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The nomenclature of fungi has undergone significant changes in the last decade. Renaming of fungi has always occurred and will continue, largely due to the influence of molecular methods in taxonomy, diagnosis, and epidemiology. Recently, advancements in molecular techniques and genetic analysis have have led to changes in the classification of dermatophytes that could potentially affect the clinical diagnosis and management of the disease. Fungal culture is still considered the "gold standard" for the diagnosing dermatophytosis; however, modern molecular assays have overcome the major drawbacks of culture and can be used in conjunction with it. Dermatophytes are one of the oldest groups of microorganisms known to cause disease in humans and animals. The ability of dermatophytes to secrete keratinolytic enzymes in vitro has led to the study of secretory proteases. Studies on the quantity and function of keratinolytic proteases produced by dermatophytes are increasing, and the enzymatic properties of these compounds have been investigated. Because of the importance of this group of fungi in public health, this review focuses on the new classification of dermatophytes, including the renaming of important medical species, as well as the pathogenic factors associated with them. The search strategy used was a narrative review, using national and international sources such as PubMed, Web of Science, Google Scholar, Scopus, and the country's periodicals database to collect information. In the proposed new classification, Arthroderma includes 21 species, Trichophyton 16, Nanzia 9, Microsporum 3, Paraphyton 3, Epidermophyton and Lophophyton one each. However, more detailed studies are needed to determine the boundaries of these species. Based on previous studies, it can be concluded that the current classification of common anthropophilic dermatophytes is fairly stable and is unlikely to change drastically in the future. While the study of geophilic dermatophytes has been inadequate compared to many zoophilic species, further taxonomic innovation in these groups is expected. In addition, research into the pathogenic mechanisms of dermatophytes has shown that endopeptidases and exopeptidases play an important role in the pathogenesis of dermatophytes by degrading keratin and converting it into free peptides and smaller amino acids. The most important endopeptidases are from the fungilysin family (M36) and two genes encoding MEP1 and MEP5, while the most important extracellular aminopeptidases are the leucine aminopeptidases Lap1 and Lap2 (M28A family) and the dipeptidyl peptidases DppIV and DppV, which are considered to be dermatophyte exopeptidases. The results of previous studies suggest that by identifying these major proteases, it is possible to isolate and identify dermatophyte species. Based on the results of this study, many factors related to the classification and pathogenesis of dermatophytes remain inadequately expressed in existing research. However, the factors discussed in this study serve as a guide for the identification of relevant factors.

Purpose: The increasing resistance to onychomycosis treatment globally is a major concern as it results in treatment failures. This research aimed to investigate the effectiveness of incorporating nanostructured lipid carriers (NLCs) in a gel formulation for treating mild to moderate Candida-associated onychomycosis using Zataria multiflora (Zt) essential oils. Methods: The characteristics of Zataria multiflora-loaded nanostructured lipid carriers (Zt-NLCs) were validated using a carbopol gel (1 % w/w). In vitro tests were conducted using CLSI M60 standards to evaluate the susceptibility of ten commonly encountered dermatophyte species to antifungal agents. A clinical study was conducted involving 40 volunteers randomly assigned to two groups: one receiving the Zt-NLCs gel and the other receiving a placebo, using a double-blind, placebo-controlled design. The objective was to assess clinical manifestations and mycological findings after topical application for 2 and 4 weeks. The causative agents' species were identified using a PCR-RFLP method. Results: The preparation of Zt-NLCs gel resulted in a uniform suspension of spherical nanoparticles with favourable characteristics and no cytotoxic effects. Zt-NLCs demonstrated significant inhibitory effects on fungal growth and effectively improved clinical and mycological criteria in comparison to the placebo group (p

Tinea capitis is a chronic fungal infection of the scalp occurring commonly in children of school age, especially in developing countries. It is caused primarily by the dermatophyte members of genera Microsporum and Trichophyton. Here we report presence of free-living mycelial stage of dermatophytes, a stage of fungal growth which form in culture medias, around affected hairs and skin scrapings of scalp lesions in a 3-year-old boy presenting with alopecia and multiple scaly, non-erythematous plaques. On direct microscopy examination using 10 % potassium hydroxide, the fungal hyphae and arthrospores were detected in ectothrix form. In addition, we also observed numerous multicellular, thick-walled spindle-shaped macroconidia around hairs and skin scrapings. To our knowledge this is the first study reporting dermatophyte's macroconidia directly seen on clinical samples. Species level identification of the dermatophyte isolate growing on Mycosel™ agar was confirmed by PCR-sequencing of internal transcribed spacer of ribosomal RNA as Microsporum ferrugineum. The patient was successfully treated with systemic itraconazole combined with topical ketoconazole shampoo.

Background and purpose: Among different clinical entities of dermatophytosis, tinea capitis (TC) is considered a major public health challenge in the world, especially in regions with poor health and low income. Therefore, this study aimed to provide a retrospective analysis of the patients suspected of TC who were referred to the medical mycology laboratory of Mazandaran, a northern province of Iran.Materials and Methods: A retrospective analysis was performed on the patients suspected of TC who were referred to the medical mycology laboratory from July 2009 to April 2022. Hair roots and skin scrapings were collected from the participants. The laboratory diagnosis was confirmed by direct microscopic examination and culture. Finally, 921 out of 11095 (8.3%) patients were suspected of TC.Results: Based on the findings, TC was confirmed in 209 out of 921 patients (22.7%). In terms of gender, 209 TC patients (75.1%) were male. Moreover, the male to female ratio of TC patients was 1:3.0. Trichophyton tonsurans (146/174, 83.91%) was the most etiological agent,followed by T. mentagrophytes (13/174, 7.47%), T. violaceum (9/174, 5.17%), Microsporum canis (3/174, 1.71%), T. verrucosum (2/174, 1.15%) and T. rubrum (1/174, 0.57%). Besides, endothrix (77.0%) was the most prevalent type of hair invasion.Conclusion: The results revealed the predominance of T. tonsurans, as a causative agent of TC. Despite the prevalence of TC, the absence of appropriate consideration highlights that it is a neglected complication among children.

Abstract Background Dermatophytes have the ability to invade the keratin layer of humans and cause infections. The aims of this study were the accurate identification of dermatophytes by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism method and sequencing and comparison between the in vitro activities of newer and established antifungal agents against them. Methods Clinical specimens of patients from five Iranian university laboratories were entered in this study. Samples were cultured on sabouraud dextrose agar medium. For molecular identification, extracted DNAs were amplified by the universal fungal primers ITS1 and ITS4, and digested with MvaI enzymes. The antifungal susceptibility test for each isolate to terbinafine, griseofulvin, caspofungin, fluconazole, itraconazole, luliconazole, and isavuconazole was performed, according to the microdilution CLSI M38-A2 and CLSI M61 standard methods. Results Two hundred and seven fungi species similar to dermatophytes were isolated of which 198 (95.6%) were dermatophytes by molecular assay. The most commonly isolated were Trichophyton mentagrophytes (76/198), followed by Trichophyton interdigitale (57/198), Trichophyton rubrum (34/198), Trichophyton tonsurans (12/198), Microsporum canis (10/198), Trichophyton simii (3/198), Epidermophyton floccosum (3/198), Trichophyton violaceum (2/198), and Trichophyton benhamiae (1/198). The GM MIC and MIC90 values for all the isolates were as follows: terbinafine (0.091 and 1 μg/ml), griseofulvin (1.01 and 4 μg/ml), caspofungin (0.06 and 4 μg/ml), fluconazole (16.52 and 32 μg/ml), itraconazole (0.861 and 8 μg/ml), isavuconazole (0.074 and 2 μg/ml), and luliconazole (0.018 and 0.25 μg/ml). Conclusion Trichophyton mentagrophytes, Trichophyton interdigitale, and Trichophyton rubrum were the most common fungal species isolated from the patients. luliconazole, terbinafine, and isavuconazole in vitro were revealed to be the most effective antifungal agents against all dermatophyte isolates.

Introduction: This study was proposed to assess the potential role of efflux transporters in reversing fluconazole resistance in Candida glabrata isolates treated with fluconazole loaded nanostructured lipid carriers (FLZ-NLCs). Methods: The ultrasound technique was used to synthesize the FLZ-NLCs. Four fluconazole-resistant, as well as one susceptible standard C. glabrata isolates, were applied and exposed to FLZ/ FLZ-NLCs for 20 h at 37°C. Real-time PCRs were done to estimate the likely changes in ATP-binding cassette transporter genes. Results: Similar to the FLZ-exposed-susceptible standard strain which showed no alteration, the genes were not up-regulated significantly under the FLZ-NLCs treated condition. While they were over-expressed when the yeasts were treated with fluconazole. Conclusion: It is highly suggested that due to the nature of the NLCs which shields the whole conformation of the drug, FLZ is not recognized by the efflux transporter subunits and consequently the translocation would not happen.

