Epidemiology and Antifungal Susceptibility of Candida Species Isolated from 10 Tertiary Care Hospitals in Iran.

In recent decades, the incidence of Candida infections has increased in immunocompromised patients. This multicenter study aimed to evaluate in vitro antifungal activities of 8 antifungal agents against the Candida species isolated from 10 university hospitals in Iran. During the period from Dec 2019 to Dec 2021, Candida species were collected from clinical samples of patients. The isolates were identified by PCR restriction fragment length polymorphism and sequencing methods. The antifungal susceptibility tests of each isolate to eight antifungal agents were performed according to the microdilution CLSI M27, M59, and M60 standard methods. A total of 598 Candida strains were isolated from clinical samples. The most commonly isolated Candida species was C. albicans, followed by C. glabrata, C. parapsilosis, Debaryomyces hansenii (Candida famata), C. tropicalis, Pichia kudriavzevii (Candida krusei), C. orthopsilosis, Meyerozyma guilliermondii (Candida guilliermondii), Kluyveromyces marxianus (Candida kefyr), and Clavispora lusitaniae (Candida lusitaniae). MIC90 values in all Candida species were as follows: 0.25 µg/mL for caspofungin and voriconazole; 0.5 µg/mL for amphotericin B and isavuconazole; 2 µg/mL for itraconazole, luliconazole, and posaconazole; and 16 µg/mL for fluconazole. Although 30/285 C. albicans, 15/31 C. hansenii, 3/12 M. guilliermondii, 67/125 C. glabrata, 5/15 P. kudriavzevii, 6/60 C. parapsilosis, and 5/23 C. tropicalis isolates were multiazole resistant with resistance to 2 to 4 azoles, pan-azole resistance was not observed. According to our data, Candida albicans and C. glabrata were the most frequent species isolated from clinical samples in Iran. Caspofungin and voriconazole, with lower MIC90 values, are the most effective than other antifungal agents for the treatment of Candida infections in this region. IMPORTANCE Candida species cause severe invasive infections of the heart, brain, eyes, bones, and other parts of the body. Knowledge of regional distributions of causative Candida agents and their antifungal susceptibility patterns can help to monitor resistance to antifungal agents of various species and support local and national surveillance programs. In the present study, C. albicans and C. glabrata were the most frequently isolated species from clinical samples in Iran. Increasing rates of non-albicans Candida isolates from the Iranian population should be looked at as alarming due to various levels of intrinsic MIC values or resistance to various antifungal drugs. Caspofungin and voriconazole are recommended over fluconazole for the treatment of Candida infections in the study region. However, amphotericin B and isavuconazole are also active against the most common Candida species isolated from patients. Pan azole-resistant Candida species were not observed in the present study.

WITHDRAWN: Evaluating the Antifungal Activity of Rumex acetosella, Teucrium polium, and Glycyrrihize globra var. violacca on Pathogenic Dermatophytes and Determining Phenolic Compounds

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Inhibitory Effect of Standardized Extract and Fractions of Nigella sativa L. on Nystatin Susceptible and Clinically Nystatin Resistant Candida albicans.

INTRODUCTION: Candidiasis infection is caused by different species of Candida, which are characterized by host immunologic weakness. Black cumin seeds (Nigella sativa) have shown an inhibitory effect against Candida albicans. In this work, the inhibitory effect of standardized extract and different fractions of Nigella sativa seeds has been evaluated on both nystatin-susceptible and resistant strain of C. albicans. MATERIALS AND METHODS: Canadida albicans (NSCA) with ATCC 76645 and nystatin-resistant Candida albicans (NRCA) were prepared from oral samples of HIV individuals. Total extract and different fractions of N. sativa were prepared using maceration and sonication methods. Thymoquinone (TQ) content of the plant was determined by spectrophotometric method. Total extract (TTE) and the fractions along with TQ were evaluated on NSCA and NRCA by the microdilution method. RESULTS: TQ content of the plant was 0.92 ± 0.37g/100g dried extract. The least MIC and MFC (62.5 and 125 µg/ml, respectively) were due to petroleum ether fraction (PEF) against both NSCA and NRCA, followed by chloroform fraction (CHF) with MIC and MFC of 125 and 250 µg/ml, respectively. TQ exhibited MIC of 0.78 and 3.12 µg/ml against NSCA and NRCA, stronger than nystatin (MIC of 2 and 16 µg/ml, respectively). Thymoquinone was detected in the PEF and CHF. CONCLUSION: Considering more inhibitory effects of PEF and CHF than TTE, we can conclude that active components of the plant belong to non-polar compounds. PEF showed identical inhibitory effects on NRCA and NSCA, which is a valuable result for finding novel medicaments against NRCA infections.

