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Introduction: The skin plays a crucial role as a protective barrier against external factors, but disruptions to its integrity can lead to wound formation and hinder the natural healing process. Scar formation and delayed wound healing present significant challenges in skin injury treatment. While alternative approaches such as skin substitutes and tissue engineering exist, they are often limited in accessibility and cost. Exosomes have emerged as a potential solution for wound healing due to their regenerative properties. Methods: In this study, exosomes were isolated from human blood serum using a kit. The exosomes were characterized, and their effects on cell migration were assessed in vitro. Additionally, the wound healing capacity of exosomes was evaluated in vivo using a rat full-thickness wound model. Results: Our in vitro findings revealed that exosomes significantly promoted cell migration. In vivo experiments demonstrated that the injection of exosomes at different areas of the wound accelerated the wound healing process, resulting in wound closure, collagen synthesis, vessel formation, and angiogenesis in the wound area. These results suggest that exosomes have a promising therapeutic potential for expediting wound healing and minimizing scar formation. Conclusions: The findings of this study highlight the potential of exosomes as a novel approach for enhancing wound healing. Exosomes showed positive effects on both cell migration and wound closure in in vitro and in vivo studies, suggesting their potential use as a regenerative therapy for skin injuries. Further research is needed to fully understand the mechanisms underlying the beneficial effects of exosomes on wound healing and to optimize their application in clinical settings.

BackgroundIdentification of the opportunistic fungus Pneumocystis jirovecii in respiratory specimens presents challenges, particularly in differentiating between colonization and active infection. The present study assessed a probe-based real time PCR (qPCR) diagnostic effectiveness in patients with diverse underlying conditions, particularly those with COVID-19 and pulmonary insufficiency.MethodsTo set up the qPCR, clinical samples from 281 patients with respiratory ailments were tested. Subsequently, a descriptive study was conducted on 112 patients with pulmonary insufficiency with and without COVID-19 suspected of P. jirovecii infection. All specimens were subjected to DNA extraction followed by nested PCR and qPCR targeting the mitochondrial large subunit (mtLSU)-rRNA gene.ResultsBased on nested PCR and qPCR, P. jirovecii was identified in 40 out of 281 patients, with slight variations in positive samples observed across dilutions. Three patients who tested positive in nested PCR yielded negative results in probe-based qPCR. Conversely, three patients who tested positive in probe-based qPCR yielded negative results in nested PCR. Considering nested PCR as the golden standard, probe-based qPCR demonstrated good diagnostic performance, with 92.5% sensitivity and 98.7% specificity. Based on cycle threshold (Ct) values, the positive cases were categorized: ≤32 as infection, >35 as colonization, and a grey zone between these values (32 < X ≤ 35). The analysis of 112 PCP-suspected patients revealed a prevalence ranging from 6.25% (nested PCR) to 7% (probe-based qPCR).ConclusionsThis study suggested Ct values to differentiate Pneumocystis pneumonia/colonization in immunocompromised patients. To further augment the diagnostic sensitivity, it is recommended to integrate qPCR results with clinical parameters and biomarkers to offer a more precise understanding of Pneumocystis-related conditions.

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly contagious virus that uses angiotensin converting enzyme 2 (ACE2), a pivotal member of the renin–angiotensin system (RAS), as its cell-entry receptor. Another member of the RAS, angiotensin II (Ang II), is the major biologically active component in this system. There is growing evidence suggesting that serum miRNAs could serve as prognostic biomarkers for SARS-CoV-2 infection and regulate ACE2 expression. Therefore, the aim of this study is to evaluate the changes in the serum levels of sACE2 and Ang II, as well as the expression level of miR-141-3p and miR-421 in SARS-CoV-2 positive and negative subjects. Methods In the present study, the serum levels of sACE2 and Ang II were measured in 94 SARS-CoV-2 positive patients and 94 SARS-CoV-2 negative subjects with some symptoms similar to those of SARS-CoV-2 positive patients using the ELISA method. In addition, the expression level of miR-141-3p and miR-421 as ACE2 regulators and biomarkers was evaluated using quantitative real-time PCR (qRT-PCR) method. Results The mean serum sACE2 concentration in the SARS-CoV-2-positive group was 3.268 ± 0.410 ng/ml, whereas in the SARS-CoV-2 negative group, it was 3.564 ± 0.437 ng/ml. Additionally, the mean serum Ang II level in the SARS-CoV-2 positive and negative groups were 60.67 ± 6.192 ng/L and 67.97 ± 6.837 ng/L, respectively. However, there was no significant difference in the serum levels of sACE2 (P value: 0.516) and Ang II (P value: 0.134) between the SARS-CoV-2 positive and negative groups. Meanwhile, our findings indicated that the expression levels of miR-141-3p and miR-421 in SARS-CoV-2 positive group were significantly lower and higher than SARS-CoV-2 negative group, respectively (P value

IntroductionAlthough the existence of Candida species in the respiratory tract is often considered commensal, it is crucial to recognize the significance of Candida colonization in immunocompromised or COVID-19 patients. The emergence of Candida auris as an emerging pathogen further emphasizes the importance of monitoring yeast infection/colonization, particularly in COVID-19 patients.MethodsIn this study, respiratory samples mainly from COVID-19 patients, primarily those suspected of having a fungal infection, were cultured on Sabouraud dextrose agar plates and the yeast colonies were identified using a two-step multiplex PCR method. The samples suspected of C. auris underwent specific nested PCR followed by sequence analysis.ResultsA total of 199 respiratory samples were collected from 73 women and 126 men, ranging in age from 1.6 to 88 years. Among the patients, 141 had COVID-19, 32 had cancer, 5 were hospitalized in ICU, 2 had chronic obstructive pulmonary disease)COPD(, and others were patients with combination diseases. From these samples, a total of 334 yeast strains were identified. C. albicans (n=132, 39.52%) was the most common species, followed by C. tropicalis (n=67, 20%), C. glabrata (n=56, 16.76%), C. krusei (n=18, 5.4%), C. parapsilosis (n=17, 5.08%), Saccharomyces cerevisiae (n=10, 3%), C. kefyr (n=9, 2.6%), C. dubliniensis (n=7, 2.1%), C. lusitaniae (n=5, 1.5%), C. auris (n=3, 0.9%), C. guilliermondii (n=2, 0.6%), C. rugosa (n=1, 0.3%), C. intermedia (n=1, 0.3%), and Trichosporon spp. (n=1, 0.3%). C. auris was detected in a patient in ICU and two COVID-19 patients. While its presence was confirmed through sequence analysis, our extensive efforts to isolate C. auris were unsuccessful.ConclusionWhile C. albicans colonization remains prevalent, our study found no evidence of Candida lung infection. Since the role of Candida colonization in airway secretions remains ambiguous due to limited research, further studies are imperative to shed light on this matter.

