Isolation of keratinophilic fungi from the soil of Greater Tunb, Abu-Musa, and Sirri islands in the Persian Gulf, Iran.

BACKGROUND AND PURPOSE: Keratinophilic fungi are among the important groups of fungi living in the soil. This study aimed to isolate and identify keratinophilic fungi from the soil of three Iranian islands, namely Greater Tunb, Abu Musa, and Sirri, located in the Persian Gulf using morphological and molecular (polymerase chain reaction) methods. MATERIALS AND METHODS: In this study, a total of 60 soil samples were collected from the three islands of Greater Tunb, Abu Musa, and Sirri. The samples were analyzed for the presence of the keratinophilic fungi using a hair baiting technique. Furthermore, the identification of keratinophilic fungi was accomplished through the employment of molecular and sequencing techniques. RESULTS: A total of 130 fungal isolates, including 11 genera with 24 species, were collected. Accordingly, Chrysosporium tropicum (24;18.5%), C. keratinophilum (17; 13.1%), Chrysosporium species (15; 11.5%), Aspergillus species ( 8;6.1%), Aspergillus flavus (8; 6.1%), Penicillium species (8;6.1%), Alternaria spp ( 6; 4.6%), Phoma species (5; 3.8%), Aphanoascus verrucosus (4;3.1%), Fusarium chlamydosporum (4; 3.1%), Aspergillustrreus (4;3.1%), Acremonium species (4; 3.1%), and other fungi( 23; 17.8 %) isolates were identified . All isolates of keratinophilic fungi were isolated from the soils with the pH range of 7-9. CONCLUSION: The results of this study contributed towards a better conceptualization of the incidence pattern of keratinophilic fungi in the regions of Iran. Given that no study has investigated this issue, the findings of the present study can be beneficial for the management of public health surveillance, physicians, and epidemiologists.

The antagonistic effects of Candida parapsilosis on the growth of Fusarium species and fumonisin production.

BACKGROUND AND PURPOSE: Fusarium species are avid producers of secondary toxic and carcinogenic metabolites such as fumonisin. Contamination of food and feed products with fumonisin can be hazardous to the health of humans and animals and may lead to agricultural loss. Accordingly, in this study, we aimed to evaluate the effects of Candida parapsilosis on the growth and fumonisin production of Fusarium species. MATERIALS AND METHODS: Mycelial growth rate of 26 Fusarium isolates, including F. verticillioides (n=6), F. proliferatum (n=18), F. solani (n=1), and F. oxysporum (n=1), in the presence of 42 C. parapsilosis strains was investigated by pour-plate method. The decline in fumonisin production was measured in co-cultured fungi in coarsely ground maize after four weeks of incubation in the dark at 22°C, using ELISA technique. For data analysis, paired t-test was performed, using SPSS version 20. RESULTS: The mycelial growth and fumonisin production of Fusarium isolates significantly decreased in the presence of C. parapsilosis in comparison with the control cultures (P<0.05). The percentage of mycelial growth inhibition ranged from 56.36% to 74.54%. The minimum and maximum decline in total fumonisin production was 12% and 78%, respectively. F. oxysporum and F. solani were found to be minor fumonisin producers among the studied Fusarium species. On the other hand, a decline was reported in the growth of Fusarium species and fumonisin production in the presence of C. parapsilosis. CONCLUSION: C. parapsilosis showed notable inhibitory activities against Fusarium isolates. Therefore, this fungal species could be considered as a biocontrol agent against the growth and fumonisin production of toxigenic Fusarium species in the future.

Effects of thiamine on growth, aflatoxin production, and aflr gene expression in A.parasiticus.

BACKGROUND AND PURPOSE: Mycotoxins are secondary fungal metabolites with a very high diversity that are produced by some species of Aspergillus which frequently leads to contaminate food and agricultural products. Recently, elimination of aflatoxin contamination in food and feed has been considered by scientists worldwide. Although, the antibacterial and antifungal effects of vitamins as natural compounds have been proven, the mechanism of vitamins effect on Aspergillus parasiticus growth and aflatoxin production is not yet clear. In this study, the effect of thiamine (vitamin B1) was studied on Aspergillus parasiticus growth, aflatoxins production and the afIR gene expression. MATERIALS AND METHODS: A standard strain of Aspergillus parasiticus was applied for performing antifungal susceptibility test in different concentrations of thiamine. Antifungal susceptibility test was performed according to CLSI M38-A2 document. The concentration of aflatoxin was determined by HPLC. Moreover, the quantitative changes in the aflR gene expression were analyzed by Real Time PCR method. RESULTS: The minimum inhibitory concentration was yielded as > 500 mg/ml. However, HPLC analysis results showed that aflatoxin production reduced in samples treated with 500 mg/ml of thiamine. In addition, the level of afIR gene expression was significantly reduced after treating with 500 and 250 mg/ml of vitamin B1. CONCLUSION: Based on the obtained results, thiamine could not inhibit the fungal growth completely. However, the rate of afIR gene expression and aflatoxin production was significantly reduced after fungal treating with thiamine. Consequently, using natural compounds such as vitamins may be regarded as potential antitoxic agent in food industry and the industries related to agriculture.