Background and Purpose: This study aimed to evaluate the species distribution and susceptibility pattern of the strains isolated from Candida colonization in pediatric patients staying at pediatric intensive care unit (ICU) and infant ICU of Children’s Medical Center in Tehran, Iran.Materials and Methods: This study was conducted in the Children’s Medical Center in Tehran, Iran. In total, 440 samples from 56 patients with oral cavity, skin surrounded catheters, and ear, throat, nasal, and urine cultures were collected. All patients were evaluated in terms of Candida colonization on the admission day as well as the days 7, 14, and 28 according to the previous studies. CHROMagar Candida medium was applied for primary/multiple species identification and the isolates were identified by using polymerase chain reaction-based methods to the species-specific complex level. The antifungal susceptibility test was performed according to the Clinical and Laboratory Standards protocol published as M27-A3 and M60 documents.Results: In total, 136 yeast samples from 26 individuals (30.9%) out of 440 samples were considered colonization. The most prevalent species in IICU was C. albicans (27%,n=20) followed by C. krusei (24 %, n=18) and C. parapsilosis (16%, n=12). In PICU, the predominant species was C. krusei (40%, n=24) followed by C. parapsilosis (18%, n=11) and C. dubliniensis (16%, n=10). Among the 40 tested isolates from both units, fluconazole-resistant isolates (n=11, 8.15%) were determined according to the new breakpoints. In the case of echinocandins, 2 isolates, including C. albicans (n=1) and C. krusei (n=1) were resistant against both caspofungin and anidulafungin (totally 1.48%).Conclusion: In the present study, since C. krusei is intrinsically-resistance against fluconazole, emphasizing the importance of species-level identification of Candida isolates is outstanding. However, according to the antifungal susceptibility testingresults, only 7.2% of the strains were resistant to fluconazole. It would be beneficial tomonitor the ICU patients who are at high risk of invasive Candida infection.

Background and Purpose: Invasive mucormycosis is a rare mycosis that affects most cases of uncontrolled diabetes and has a high mortality rate. Patients with COVID-19 are at high risk of developing invasive mucormycosis due to the consumption of anti inflammatory drugs such as corticosteroids and dexamethasone. Rhizopus species followed by Rhizomucor spp. and Mucor spp. are the main common etiological agents of rhino-orbital mucormycosis. Therefore, this study aimed to present a case of mucormycosis due to Syncephalastrum racemosum in a diabetic patient with COVID-19 for the first time in Iran.Case report: A 73-year-old diabetic female was referred to Ayatollah Rouhani Hospital in Babol, Iran, with a confirmed COVID-19 diagnosis, based on positive RT-PCR and computed tomography of the lungs. She has received methylprednisolone due to severe lung complications. Nasal involvement and left orbital swelling were observed 20 days after the hospitalization. By sinus endoscopic surgery, debridement was done and histopathology indicated wide hyphae (without septa). The sequenced PCR products displayed Syncephalastrum racemosum. In the antifungal susceptibility test, amphotericin B showed good activity against S. racemosum and the patient survived with timely treatment.Conclusion: This is the first case report of rhino-orbital mucormycosis due to S. racemosum in COVID-19 patient; therefore, S. racemosum can be considered one of the etiological factors of rhino-orbital mucormycosis in COVID-19 cases.

Coronavirus disease 2019 (COVID-19) has seriously spread worldwide. One of the problems of COVID-19 patients is co-infection with other microbial infections such as fungal infections that are severely life-threatening. Mucormycosis, one of these opportunistic fungal diseases that is mistakenly known as black fungal disease, has been reported in Iran and the world. This non-systematic review is intended to review the latest studies in the world and Iran about the incidence of mucormycosis in COVID-19 patients to better understand the disease and its predisposing factors. In general, patients with COVID-19 with underlying diseases, diabetic patients, those treated with immunosuppressive drugs, ICU patients, and patients under invasive mechanical ventilation are at higher risk for mucormycosis. Therefore, early diagnosis, treatment, and prevention methods can be effective in reducing the risk of mucormycosis in COVID-19 patients.

Background and Purpose: Scedosporium spp. is a saprophytic fungus that may cause invasive pulmonary infection due to the aspiration of contaminated water in both immunosuppressed and immunocompetent hosts.Case report: Herein, we report a fatal case of pulmonary infection caused by Scedosporium species associated with a car crash and near-drowning in a sewage canal.Scedosporium aurantiacum isolated from bronchoalveolar lavage was identified by PCR-sequencing of β-tubulin genes. The minimum inhibitory concentration values for amphotericin B, itraconazole, posaconazole, isavuconazole were >16 µg/ml, and >8 µg/ml for anidulafungin, micafungin, and caspofungin. Voriconazole was found to be the most active agent with a MIC of 1 µg/ml.Conclusion: This report, as the first case of pulmonary scedosporiosis after near-drowningin Iran, highlights the importance of high suspicion in near-drowning victims, promptidentification of Scedosporium spp., and early initiation of appropriate antifungal therapy.

Background and purpose: Invasive candidiasis in infants has high mortality rate and is often associated with colonization on medical devices such as catheters. Candida colonization is considered as a predisposing factor for Candida related infection. The aim of this study was to evaluate the rate of catheters infectivity by Candida species. Materials and methods: In this descriptive cross-sectional study, sampling was done from catheters of hospitalized children (0-18 years old) in pediatric intensive care unit (PICU) in Mashhad Dr. Sheikh Hospital, June-November 2018. All catheters were evaluated to determine the extent of contamination and early identification of isolates was done by morphological and molecular methods. Results: The types of catheter infectivity for 114 samples included colonization or infective (n=25 isolates, 21.92%), contamination (n=4, 3.5%), and no growth (n=85 isolates, 56.74%). The most isolated species was C. albicans (n=18, 13.39%), followed by C. glabrata (n=9, 19.56%), C. krusei and C. parapsilosis (n=7, 15.21%, each isolate), C. tropicalis (n=3, 6.25%), and C. lusitaniae and C. kefyr (n=1, 2.17% each isolate). Conclusion: Isolation of Candida species in 25% of catheters raised the possibility of colonization, considering the catheters as an exogenous potential source of candidiasis which is very important in PICU.

Background and Purpose: Coronavirus disease 2019 (COVID-19) and Candida prostatic valve endocarditis present various clinical manifestations which may overlap;hence, discrimination between them is extremely difficult.Case report: The case was a 66-year-old man with a past medical history of mitral and aortic valves replacement one year before COVID-19 co-infection. He was admitted with fever (for 7 days), shortness of breath, cough, seizure, lethargy,headache, and 85% oxygen saturation. Transesophageal echocardiography revealed multiple large-sized, highly mobile masses on both sides of the mechanical mitral valve highly suggestive of vegetation. Chest computed tomography scanning showed simulating scattered COVID-19 peripheral ground-glass opacities confirmed by reverse-transcription polymerase chain reaction. The set of blood cultures yieldedyeast colonies that were identified as Candida tropicalis. The patient died of septic shock shortly after receiving antifungal therapy.Conclusion: This case emphasized the importance of early diagnosis andimplementation of antifungal treatment, particularly in patients with prosthetic cardiacvalves, to reduce their unfavorable outcomes in COVID-19 patients

ABSTRACT Aspergillus species are a major cause of life-threatening invasive infections and noninvasive diseases. This study seeks to investigate the frequency of Aspergillus species among Iranian patients and their susceptibility to seven antifungals. In a cross-sectional study, 233 Aspergillus isolates were collected from 11 university hospitals in Iran between 2018 and 2021. Aspergillus isolates were identified based on colony morphology, microscopic characteristics, PCR-restriction fragment length polymorphism (RFLP), and sequencing of the beta-tubulin gene. The CLSI M38-A2 reference methodology was used for antifungal susceptibility testing of amphotericin B, voriconazole, posaconazole, itraconazole, luliconazole, isavuconazole, and caspofungin. Members of Aspergillus section Flavi (117/233, 50.2%), Aspergillus section Nigri (77/233, 33.1%), Aspergillus section Fumigati (21/233, 9%), Aspergillus section Terrei (14/233, 6%), Aspergillus pseudodeflectus (2/233, 0.85%), and Aspergillus melleus (2/233, 0.85%) were isolated from the samples. The lowest 0.25 MIC90 values for all isolates tested were for luliconazole (0.016 μg/mL) and isavuconazole (0.250 μg/mL), and the highest value was observed for itraconazole (≥ 8μg/mL). The 90% minimum effective concentration (MEC90) value for caspofungin was 0.125 μg/mL. MIC90 values for voriconazole, amphotericin B, and posaconazole were 1, 2, and 2 μg/mL, respectively. The non-wild-type species were presented for amphotericin B (3%), voriconazole (1.3%), posaconazole (2.6%), luliconazole (1.3%), isavuconazole (1.7%), and caspofungin (4.7%). Positive correlations in the MIC values of azole antifungals were observed, and using one azole increases the MIC value rates of other ones. None of the species were pan-azole resistant. Species of Aspergillus section Flavi were the most common Aspergillus species isolated from Iranian samples. Luliconazole, caspofungin, and isavuconazole present the most effective antifungal agents for treatment of infection due to Aspergillus species. Susceptibility tests should be performed frequently in each region for the best management of patients. IMPORTANCE Aspergillus species are the leading cause of invasive aspergillosis in immunocompromised hosts. The susceptibility of Aspergillus species to antifungal agents might be different. Azole-resistant species have emerged worldwide. Performing susceptibility testing in each region can help in the best management of patients. Here, we show the epidemiology and distribution of Aspergillus species in Iran and their susceptibility patterns for seven antifungal agents. The significant points of the present study are that species of Aspergillus section Flavi are the most prevalent Aspergillus species isolated from 11 university hospitals. Luliconazole, caspofungin, and isavuconazole were effective antifungal agents against all Aspergillus species.