Multicenter Study of Susceptibility of Aspergillus Species Isolated from Iranian University Hospitals to Seven Antifungal Agents.

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.

Assessment of Aflatoxin M1 in human breast milk in Rafsanjan, Iran.

BACKGROUND AND PURPOSE: Aflatoxins (AFs) are a group of highly toxic mycotoxins present both in the environment and in foodstuffs. The food of infants should be safe and free of various pollutants, including breast milk mycotoxins. This study aimed to measure the mycotoxin of Aflatoxin M1 (AFM1) in human milk samples obtained from lactating mothers living in Rafsanjan city, Iran. MATERIALS AND METHODS: In the current cross-sectional study, breast milk samples were collected from 150 lactating mothers in Rafsanjan city from September 2015 to April 2016 using the structured food-frequency questionnaire. The AFM1 was measured by employing enzyme-linked immunosorbent assay specific kits. The statistical analysis was performed in SPSS software (version 16). RESULTS: The AFM1 was detected in 98 mothers (65%) with a mean concentration of 14.69±8.15 ng/kg, ranging from 5.02 to 41.25 ng/kg. The AFM1 concentration exceeded the tolerable and accepted limit promulgated by the European Union and the USA (25 ng/kg) in only 10 milk samples. Moreover, in 59 milk samples, the AFM1 concentration exceeded the limit recommended by Australia and Switzerland (10 ng/kg). CONCLUSION: According to the results of the present study, lactating mothers and their infants are at risk of AFM1 exposure in southern Iran. Accordingly, the examination of AFM1 concentrations in lactating mothers, as a critical postnatal exposure marker of infants to this carcinogenic compound, requires further studies in various seasonal periods and different parts of Iran.

Trachyspermum ammi aromatic water: A traditional drink with considerable anti-Candida activity.

BACKGROUND AND PURPOSE: Aromatic waters (AWs) are therapeutic distillates, which harbor both essential oil and water-soluble components of a plant. Due to the dispersion of the light amount of essence through the AWs, they have their specific pleasant smell, taste, and medicinal properties. In Iranian traditional medicine, Trachyspermum ammi AW is used to treat gastrointestinal disorders. The present study was conducted to determine the chemical composition of the essential oil extracted from T. ammi AW and its antifungal activities against Candida species. MATERIALS AND METHODS: The composition of the essential oil extracted from T. ammi AW was analyzed by gas chromatography-mass spectrometry. In addition, the evaluation of the antifungal activity of AW against Candida species was performed using broth microdilution methods as recommended by the Clinical Laboratory Standard Institute. Moreover, the biofilm formation inhibition, antioxidant properties, and experimental activity of AW were determined in an animal model. RESULTS: According to the results, thymol (78.08%) was the major compound of EO, followed by carvacrol (8.20%) and carvotanacetone (6.50%). Furthermore, T. ammi AW exhibited antifungal activities against the examined fungi and inhibited the biofilm formation of C. albicans at a concentration of up to 0.25 V/V. Histopathological analyses revealed that Candida colonization declined in the mice following the administration ofT. ammi AW in a therapeutic trial. CONCLUSION: It seems that the presence of phenolic monoterpenes in AW has resulted in antifungal effects. Pleasant odor and antioxidant properties are extra bonuses to the antimicrobial effects of this plant. Based on the findings, AW might have the potential to be used in the management of alimentary candidiasis or oral hygienic products.

Epidemiology of dermatomycoses in Kerman province, southeast of Iran: A 10-years retrospective study (2004-2014).

OBJECTIVES: Superficial and cutaneous fungal infections (SCFIs) are an important public health problem and are common in tropical and subtropical countries. Pityriasis versicolor, dermatophytosis, erythrasma, onychomycosis, and otomycosis are the major diseases associated with SCFIs. The aim of this study was to evaluate the prevalence and causative agents of dermatomycoses over a period of 10 years in Kerman province, southeast of Iran. METHODS: A number of 1782 subjects, including 1096 females and 686 males, with cutaneous disorders in their skin, nail, and hair suspected to have SCFIs participated in this study. The collected specimens were examined using direct microscopy examination, staining, culture on specific media and PCR-RFLP technique. RESULTS: In total, 617 (34.62%) subjects had SCFIs, of whom 290 (47%) were female and 327 (53%) were male. Identified SCFIs included yeast infections, dermatophytosis, saprophyte onychomycosis, erythrasma, and otomycosis due to non-dermatophytic molds (NDMs). The highest prevalence of dermatomycoses was found among the 41-50-year and 31-40-year age groups. Tinea unguium was the most common clinical pattern of dermatomycoses, and T. mentagrophytes was the predominant agent. Also, Aspergillus spp. were the most common NDMs agents of onychomycosis and otomycosis. CONCLUSIONS: This study summarized the epidemiological trends and etiologic agents of SCFIs in a 10-year period in Kerman, southeast of Iran. Consideration of the current epidemiologic trends in the prevalence and knowledge of the exact causative agents of SCFIs may play an important key role towards further investigations, diagnosis, and modification of current treatments.

