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Current Medical Mycology - Volume:5 Issue: 2, Jun 2019

Current Medical Mycology
Volume:5 Issue: 2, Jun 2019

  • تاریخ انتشار: 1398/03/11
  • تعداد عناوین: 9
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  • Abdelnasser Mohammadi*, Seifollah Bahramikia Pages 1-8
    Background and Purpose
    Alternaria is one of the most abundant fungi that exists in numerous places around the world. This saprophytic fungus causes diseases in plants and accounts for the spoilage of cereals in warehouses. The aim of this study was to identify Alternaria isolates based on their morphological characteristics and internal transcribed spacer ribosomal RNA (ITS rRNA) sequencing method. To this end, genetic diversity in the isolates was also examined using inter simple sequence repeat (ISSR) markers.

    Materials and Methods
    To conduct this research, a total of 60 tomato samples with black spots were collected from supermarkets in Khorramabad City, Iran, in the winter of 2017. The specimens were cultured on a potato dextrose agar medium. After the purification of the fungus by the single-spore method, the identification of the species was carried out using morphological characteristics and ITS rRNA sequencing by polymerase chain reaction. The genetic diversity of the identified species with four primers was evaluated using the ISSR marker.

    Results
    Based on the sequencing of the ITS1 region, all the isolates were identified as A. alternata. Cluster diagrams for the ISSR marker were classified into six distinct groups. The mean polymorphism information content was obtained as 0.35, indicating the effectiveness of the primers in the separation of the isolates.

    Conclusion
    The sequencing of ITS1 led to the identification of Alternaria species that are morphologically similar. The production of various mycotoxins by A. Alternata species leads to the contamination of livestock and human food. Regarding this, the investigation of the genetic diversity of A. alternata species using the ISSR marker would facilitate the identification of suitable and effective strategies for controlling the fungal and mycotoxin contamination of human nutrition.
    Keywords: Alternaria alternata, Inter simple sequence repeat methods, ITS rRNA, Tomato
  • Parisa Aryamloo, Hossein Asgarian, Omran, Narges Aslani, Hadi Hossein, Nataj, Tahereh Shokohi, Hamid Badali, Mojtaba Nabili, Atefeh Abdollahi Gohar, Maryam Moazeni* Pages 9-15
    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<0.05). In addition, the rate of necrosis was significantly greater than that of apoptosis in response to caspofungin. Accordingly, necrosis occurred in 71.26%, 71.26%, and 61.26% of the cells at the caspofungin concentrations of 0.25, 0.5, and 1 μg/mL, respectively (P<0.05). The analysis of the data in the REST software demonstrated a significant increase in the expression of MCA1 and NUC1 genes (P<0.05).

    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.
    Keywords: Candida glabrata, Caspofungin, Flow cytometry, MCA1, NUC1
  • Maryam Ebrahimi, Hossein Zarrinfar*, Ali Naseri, Mohammad Javad Najafzadeh, Abdolmajid Fata, Mahmoud Parian, Imaneh Khorsand, Monika Novak Babič Pages 16-21
    Background and Purpose
    Dermatophytes as the causative agents of dermatophytosis (ringworm) are widely spread around the world. Accurate identification of dermatophytes in one area can be particularly important for epidemiological studies. Regarding this, the aim of the present study was to describe the species spectrum of dermatophytes, isolated from patients in Mashhad city, Iran, using the molecular-based method.

    Materials and Methods
    This study was conducted on 79 dermatophyte isolates obtained from the human skin, hair, and nail specimens. Species identification was performed by the polymerase chain reaction-restriction fragment length polymorphism analysis of ribosomal DNA internal transcribed spacer regions using MvaI restriction enzyme.

    Results
    The identified species included Trichophyton mentagrophytes/T. interdigitale species complex (n=37, 46.8%), Epidermophyton floccosum (n=12, 15.2%), T. rubrum (n=8, 10.1%), Microsporum canis (n=8, 10.1%), T. violaceum (n=5, 6.3%), T. tonsurans (n=4, 5.1%), Nannizzia gypsea (n=3, 3.8%), T. benhamiae (n=1, 1.3%), and T. verrucosum (n=1, 1.3%). The clinical forms of infection were tinea corporis (n=26, 32.8%), tinea cruris (n=22, 27.8%), tinea capitis (n=10, 12.6%), tinea unguium (n=7, 9%), tinea manuum (n=6, 8%), tinea pedis (n=5, 6.3%), and tinea faciei (n=3, 3.5%).