Background and Purpose: Radiation therapy in patients with head and neck malignancies predisposes them to oral Candida colonization and infection due to damage of oral mucosa and destruction of the salivary gland. This study aimed to determine the prevalence of oropharyngeal candidiasis (OPC) in patients with head and neck cancer (HNC) undergoing radiotherapy (RT), identify the yeasts isolated from them, and determine their antifungal susceptibility. Materials and Methods: This cross-sectional study was conducted from December 2018 to June 2019 at two referral radiotherapy centers in northern Iran. Yeast strains that were isolated from patients with HNC were identified using conventional and molecular methods. The in vitro activities of eight common antifungal drugs against 55 isolates were investigated according to the guidelines of the Clinical and Laboratory Standard Institute (M27-A3 and M27-S4) broth microdilution document. Results: Among 59 patients receiving RT, the prevalence of OPC was 21 (35.59%) and 15 (25.42%) patients were diagnosed with colonization. The mean age of the patients was 55.32±13.3 years (within the range of 27-87 years). In this study, the pseudomembranous form was reported as the most clinical type of OPC. Candida albicans with the frequency of 60% was the most common type of Candida spp. that was observed in this study, although non-albicans Candida spp., such as C. glabrata (27.27%), C. tropicalis (5.45%), C. parapsilosis (3.63%), C. krusei (1.83%), and C. kefyr (1.83%) were also isolated. Considering the low minimum inhibitory concentration range of amphotericin B, compared to fluconazole, administration of this agent is a more suitable antifungal drug for extensive oral candidiasis in these patients. Among azoles,clotrimazole had low efficacy and several studied isolates (65.5%) showed resistance. Conclusion: Correct diagnosis as well as determining drug sensitivity and risk factors are the effective steps in reducing the complications related to oral candidiasis in people undergoing RT.

Background and Purpose: Because of the growing incidence of Aspergillus infection,typing methods of Aspergillus species are increasingly being used. Accordingly,studying the spread and population dynamics of strains isolating from clinical and environment, from a single host to large scale ecosystems is definitely needed. In the current study, we carried out a genetic analysis of nine microsatellite loci in isolates from different regions of Iran to compare and explore the genetic diversity between environmental and clinical A. fumigatus strains. Materials and Methods: Sixty-six clinical (n=43) and environmental (n= 23) isolates of A. fumigatus, have collected from six cities of Iran. All A. fumigatus isolates identified based on macroscopic and microscopic characters, the ability to grow at above 45°C, and confirmed using DNA sequencing of the partial b-tubulin gene.Sixty-six A. fumigatus isolates were subjected by microsatellite typing using three separate multiplex PCRs with a panel of nine short tandem repeats (STR) to evaluate the genetic relatedness. Results: The STR typing of 66 A. fumigatus isolates revealed 38 distinct genotypes distributed among environmental and clinical isolates. We identified 12 clones including 40 different isolates representing 60% of all isolates tested, which each clone included 2–7 isolates. Conclusion: The STR typing is considered as a valuable tool with excellent discriminatory power to study the molecular epidemiology and genotypic diversity of A.fumigatus isolates. These findings show that the high genetic diversity observed of Iranian A. fumigatus isolates with those outside Iran and formed a separate cluster.

Background and Purpose: Candida auris, as a new characterized pathogenic yeast,has attracted remarkable attention in the recent decade due to its rapid global emergence and multidrug resistance traits. This unique species is able to cause nosocomial outbreaks and tolerate adverse conditions; however, it has been mostly misidentified by conventional methods. Case report: This report aimed to describe the first fluconazole-resistant case of C.auris otitis in an immunocompetent patient in Iran. The isolate showed minimum inhibitory concentration of ≥ 32 μg/ml for fluconazole; however, the patient was treated with topical clotrimazole and miconazole with no recurrence. Conclusion: This was the second strain of C. auris isolated from otitis in Iran which was fluconazole-resistant, unlike the first Iranian isolate.

Several prolonged and significant outbreaks of dermatophytosis caused by Trichophyton indotineae, a new emerging terbinafine-resistant species, have been ongoing in India in recent years, and have since spread to various countries outside Asia. Miltefosine, an alkylphosphocholine, is the most recently approved drug for the treatment of both visceral and cutaneous leishmaniasis. Miltefosine in vitro activity against terbinafine-resistant and susceptible T. mentagrophytes/T. interdigitale species complex, including T. indotineae, is limited. The current study aimed to assess miltefosine’s in vitro activity against dermatophyte isolates, which are the most common causes of dermatophytosis. Miltefosine, terbinafine, butenafine, tolnaftate, and itraconazole susceptibility testing was performed using Clinical and Laboratory Standards Institute broth microdilution methods (CLSI M38-A3) against 40 terbinafine-resistant T. indotineae isolates and 40 terbinafine-susceptible T. mentagrophytes/T. interdigitale species complex isolates. Miltefosine had MIC ranges of 0.063–0.5 µg/mL and 0.125–0.25 µg/mL against both terbinafine-resistant and susceptible isolates. In terbinafine-resistant isolates, the MIC50 and MIC90 were 0.125 µg/mL and 0.25 µg/mL, respectively, and 0.25 µg/mL in susceptible isolates. Miltefosine had statistically significant differences in MIC results when compared to other antifungal agents (p-value 0.05) in terbinafine-resistant strains. Accordingly, the findings suggest that miltefosine has a potential activity for treating infections caused by terbinafine-resistant T. indotineae. However, further studies are needed to determine how well this in vitro activity translates into in vivo efficacy.

Background and Purpose:Cryptococcalmeningitis (CM) is a serious fungal infection that especially affectspatients with human immunodeficiency virus (HIV). In this regard,the presentretrospective study aimed to analyze the clinical and laboratory features and therapeutic outcomes of patients with CM admitted totwo teaching referral centersin the north of Iran during 2011-19. Materials and Methods:This studywas performed onall the hospitalized patients diagnosedwith CM in two therapeutic centers of infectious diseases in the north of Iran. The required data,such as demographic characteristicsandclinical and paraclinical featuresof patients,were extracted and entered in the information forms. Finally, the collected data were analyzed inSPSSsoftware(version16). Results:For the purpose of the study, records of 12confirmed CM patients were evaluatedin this research. Based onthe results,75% ofthepatients were male.Moreover,the average age of the subjectswas 40.33± 8.93 years oldand 66.6%ofthem(n=8) were HIV-positive. Other underlying diseases among HIV-positive patients includedinfection with hepatitis C virus (25%) and a history of tuberculosis (25%). In total, threeHIV-negative patients suffered from Hodgkin lymphoma (25%), sarcoidosis (25%),and asthma (25%) and one patient (25%) had no underlying disease. Headache (75%), weakness,and fatigue (75%) were the most common symptomsamong the participants. The cluster of differentiation 4count in all HIV-positive patients was less than 100 cells/μl. There was no significant difference between symptoms in HIV-positive and HIV-negative patients. Besides, no significant differencewas observedbetween the groups of HIV-positive and HIV-negative patientsregardingthe period betweentheonset of symptoms and diagnosisof CM,the length of hospital stay,and the duration of antifungal medicationconsumption. In total,threepatients (25%) expired,and six patients recovered. The CM recurred in two HIV-negative and oneHIV-positivesubjects;the two HIV-negativepatientswere treated,whilethe HIV-positivepatientexpired due to thisrecurrence. Conclusion:Clinical features and cerebrospinal fluid parameters were not different in HIV-positive and HIV-negative participants. Despite the fact thatCM is not commoninIran, due to the increasing number of immunosuppressive patients, the differential diagnosis of CM should be considered forpatients with signs and symptoms of infection in the central nervous system.