In vitro antifungal activities of Euphorbia macroclada and fluconazole against pathogenic Candida species.

BACKGROUND AND PURPOSE: Candida species constitute an important group of opportunistic fungi, which cause various clinical diseases. Considering the resistance of some Candida species to conventional antifungal agents, treatment of such cases may be challenging and complicated. The purpose of this study was to evaluate and compare the antifungal activities of Euphorbia macroclada latex and fluconazole against different Candida species. MATERIALS AND METHODS: A total of 150 Candida isolates including C. albicans (n=77), C. glabrata (n=28), C. parapsilosis (n=23), C. tropicalis (n=15), C. krusei (n=4), C. famata (n=1), C. kefyr (n=1) and C. inconspicua (n=1) were included in this study. In vitro antifungal activities of Euphorbia macroclada latex and fluconazole against these Candida species were evaluated, according to M27-A2 protocol on broth macrodilution method by the Clinical and Laboratory Standards Institute (CLSI). RESULTS: Among 150 Candida isolates, 98 isolates (65.33%), i.e., C. albicans (n=41), C. glabrata (n=23), C. tropicalis (n=12) and C. parapsilosis (n=22) with minimal inhibitory concentration (MIC) ≤ 8 µg/ml were susceptible to fluconazole. Resistance to fluconazole was noted in 15 isolates, i.e., C. albicans (n=10), C. glabrata (n=2), C. krusei (n=1), C. kefyr (n=1), and C. inconspicua (n=1), with MICs of 64 µg/ml. The remaining isolates (n=37) including C. albicans (n=26), C. glabrata (n=3), C. tropicalis (n=3), C. parapsilosis (n=1), C. krusei (n=3) and C. famata (n=1) with MIC= 16-32 µg/ml showed dose-dependent susceptibility. The latex of Euphorbia macroclada was able to inhibit the growth of 30 out of 150 tested Candida isolates with MIC range of 128-512 µg/ml. These isolates were as follows: C. albicans (n=2), C. glabrata (n=4), C. parapsilosis (n=19), C. krusei (n=2) and C. tropicalis (n=3). Compared to other isolates, higher MIC values were noted for C. albicans and C. glabrata (512 µg/ml), respectively. CONCLUSION: The latex of Euphorbia macroclada showed notable antifungal activities against some pathogenic Candida species. Therefore, it can be potentially used as an alternative antifungal agent in future. However, further research is required to identify its active components.

A survey of the etiological agents of scalp and nail dermatophytosis in Yazd, Iran in 2014-2015.

BACKGROUND AND PURPOSE: Tinea capitis and tinea unguium are regarded as global public health concerns. The purpose of the present study was to identify the etiological agents of tinea capitis and tinea unguium in patients, referring to the Central Laboratory of Yazd University of Medical Sciences, Yazd, Iran. MATERIALS AND METHODS: This study was conducted during 2014-2015. Skin scraping, scalp hair, and nail clipping specimens were collected from 134 patients (80 males and 54 females) with clinical features suggesting fungal involvement. Direct microscopic examinations were carried out, using potassium hydroxide 10%, while culture studies were performed on Sabouraud dextrose agar, containing chloramphenicol and cycloheximide at 28°C for four weeks. Fungal colonies were identified based on their macroscopic and microscopic characteristics, as well as supplementary diagnostic tests. RESULTS: Among 134 patients, 12 cases showed positive results on direct examination and culture studies. The frequency of infections was equal among male and female subjects. Among 12 affected cases, the frequency of tinea capitis and tinea unguium was 91.6% and 8.4%, respectively. Microsporum canis (50%) was the most prevalent species, followed by Trichophyton verrucosum (25%) and Trichophyton mentagrophytes (25%). Also, tinea unguium, caused by T. mentagrophytes, was found in a female patient. CONCLUSION: The etiological agents of scalp and nail dermatophytosis have changed in Yazd over the past 13 years. In the present study, replacement of anthropophilic dermatophytes by zoophilic species was noteworthy, highlighting the necessity of efficient surveillance for the management and prevention of infections.