    Conclusion
    As the findings indicated, T. mentagrophytes/T. interdigitale species complex had the highest prevalence, and T. benhamiae appeared to be a new emerging agent of dermatophytosis in Mashhad, northeastern Iran.
    Keywords: Dermatophyte, Dermatophytosis, Mashhad, PCR-RFLP, Subtropical
  • Mohammad Taghi. Hedayati, Saham Ansari, Bahram Ahmadi, Mojtaba Taghizadeh Armaki, Tahreh Shokohi, Mahdi Abastabar, Halil Er, Betil Özhak, Dilara Öğünç, Macit Ilkit*, Seyedmojtaba Seyedmousavi Pages 22-26
    Background and Purpose
    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is widely used to discriminate among pathogenic microorganisms in clinical laboratories. The aim of this study was to assess the utility of MALDI-TOF MS in the routine identification of clinical dermatophyte isolates obtained from various geographical regions of Iran.

    Materials and Methods
    A total of 94 isolates, including Trichophyton interdigitale (n=44), T. rubrum (n=40), T. tonsurans (n=4), Microsporum canis (n=4), and Epidermophyton floccosum (n=1), were analyzed in this study. The identity of each isolate was determined by polymerase chani reaction amplification and sequencing of the internal transcribed spacer (ITS) region of nuclear-encoded ribosomal DNA and also MALDI-TOF MS. The obtained data by molecular approach were compared with MALDI-TOF MS.

    Results
    The MALDI-TOF MS led to the identification of 44 (47%) isolates at the species level by generating the spectral score values of ≥ 2.0. However, there was not sufficient agreement between the results of the molecular-based ITS identification methods and MALDI-TOF MS in the species identification of 16 (17%) isolates. The Bruker Daltonics database was also not able to identify protein spectra related to 12 isolates (13%), including T. interdigitale (n=5), T. rubrum (n=4), M. canis (n=2), and T. tonsurans (n=1).

    Conclusion
    According to the results, the utility of MALDI-TOF MS as a routine diagnostic tool for the accurate and reliable identification of dermatophytes can be justified whenever the protein spectra of a large set of worldwide clinical isolates are included in the commercial libraries. In addition, MALDI-TOF MS can be alternatively used to construct an in-house reference database.
    Keywords: Dermatophytes, Iran, ITS phylogeny, MALDI-TOF MS
  • Maryam Akbari Dana, Seyed Jamal Hashemi, Roshanak Daei Ghazvini, Sadegh Khodavesi, Mona Modiri, Ladan Nazemi, Sima Darabian, Sasan Rezaie* Pages 27-32
    Background and Purpose
    Aspergillus flavus is an important pathogen in immunodeficient patients. Due to the abundance of this fungus in nature, fungicides are commonly used to preserve and maintain agricultural products. Long-term exposure to these pesticides can lead to the induction of drug resistance in this fungus.

    Materials and Methods
    For the purpose of the study, 10 strains of A. flavus ATCC 204304 were cultured in benomyl and diazinon pesticides at the concentrations of 62.5, 125, 250.500, 750, 1000, 1500, 2000, and 2500 mg/L in nine steps. Morphological changes and resistance to voriconazole, itraconazole, and amphotericin B were evaluated at the end of each step. Subsequently, changes in the expression of mdr1 and cyp51C genes were studied in the strains showing drug resistance.

    Results
    The results showed that during the nine stages of the adjacency of strains with benomyl and diazinon at different concentrations, resistance to voriconazole, itraconazole, and amphotericin B in these toxins increased by 30% and 10%, respectively. In addition, the microscopic examination of resistant strains revealed the absence of sporulation, and only mycelium was found. Macroscopically, the color of the colonies changed from green to white. Furthermore, the investigation of the expression of mdr1 and cyp51c genes in these strains showed a decrease and increase in adjacency with diazinon and benomyl, respectively.

    Conclusion
    As the findings indicated, exposure to agricultural pesticides can lead to the incidence of morphological changes and resistance to amphotericin B, itraconazole, and voriconazole in the sensitive species of A. flavus by altering the expression of genes involved in drug resistance.
    Keywords: Aspergillus flavus, Azole resistant, Gene expression, Pesticides
  • Amin Gharanfoli, Elaheh Mahmoudi*, Roya Torabizadeh, Farzad Katiraii, Saeid Faraji Pages 33-36
    Background and Purpose
    Candida species are reportedly the most common human fungal pathogens. The incidence of urinary tract infections (UTIs) caused by Candida pathogens has increased in recent decades. However, such infections rarely occur in the absence of any predisposing factors. Regarding this, the aim of the present study was to identify the Candida species causing UTIs and determine the predisposing factors for candiduria.

    Materials and Methods
    The current study was conducted on 1,450 urine samples obtained from patients suspected of UTI. Out of this number, 19 cases were candidiasis, and 2 cases were mixed infections caused by bacteria and fungi. Candida species were diagnosed differentially using the germ tube test, colony staining on CHROMagar medium, intracellular beta-glucosidase enzyme activity, and glucose absorption pattern. Then, the colonies with the same morphology were confirmed by the DNA sequencing of internal transcribed spacer regions.