The world is involved with a pandemic coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2. The clinical manifestations of reported COVID-19-associated pulmonary impairments range from asymptomatic infections to a pneumonia-induced acute respiratory distress syndrome that requires mechanical ventilation. Fungal superinfections complicating the clinical course remain underexplored. Angiotensin-converting enzyme 2, the receptor for COVID-19 that is mainly expressed in airway epithelia and lung parenchyma, is considered an important regulator of innate immunity. With regard to the viral-cell interaction, imbalanced immune regulation between protective and altered responses caused by the exacerbation of inflammatory responses should be considered a major contributor to secondary pulmonary aspergillosis. In addition, the complex inherited factors, age-related changes, and lifestyle may also affect immune responses. The complication and persistence of invasive aspergillosis have been well described in patients with severe influenza or COVID-19. However, there is a scarcity of information about the immunological mechanisms predisposing patients with COVID-19 to fungal co-infections. Therefore, this study was conducted to investigate the aforementioned domain.

Background and Purpose: The present study aimed to evaluate the effect of cyproconazole, the most used fungicide in Iranian wheat farms, on the induction of voriconazole resistance in Aspergillus fumigatus isolates. Materials and Methods: A collection of 20 clinical and environmental isolates were selected for investigation of the in vitro activity of fungicides. The minimum inhibitory concentrations (MICs) were determined by the documented broth microdilution method M38-A2 (CLSI, 2008). Induction experiments were performed and the possibly induced isolate(s) were subjected to antifungal susceptibility testing, sequencing of the CYP51A promoter, and full coding gene. Furthermore, CYP51-protein homology modeling and docking modes were evaluated using SWISS-MODEL (https://swissmodel.expasy.org/) and SEESAR software (version 9.1). Results: Among 10 susceptible isolates, only one strain showed a high MIC value against voriconazole (MIC=4μg/ml) after 25 passages. Nevertheless, sequencing of the CYP51A promoter and full coding gene did not reveal any mutations. Cyproconazole, which has three nitrogen atoms in the aromatic ring, coordinated to the iron atom of heme through a hydrogen bond contact to residue Lys147 present in the active site of the A. fumigates Cyp51 homology model. Conclusion: Cyproconazole is being applied extensively in wheat farms in Iran. According to the results, cyproconazole may not play a key role in the induction of azole resistance in the isolates through the environmental route. However, the potential ability of the fungicide to induce medically triazole-resistant strains over a long period of application should not be neglected.

Objectives: The rate of resistance of Candida parapsilosis to echinocandins remains unexplored in Iran. The main aims of this study were to investigate the susceptibility patterns and possible mechanisms of echinocandin resistance in echinocandin-resistant clinical C. parapsilosis isolates in Iran. Methods: A total of 105 isolates of C. parapsilosis sensu stricto underwent antifungal susceptibility testing to echinocandins by the broth microdilution reference method. Sequences of the CpERG3 and CpFKS1 genes were analysed using MEGA6 software, and alterations in CHS3, FKS1 and Rho gene expression were evaluated by quantitative reverse transcription (RT-qPCR). REST® software was used to analyse the results. Results: The rate of echinocandin cross-resistance was 2.9% (3/105). No substitutions were detected in Fks1p except for the naturally occurring P660A amino acid substitution observed in isolates both with high and low minimum inhibitory concentrations (MICs). Moreover, the G111R amino acid substitution was not found in Erg3p. Following echinocandin exposure, expression of Rho and FKS1 genes was significantly increased in resistant isolates, whilst the CHS3 gene showed no change. Conclusion: Alterations in the expression of some key genes may be responsible for echinocandin resistance among C. parapsilosis isolates. Understanding the mechanisms responsible for drug resistance in C. parapsilosis is not only crucial for the development of new antifungals but is also important in choosing appropriate antifungals for patient treatment at the earliest stage.

Background and purpose: It is estimated that 70–75% of women experience one episode of vulvovaginal candidiasis (VVC) during childbearing age and 40-50% suffer from recurrent VVC. Because of the resistance of Candida species to certain drugs, such as azoles, physicians tend to use complementary or alternative medicine such as vinegar in treatment of VVC. The current study aimed to determine the therapeutic effects of acetic acid vaginal gel on VVC. Materials and methods: A double-blind randomized clinical trial was done in women (n=41) with VVC in Sari, Iran. The patients were allocated into two treatment groups to receive either 2% acetic acid vaginal gel + 2% acetic acid intravaginal wash + clotrimazole vaginal cream (n=20) or 2% acetic acid intravaginal wash + clotrimazole vaginal cream (n=21) for 10 days. After treatment, clinical symptoms and mycological cure rate were assessed by culture of vaginal secretions. All yeast isolates were identified by molecular method. Results: Before the intervention, there were no significant differences in signs and symptoms (P= 0.703), fungal burden rate (low, medium, heavy) (P= 0.092), and causative Candida species (P= 0.188) between the two groups. After treatment, an improvement in symptoms was seen in case group but the difference was not significant (P =0.3512). A significant difference was found between the two groups in mycological cure rate (P= 0.0377). Candida albicans were the most common species causing the disease. Conclusion: The study showed that acetic acid vaginal gel could have a significant effect on the definitive cure of VVC. (Clinical Trials Registry Number: IRCT20190309042978N3)

Background and Purpose: The aim of the current study was to investigate the epidemiology of vulvovaginal candidiasis (VVC) and recurrent VVC (RVVC), as well as the antifungal susceptibility patterns of Candida species isolates. Materials and Methods: A cross-sectional study was carried out on 260 women suspected of VVC from February 2017 to January 2018. In order to identify Candida species isolated from the genital tracts, the isolates were subjected to polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) using enzymes Msp I and sequencing. Moreover, antifungal susceptibility testing was performed according to the Clinical and Laboratory Standards Institute guidelines (M27-A3). Results: Out of 250 subjects, 75 (28.8%) patients were affected by VVC, out of whom 15 (20%) cases had RVVC. Among the Candida species, C. albicans was the most common species (42/95; 44.21%), followed by C. lusitaniae (18/95; 18.95%), C. parapsilosis (13/95; 13.69%), C. glabrata (8/95; 8.42%), C. kefyr (6/95; 6.31%), C. famata (5/95; 5.26%), C. africana (2/95; 2.11%), and C. orthopsilosis (1/95; 1.05%), respectively. Multiple Candida species were observed in 28% (21/75) of the patients. Nystatin showed the narrowest range of minimum inhibitory concentration (MIC) (0.25-16 μg/ml) against all Candida strains, whereas fluconazole (0.063-64 μg/ml) demonstrated the widest MIC range. In the current study, C. lusitaniae, as the second most common causative agent of VVC, was susceptible to all antifungal agents. Furthermore, 61.1% of C. lusitaniae isolates were inhibited at a concentration of ≤ 2 μg/ml, while38.9% (n=7)of them exhibited fluconazole MICs above the epidemiologic cutoff values (ECV). Candida species showed the highest overall resistance against fluconazole (61.3%), followed by itraconazole (45.2%) and caspofungin (23.7%). All of C. albicans strains were resistant to itraconazole with a MIC value of ≥ 1 μg/ml; in addition, 87.5% of them were resistant to fluconazole. Moreover, 100% and 87.5% of C. glabrata strains were resistant to caspofungin and fluconazole, respectively. Conclusion: As the findings revealed, the majority of VVC cases were caused by non-albicans Candida species which were often more resistant to antifungal agents. Candida lusitaniae generally had fluconazole MICs above the ECV. Given the propensity of C. lusitaniae to develop resistance under drug pressure, antifungals should be administered with caution. The emergence of these species justify the epidemiological surveillance surveys to watch out the distribution of yeast species.