Candida auris has emerged globally as a multidrug-resistant pathogen causing outbreaks in health care facilities. Whole genome sequencing (WGS) analysis has identified four major clades, while earlier WGS data from a single Iranian isolate suggested the existence of a potential fifth clade. Here, we confirm the existence of this fifth clade by providing WGS data of another four Iranian isolates. These clade V isolates differed less than 100 single-nucleotide polymorphisms (SNPs) between each other, while they were separated from the other clades by more than 200,000 SNPs. Two of these isolates were resistant to fluconazole and were found to harbour mutations in the TAC1b and ERG11 genes.

Candidemia caused by rare and uncommon Candida species is becoming more prevalent in pediatric healthcare settings, resulting in significant morbidity and mortality. One such species, Candida palmioleophila, is resistant to fluconazole but highly susceptible to echinocandins. Here, we report the first documented case of C. palmioleophila candidemia in Iran that occurred in a male infant with biliary atresia who had been hospitalized for 2 months. The patient’s blood and urine cultures were positive for both yeast and bacterial species. Through DNA sequence analysis, the yeast isolate was identified as C. palmioleophila. In vitro antifungal susceptibility testing of the isolate against amphotericin B, fluconazole, itraconazole, voriconazole, isavuconazole, posaconazole, and nystatin revealed MIC values of 2, 16, 0.25, 0.0625, 0.125, 0.25, and 4 μg/mL, respectively, and minimum effective concentration for caspofungin was 0.031 μg/mL. Despite receiving antibacterial and antifungal therapies, the patient unfortunately expired due to bradycardia and hypoxemia. Proper identification and epidemiological surveillance studies are needed to understand the exact prevalence of these emerging yeast pathogens. Previously reported cases of C. palmioleophila infection, primarily associated with bloodstream infections and catheter-related candidemia, were reviewed.

BackgroundDeveloping more sensitive methods for the diagnosis of echinococcosis is essential. In this study PCR assay for sensitive detection of specific cell-free DNA (cfDNA) of Echinococcus granulosus sensu lato in the sera of the sheep naturally infected with echinococcosis was investigated.MethodsTo extract cfDNA from 35 infected sheep, the modified phenol-chloroform method was used for two different volumes (0.5 and 2 ml) of serum samples. From each extracted sample, two DNA volumes (5 and 10 μl) were amplified using both standard PCR and semi-nested PCR targeting NADH dehydrogenase subunit I.ResultsStandard and semi-nested PCR on 0.5 ml of serum samples detected Echinococcus DNA in 8 and 12 out of 35 sheep, respectively; however, using 2 ml of serum samples, they detected 24 and 27 samples. By increasing the volume of template DNA, the PCRs could detect 29 and 33 out of 35 samples. The results were confirmed by sequencing of randomly selected PCR amplicons and comparing them with GenBank databases.ConclusionsLarger volumes of serum for DNA extraction, greater volumes of DNA template for PCR, and employing a semi-nested PCR protocol, increased the sensitivity of PCR to 95%. This approach can also be applied to the diagnosis of echinococcosis in humans.

Invasive fungal rhinosinusitis (IFRS) is a life-threatening infection that can occur in immunocompromised patients, including those with COVID-19. Although Mucorales and Aspergillus species are the most common causes of IFRS, infections caused by other fungi such as Fusarium are rare. In this report, we present three cases of proven rhinosinusitis fusariosis that occurred during or after COVID-19 infection. The diagnosis was confirmed through microscopy, pathology, and culture, and species identification of the isolates was performed by DNA sequencing the entire ITS1-5.8 rRNA-ITS2 region and translation elongation factor 1-alpha (TEF-1α). Antifungal susceptibility testing was conducted according to CLSI guidelines. The causative agents were identified as Fusarium proliferatum, F. oxysporum + Aspergillus flavus, and F. solani/falciforme. Treatment involved the administration of antifungal medication and endoscopic sinus surgery to remove the affected mucosa, leading to the successful resolution of the infections. However, one patient experienced a recurrence of IFRS caused by A. flavus 15 months later. Early diagnosis and timely medical and surgical treatment are crucial in reducing mortality rates associated with invasive fusariosis. Additionally, the cautious use of corticosteroids in COVID-19 patients is highly recommended.

Candida auris, a multidrug-resistant yeast, can cause primary or secondary infections in a wide range of patients, including those diagnosed with the new coronavirus to even healthy individuals. The fungus has been reported in less than a decade on all six continents and in more than 45 countries. Ease of distribution, long shelf life, and resistance to several antifungal drugs have raised concerns about the prevention and management of patients with C. auris infection. Recent reports indicate serious challenges in identifying, understanding the mechanism of drug resistance, and preventing mortality from the infection with this microorganism. Given the prevalence of COVID-19 infection, it is important to identify patients colonized with C. auris correctly and at the early stages, to control and prevent a possible outbreak. In this article, the widespread occurrence of infections due to C. auris in the world and Iran, its clinical manifestations, risk factors, pathogenic mechanisms, diagnostic enhancements and challenges, drug resistance, treatment options, prevention, and control as well as concomitant C. auris infections in patients with COVID-19 virus, are reviewed.

A high-quality genome for the parasitic tapeworm, Echinococcus granulosus, provides further insight into the EG95 vaccine target for cystic echinococcosis.

IntroductionEarly detection of Pneumocystis jirovecii as an opportunistic pathogen that may endanger predisposed persons, including COVID-19 patients, may help to choose the optimal management.MethodsIn this study, 585, including 530 COVID-19 patients, with clinical and radiological evidence of respiratory diseases, were investigated for P. jirovecii screening. Clinical specimens were examined by direct microscopy and PCR, and randomly selected positive PCR products were confirmed through DNA sequence analysis.ResultsThirty-one (5.3%) samples were positive in P. jirovecii-specific nested-PCR, while by direct microscopic tests, Pneumocystis was observed in 22 (3.76%) samples. Males (61.7%) and patients over 50 years old (75.6%) were more commonly affected than others, and malaise and fatigue (84%), and wheezing (75%) were the most common symptoms, followed by fever (40.48%) and dyspnea (39.51%). Among the Pneumocystis-positive patients, three cases had coinfection with Aspergillus fumigatus, A. flavus, and A. niger (each n = 1), as documented by direct microscopy, culture, and species identification by PCR-sequencing.ConclusionPneumocystis pneumonia is still a diagnostic challenge; therefore, additional large-scale studies are needed to clarify the epidemiology of the disease in immunocompromised or COVID-19 patients.