Plasma Levels of IFN-γ, IL-4, IL-6 and IL-17 in HIV-Positive Patients With Oral Candidiasis.

BACKGROUND: Cell-mediated immunity (CMI) by CD4 + Th (T helper)-type cells is the predominant host defense mechanism against Oral Candidiasis (OC) in HIV-infected individuals. Weakened CMI and depletion of CD4 + T cells are the main factor contributing to the output of OC in HIV-positive individuals. The cytokines produced by Th1, Th2 and Th17 cells play a role in mediating an increased susceptibility to OC during HIV infection. OBJECTIVES: The present study investigated plasma concentration of IFN-γ, IL-4, IL-6 and IL-17 in HIV-1 patients suffering from OC. PATIENTS AND METHODS: In total, 98 samples in four groups (HIV-positive and HIV-negative persons with and without OC) were obtained from the oral cavities and cultured on Sabouraud's dextrose agar and CHROMagar. Also blood samples were obtained to assess plasma level of IFN-γ, IL-4, IL-6 and IL-17 using ELISA technique. RESULTS: There was a statistically significant difference in the plasma concentration of IFN-γ, IL-6 and IL-17 but not about IL-4. Our findings suggest a significant interaction between fungal infection and HIV on expression of assessed cytokines. CONCLUSIONS: Fungal infection and HIV alone and together could seriously alter immune system function as assessed by measuring the levels of the plasma cytokines. Therefore, these results provide important new information relative to the putative immune-based factors associated with resistance and/or susceptibility to OC in HIV-positive persons.

The Effects of Opium Addiction on the Immune System Function in Patients with Fungal Infection.

BACKGROUND: The use of narcotics such as opium exposes addicts as susceptible targets of different diseases so that they might easily be exposed to different diseases such as fungal infections. The present study aimed to investigate the effects of addiction to opium and fungal infection on plasma levels of certain cytokines including interleukin-4 (IL-4), IL-6, IL-17, Interferon gamma (IFN-γ) and transforming growth factor-ß (TGF-ß). METHODS: Present study included 72 individuals who were divided into 4 groups: 1) opium-addicted with fungal infection; 2) opium-addicted without fungal infection; 3) non-opium-addicted with fungal infection; and 4) normal individuals (non-opium-addicted and non-fungal infection). The fungal samples, after being detected and confirmed by a physician, were prepared based on clinical symptoms and then analyzed by direct smear and culture method. The measurement of the plasma level of cytokines was done by enzyme-linked immunosorbent assay (ELISA) method. FINDINGS: The comparison of the mean of the plasma level of cytokines showed that addiction to opium and fungal infection had significant effect on the plasma levels of IL-17, IFN-γ, TGF-ß cytokines in all studied groups. The interaction of addiction to opium and fungal infection was only significant in the case of plasma level of IL-6. CONCLUSION: Addiction to opium and fungal infection, either separately or simultaneously, poses significant effect on the immune system and causes disorders in the cytokine network and the immune system and also provides a suitable environment for fungal infection.

Etiologic Agents of Otomycosis in the North-Western Area of Iran.

BACKGROUND: Otomycosis is a superficial fungal infection often involves the pinna and external auditory canal. It is a pathologic condition, with Candida and Aspergillus, the most common fungal species. It is common worldwide but more prevalent in tropical and subtropical countries. OBJECTIVES: The aim of this study was to determine the etiologic agents and local epidemiologic pattern of otomycosis in northwest Iran. PATIENTS AND METHODS: A series of 140 patients with clinically symptomatic otomycosis were studied in 21 cities, towns, and villages throughout northwest Iran between 2009 and 2011. Clinical samples were collected by swabs and then assessed by mycological investigation. RESULTS: Otomycosis was diagnosed in 129 cases (92%, 76 male, 53 female) with the highest prevalence of cases occurring in males between 21 - 40 years of age. From an etiological point of view, 116 patients (90%, 21 - 40 years old) were infected by saprophytic mold and 9 patients (7%) were infected by yeast. Three cases (2%) involved dermatophytes, and in one case (1%) the subject was infected with Eurotium (the perfect stage of Aspergillus fumigatus). Aspergillus niger was the most common mold that was isolated, followed by A. flavus, A. fumigatus, Penicillium spp., Fusarium spp., and Rhizopus spp. A total of 2 yeasts belonging to genus Candida, C. albicans and C. tropicalis, were isolated. CONCLUSIONS: Our study showed a high prevalence of otomycosis in the northwestern area of Iran. As such, proper diagnosis and treatment by aseptic techniques for this disease is urgently needed.