Background and Purpose: Incidence of fungal infections caused by opportunistic fungal pathogens, such as yeasts and yeast-like species, has undergone an increase in otherwise healthy individuals. These pathogens account for high mortality and show reduced susceptibility to the routine antifungal drugs. Accordingly, antifungal susceptibility testing is an urgent need in the determination of the susceptibility spectrum of antifungals and selection of appropriate antifungal agents for the management of patients with fungal infection. Materials and Methods: The present study was conducted on 110 yeast strains belonging to 15 species recovered from clinical specimens. Susceptibility of the isolates to four antifungal drugs (i.e., fluconazole, itraconazole, voriconazole, and posaconazole) was tested according to the Clinical and Laboratory Standards Institute guidelines M27-A3 and M27-S4. Results: Fluconazole exhibited no activity against 4.3% (n=2) of C. albicans isolates, whereas the remaining 44 isolates had a minimum inhibitory concentration (MIC) range of 0.125-4 μg/ml. Voriconazole had the lowest geometric mean MIC (0.03 μg/ml) against all isolated yeast species, followed by posaconazole (0.07 μg/ml), itraconazole (0.10 μg/ml), and fluconazole (0.60 μg/ml). Overall, all of the isolates had reduced voriconazole MICs with a MIC range of 0.016-0.5 μg/ml, except for one isolate of C. albicans that had a MIC of 1 μg/ml. Candida haemulonii as a multidrug-resistant fungus showed a fluconazole MIC of > 64 μg/ml. Conclusion: The current study provides insight into the antifungal susceptibility profiles of clinically common and uncommon yeast species to four triazole antifungal agents. According to our findings, voriconazole was the most active agent. Awareness about antifungal susceptibility patterns is highly helpful in the selection of appropriate antifungal drugs and identification of the efficiency of the currently used agents.

Background and Purpose: Emergence and development of antifungal drug resistance in Candida species constitute a serious concern. Candida auris as an emerging multidrug-resistant fungus is the most important public health threat with high levels of mortality and morbidity. Almost all C. auris isolates are resistant to fluconazole, and there have been reports of elevated minimum inhibitory concentrations (MICs) to amphotericin B and echinocandins. To overcome the growing challenge of antifungal resistance, a valuable alternative option would be the use of drug combination. Materials and Methods: The present study evaluated the in vitro combination of nonsteroidal anti-inflammatory drugs (NSAIDs), such as ibuprofen, diclofenac and aspirin with fluconazole against fluconazole-resistant C. auris in comparison to other fluconazole-resistant Candida species, including C. albicans, C. glabrata, C. parapsilosis, C. tropicalis, and C. krusei originating from patients with candidiasis. Results: The MIC ranges of fluconazole-ibuprofen and fluconazole-diclofenac decreased from 32-256 to 32-128 and 16-256 μg/ml, respectively and remained the same for fluconazole-aspirin against C. auris. However, the combination of fluconazole with ibuprofen resulted in a synergistic effect for 5 strains, including C. albicans (n=2), C. tropicalis (n=1), C. glabrata (n=1), and C. krusei (n=1), by decreasing the MIC of fluconazole by 2-3 log2 dilutions. Conclusion: Although the interaction of NSAIDs with fluconazole was not synergistic against fluconazole-resistant C. auris isolates, no antagonism was observed for any combinations. Therefore, combination with newer azole agents needs to be conducted.

Background and Purpose: Pneumocystis jirovecii colonization plays a key role in the progression of pulmonary infection. However, there are limited data regarding the colonization of these fungi in the patients residing in different regions of Iran. Regarding this, the present study was conducted to evaluate the prevalence of P. jirovecii colonization in non-HIV-infected patients with respiratory failure introduced by physicians using nested polymerase chain reaction (PCR). Materials and Methods: This study was conducted on 136 samples obtained from 136 patients with respiratory disorders referring to different hospitals in the capital and north of Iran during 2013-2015. The samples were collected using bronchoalveolar lavage (BAL; n=121) and sputum induction (n=15). Nested PCR method targeting mtLSU rRNA gene was used for the detection of P. jirovecii DNA in the specimens. Results: The nested PCR analysis resulted in the detection of P. jirovecii DNA in 32 (23.5%) patients. The mean age of the participants was 49.04±11.94 years (age range: 14-90 years). The results revealed no correlation between Pneumocystis colonization and gender. The studied patients were divided into two groups of immunocompromised and immunocompetent patients. In the regard, 25.4% of the patients with detectable P. jirovecii DNA were immunocompromised and had cancer, organ transplantation, asthma, sarcoidosis, dermatomyositis, chronic obstructive pulmonary disease, bronchiectasis, and pulmonary vasculitis. On the other hand, Pneumocystis DNA was detected in 21.8% of the immunocompetent patients. Frequencies of P. jirovecii DNA detection in the patients with tuberculosis, hydatid cyst, and unknown underlying diseases were obtained as 20.8%, 25%, and 22%, respectively. The prevalence of Pneumocystis colonization varied based on age. In this regard, P. jirovecii colonization was more prevalent in patients aged above 70 years. Conclusion: As the findings indicated, non-HIV-infected patients, especially the elderly, had a high prevalence of P. jirovecii colonization. Therefore, these patients are probably a potential source of infection for others. Regarding this, it is of paramount importance to adopt monitoring and prophylactic measures to reduce this infection.

Background and Purpose: Although the mechanism of action for echinocandins is known, the physiological mechanisms by which these antifungal agents cause cell death via the classical apoptotic pathways are not well-defined yet. Regarding this, the present study aimed to evaluate the mechanisms of caspofungin-induced Candida glabrata cell death. Materials and Methods: For the purpose of the study, the minimum inhibitory concentration (MIC) of caspofungin against C. glabrata (ATCC 90030) was determined using the broth microdilution reference method (CLSI M27-A2 and M27-S4). The annexin V and propidium iodide staining was performed to determine the way through which caspofungin acts against C. glabrata (i.e., through the induction of apoptosis and/or necrosis). Additionally, the possible effect of caspofungin on inducing the expression of two apoptotic genes, namely MCA1 and NUC, was studied using the real-time polymerase chain reaction assay. Results: According to the obtained MIC value (0.5 μg/mL), C. glabrata, exposed to 0.25, 0.5, and 1 μg/mL of caspofungin, exhibited the features of late apoptosis/necrosis after 18 h of incubation. Furthermore, the use of 0.25, 0.5, and 1 μg/ml caspofungin induced apoptosis (early/late) in 14.67%, 17.04%, and 15.89% of the cells, respectively. The results showed a significant difference between the percentages of early-apoptotic cells at the three concentrations (p Conclusion: As the findings of the present study indicated, caspofungin promoted both necrosis and apoptosis of C. glabrata cells at concentrations higher than or equal to the MIC value.

Background and Purpose: Rhinocerebral mucormycosis is a rare fatal fungal infection which is on a growing trend, particularly among immunocompromised patients. Immunosuppressive drugs, including corticosteroids and antimetabolites, increase the risk of this infection. Herein, we reported the case of fulminant rhinocerebral mucormycosis in a patient with ulcerative colitis receiving azathioprine and corticosteroid. Case report: A 58-year-old woman was admitted to the hospital in a state of coma with an extensive necrosis in her nose. She was afflicted with intestinal bleeding after 1 month of fasting and was treated with azathioprine and a high dose of prednisolone for ulcerative colitis 2 months prior to hospital admission. The direct microscopic examination of the necrotic tissues of the paranasal sinuses showed several non-septate hyphae consistent with Mucorales. Culture media yielded Rhizopus species, which was identified as Rhizopus oryzae by internal transcribed spacer polymerase chain reaction sequencing. Despite the implementation of surgical and pharmaceutical (liposomal amphotericin B) treatments, the patient expired after 2 weeks of admission. Conclusion: The gastroenterologists should be aware of the adverse effect of immunosuppressive drugs they prescribe for the treatment of inflammatory bowel disease.

Background and Purpose: Candidemia is a life-threatening fungal infection with significant mortality and morbidity in neutropenic individuals, immunosuppressive chemotherapy recipients, and broad-spectrum antibiotics consumers. The epidemiology and antifungal susceptibility testing of non-albicans Candida species have been poorly studied. These species are characterized by low susceptibility to azoles and echinocandins. Herein, we report the first pediatric case of candidemia due to C. guilliermondii in Iran and review the literature on fungemia caused by C. guilliermondii. Case report: We presented the first candidemia case due to Candida guilliermondii in a 4-month-old male infant with neuroblastoma in Iran. This study also involves a comprehensive literature review on fungemia caused by C. guilliermondii during a period of 18 years (i.e., 2000-2018) to discuss the epidemiology, clinical features, and treatment of this disease. The literature review resulted in the identification of 501 cases of candidemia caused by C. guilliermondii. Most of the patients were adults and had multiple risk factors. However, the main risk factors were significantly related to cancer chemotherapy, followed by central venous catheter use and Intensive Care Unit admission. Mortality rate due to this disease had a range of 3.4-66.6%; in this regard, the patients with cancer had the highest mortality rate. Conclusion: Given the high mortality of candidemia, the early diagnosis of this infection and timely initiation of antifungal therapy significantly improve the patients’ survival rate and result in better outcomes. Consequently, it is highly recommended to monitor the local epidemiology of this life-threatening infection and raise awareness in this regard.