Along with the pandemic COVID-19 spreads, new clinical challenges have emerged in the health care settings, among which there is a high risk of secondary invasive fungal infections with significant mortality. Here, we report a case of invasive fungal rhino orbital sinusitis due to the simultaneous co-infection by Rhizopus oryzae and Lomentospora prolificans, both identified by sequencing, in a 70-year-old Afghanistanian female with COVID-19. The patient was subjected to surgical debridement as well as taking liposomal amphotericin B, voriconazole, and on discharge, her condition was good. As far as we know, this is the first case of co-infection of COVID-19-associated mucormycosis (CAM) and Lomentospora prolificans infection. Multiple fungal co-infections in COVID-19 patients are reviewed.

Abstract Harmless commensal Candida species, especially uncommon and rare ones may rarely cause a serious infection. Candida metapsilosis is a recently described yeast that is phenotypically indistinguishable from Candida parapsilosis and molecular methods are essential for its identification. We report the first case of Candida conjunctivitis due to C. metapsilosis obtained from the eye discharge of a 40‐day‐old girl with congenital heart disease admitted to the cardiac intensive care unit (CICU). The yeast isolate was identified by sequencing the entire ITS1‐5.8 rRNA‐ITS2 region. Antifungal susceptibility testing performed according to the CLSI M27‐A3 showed that the isolate was susceptible to amphotericin B, fluconazole, itraconazole, voriconazole, clotrimazole, nystatin, terbinafine, 5‐fluorocytosine, and caspofungin. Differentiation of the fungal new species allows us the accurate diagnosis and treatment, and a better understanding the microbial epidemiology.

During replication, some mutations occur in SARS-CoV-2, the causal agent of COVID-19, leading to the emergence of different variants of the virus. The mutations that accrue in different variants of the virus, influence the virus' ability to bind to human cell receptors and ability to evade the human immune system, the rate of viral transmission, and effectiveness of vaccines. Some of these mutations occur in the receptor binding domain (RBD) of the spike protein that may change the affinity of the virus for the ACE2 receptor. In this study, several in silico techniques, such as MD and SMD simulations, were used to perform comparative studies to deeply understand the effect of mutation on structural and functional details of the interaction of the spike glycoprotein of SARS-CoV-2, with the ACE2 receptor. According to our results, the mutation in the RBD associated with the Omicron variant increase binding affinity of the virus to ACE2 when compared to wild type and Delta variants. We also observed that the flexibility of the spike protein of the Omicron variant was lower than in comparison to other variants. In summary, different mutations in variants of the virus can have an effect on the binding mechanism of the receptor binding domain of the virus with ACE2.

Overcoming drug resistance and specifically targeting cancer stem cells (CSCs) are critical challenges in improving cancer therapy. Nowadays, the use of novel and native medicinal plants can provide new sources for further investigations for this purpose. The aim of this study was to assess the potential of S. bachtiarica, an endemic plant with diverse medicinal applications, in suppressing and targeting cancer and cancer stem cells in glioblastoma and breast cancer. The effect of S. bachtiarica on viability, migration, invasion, and clonogenic potential of MDAMB-231 and U87-MG cells was assessed in both two- and three-dimensional cell culture models. Additionally, we evaluated its effects on the self-renewal capacity of mammospheres. The experimental outcomes indicated that S. bachtiarica decreased the viability and growth rate of cells and spheroids by inducing apoptosis and inhibited colony formation, migration, and invasion of cells and spheroids. Additionally, colony and sphere-forming ability, as well as the expression of genes associated with EMT and stemness were reduced in mammospheres treated with S. bachtiarica. In conclusion, this study provided valuable insights into the anti-cancer effects of S. bachtiarica, particularly in relation to breast CSCs. Therefore, S. bachtiarica may be a potential adjuvant for the treatment of cancer.

Background and Purpose: Given the important role of Malassezia spp. in skin diseasesand other associated infections in neonates, this study aimed to investigate the presenceand frequency of Malassezia spp. in the skin of neonates hospitalized in neonatalintensive care units and their mothers using culture and accurate molecular-basedmethods.Materials and Methods: In total, 205 samples were collected from 130 neonates (>4-day-old) and 75 mothers. Isolation of Malassezia spp. from the skin was performed usingLeeming-Notman agar and modified Dixon agar media. To compare the Malasseziamicroflora on the skin of the neonates and their mothers, a polymerase chain reactionsequencing method was performed for spp. identification of 92 isolates obtained fromneonates and their mothers. Moreover, possible associated risk factors for thecolonization of Malassezia spp. on the skin were recorded.Results: Cultures from 62.3% of neonates and 77.3% of mothers were positive forMalassezia spp. growth. Malassezia globosa was the most prevalent isolated spp. foundin the skin of the study population. It is noteworthy that a rare Malassezia spp.,Malassezia arunalokei, was isolated from the skin of one neonate. There was a 76%similarity between the mother-neonate isolate sequences results. The statistical analysisshowed that the type of feeding is a significant (P

Background and Purpose: The frequency and genetic diversity of black fungi in environmental and clinical settings have not been fully studied in Iran. This study aimed to identify and evaluate intra- and inter-species DNA sequence variation and also understand the phylogenetic relationships of melanized fungi and relatives isolated from different geographical regions of Iran.Materials and Methods: In total, 111 clinical and environmental strains of dematiaceous fungi were isolated, and their internal transcribed spacer ribosomal DNA(rDNA) regions were sequenced and analyzed.Results: An inter-species nucleotide sequence diversity rate of 1 to 464 nucleotides was observed between the species. Intra-species differences were found in the strains of Alternaria alternata, Cladosporium cladosporioides, Alternaria tenuissima, Curvularia spicifera, Aureobasidium pullulans, Curvularia hawaiiensis, Neoscytalidium dimidiatum,Alternaria terricola, Alternaria chlamydospora, Didymella glomerata, and Drechslera dematioidea by 0–59, 0–22, 0–4, 0–4, 0–3, 0–2, 0–2, 0–2, 0–2, 0–1, and 0–1 nt, respectively.Conclusion: The internal transcribed spacer rDNA is useful for the discrimination of several taxa of dematiaceous fungi. However, a better understanding of the taxonomy of species of Alternaria requires a larger rDNA region or a library of other gene sequences.