A study on etiologic agents and clinical manifestations of dermatophytosis in Yazd, Iran.

BACKGROUND AND PURPOSE: Dermatophytosis is one of the most common infections of skin, hair, and nails, caused by a group of keratinophilic fungi known as dermatophytes. Species identification of these fungi is of great significance from epidemiological and therapeutic points of view. The objective of the present study was to investigate dermatophytosis and its causative agents in patients, referring to the Central Mycology Laboratory of Yazd University of Medical Sciences, Yazd, Iran. MATERIALS AND METHODS: In total, 139 clinically suspected cases of dermatophytosis were examined during 12 months from February 2014 to February 2015. Skin scrapings were assessed through direct microscopic examinations and culture studies. Dermatophyte isolates were identified based on colony morphology on potato dextrose agar and dermatophyte test medium, nutritional requirements, urease and hair perforation tests, and microscopic characteristics on slide cultures. RESULTS: Dermatophytosis was mycologically confirmed in 26 (18.70%) out of 139 cases. Although there was a statistically insignificant difference between male and female subjects, men were dominantly affected. Infection was significantly common in the age group of ≤ 29 years (P<0.043). The most common clinical manifestation of dermatophytosis was tinea corporis (69.2%), followed by tinea cruris (15.4%), tinea manuum (11.5%), and tinea pedis (3.8%). Trichophyton mentagrophytes complex was the main etiologic agent (38.5%), followed by T. rubrum (23%), T. violaceum (15.5%), T. verrucosum (11.5%), Microsporum canis (7.7%), and Epidermophyton floccosum (3.8%). CONCLUSION: In comparison with previous research, epidemiology of dermatophytosis has changed in Yazd over the past decades. Therefore, periodical investigations on the epidemiological aspects of this infection are required for efficient control and prevention of this cutaneous dermatophytic disease.

Development of RFLP-PCR method for the identification of medically important Aspergillus species using single restriction enzyme MwoI.

In this study we attempted to modify the PCR-RFLP method using restriction enzyme MwoI for the identification of medically important Aspergillus species. Our subjects included nine standard Aspergillus species and 205 Aspergillus isolates of approved hospital acquired infections and hospital indoor sources. First of all, Aspergillus isolates were identified in the level of species by using morphologic method. A twenty four hours culture was performed for each isolates to harvest Aspergillus mycelia and then genomic DNA was extracted using Phenol-Chloroform method. PCR-RFLP using single restriction enzyme MwoI was performed in ITS regions of rDNA gene. The electrophoresis data were analyzed and compared with those of morphologic identifications. Total of 205 Aspergillus isolates included 153 (75%) environmental and 52 (25%) clinical isolates. A. flavus was the most frequently isolate in our study (55%), followed by A. niger 65(31.7%), A. fumigatus 18(8.7%), A. nidulans and A. parasiticus 2(1% each). MwoI enabled us to discriminate eight medically important Aspergillus species including A. fumigatus, A. niger, A. flavus as the most common isolated species. PCR-RFLP method using the restriction enzyme MwoI is a rapid and reliable test for identification of at least the most medically important Aspergillus species.

Development of RFLP-PCR method for the identification of medically important Aspergillus species using single restriction enzyme MwoI

In this study we attempted to modify the PCR-RFLP method using restriction enzyme MwoI for the identification of medically important Aspergillus species. Our subjects included nine standard Aspergillus species and 205 Aspergillus isolates of approved hospital acquired infections and hospital indoor sources. First of all, Aspergillus isolates were identified in the level of species by using morphologic method. A twenty four hours culture was performed for each isolates to harvest Aspergillus mycelia and then genomic DNA was extracted using Phenol-Chloroform method. PCR-RFLP using single restriction enzyme MwoI was performed in ITS regions of rDNA gene. The electrophoresis data were analyzed and compared with those of morphologic identifications. Total of 205 Aspergillus isolates included 153 (75%) environmental and 52 (25%) clinical isolates. A. flavus was the most frequently isolate in our study (55%), followed by A. niger 65(31.7%), A. fumigatus 18(8.7%), A. nidulans and A. parasiticus 2(1% each). MwoI enabled us to discriminate eight medically important Aspergillus species including A. fumigatus, A. niger, A. flavus as the most common isolated species. PCR-RFLP method using the restriction enzyme MwoI is a rapid and reliable test for identification of at least the most medically important Aspergillus species.

Mycological Microscopic and Culture Examination of 400 Bronchoalveolar Lavage (BAL) Samples.