Fusarium species are filamentous fungi that cause a variety of infections in humans. Because they are commonly resistant to many antifungal drugs currently available in clinical settings, research into alternative targets in fungal cells and therapeutic approaches is required. The antifungal activity of miltefosine and four comparators, amphotericin B, voriconazole, itraconazole, and caspofungin, were tested in vitro against a collection of susceptible and resistant clinical (n = 68) and environmental (n = 42) Fusarium isolates. Amphotericin B (0.8 μg/mL) had the lowest geometric mean (GM) MICs/MECs values followed by miltefosine (1.44 μg/mL), voriconazole (2.15 μg/mL), caspofungin (7.23 μg/mL), and itraconazole (14.19 μg/mL). Miltefosine was the most effective agent against Fusarium isolates after amphotericin B indicating that miltefosine has the potential to be studied as a novel treatment for Fusarium infections.

Background and Purpose: Candida endocarditis is an infrequent disease with a high mortality rate, which commonly occurs in immunosuppressed patients with cardiac valve replacement. We reported a 70-year-old woman diagnosed with Candida prosthetic valve endocarditis (PVE). This study also involved a review of all published cases of Candida PVE from 1970. Case report: Herein, we reported a 70-year-old woman with the history of severe mitral stenosis and myelodysplasia syndrome. She underwent mitral valve replacement for two times. The blood cultures were positive, and phenotypic identification of the isolates at the species level was performed based on microscopic and macroscopic characteristics. In the second prosthetic valve replacement, huge fungal white and creamy vegetation was observed which was identified as Candida albicans based on the conventional and molecular methods. Despite the administration of antifungal treatments, the patient passed away probably due to the multidrug-resistant Candida PVE. Conclusion: As PVE is a late consequence of prosthetic valve replacement, extended follow-up visits, early diagnosis, repeating valve replacement surgeries, and timely selective antifungal treatments are warranted. Keywords: Amphotericin B, Antifungal resistant, Azoles, Candida endocarditis, Multi-drug resistant, Myelodysplasia syndrome, Prosthetic valve replacement

Cryptococcosis is a serious fungal infection in both huaman and animals caused by members of the Cryptococcus neoformans/ Cryptococcus gattii species complex. The main route of entry is respiratory tract. Meningitis is the most common clinical manifestation of cryptococcosis in immunocompromised patients and considered as one of AIDS defining illnesses. The known natural sources of the fungus are avian (pigeon, chicken, and other birds) excreta, dead plants and trees like Eucalyptus, Pine, and Fig species. Cryptococcus neoformans/ Cryptococcus gattii species complex have well defined virulence factors that help them to survive and multiply in macrophages and monocytes, escape into blood and disseminate throughut the body. Laboratory diagnosis of cryptococcosis is based on direct microscopy examination, fungal isolation, serology and molecular analysis of serum and cerebrospinal fluid. Amphhotercin B combined with other antifungals including 5-flourocytosisne and fluconazole monotherapy are used for treatment of cryptococcosis. Treatment in HIV infected patients consist of three phases including induction, consolidation and maintenance therapy. The preferred therapeutic regimen is combination therapy of antifungal and highly active antiretroviral therapy. In this article, we summarized recent studies on pathogenesis, diagnosis, and treatment of cryptococcosis in HIV infected patients.

Background and Purpose: Catheter-related blood circulation infection is the most dangerous and serious side-effects of vascular catheters, which leads to the enhancement of the costs, mortality, and hospital stay duration, especially in the Intensive Care Unit. Regarding this, the aim of the current study was to identify the prevalence of catheter-induced candidemia in the Tehran Heart Center, a heart hospital in Tehran, Iran. Materials and Methods: This study was conducted on patients admitted to Tehran Heart Center for a minimum of 7 days during 18 months. To detect the fungal elements, blood culture and catheter culture were performed in the patients receiving central or peripheral venous catheter. Then, the polymerase chain reaction (PCR) was applied to determine the possible diagnosis. Results: The investigation of 223 samples led to the identification of a total of 15 (6.7%) yeast isolates obtained from 9 (60%), 4 (26.6 %), and 2 (13.4%) catheter, blood, and skin (of the catheter insertion areas) cultures, respectively. Out of nine Candida isolates obtained from the catheter samples, 1 (11.1%), 1 (11.1%), 2 (22.2%), and 5 (55.6%) cases were identified as C. tropicalis, C. membranifaciens, C. glabrata, and C. albicans, respectively, using the internal transcribed spacer region sequencing. Furthermore, the four yeasts isolated from the blood culture included C. tropicalis, C. carpophila, C. membranifaciens, and Cryptococcus albidus. Additionally, one case of C. glabrata and one case of C. albicans were isolated from the skin culture of the catheter insertion areas in patients with positive catheter culture. We reported two cases of catheter-related candidemia caused by C. membranifaciens and C. tropicalis on the basis of the genetic similarity of the species isolated from the blood and catheter. These cases were treated successfully with intravenous fluconazole and catheter removal. Conclusion: There is some evidence indicating the growing prevalence of non-albicans Candida infections. Many risk factors, including prior antibiotic therapy, use of a central venous catheter, surgery, and parenteral nutrition, are considered to be associated with candidemia in hospitalized heart failure patients. The identification of the route of infection in candidemia is difficult. In the current study, the positive blood and catheter cultures for Candida isolates and the similarity of the ITS region of ribosomal DNA sequence of Candida isolated from two patients confirmed the diagnosis of intravenous catheter-related candidemia.

In this research a novel and economic optical sensor for determination of benzaldehyde (BA), which is a dangerous oxidation product of benzyl alchol in injection pharmaceuticals, is introduced. The optode is based on immobilization of 2, 4- dinitrophenylhydrazine in a PVC membrane. The response of the proposed optode was based on the nucleophilic addition of 2, 4- dinitrophenylhydrazine to BA that cause an increase in absorbance at 366 nm. Different variables affect the optical signal such as pH of solution, compositions of the membrane components and type of plasticizer were optimized. A universal buffer solution with pH 2 and dibutylphthalate were used as optimum measurement media and plasticizer, respectively. Under the optimum conditions, the optode has a response time of 6 min, linear concentration range of 4.0 − 100.0 ppm of BA with a detection limit of 0.6 ppm. The influence of interfering compounds possibly found in injection formulations on the BA determination was studied. The results showed that the prepared optical sensor has good selectivity to BA. The proposed optode has a reversible response and good repeatability. This optode was successfully applied to assay benzaldehyde in sodium diclofenac and piroxicam ampoules.

Background and Purpose: Patulin is a mycotoxin produced by some molds,especially Aspergillus and Penicilium, and is responsible for mycotoxicosis in animals and humans.There is still not very detailed data about the anti-cancer potency of patulin, but some reports demonstrated that it induces cellular apoptosis and toxicity. Materials and Methods: To determine the efficacy of patulin as a therapeutic strategy for cervical and colorectal cancers, we investigated its effects on HeLa,SW-48, and MRC-5 cell lines. Cell lines were exposed to various concentrations of patulin (i.e., 0.5, 1, 2, and 4 µM), then using methyl thiazolyl tetrazolium (MTT) and bromo-2′-deoxyuridine (BrdU) assays, the rates of apoptosis and cell viability were determined. Results: The obtained results showed a significant reduction in cell viability and apoptosis induction in a dose-dependent manner. Among all the cell lines, the highest growth inhibition rate was obtained at the 4 μM concentration of patulin.Conclusion: Our results suggested that patulin could significantly decrease tumor growth in human cervical and colorectal cancer models.

Background and Purpose: Rhino-orbito-cerebral mucormycosis (ROCM) is a rare disease with acute and fulminant manifestation. This infection is associated with high morbidity and mortality rates. Herein, we reviewed the manifestations, underlying conditions, medical treatments, and surgical interventions in ROCM patients admitted to a tertiary referral center in northern Iran over a seven-year period. Materials and Methods: In a retrospective analysis, 15 cases of ROCM were identified from 2007 to 2013 in Bu Ali Sina Hospital, Sari, Iran. All the ROCM cases were clinically diagnosed and confirmed by histopathological and/or mycological examination. The relevant demographic data, clinical, ophthalmic, and neurologic manifestations, underlying conditions, medical treatments, and surgical interventions were recorded and analyzed. Results: The mean age of the patients was 54±11 years (age range: 28–70 years); 26.7% of the patients were male and 73.3% female (male: female ratio of 1: 2.7). Uncontrolled diabetes was noted in at least 86.7% (13/15) of the cases. The maxillary sinuses were the most frequently involved sites (66.7% of the cases) followed by the ethmoid sinus. Amphotericin B in combination with surgical debridement was used in the treatment of 80% of the cases. Furthermore,73.3% of the patients who were diagnosed early and underwent medical and extensive surgical debridement of the infected tissues survived. Conclusion: Uncontrolled diabetes mellitus is considered to be the main predisposing factor for ROCM. To prevent and reduce mortality rate of this acute disease, early diagnosis based on clinical findings and biopsy is recommended.