A fatal case of COVID-19-associated mucormycosis (CAM) affected a 40-year-old woman who was initially admitted to our hospital due to a SARS-CoV-2 infection. Her clinical condition worsened, and she finally died because of respiratory failure, hemodynamic instability, and mucormycosis with invasion into the orbit and probably the brain. According to DNA sequence analysis of the fungus isolated from the patient, Apophysomyces variabilis was involved. This is the first published case of CAM and the third case of mucormycosis due to this mold.

Background and Purpose: The most common etiological agents of human dermatophytosis in various parts of the world are Trichophyton rubrum, Trichophyton interdigitale, and Epidermophyton floccosum. The main aim of this study was to design and evaluate a simple and straightforward multiplex polymerase chain reaction (PCR)assay for reliable identification/differentiation of these species in clinical isolates.Materials and Methods: The reliable sequences of several molecular targets of dermatophytes species were used to design a multiplex PCR for the identification of common pathogenic dermatophytes. The isolates and clinical specimens examined in this study included seven standard strains of dermatophytes, 101 isolates of dermatophytes and non-dermatophyte molds/yeasts which had already been identified by sequencing or PCR-restriction fragment length polymorphism (RFLP), and 155 clinical samples from patients suspected of cutaneous mycoses.Results: Species-specific primer pairs for T. rubrum and T. interdigitale/T. mentagrophytes were designed based on the sequence data of the translation elongation factor 1-alpha gene, and the primers for E. floccosum targeted the specific sequence of the internal transcribed spacer region (ITS). The multiplex PCR successfully detected T.rubrum, T. interdigitale/T. mentagrophytes, and E. floccosum strains that were identified by sequencing or PCR-RFLP. However, the primer pairs selected for T. interdigitale/T. mentagrophytes cross-reacted with Trichophyton tonsurans. In testing the PCR system directly for clinical samples, the proportion of positive multiplex PCR was higher than positive culture (68.1% vs. 55.4%, respectively).Conclusion: The multiplex assay could detect three common agents out of several causal agents of dermatophytosis, namely T. rubrum, T. interdigitale, and E. floccosum.Therefore, by adding pan-dermatophyte primers it can be used as a comprehensive detection/identification test.

Background and Purpose: The taxonomy of Candida is controversial and has undergone changes due to the investigation of the novel species. Candida africana and Candida dubliniensis are new members of C. albicans complex that are currently gaining both clinical and epidemiologic significance. This study reports the prevalence of C. africana among the strains isolated from patients, by using HWP1 gene size polymorphism.Materials and Methods: A total of 235 yeasts confirmed as C. albicans complex based on chromogenic media and ITS-sequencing isolated from various clinical forms of invasive and non-invasive candidiasis mainly candidemia, were re-identified based on HWP1 gene polymorphisms. The Hwp1-PCR amplicons were re-confirmed by sequencing and BLAST analysis.Results: Based on the HWP1 gene size polymorphism, 223 strains were identified as C. albicans (94.89%) from which 7 isolates produced two DNA fragments (850 and 941 bp). C. dubliniensis (n=4, 1.7%), C. africana (n=1, 0.42%) and mix of C. albicans and C. africana (n=7, 2.97%) were also identified. Conclusion: Although C. albicans remains the most common Candida species, C. dubliniensis and C. africana are rarely found among the patient isolates. Due to limited information on the molecular epidemiology of this novel yeast, more studies using molecular methods are recommended.

The use of by-products from the agri-food industry is a promising approach for production of value-added, polyphenol-rich dietary supplements or natural pharmaceutical preparations. During pistachio nut processing, a great amount of husk is removed, leaving large biomass for potential re-use. The present study compares antiglycative, antioxidant, and antifungal activities as well as nutritional values of 12 genotypes belonging to four pistachio cultivars. Antioxidant activity was measured using DPPH and ABTS assays. Antiglycative activity was evaluated as inhibition of advanced glycation end product (AGE) formation in the bovine serum albumin/methylglyoxal model. HPLC analysis was performed to determine the major phenolic compounds. Cyanidin-3-O-galactoside (120.81–181.94 mg/100 g DW), gallic acid (27.89–45.25), catechin (7.2–11.01), and eriodictyol-7-O-glucoside (7.23–16.02) were the major components. Among genotypes, the highest total flavonol content (14.8 mg quercetin equivalents/g DW) and total phenolic content (262 mg tannic acid equivalent/g DW) were in KAL1 (Kaleghouchi) and FAN2 (Fandoghi), respectively. The highest antioxidant (EC50 = 375 μg/mL) and anti-glycative activities were obtained for Fan1. Furthermore, potent inhibitory activity against Candida species was recorded with MIC values of 3.12–12.5 µg/mL. The oil content ranged from 5.4% in Fan2 to 7.6% in Akb1. The nutritional parameters of the tested cultivars were highly variable: crude protein (9.8–15.8%), ADF (acid detergent fiber 11.9–18.2%), NDF (neutral detergent fiber, 14.8–25.6%), and condensed tannins (1.74–2.86%). Finally, cyanidin-3-O-galactoside was considered an effective compound responsible for antioxidant and anti-glycative activities.

مقدمه: شیوع بیماری Coronavirus disease-2019 (COVID-19) ناشی از ویروس Severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) دسته‌ی کروناویروس‌ها موسوم به 2019-nCoV، از اواخر سال ۲۰۱۹ میلادی در چین و در پی آن در دیگر کشورها، هنوز موضوع اصلی و مورد توجه در همه‌ی مجامع علمی و اجتماعی دنیا می‌باشد. با توجه به نوظهور بودن این ویروس، کسب اطلاعاتی درمورد تشابه یا تفاوت‌های آن با سایر ویروس‌های این خانواده، ویژگی‌های ژنتیک، چرخه‌ی بیولوژیک و چگونگی عملکرد پاتولوژیک آن، می‌تواند کمک شایانی در اتخاذ راهبردهای دقیق‌تر و بازدهی بالاتر در شناخت، کنترل و درمان بیماری حاصل از آن داشته باشد. روش‌ها: در این مطالعه‌ی مروری نقلی، تلاش شده است تا با جمع‌بندی مختصری از گزارش‌های متعدد علمی از دسامبر 2019 تا آپریل 2020 در زمینه‌ی مشخصات ویروس، ویژگی‌های ژنوم آن و ارزیابی با شواهد علمی از پیش موجود در زمینه‌ی کروناویروس‌ها، راهکارهای پیش رو در پاتوژنز، درمان و طراحی و تولید واکسن علیه آن ارایه شود. یافته‌ها: واکاوی و مقایسه‌ی ژنوم کروناویروس جدید با دیگر اعضای این خانواده، نشان دهنده‌ی شباهت زیاد بخش‌های مختلف ژنوم با انواع بیماری‌زای قبلی SARS و (Middle East Respiratory Syndrome) MERS می‌باشد و منشأگیری آن از گونه‌هایی با میزبان خفاش را نشان می‌دهد. نتیجه‌گیری: با توجه به اضطرار شناخت و معرفی راهکارهای درمانی و پیش‌گیرانه در پاندمی حاضر، شناخت مکانیسم‌های مولکولی درگیر در تکثیر و عفونت‌زایی ویروس بر پایه‌ی شباهت ژنوم آن با دیگر هم‌خانواده‌های کرونا ویروسی و همچنین، ویروس‌های دیگر، امکان پیش‌بینی راهبردهای فوری و امیدبخش را در مواجهه با ویروس فراهم می‌آورد.