Antifungal Activity of Selenium Nanoparticles Synthesized by Bacillus species Msh-1 Against Aspergillus fumigatus and Candida albicans.

BACKGROUND: Fungal infections affect various parts of the body and can be difficult to treat. Aspergillus infection causes a spectrum of diverse diseases particularly in lung according to host immunity. The two major entities are invasive pulmonary aspergillosis and chronic pulmonary aspergillosis. Candida infections can be superficial or invasive. Superficial infections often affect the skin or mucous membranes. However, invasive fungal infections are often life-threatening. Advances in nanotechnology have opened new horizons in nanomedicine, allowing the synthesis of nanoparticles that can be assembled into complex architectures. Novel studies and technologies are devoted to understanding the mechanisms of disease for the design of new drugs. OBJECTIVES: In the present study, the antifungal activity of biogenic selenium nanoparticles (Se NPs) against Aspergillus fumigatus and Candida albicans was investigated. MATERIALS AND METHODS: Se-reducing bacteria previously identified as Bacillus sp. MSh-1 were used for the intracellular biosynthesis of elemental Se NPs. The shape, size, and purity of the extracted NPs were determined with various instrumental techniques. The nanoparticles antifungal characterization mainly derives from the following pathways: (i) to generate sustained flux of nano-ions from the compounds that deposited on special substrates or imbedded in colloidal or semisolid matrices. (ii) To transport active those ions to sensitive targets on plasma membrane of fungi. RESULTS: The results of energy-dispersive X-ray demonstrated that the purified NPs consisted of only Se. In addition, transmission electron micrographs showed that 120- to 140-nm spherical Se NPs were the most common. An antifungal assay was performed with a standard Clinical and Laboratory Standards Institute broth microdilution method. Minimum inhibitory concentration (MIC) measurements of the antifungal activity of the Se NPs against C. albicans (70 µg/mL) and A. fumigatus (100 µg/mL) showed that yeast cells were more sensitive than mold cells. CONCLUSIONS: The MICs against A. fumigatus (100 µg/mL) and C. albicans (70 µg/mL) showed that biogenic Se NPs are useful antifungal agents.

In Vitro Evaluation of Enzymatic and Antifungal Activities of Soil-Actinomycetes Isolates and Their Molecular Identification by PCR.

BACKGROUND: Human cutaneous infection caused by a homogeneous group of keratinophilic fungi called dermatophytes. These fungi are the most common infectious agents in humans that are free of any population and geographic area. Microsporum canis is a cause of dermatophytosis (Tinea) in recent years in Iran and atypical strain has been isolated in Iran. Its cases occur sporadically due to M. canis transmission from puppies and cats to humans. Since this pathogenic dermatophyte is eukaryotes, chemical treatment with antifungal drugs may also affect host tissue cells. OBJECTIVES: The aim of the current study was to find a new antifungal agent of soil-Actinomycetes from Kerman province against M. canis and Actinomycete isolates were identified by PCR. MATERIALS AND METHODS: A number of hundred Actinomycete isolated strains were evaluated from soil of Kerman province, for their antagonistic activity against the M. canis. M. canis of the Persian Type Culture Collection (PTCC) was obtained from the Iranian Research Organization for Science and Technology (IROST). Electron microscope studies of these isolates were performed based on the physiological properties of these antagonists including lipase, amylase, protease and chitinase activities according to the relevant protocols and were identified using gene 16SrDNA. RESULTS: In this study the most antagonist of Actinomycete isolates with antifungal activity against M. canis isolates of L1, D5, Ks1m, Km2, Kn1, Ks8 and Ks1 were shown in vitro. Electron microscopic studies showed that some fungal strains form spores, mycelia and spore chain. Nucleotide analysis showed that Ks8 had maximum homology (98%) to Streptomyces zaomyceticus strain xsd08149 and L1 displayed 100% homology to Streptomyces sp. HVG6 using 16SrDNA studies. CONCLUSIONS: Our findings showed that Streptomyces has antifungal effects against M. canis.