Background and Purpose: Hyphal wall protein 1 (HWP1) is an important adhesin which usually is expressed on the germ tube and hyphal surface produced by different Candida species. The hyphal wall protein-coding gene (HWP1) was evaluated as a novel identification and phylogenetic marker in Candida tropicalis, C. orthopsilosis, C. parapsilosis and C. glabrata. Materials and Methods: Initially, four specific primer pairs were designed, and the target was amplified and finally sequenced. A total of 77 Candida isolates from four different species were included in the study. Consensus sequences were used for the evaluation of phylogenetic tree using the CLC Genome Workbench, GENEIOUS, and MEGA softwares and the levels of nucleotide and amino acid polymorphism were assessed. Results: According to the results, the specific amplified fragments of HWP1 gene were useful for the differentiation of four species. Intra-species variation was observed only in C. tropicalis with two DNA types. The phylogenetic tree of Candida species based on the HWP1 gene showed consistency in topology with those inferred from other gene sequences. Conclusion: We found that HWP1 gene was an excellent marker for the identification of non-albicans Candida species as well as the phylogenetic analysis of the most clinically significant Candida species.

Background and Purpose: Candida albicans is the most common causative agent of candidiasis. Candidiasis management is dependent on the immune status of the host, severity of disease, and the choice of antifungal drug. Antifungals, specifically triazoles, are widely administered for the treatment of invasive fungal infections. Herein, we aimed to evaluate the in vitro susceptibility of C. albicans isolates to fluconazole (FLZ), itraconazole (ITZ), voriconazole (VRZ), amphotericin B (AMB), and Caspofungin (CAS). Materials and Methods: A total of 44 clinical strains of C. albicans were collected from 36 patients admitted to four hospitals in Mazandaran Province, Iran. The in vitro antifungal susceptibility testing was performed based on the Clinical and Laboratory Standards Institute methods. Results: Generally, 34 isolates were susceptible to all the five antifungal drugs, while four isolates were susceptible or susceptible dose-dependent (SDD) and six isolates were SDD or resistant to these antifungals. The lowest minimum inhibitory concentration (MIC; 0.016 μg/ml) belonged to AMB and the highest MIC was for FLZ (16 μg/ml). The lowest MIC50 (0.063 μg/ml) was related to ITZ and the lowest MIC90 (0.25 μg/ml) pertained to CAS; in addition, the highest MIC50 (1 μg/ml) and MIC90 (4 μg/ml) were for FLZ. Four of the isolates showed resistance to both FLZ and VRZ, separately, and five isolates were resistant to ITZ. Caspofungin showed potent activity against more than 95% of the C. albicans isolates. Conclusion: Overall, we reported 9.1% resistance to FLZ and VRZ, 11.3% resistance to ITZ and AMB, and 4.6% resistance to caspofungin. Our finding is in agreement with previous observations proposing that C. albicans isolates develop resistance to some antifungal drugs such as FLZ since they are widely used as prophylaxis.

Background and Purpose: Fungal keratitis is a suppurative, ulcerative, and sight-threatening infection of the cornea that sometimes leads to blindness. The aims of this study were: recuperating facilities for laboratory diagnosis, determining the causative microorganisms, and comparing conventional laboratory diagnostic tools and semi-nested PCR. Materials and Methods: Sampling was conducted in patients with suspected fungal keratitis. Two corneal scrapings specimens, one for direct smear and culture and the other for semi- nested PCR were obtained. Results: Of the 40 expected cases of mycotic keratitis, calcofluor white staining showed positivity in 25%, culture in 17.5%, KOH in 10%, and semi-nested PCR in 27.5%. The sensitivities of semi-nested PCR, KOH, and CFW were 57.1%, 28.5%, and 42% while the specificities were 78.7%, 94%, and 78.7%, respectively. The time taken for PCR assay was 4 to 8 hours, whereas positive fungal cultures took at least 5 to 7 days. Conclusion: Due to the increasing incidence of fungal infections in people with weakened immune systems, uninformed using of topical corticosteroids and improper use of contact lens, fast diagnosis and accurate treatment of keratomycosis seems to be essential. Therefore, according to the current study, molecular methods can detect mycotic keratitis early and correctly leading to appropriate treatment.

Background and Purpose: Candida species are the most common organisms involved in superficial fungal infections, worldwide. Although econazole is among the most frequently used topical formulations for the treatment of candidiasis, no information is available regarding the susceptibility profiles of Candida species in Iran. Materials and Methods: In vitro susceptibility of 100 clinical Candida isolates belonging to 6 species from superficial candidiasis of Iran towards to econazole was compared with three other common antifungal agents including itraconazole, fluconazole, and miconazole. Minimum inhibitory concentrations (MICs) values were analyzed according to the Clinical and Laboratory Standards Institute (CLSI) M38-A3 document. All isolates were previously identified to the species level, using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) on ITS region. Results: The MIC of econazole, itraconazole, miconazole, and fluconazole were within the range of 0.016-16, 0.032-16, 0.016-16, and 0.25-64 g/ml, respectively. In general, econazole and miconazole were more active against Candida isolates, compared to the other two agents. Conclusion: The present study demonstrated that for Candida albicans isolates, miconazole and econazole had the best effect, but in non-albicans Candida species, itraconazole and miconazole displayed more activity than other antifungal agents.

Background and Purpose: Invasive candidiasis (IC) is a significant cause of morbidity and mortality in patients with hematologic disorders and bone marrow transplant recipients. Rapid, specific and sensitive test for the timely accuracy in immunocompromised patients to reduce mortality rates and prevent IC progress is necessary. We established a real-time PCR assay on blood for the diagnosis and differentiation of the causative Candida species. Materials and Methods: Whole blood samples were collected twice, from 72 patients for Real Time PCR and blood culture assays. The primers and hybridization probes were designed to potentiate the specific sequence of 18S rRNA genes using Light Cycler system and Fluorescence Resonance Energy Transfer (FERT). The patients with hematologic malignancies and bone marrow transplant recipients were evaluated for IC based on the revised European Organization for Research and Treatment of Cancer/ Mycoses Study Group (EORTC/MSG) criteria. Results: From 2009 to 2011, 72 patients with hematologic malignancies and bone marrow transplant recipients were evaluated for IC. The female to male ratio was 27:45; the mean age was 32.1 years. The most common malignancy in this patient was acute myeloid leukemia (AML) (27.8%) and acute lymphoblastic leukemia (ALL) (26.4%). Out of 72 patients, 11 patients (15.3%) had positive real time PCR /probe results. Based on the melting temperature (Tm) analysis, 5 (45.4%) C. krusei, 3 (27.2%) C. tropicalis, 2 (18.1%) C. parapsilosis and 1 C. albicans (9%) were identified. According to the revised EORTC / MSG, 1 patient (9%) and 10 patients (91%) were defined as proven and possible groups of IC, respectively. The mortality rate in proven and possible IC patient was found 54.5%. Conclusion: The established Real-time PCR/FRET probe assay is an appropriate diagnostic tool for the detection of Candida species DNA and the management of patients suffering from hematologic malignancies and bone marrow recipient are at risk for IC.

We report an unusual case of co-infection of invasive pulmonary aspergillosis (IPA) and fusarial skin infection in a patient with classic pyoderma gangrenosum with unknown causes, which were previously controlled with oral prednisolone, cyclosporine. The diagnosis was made on direct microscopy and culture of endobronchial washing, bronchoalveolar lavage and skin lesion biopsy. The treatment failed, and the patient expired 12 days following hospitalization. This report highlights the rarity of coexistence of IPA and a chronic fusarial skin infection and thereby reinforcing the physician′s attention toward the possibility of invasive fungal infection in the immunosuppressed patients.

Background: Candida albicans (C. albicans) is a major cause of candidaemia in people with impaired immunity. Blood culture is a “gold standard” for candidaemia detection but is time-consuming and relatively insensitive. We established a real-time PCR assay for C. albicans detection in blood by LightCycler PCR and melting curve analysis. Methods: Five milliliter blood samples from healthy volunteers were spiked with 100-106 C. albicans cells to determine the detection limit of our method. DNA was extracted from whole blood using glass beads and the QIAamp DNA Blood Mini Kit (Qiagen, Hilden Germany). DNA from C. albicans isolates were amplified with primers and inserted into Escherichia coli (E. coli) DH5α.1 cells with the TA cloning vector (Invitrogen). The plasmid was used for standardization and optimization. A quantitative PCR assay with the LightCycler amplification and detection system based on fluorescence resonance energy transfer (FRET) with two different specific probes was established. To assess the precision and reproducibility of real-time PCR the intra-assay precision was determined in six consecutive assays. Results: No cross-reactivity of the hybridization probes with the DNA of non-C. albicans species or human genomic DNA was observed, which confirmed its 100% specificity. The minimum limit detected was one C. albicans cell or 100 CFU/ml (10 fg) per PCR reaction. The real-time PCR efficiency rate for Candida was high (E = 1.95). Melting curve analysis of C. albicans showed a specific melting peak temperature of 65.76 °C. Conclusion: The real-time PCR assay we developed is highly specific and sufficiently sensitive to detect the fungal load for early diagnosis of invasive candidiasis.