Background: Candida species cause a wide spectrum of disease entities. Candida africana and Candida dubliniensis–members of Candida albicans complex-are currently gaining both clinical and epidemiologic significance. Materials and methods: Totally, 150 pediatric isolates that had previously been identified as C. albicans species complex based on a positive germ tube test were included. The isolates were cultured on CHROMagar Candida medium to ensure their purity and the results of germ tube test. For definitive speciation, PCR amplification and size polymorphism of the hyphal wall protein 1 (HWP1) gene was used. The results of HWP1-PCR were confirmed by sequencing the amplified fragments for randomly selected isolates of C. africana and C. dubliniensis. Results: All 150 isolates included in this study were reconfirmed as C. albicans complex on chromogenic media. Based on the HWP1 gene size polymorphism, 141 (94%) isolates were identified as C. albicans, 2 (1.33%) as C. africana, and 1 (0.67%) as C. dubliniensis. The remaining 6 (4%) C. albicans complex isolates were a mix of C. albicans and C. africana. All isolates of C. africana and C. dubliniensis have been recovered from cases of candiduria. Conclusion: C. africana, either alone or mixed with C. albicans, could be a cause of candiduria among pediatric patients and should not be ignored.

Background and Purpose: Invasive aspergillosis (IA) of the central nervous system (CNS) is a devastating complication which is rarely reported in immunocompromised children. In this case presentation, we reported a rare and fatal IA with spinal cord involvement in a 10-year-old child with X-linked chronic granulomatosis disease (CGD). Case report: The child had a previous history of pulmonary tuberculosis. A cervical spine X-ray revealed the involvement of cervical vertebrae (T4/T5) and ribs causing spinal cord compression and epidural abscess. The patient underwent a decompressive laminectomy and mass removal. The histopathology and culture results suggested IA. Despite the aggressive and prolonged therapy, he died within one year. Aspergillus nidulans was identified as the causative agent based on morphological and molecular studies. Conclusion: This synopsis represents the aggressive behavior of infection caused by A. nidulans in the CGD patient.

Background and Purpose: Culture-based identification methods have been the gold standard for the diagnosis of candidal onychomycosis. Molecular technologies, such as polymerase chain reaction (PCR) assays, can provide an alternative for the rapid detection of Candida species. The present study was conducted to investigate a pan-Candida PCR assay based on the translation elongation factor 1-alpha (TEF-1α) gene for the detection of the most prevalent pathogenic Candida species. Materials and Methods: For the purpose of the study, an optimized pan-Candida PCR primer pair was designed, and the target was amplified and sequenced. The analytical and clinical diagnostic performance of the designed primers was tested using 17 reference strains, 137 nail scrapings suspected of onychomycosis, and 10 healthy nail specimens. Results: The use of the universal pan-Candida primers designed on TEF-1α gene resulted in the successful amplification of a 270-base pair fragment in all Candida species tested, except for C. glabrata, and reacted neither with other fungi nor with E. coli. The sequence difference count matrix showed poor insertion/deletion differences (0-2 nt) among Candida species. Among 137 nail specimens, 35% (n=48), 30.7% (n=42), and 40.1% (n=55) of the samples were found to be positive by direct microscopy, culture, and pan-Candida PCR, respectively. Conclusion: Based on the findings, the PCR-based detection targeting the DNA TEF-1α gene is a rapid and simple procedure for the diagnosis of candidal onychomycosis directly from nail sample.

Background: Dermatophytes create the most common fungal disease in humans, called dermatophytosis. The two species of Trichophyton rubrum and Trichophyton interdigital are responsible for over 80% of types of dermatophytosis. So far, several morphological and physiological methods have been used to differentiate these very similar species, but these methods are generally time-consuming and have low specificity. The purpose of this study was to introduce a simple and rapid duplex polymerase chain reaction (PCR) reaction to differentiate these two species from each other. Methods: This research was an analytical and experimental study that was carried out from 2017 to 2018 in the Medical Mycology Laboratory, School of Public Health, Tehran University of Medical Sciences, Iran. For this purpose, the nucleotide sequences of the 4 regions of internal transcribed spacer (ITS), beta-tubulin, elongation factor 1 alpha and calmodulin in the two considered species of fungi were conducted bioinformatics analysis. The differences and similarities of nucleotides between two species in each of these genes were studied for selecting the primer. The specificity of selected primers was tested for duplex PCR reaction against sequenced isolates of dermatophyte species. Results: According to the total data, the specific primers were selected from elongation factor 1 alpha gene. These primers produced a product of 173 and 384 bp, in Trichophyton rubrum and Trichophyton interdigital, respectively. They had high specificity in the face of various dermatophytes. The length of nucleotide sequences found in the genebank of this gene in the two species is between 700 and 770 bp. The similarity of the two species in this region is 94.6% and differs by 78 bp. Of the 107 extracted DNAs from clinical dermatophyte isolates, in duplex PCR 24 isolates were positive with Trichophyton interdigital primer and 71 isolates against Trichophyton rubrum. The remaining isolates, which included 6, were negative in this reaction, which included other dermatophyte species. Conclusion: This method is a specific and fast differential method compared to conventional methods for identifying Trichophyton rubrum and Trichophyton interdigital from each other.

