. erinacei, T. violaceum, T. schoenleinii, and a very rare species T. eriotrephon (each one 0.25%). The two strains of T. eriotrephon isolated from tinea manuum and tinea faciei are the second and third reported cases worldwide. Application of DNA-based methods is an important aid in monitoring trends in dermatophytosis in the community.

Candida species in cutaneous candidiasis patients in the Guilan province in Iran; identified by PCR-RFLP method.

Due to the epidemiological alteration in distribution of Candida species as well as significant increasing trend of either intrinsic or acquired in resistance of some of these fungi, the precise identification of Candida species is necessary for effective antifungal therapy and also for prevention of nosocomial infections. PCR-RFLP method is indicated to be a reliable, rapid and simple technique which is able to differentiate the Candida species. In the present study, we applied this method to evaluate the distribution of Candida species in patients affected with cutaneous candidiasis in the Guilan province. 896 clinical cutaneous samples were collected from different parts of skin and nail of suspected patients referred to clinical centers all over the Guilan province during 24 months. Samples were examined directly with 15% KOH and cultured on fungal specific media. Genomic DNA was extracted and the restriction enzyme Msp1 was applied for polymorphism analysis. Totally, 47 yeast strains were successfully isolated from different clinical samples and identified by conventional as well as PCR-RFLP methods. The results indicated that Candida albicans (36.17%) was the most frequent species followed by C. parapsilosis (25.53%), C. tropicalis (19.14%), C. guilliermondii (14.89%), C. famata (2.12%) and C. krusei (2.12%). Female finger nails were the most common location to be affected by Candida species. In conclusion, PCR-RFLP method was successfully used for recognition of clinical Candida species within the Guilan province and obtained results revealed C. albicans as the predominant causative agent of cutaneous candidiasis. However, distribution of other Candida species did not completely consist with the reported distribution of Candida species in other parts of Iran with different climate to the Guilan province.

In vitro susceptibility of aflatoxigenic and non-aflatoxigenic Aspergillus flavus strains to conventional antifungal agents.

Presently appearance of resistance to antifungal agents among Aspergillus species is dramatically increasing. The objective of this study was to look at the in vitro activities of antifungal drugs against Iranian clinical (from nail, bronchoalveolar lavage, paranasal sinus) isolated A. flavus strains. The susceptibility of 45 aflatoxigenic and non-aflatoxigenic Aspergillus flavus strains were evaluated to six antifungal agents (caspofungin, itraconazole, amphotericin B, ketoconazole, fluconazole, nystatin) using CLSI M38-A2 broth microdilution method. The results indicated that 57.1%, 28.6% of aflatoxigenic and 25.8%, 6.5% of non-aflatoxigenic isolates were susceptible to caspofungin, amphotericin B respectively. All isolates but one aflatoxigenic strain were sensitive to ketoconazole. All 45 strains showed to be resistant to nystatin. Also 64.28%, 92.9% of aflatoxigenic and 64.51%, 100% of non-aflatoxigenic isolates were resistant to fluconazole and itraconazole in ranking order. There was no statistically significant difference between the susceptibilities of aflatoxigenic and non-aflatoxigenic strains of A. flavus to tested antifungal agents.

A case of onychomycosis caused by Aspergillus candidus.

Based on epidemiological studies, Aspergillus candidus has been demonstrated as an emerging fungal agent of toenail onychomycosis. Here we report a case of a toenail infection caused by A. candidus in a healthy 60-year-old woman. Based on macroscopic and microscopic characteristics of the culture as well as nucleotide sequencing of 28S region, the causative agent was identified as A. candidus.

A case report of tinea pedis caused by Trichosporon faecale in Iran.

Trichosporon species are known as the causative agents of cutaneous infections and are involved in systemic, localized, as well as disseminated mycoses particularly in immunocompromised patients. Here we report a case of tinea pedis infection caused by Trichosporon faecale in a healthy 29-year-old woman in the north of Iran. Macroscopic and microscopic characteristics using direct examination as well as culture method revealed the causative agent as Trichosporon species. Molecular analysis of the internal transcribed spacer region validated the initial result and indicated that this case of tinea pedis was caused by T. faecale. The patient was recovered after treatment with topical myconazole accompanied with oral fluconazole.

Partial purification and characterization of a 37 kDa extracellular proteinase from Trichophyton vanbreuseghemii.

An exocellular proteinase synthesized by the geophilic dermatophyte Trichophyton vanbreuseghemii has been purified and characterized. The fungus obtained from soil in Iran was cultivated in modified Czapek-Dox liquid medium containing 0.1% bacteriological peptone and 1% glucose as the nitrogen and carbon sources. Partial purification of the proteinase was accomplished by (NH(4))(2)SO(4) precipitation, followed by ion exchange chromatography. Analysis of the enzyme by SDS-PAGE revealed a single polypeptide chain with an apparent molecular mass of 37 kDa. Proteinase activity was optimum at pH 8, but remained high in the range of pH 7-11. Moreover, the partially purified enzyme presented a keratinolytic activity as evidenced by the keratin azure test. The inhibition profile and the good activity of the enzyme towards the synthetic substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide suggested that it belonged to the chymotrypsin/subtilisin group of serine proteinases. The keratinolytic properties of T. vanbreuseghemii suggest that this fungus may be an alternative for the recycling of industrial keratinic wastes.