Objective: The invasive aspergillosis (IA) is a serious opportunistic infection caused by various species of Aspergillus in immunocompromised individuals. Basically, rapid and early diagnosis prevents IA progression. In this study we performed a Real Time PCR/ Fluorescence Resonance Energy Transfer (FRET) for diagnosis of IA in hematologic malignancies and bone marrow transplant recipients. Materials and Methods: Sixty two patients with hematologic malignancies and marrow transplant recipients were evaluated for IA in Sari and Tehran from 2009 to 2010. The primer and hybridization probe were designed to amplify the specific sequence of 18S rRNA genes using Light Cycler system and FRET. Galactomannan (GM) assay was performed on serums which obtained from selected patients using the Platelia Aspergillus kit. Results: According to the criteria defined by the European Organization for Research and Treatment of Cancer and Mycoses Study Group (EORTC/MSG) for IA, 18 (29%) patients out of 62 patients were stratified into probable and possible groups. The female-to-male ratio was 1:2; the mean age of the patients was 36 years. The most common malignancies in these patients were acute lymphoblastic leukemia (38.9%). The minimum detection limit was 10 conidia (10 1 CFU/ml) equivalents (100 fg) per PCR reaction. GM assay was positive in 20.9% and real-time PCR probe set assay were positive in 17.7% patients who had clinical signs and host factor according to the mentioned criteria. Conclusion: Using the Real-Time PCR/FRET assay in whole blood specimens seems to be a promising method for diagnosis of IA, especially when used in combination with the GM detection test.

Introduction: Recently, the incidence of fungal infections dramatically increased with an appearance of many novel species due to different criteria. Therefore, several molecular methods have been established for identification of these agents which cause human and animal disorder based on genomic DNA. Among these methods, RAPD-PCR technique is a powerful discriminative method based on amplifies target genomic DNA sequence by short random primers (arbitrary short primers) with low annealing temperature (36˚C) for discrimination and identification in the species level. Materials and Methods: All clinical strains were originated oropharyngeal lesions of cancer patients from four Mazandaran University Hospitals in Iran. These strains were previously identified by phenotypic methods such as colony on CHROM-agar Candida medium, germ-tube formation in horse serum and chlamydospore formation. In this study RAPD-PCR technique was used to amplify hyper variable inter-repeat DNA sequences using oligonucleotide primers specific microsatellite (GACA) 4 for identifying, clustering and take into account the genetic correlation of 30 clinical isolates. Results: The results in this study showed that the RAPD-PCR by using of arbitrary short primer was able to amplify hyper variable inter repeated DNA sequences with classifying the isolates. RAPD patterns showed genetically inter speciation relationship in the best possible way and PCR-fingerprinting with primer (GACA) 4, was able to discriminate both C. albicans with other Candida species based on size and number of bands. Conclusion: We concluded that, regarding to the previous studies which have been reported misidentification by conventional mycological method for identifying Candida species, RAPD-DNA method is able to discriminate Candida species by using of a single primer. However, determination of differences and accurate assessment of genetic distances in the RAPD technique was generally limited.

"nIntroduction: Candida dubliniensis and C. albicans are very similar in morphology and phenotypic characteristics. Approximation of this yeast species has caused major problems in identifying these two correctly. "nMaterials and Methods: To distinguish among sixty yeast clinical isolates from patients with cancer, polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) was done. "nResults: PCR-RFLP of the ITS region showed different patterns between Candida dubliniensis and C. albicans after digestion with enzymes BlnI. All of the Clinical isolates were diagnosed as C .albicans. The results were confirmed by sequencing and RAPD-PCR. "nConclusion: PCR-RFLP would be a useful and applicable technique in clinical laboratories for discrimination of C. albicans and C. dubliniensis.

(Received 20 May, 2009 ; Accepted 10 August, 2009)AbstractBackground and purpose: Onychomycosis is a nail fungal infection caused by various species of dermatophytes, yeasts and non-dermatophytic molds and represents about 30% of cutaneous mycotic infections. The goal of this study was to investigate the frequency of onychomycosis and its associated factors in patients referred to Boali Sina Hospital and Toba dermatology outpatient clinics, Sari.Materials and methods: In this cross-sectional study, nail specimens were collected from 101 patients suspected of onychomycosis during a 14 month period. Nail specimens were examined by direct microscopy, using potassium hydroxide (KOH) 20%, KOH + CFW, KONCPA (KOH treated nail clipping+PAS) and also culturing on sabouraud’s dextrose agar, containing chloramphenicol (SC), and sabouraud’s dextrose agar containing cyclohexamide and chloramphenicol (SCC) mediums.Results: In this study, 79 (78%) of patients were female and 22 (22%) were male. Yeasts were isolated in 30% cases of onychomycosis, mainly from fingernails. Candida albicans, and C.krusei was the most prevalent species. Non-dermatophytic filamentous fungi were yielded at 24%, especially from toenails, with Aspergillus terreus being the most prevalent species. Dermatophytes were found in 7% of the samples, especially from toenails. Trichophyton mentagrophytes was the predominant species. Unknown filamentous fungi were identified in 19% of samples, while mixed infections were identified in 20% of samples. The highest prevalence rate of onychomycosis was identified in the patients within the 30 to 49 years of age group. Distal and lateral subfungual onychomycosis (DLSO) was the most prevalent clinical types with (88%), followed by WSO (5%), PSO (5%), TDO (1%) and DLSO + PSO (1%).Conclusion: The results of our study showed that onychomycosis is one of the most prevalent infections in patients who are suffering from nail disorders. Yeast and yeast like organisms cause these infections more than other fungus in this region. Women are more infected, as they are in daily contact with detergents and moisturJ Mazand Univ Med Sci 2009; 19(71): 33-43 (Persian)

, , , , , (Received 12 Aug, 2008; Accepted 3 Dec, 2008) Abstract Background and purpose: Grown fungi on books can be a risk factor for occupants as well as its known agents of bio-deterioration. Therefore, in this study, we surveyed the myco-flora of air, book and cabinets at Mazandaran University of Medical Sciences Libraries.Material and methods: Opened plates (containing Sabouraud’s dextrose agar with chlor-amphenicole media (SC) were used for the isolation of fungi in the air of indoor environment of libraries. Pleated carpet sterile fragments were used for survey of cabinets and books contamination. Then, these fragments were cultured on SC in laboratory.Results: A total of 939 colonies with 17 genera of fungi were identified from the environment of 4 school libraries at the Mazandaran University of Medical Sciences. The most common fungi isolated were: Penicillium (62.0%), Yeast (13.6%), sterile hyphae (7.6%) and Candida (5.6%). The most number of colonies were isolated from the air. Conclusion: Penicillium, Aspergillus, Alternaria and Stachybotrys were isolated from the libraries. They are considered toxigenic, allergenic, infective and also, as book deterioration agents. J Mazand Univ Med Sci 2008; 18(67):107-110 (Persian)

Background: As regards multilocus sequence typing (MLST) method directly analyze the polymorphism within DNA sequences; we performed the first nationwide study on the genotypic relationships of Candida albicans strains obtained from oropharynx and bronchoalveolar lavage (BAL) samples from immunocompromised patients. Methods: Fourteen epidemiologically unrelated clinical strains of C. albicans were obtained from three hospitals in Mazandaran Province, Iran (2006 to 2012) from seven patients with pulmonary infections and the rest with oropharyngeal samples of immunocompromised patients. Seven loci of housekeeping genes were sequenced for all fourteen isolates. Results: MLST was applied to a subset of 14 unrelated isolates. Seventy-one (2.5%) nucleotide sites were found to be variable. Accordingly, 60 different alleles were identified in seven loci among the isolates, among which two new alleles were obtained. Furthermore, 12 independent diploid sequence types (DSTs) including five novel DSTs were identified. The fourteen unrelated isolates were placed in 10 clonal clusters (CC) while two isolates were singletons, by eBURST analysis. Most of the isolates belonged to CC461 of eBURST analysis from the clade 11 and two isolates assigned to CC172 from the clade 15. Conclusion: Pathogen distribution and relatedness for determining the epidemiology of nosocomial infections is highly recommended for pathogen control methods.