BackgroundConsidering the importance of differential diagnosis of infectious otitis externa (OE), a stepwise PCR-based assay using universal and genus- or species-specific primers for the detection/identification of the most prevalent bacterial and fungal OE was developed and evaluated on the ear aspiration specimens of clinically suspected patients.Methods and MaterialsA total of 120 ear aspiration specimens with otomycosis suspicion were subjected to manual DNA extraction using phenol–chloroform extraction after tissue digestion with a lysis buffer. The multiplex PCR was initially performed using pan-fungal and bacterial homemade primers. Pseudomonas and Staphylococcus specific primers were simultaneously used in one reaction mixture to identify the bacterial genera. Furthermore, for the identification of fungal agents, Candida species-specific multiplex primers targeting the most clinically important Candida species causing OE (i.e., C. albicans, C. parapsilosis, and C. auris), as well as Aspergillus related multiplex PCR identifying the most prevalent Aspergillus species were used in two separate reaction mixtures. All the results of multiplex PCR were interpreted based on the amplicon size.ResultsThe overall multiplex PCR-based detection rate of bacterial (n = 88; 73.3%) and fungal (n = 97; 81%) OE was documented to be 100% along with and complete consistency with the results of direct examination and Giemsa staining. Double amplicon bands of bacterial and fungal pathogens were evidenced in 76 specimens (63.3%). Moreover, the positivity rate of pan-fungal PCR was higher than that of the culture result. Out of 88 pan-bacterial positive PCR specimens, 66 and 47 ones were positive for Staphylococcus and Pseudomonas, respectively. In addition, 30 samples exhibited mixed infection of both, and five specimens remained negative. Out of 97 pan-fungal positive PCR specimens, 67 and 51 ones contained Candida and Aspergillus species, respectively. It should be noted that dual amplicon bands of Candida and Aspergillus-related multiplex PCR were yielded in 30 specimens.ConclusionThe stepwise multiplex PCR assay proved to be more sensitive, more rapid, as well as less cumbersome in detection and identification of fungal and bacterial OE, compared to culture.

Background: Leishmaniasis is an expanding neglected tropical disease in the world reporting from 98 countries including Iran. This study focused on eco-epidemiological determinants of the disease following a rapid and unexpected increase of leishmaniasis incidence in a strategic residential district in North-East of Isfahan County, Iran. Methods: This study was accomplished from Apr 2012 to Jan 2014 in a strategic residential zone in North-East of Isfahan County, Esfahan, Iran. Monthly activity, parity, Leishmania infection and susceptibility tests, were determined on sand flies. Some portion of inhabitants and school children were surveyed to find active or passive cases of leishmaniasis and also wild rodents were collected to determine reservoir host. Results: Totally 5223 sand flies belonging to Phlebotomus and Sergentomyia genus were collected and identified; Ph. papatasi was the dominant species and started to appear in May and disappeared in Oct. The majority of living dissected sand flies were unfed and parous. Ph. papatasi showed 4.6% Leishmania infection through direct examination and 39.54% by nested-PCR respectively. Phlebotomus papatasi was susceptible against deltametrin 0.05%. Totally 2149 people were surveyed and incidence and prevalence of zoonotic cutaneous leishmaniasis estimated as 45.39 and 314.40 per 1000 population. Rodents showed 73.91% and 80% Leishmania infection by direct examination and nested-PCR respectively. Conclusion: Cutaneous leishmaniasis due to L. major has been established in this area. Rodent control operation and personal protection are highly recommended to control the disease in this focus.

Abstract Background and Purpose: Candida parapsilosis is a common cause of candidemia in children and onco-hematologic patients as well as in patients with septic arthritis, peritonitis, vaginitis, and nail and skin infections. Here, we evaluated intra- and inter-species beta tubulin DNA sequence variation within the C. parapsilosis complex with a view to establishing its utility in the identification and phylogenetic analysis of the species. Materials and Methods: The novel primers successfully amplified the target for all 23 strains included in the study. Results and Conclusion: Multiple alignment of 623–629 bp-long nucleotide (nt) sequences reflecting the beta tubulin gene indicated an inter-species divergence ranging from 0 to 68 nt in C. parapsilosis, C. orthopsilosis and C. metapsilosis with a mean similarity of 84.7% among the species. Meanwhile, intra-species differences of 0–20 and 0–6 nt were found within the strains of C. parapsilosis and C. orthopsilosis, respectively. The phylogenetic tree topology was characterized by a clade made up by C. parapsilosis and C. orthopsilosis, while C. metapsilosis formed a related but separate lineage. Our data provide the basis for further discoveries of the relationship between the species belonging to the C. parapsilosis complex and the value of beta tubulin DNA sequence data in the identification and taxonomy of C. parapsilosis and other pathogenic yeasts.

Candida parapsilosis is the second most common cause of candidemia in some geographical areas and in children in particular. Yet, the proportion among children varies, for example, from 10.4% in Denmark to 24.7% in Tehran, Iran. As this species is also known to cause hospital outbreaks, we explored if the relatively high number of C. parapsilosis pediatric cases in Tehran could in part be explained by undiscovered clonal outbreaks. Among 56 C. parapsilosis complex isolates, 50 C. parapsilosis were genotyped by Amplified Fragment Length Polymorphism (AFLP) fingerprinting and microsatellite typing and analyzed for nucleotide polymorphisms by FKS1 and ERG11 sequencing. AFLP fingerprinting grouped Iranian isolates in two main clusters. Microsatellite typing separated the isolates into five clonal lineages, of which four were shared with Danish isolates, and with no correlation to the AFLP patterns. ERG11 and FKS1 sequencing revealed few polymorphisms in ERG11 leading to amino-acid substitutions (D133Y, Q250K, I302T, and R398I), with no influence on azole-susceptibilities. Collectively, this study demonstrated that there were no clonal outbreaks at the Iranian pediatric ward. Although possible transmission of a diverse C. parapsilosis community within the hospital cannot be ruled out, the study also emphasizes the necessity of applying appropriately discriminatory methods for outbreak investigation.

Background: Since December 2019, the world is struggling with an outbreak of coronavirus disease-2019 (COVID-19) infection mostly represented as an acute respiratory distress syndrome and has turned into the most critical health issue worldwide. Limited information is available about the association between dynamic changes in the naso/oropharyngeal viral shedding in infected patients and biomarkers, aiming to be assessed in the current study. Materials and Methods: This quasi-cohort study was conducted on 31 patients with moderate severity of COVID-19 manifestations, whose real-time polymerase chain reaction (RT-PCR) test was positive for severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) RNA at baseline. RT-PCR was rechecked for patients every 3–4 days until achieving two negative ones. In parallel, biomarkers, including lymphocyte count, lactate dehydrogenase (LDH), and C-reactive protein (CRP), were assessed every other day, as well. Viral shedding also was assessed. Results: Spearman's correlation test revealed a significant direct correlation between the viral shedding from the symptom onset and the time, in which CRP (P = 0.0015, r = 0.54) and LDH (P = 0.001, r = 0.6207) return to normal levels after symptom onset, but not for lymphocyte count (P = 0.068, r = 0.34). Conclusion: Based on the current study's findings, the duration of SARS-CoV-2 RNA shedding was directly correlated with the required time for LDH and CRP return to normal levels. Therefore, these factors can be considered the determinants for patients' discharge, isolation, and return to social activities; however, further investigations are required to generalize the outcomes.

