Background and aims: Mucorales are fungi belonging to the category of Zygomycetes, found much in nature. Culture-based methods for clinical samples are often negative, difficult and time-consuming and mainly identify isolates to the genus level, and sometimes only as Mucorales. Therefore, applying fast and accurate diagnosis methods such as molecular approaches seems necessary. This study aims at isolating Mucorales for determination of Rhizopus genus between the isolates using molecular methods. Methods: In this descriptive observational study, a total of 500 samples were collected from air and different surfaces and inoculated on Sabouraud Dextrose Agar supplemented with chloramphenicol. Then, the fungi belonging to Mucorales were identified and their pure culture was provided. DNA extraction was done using extraction kit and the chloroform method. After amplification, the samples belonging to Mucorales were identified by observing 830 bp bands. For enzymatic digestion, enzyme BmgB1 was applied for identification of Rhizopus species by formation of 593 and 235 bp segments. Results: One hundred pure colonies belonging to Mucorales were identified using molecular methods and after enzymatic digestion, 21 isolates were determined as Rhizopus species. The sequencing of PCR products and macroscopic and microscopic studies confirmed the existence of R. stolonifera, R. oryzae and R. caespitosus in the samples. Conclusion: Generally, developing a reliable method for determining Zygomycete species can be a useful tool for better understanding of the epidemiology of mucoromycosis.

Background: Trichostrongylus is an intestinal parasite that is highly prevalent in humans and livestock worldwide. There is limited information about the prevalence and epidemiology of Trichostrongylus species among the infected livestock in Mazandaran Province, northern Iran. This study aimed to identify Trichostrongylus spp. among small ruminants using morphometric and molecular methods. Materials and Methods: Small intestinal organs of sheep and goats, slaughtered in Mazandaran Province, were examined for infectivity with Trichostrongylus parasites. Primary species identification was conducted based on the morphological characterization of the male worms. The internal transcribed spacer (ITS) II regions of the ribosomal DNA of the worm tissues were amplified using the polymerase chain reaction (PCR) assay and then the product was subjected to sequencing. Subsequently, the PCR products of the ITS II region were subjected to digestion by HinfI and DraI restriction enzymes using the PCR-restriction fragment length polymorphism (RFLP). Results: Of 180 samples, 98 (54. 44%) were confirmed positive for Trichostrongylus based on the conventional PCR. The digestion of the PCR products with HinfI and DraI facilitated the identification of three Trichostrongylus species, namely Trichostrongylus colubriformis (35%, 90. 81%), Trichostrongylus axei (4%, 4. 08%), and Trichostrongylus vitrinus (5%, 5. 1%). Both morphometric and RFLP techniques resulted in the differentiation of the three Trichostrongylus species. Conclusion: The present study was the 1st attempt in the last 30 years for the identification of Trichostrongylus species in small ruminants in Mazandaran Province. The findings of this study can be helpful for epidemiological and ecological studies, the establishment of effective control programs, and the management of gastrointestinal parasites in Mazandaran Province.

Background: In 2009, 3 genome-wide association studies implicated IL28B single-nucleotide polymorphisms (SNPs) as the strongest genetic pretreatment predictor of sustained virological response (SVR) in hepatitis C infection. Recently, the American Association for the Study of Liver Diseases (AASLD) and the European Association for the Study of the Liver (EASL) included IL28B testing in their guidelines.Objectives: The main aim of this study was to develop and validate a simple, rapid, and inexpensive polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for genotyping of common IL28B polymorphisms (rs12979860 and rs8099917).Patients and Methods: Two methods were developed to genotype common IL28B polymorphisms: 1) PCR-sequencing as a reference method and 2) PCR-RFLP as a rapid and inexpensive method. Both polymorphisms were genotyped in 104 Iranian hepatitis C patients by both methods simultaneously. To validate the PCR-RFLP method, the PCR-RFLP genotyping results should be 100% concordant with the PCR-sequencing results.Results: Genotyping of rs12979860 and rs8099917 by PCR-RFLP was concordant with PCR-sequencing in 104 (100%) individuals. The analytical sensitivity and specificity of the PCR-RFLP method for genotyping of both SNPs are 100%. Among these 104 patients with chronic hepatitis C, the frequency of the rs12979860 CC, CT and TT genotypes were 40.4%, 47.1% and 12.5% and the frequency of the rs8099917 TT, GT and GG genotypes were 59.6%, 35.6% and 4.8%, respectively. Also, three IL28B haplotypes (rs12979860-rs8099917) were found among our patients including C-T, T-G and T-T with 63.9%, 22.6% and 13.5% frequency, respectively. C-G haplotype was absent in all of our patients.Conclusions: We have developed a validated, fast, and simple PCR-RFLP method for genotyping of common IL28B SNPs that is more cost-effective than sequencing.

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 μ g/ml) against all Candida strains, whereas fluconazole (0. 063-64 μ g/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 μ g/ml, while 38. 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 μ g/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 nonalbicans 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.