BACKGROUND: The frequency of invasive opportunistic mycoses has increased significantly over the past decades especially in immunocompromised patients. Invasive aspergillosis (IA) has become a major cause of morbidity and mortality among these patients. As bronchoalveolar lavage (BAL) fluid samples are generally useful specimens in the diagnosis of invasive pulmonary aspergillosis (IPA), this study was designed to evaluate the incidence of fungal elements in at-risk patients by direct microscopy and culture of BAL samples. METHODS: In a 16-month period, 400 BAL samples were obtained from several groups of different patients with pulmonary and respiratory disorders and examined by using both direct microscopy and culture. RESULTS: Of the 400 samples, 16 (4%) were positive direct examination with branching septate hyphae and 46 (11.5%) were positive culture: 25 (54%) Aspergillus flavus, 6 (13%) A. fumigatus, 5 (10.9%) A. niger, 1 (2.2%) A. terreus, 3 (6.5%) Penicillium spp. and 6 (13%) mixed A. flavus/A. niger. A. flavus was the most common cause of Aspergillus infection or colonization. Bone marrow transplant (BMT) recipients were the most susceptible group to fungal infection and/or colonization. CONCLUSION: Among Aspergillus species, A. flavus was the most common isolate in both infections and colonization in Iran. More studies are needed to clarify the epidemiological aspect of aspergillosis in Iran.

Candida parapsilosis as a Potent Biocontrol Agent against Growth and Aflatoxin Production by Aspergillus Species.

BACKGROUND: Aflatoxin contamination of food and feed stuff is a serious health problem and significant economic concerns. In the present study, the inhibitory effect of Candida parapsilosis IP1698 on mycelial growth and aflatoxin production in aflatoxigenic strains of Aspergillus species was investigated. METHODS: Mycelial growth inhibitions of nine strains of aflatoxigenic and non-aflatoxigenic Aspergillus species in the presence of C. parapsilosis investigated by pour plate technique at different pH, temperature and time of incubation. Reduction of aflatoxin was evaluated in co-cultured fungi in yeast extract sucrose broth after seven days of incubation using HPLC method. The data were analyzed by SPSS 11.5. RESULTS: The presence of the C. parapsilosis at different pH did not affect significantly the growth rate of Aspergillus isolates. On the other hand, temperature and time of incubation showed to be significantly effective when compared to controls without C. parapsilosis (P≤0.05). In aflatoxigenic strains, minimum percentage of reductions in total aflatoxin and B1, B2, G1, G2 fractions were 92.98, 92.54, 77.48, 54.54 and 72.22 and maximum percentage of reductions were 99.59, not detectable, 94.42, and not detectable in both G1 and G2, respectively. CONCLUSION: C. parapsilosis might employ as a good biocontrol agent against growth and aflatoxin production by aflatoxigenic Aspergillus species.

Molecular detection of invasive aspergillosis in hematologic malignancies.

BACKGROUND: Aspergillus species are the most frequent causes of invasive mold infections in immunocompromised patients, particularly those who underwent chemotherapy for hematologic malignancies. The aim of this study was to determine the incidence and efficiency of the PCR-enzyme linked immunosorbent assay method (PCR-ELISA) for early detection of Aspergillus species in patients with hematologic malignancies. PATIENTS AND METHODS: From 2004 to 2006, 194 patients with hematologic malignancies (who received chemotherapy) were evaluated for invasive aspergillosis (IA) in Shiraz, southern Iran. Ethylenediaminetetraacetic acid anticoagulant whole blood samples were collected prospectively once a week and stored at -20 degrees C until examination. All collected blood samples were assayed for the presence of the bands on ethidium bromide stained gel and for hybridization. RESULTS: The female-to-male ratio was 61:133, the mean age of patients was 33.7 years, and mean of hospitalization period was 21.2 days. PCR-ELISA was positive in 14 (7.2%) patients who exhibited clinical and radiologic signs of IA. The etiologic agents were Aspergillus flavus (11 cases) and Aspergillus fumigatus (three cases). The mean time of positivity of PCR-ELISA in the blood before the appearance of clinical signs was 12.6 days. PCR was found to be the earliest indicator of IA preceding nonspecific clinical and radiologic findings. The sensitivity, specificity, positive, and negative predictive values of PCR-ELISA to detect DNA-specific for Aspergillus species in patients with proven and probable IA were 66%, 96%, 62.5%, and 97%, respectively. In case patients were treated with antifungal drugs, and the treatment was successful, fungal PCR assay became negative after 14 days and if the treatment failed, assay was positive until death. CONCLUSIONS: We demonstrated, in the present study, the incidence of IA in leukemic patients and the usefulness of molecular assay for early diagnosis and monitoring of the treatment of IA.