Background: Early and cost-effective diagnosis and monitoring of the infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are critically important to anticipate and control the disease. We aimed to set up a SYBR Green-based one-step real-time polymerase chain reaction (PCR) as a lower-cost alternative method to detect the virus. Materials and Methods: An in-house SYBR Green-based PCR assay targeting the envelope (E) and RNA-dependent RNA polymerase (RdRp) genes, was set up to diagnose the infection, and was compared with the reference probe-based PCR method. Results: When the commercial probe-based assay was considered as the reference method, SYBR Green-based PCR had a slightly lower sensitivity (81.98% and 86.25% for E and RdRp targets, respectively) and a good specificity (100% and 94.44% for E and RdRp targets, respectively). For both gene targets, three different melting temperature (Tm) patterns were found in the PCRs of the nasopharyngeal/oropharyngeal swab samples, but no size polymorphism was seen in agarose gel electrophoresis. Conclusion: Further studies to improvement of the assay are needed to make it an inexpensive and reliable tool for the diagnosis of COVID-19.

Abstract Background Antifungal susceptibility testing is a subject of interest in the field of medical mycology. The aim of the present study were the distributions and antifungal susceptibility patterns of various Candida species isolated from colonized and infected immunocompromised patients admitted to ten university hospitals in Iran. Methods In totally, 846 Candida species were isolated from more than 4000 clinical samples and identified by the API 20 C AUX system. Antifungal susceptibility testing was performed by broth microdilution method according to CLSI. Results The most frequent Candida species isolated from all patients was Candida albicans (510/846). The epidemiological cutoff value and percentage of wild-type species for amphotericin B and fluconazole in Candida albicans, Candida tropicalis, Candida glabrata and Candida krusei were 0.5 μg/ml (95%) and 4 μg/ml (96%); 1 μg/ml (95%) and 8 μg/ml (95%); 0.5 μg/ml (99%) and 19 μg/ml (98%); and 4 μg/ml (95%) and 64 μg/ml (95%), respectively. The MIC90 and epidemiological cutoff values to posaconazole in Candida krusei were 0.5 μg/ml. There were significant differences between infecting and colonizing isolates of Candida tropicalis in MIC 90 values of amphotericin B, and isolates of Candida glabrata in values of amphotericin B, caspofungin, and voriconazole (P

Background and Purpose: The epidemiological alteration in the distribution of Candida species, as well as the significantly increasing trend of either intrinsic or acquired resistance of some of these fungi highlights the need for a reliable method for the identification of the species. Polymerase chain reaction (PCR) is one of the methods facilitating the quick and precise identification of Candida species. The aim of this study was to compare the efficiency of CHROMagar, PCR-restriction fragment length polymorphism (PCR-RFLP), and PCR-fragment size polymorphism (PCR-FSP) assays in the identification of Candida species to determine the benefits and limitations of these methods. Materials and Methods: This study was conducted on 107 Candida strains, including 20 standard strains and 87 clinical isolates. The identification of the isolates was accomplished by using CHROMagar as a conventional method. The PCR-RFLP assay was performed on the entire internal transcribed spacer (ITS) region of ribosomal DNA (rDNA), and the consequent enzymatic digestion was compared with PCR-FSP results in which ITS1 and ITS2 regions were separately PCR amplified. In both molecular assays, yeast identification was carried out through the specific electrophoretic profiles of the PCR products. Results: According to the results, the utilization of CHROMagar resulted in the identi-fication of 29 (33.3%) Candida isolates, while the PCR-RFLP and PCR-FSP facilitated the identification of 83 (95.4%) and 80 (91.9%) isolates, respectively. The obtained concordances between CHROMagar and PCR-RFLP, between CHROMagar and PCR-FSP, as well as between PCR-RFLP and PCR-FSP were 0.20, 0.23, and 0.77, respectively. Conclusion: The recognition of the benefits and limitations of PCR methods allows for the selection of the most efficient technique for a fast and correct differentiation. The PCR-RFLP and PCR-FSP assays had satisfactory concordance. The PCR-FSP provides a rapid, technically simple, and cost-effective method for the identification of Candida species. Nevertheless, to accurately differentiate among the taxonomically related species, PCR-RFLP should be implemented.

Background and Purpose: Cladosporium species are ubiquitous, saprobic,dematiaceous fungi, only infrequently associated with human and animal opportunisticinfections. Materials and Methods: Airborne samples were collected using the settle plate method, and soil samples were obtained from a depth of 5-10 cm of the superficial soil layer. Samples were cultured on Sabouraud dextrose agar (SDA) plates, incubated at 25°C, and examined daily for fungal colonies for two to three weeks. Isolates were identified as Cladosporium species according to the macroscopic and microscopic criteria. For species differentiation, DNA from 53 isolates was extracted and subjected to amplification of the internal transcribed spacer (ITS) region followed by sequencing. Results: A total of 270 samples were collected from various environmental sources, of which 79 strains of Cladosporium species were isolated. The most frequent species was C. cladosporioides (50.6%), followed by C. iridis (44.3%), C. elatum (2.5%), C. peranqestum (1.3%), and C. alicinum. (1.3%). Conclusion: The collected data can serve as baseline information for future research and may be useful in the development of preventive and educational strategies.

Background: Visceral and cutaneous leishmaniasis are common in some areas of Iran and consider as health problems. Phlebotomus alexandri has been incriminated as a suspected vector for the both form of leishmaniasis. Methods: This study was carried out in 4 western provinces of Iran. Sand flies were collected using sticky traps and light traps from indoor and outdoor resting places. Nested PCR was employed to detect Leishmania parasites among collected sand flies. Results: Seven hundred and twenty two P. alexandri females were collected and pooled in 179 batches. Results of nested PCR showed, out of 9 samples from East Azerbaijan Province, only one sample was infected by Leishmania infantum. Of 34 individual and pooled samples from Kermanshah Province, only one pooled sample was infected with Leishmania major and among 30 individual and pooled samples in Fars Province, five specimens were infected by L. major, L. infantum, Leishmania donovani and Leishmania tropica. Furthermore, out of 108 individual and pooled samples from Khuzestan Province, 10 samples showed infection with L. major and L. infantum. Conclusion: The results of this study showed that P. alexandri is more active in hot zones than in moderate zones and this species may be considered as a permissive species.