In vitro and in vivo antidermatophytic activities of some Iranian medicinal plants.

In the last decades, the number of people suffering from dermatophytoses has seriously increased, which may be due to the development of resistant strains to a range of antifungal drugs. The present study was aimed to evaluate the antidermatophytic properties of eight extracts from the selected spices and herbs, which were ethno-medicinally used in Iran against Trichophyton mentagrophytes, Trichophyton interdigitale, Microsporum canis, and Microsporum gypseum (10 strain of each). The in vitro antifungal activities of the extracts from four spices and four plants were evaluated by the broth macro dilution method against four dermatophyte strains. In addition, the in vivo therapeutic effects of Myrtus communis L. and Cinnamomum zeylanicum Blume extracts (the most active extracts) on dermatophytosis induced by M. canis and T. mentagrophytes in guinea pigs were evaluated. Results of in vitro antifungal assay revealed that all the tested extracts demonstrated both fungistatic and fungicidal activities with the geometric mean (GM) MIC ranging from 0.058 to 3.73 mg/ml and GM (MFC) ranging from 0.058 to 7.46 mg/ml, respectively. Two extracts (M. communis and C. zeylanicum) significantly inhibited the growth of all the tested dermatophytes, while other extracts demonstrated weak (MICs of >0.625 mg/ml) to moderate (MICs ranging from 100 to 0.625 mg/ml) activities. In vivo antidermatophytic assay demonstrated that clotrimazole cured T. mentagrophytes and M. canis infection on days 21 and 17, respectively, whereas M. communis and C. zeylanicum extracts significantly (p < 0.05) cured T. mentagrophytes and M. canis infection on days 9 and 13 as well as 9, 11, respectively. Phytochemical screening showed the presence of flavonoids, tannins, phenols, and alkaloids in M. communis and alkaloids, flavonoids, and tannins in C. zeylanicum. Findings of the present study also provided the scientific evidence that natural plants could be used in traditional medicine for the prevention and treatment of dermatophytic infections.

Evaluation of Antifungal Effect of Silver Nanoparticles Against Microsporum canis, Trichophyton mentagrophytes and Microsporum gypseum.

BACKGROUND: Dermatophytosis is the common cutaneous infections in humans and animals, which is caused by the keratinophylic fungus called dermatophytes. In recent years, drugs resistance in pathogenic fungi, including dermatophyte strains to the current antifungals have been increased. OBJECTIVES: The aim of this study was to evaluate the antifungal efficacy of AgNPs against Microsporum canis, Trichophyton mentagrophytes , and Microsporum gypseum. MATERIALS AND METHODS: The antifungal susceptibility of nanosilver particles compared with griseofulvin (GR). Its efficacy was investigated against three strains of dermatophytes by both agar dilution and broth microdilution test (BMD). RESULTS: The average minimum inhibitory concentration (MIC) AgNPs on M. canis, T. mentagrophytes and M. gypseum were 200, 180 and 170 µg.mL-1, respectively. Whereas these strains showed MIC of 25, 100 and 50 µg.mL-1 for GR. CONCLUSIONS: Our finding indicated that the AgNPs was less active than GR but it had anti-dermatophytic effect.

Antifungal, Antileishmanial, and Cytotoxicity Activities of Various Extracts of Berberis vulgaris (Berberidaceae) and Its Active Principle Berberine.

In this study, in vitro antidermatophytic activity against Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis, and Microsporum gypseum was studied by disk diffusion test and assessment of minimum inhibitory concentration (MIC) using CLSI broth macrodilution method (M38-A2). Moreover, antileishmanial and cytotoxicity activity of B. vulgaris and berberine against promastigotes of Leishmania major and Leishmania tropica were evaluated by colorimetric MTT assay. The findings indicated that the various extracts of B. vulgaris particularly berberine showed high potential antidermatophytic against pathogenic dermatophytes tested with MIC values varying from 0.125 to >4 mg/mL. The results revealed that B. vulgaris extracts as well as berberine were effective in inhibiting L. major and L. tropica promastigotes growth in a dose-dependent manner with IC50 (50% inhibitory concentration) values varying from 2.1 to 26.6 µ g/mL. Moreover, it could be observed that berberine as compared with B. vulgaris exhibited more cytotoxicity against murine macrophages with CC50 (cytotoxicity concentration for 50% of cells) values varying from 27.3 to 362.6 µ g/mL. Results of this investigation were the first step in the search for new antidermatophytic and antileishmanial drugs. However, further works are required to evaluate exact effect of these extracts in animal models as well as volunteer human subjects.