    Results
    According to the results, 38%, 28.6%, 14.3%, and 9.5% of the isolates were identified as C. albicans, C. glabrata, C. tropicalis, and C. kefir/C. krusei, respectively. The presence of one or more predisposing factors was proved in all patients in whom diabetes was the most prevalent predisposing factor (21.1%).

    Conclusion
    Based on the obtained results, C. albicans species was the most prevalent fungal species. In addition, urinary fungal infections were less prevalent than bacterial urinary infections.
    Keywords: Candida species, Predisposing factors, Urinary tract infections
  • Keyvan Pakshir*, Sahar Sheykhi, Kamiar Zomorodian, Hasti Nouraei, Zahra Zareshahrabadi Pages 37-40
    Background and Purpose
    Candida albicans is one of the most opportunistic yeasts around the world. This species has two heterozygous and homozygous strains at hyphal wall protein 1 (hwp1) gene locus. A simple method for the discrimination of these two strains is the amplification of HWP1 gene. Regarding this, the aim of this study was to discriminate C. albicans heterozygous and homozygous strains via the amplification of hwp1 gene and evaluation of biofilm formation between the strains.
    Materials and Methods
    A total of 60 homozygous (n=30) and heterozygous (n=30) strains were discriminated among 126 C. albicans vaginal isolates by the amplification of HWP1 gene, using specific primers. The evaluation of biofilm formation was accomplished using the visual method.
    Results
    According to the results, the homozygous and heterozygous strains produced one and two DNA fragments, respectively. The frequency of homozygous strains among the C. albicans vaginal isolates was 76.2%. Biofilm formation activity in the heterozygous strains was more than that in the homozygous strains. However, statistical analysis showed no significant difference between the strains in terms of biofilm formation.
    Conclusion
    As the findings indicated, the frequency of the heterozygous strains in C. albicans was lower than that of the homozygous strains. Both of the strains could form biofilm in the different ranges of severity. High activity of biofilm formation in heterozygous strains may set the ground for its pathogenicity.
    Keywords: Biofilm, Candida albicans, Heterozygous, Homozygous, Virulence factor
  • Ismael Maatouk*, Roger Haber, Nazim Benmihidi Pages 41-44
    Background and Purpose
     The aim of this study was to evaluate the onychoscopic patterns associated with distal lateral subungual onychomycosis (DLSO) in Lebanon.
    Materials and Methods
    The present study was conducted on 45 patients with clinical DLSO attending two dermatology clinics in Beirut, Lebanon, between January 2018 and April 2018. The patients were subjected to dermoscopy to identify the onychoscopic patterns.

    Results
    The DLSO was predominantly associated with white, yellow, and brown color changes (P<0.05). Dermoscopic patterns of longitudinal striae (n=31; 68.75%), spiked pattern (n=25; 55.5%), and jagged pattern (n=25; 55.5%) were significantly correlated with DLSO (P<0.001). Our findings are in accordance with five previous reports in which dermoscopic findings are discussed in onychomycosis.

    Conclusion
    It is recommended to perform further studies on homogeneous groups with different clinical subtypes of onychomycosis including patients with suspected traumatic onycholysis or other nail diseases. Identification of onychoscopic patterns would offer the clinicians a quick, simple, and complementary tool for the diagnosis of onychomycosis.
    Keywords: Dermatoscopy, Fungus, Lebanon, Nail, Onychomycosis, Onychoscopy
  • Mohammad Javad Najafzadeh* Pages 45-48
    Background and Purpose
    Herein, we report the first case of fungal keratitis due to Aspergillus minisclerotigenes in a 68-year-old rural woman admitted to the Ophthalmology Center of Khatam-Al-Anbia Hospital in Mashhad, northeast of Iran.

    Case report: The patient presented with severe pain, burning, foreign body sensation, and reduced vision in her right eye. She had long-term uncontrolled diabetes and was not able to close her eye due to an anatomical problem with the eyelid. The cornea smear sample was cultured, and the fungus was initially identified as Aspergillus flavus. The isolated strain was further identified by sequencing a part of the calmodulin gene as A. minisclerotigenes. The patient did not respond to any antifungal treatments (e.g., amphotericin B and voriconazole drops, and fluconazole 300 mg/day); therefore, she was eventually subjected to corneal transplantation surgery.

    Conclusion
    Fungal keratitis can be caused by the less common species. The reliable identification of the causative agents can be accomplished by the implementation of molecular methods.
    Keywords: Aspergillus minisclerotigenes, Fungal keratitis, Iran