Background: Mucormycosis is an acute and invasive fungal infection with a high mortality rate. Mucorales are less sensitive than other types of fungi to most antifungal agents. Amphotericin B (AMB) is one treatment option for this infection, but in recent studies, the antifungal activity of statins against Mucorales was shown. Therefore, therapy that combines AMB with these agents may have better effects in management of patients with mucormycosis. We evaluated the in vitro activity of AMB alone and in combination with statins, against Mucorales. Methods: Susceptibility profiles of AMB alone and in combination with two statins, atorvastatin (ATO) and lovastatin (LOV) determined against clinical (n: 15) and environmental (n: 5) Rhizopus oryzae isolates, obtained between Jan 2009 and Oct 2016 from patients with uncontrolled diabetes mellitus and cancer referred to the Department of Parasitology and Medical Mycology of Tehran University of Medical Sciences, Tehran, Iran. It was performed by microdilution method, based on the Clinical and Laboratory Standard Institute (CLSI) M38-A2 guideline. Results: All clinical and environmental isolates were susceptible to AMB (MIC≤1 µg/mL). The results of the interactions between AMB and the two statins were positive. The AMB-ATO (GM: 0.13 µg/Ml) combination produced greater activity than the AMB-LOV (GM: 0.26 µg/mL) combination. AMB, in combination with ATO and LOV, reacts positively against clinical and environmental R. oryzae isolates. Conclusion: This combination strategy may lead to more effective treatment of mucormycosis and fewer side effects using low dose of AMB.

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: Superficial mycotic infections have been only poorly described in koalas and there are no reliable mycologically confirmed data regarding clinical isolation of dermatophytes in this animal. We report an 11-yearold female koala, kept in a zoo in Tokyo, Japan, and presenting with hyperkeratotic lesions and scaly plaques on forepaw claws and pads reminiscent of fungal infection. Case Report: Direct microscopy of the scrapings was indicative of a dermatophyte infection. By culture and subsequent repeated subculturing of clinical specimens on Sabouraud dextrose agar, Mycobiotic agar, and potato dextrose agar, two distinct strains with different colony morphotypes (designed as types I and II) were identified. Macroscopic and microscopic characteristics of the strains were suggestive of three different species, i.e. Microsporum canis, M. gypseum, and M. fulvum. However, partial sequencing of internal transcribed spacer (ITS) region of rDNA, translation elongation factor-1α (Tef-1α), and beta-tubulin (BT2) genes confirmed the identity of both isolates as M. gypseum. The animal was treated with a continuous terbinafine regimen (250 mg/kg) once daily for 12 weeks. Conclusion: To the best of our knowledge, the present report is the first confirmed case of dermatophytosis in a koala. The genetics underlying a variety of phenotypic traits in most classical dermatophyte species are unknown, and further studies are needed to understand this phenomenon.

Background and Purpose: Candidiasis is a major fungal infection, and Candida albicans is the major cause of infections in humans. The Clinical and Laboratory Standards Institute (CLSI) developed new breakpoints for antifungal agents against C. albicans. In this multi-center study, we aimed to determine the drug susceptibility profile of C. albicans, isolated from Iranian population according to new species-specific CLSI. Materials and Methods: Clinical samples were cultured on Sabouraud dextrose agar and were incubated at room temperature for seven days. The isolates were transferred to Professor Alborzi Clinical Microbiology Research Center, Shiraz, Iran. C. albicans were identified by using API 20C AUX system. Broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) of amphotericin B, caspofungin, voriconazole, fluconazole, posaconazole, itraconazole, and ketoconazole, based on CLSI document M27-S4 and new breakpoints for some azoles and caspofungin. Results: Overall, 397 C. albicans were isolated from patients admitted to ten university hospitals in Iran. The MIC90 of the isolates to amphotericin B, caspofungin, voriconazole, fluconazole, posaconazole, itraconazole, and ketoconazole were 0.125, 0.125, 0.125, 1, 0.064, 0.5, and 0.125 μg/ml, and rates of resistance were 0.5%, 0.3%, 3.8%, 2.8%, and 2.5% for amphotericin B, caspofungin, voriconazole, fluconazole, and itraconazole, respectively. Conclusion: According to our data, fluconazole is the drug of choice for management of patients at risk for systemic candidiasis throughout the region, since it is cost-effective with low side effects.

Background: Candiduria in hospitalized patients may represent contamination, colonization, or urinary tract infections. On the other hand, candidemia and upper urinary tract infection could be the complications of candiduria. The aim of this study was to determine candiduria in hospitalized patients and identify isolated Candida species by conventional and molecular methods. Methods: This cross-sectional study was conducted on hospitalized patients in Imam Khomeini and Mostafa Khomeini hospitals in Ilam, western Iran from Jan to Dec 2016. Urine samples of hospitalized patients were collected during a period of 4 months for diagnosis of candiduria. Primary identification was done by conventional methods. PCR profile was carried out using phenol-chloroform method and confirmed using restriction fragment length polymorphism (PCR-RFLP) technique by MspI restriction enzyme. Results: Candiduria was diagnosed in 18 (9.2%) cases from a total of 195 patients. Isolated yeasts were identified as C. albicans (n: 13), C. glabrata (n: 5), and C. parapsilosis (n: 1) in the one case both C. albicans and C. glabrata were isolated from a urine sample. Conclusion: Candida urinary tract infection is becoming increasingly common in hospitalized patients but, differentiation fungal colonization from infection and identification of etiologic agents for optimal treatment is necessary.

Background: Toxoplasma gondii, cause severe medical complications in infants and immune-compromised individuals. As using early, sensitive and rapid technique has major in diagnosis of toxoplasmosis, the present study was aimed to detect parasite by using from repetitive element (RE) and B1genes, in blood samples of seropositive immunocompromised patients and pregnant women. Methods: A total of 110 peripheral blood samples were collected from seropositive cases with anti-T. gondii antibodies, including immunocompromised patients and pregnant women. DNA was extracted by a commercial kit and subjected to TaqMan probe-based real-time PCR assay by using primers and probes specific for RE and B1 genes, separately. The data were analyzed by Kappa test and SPSS-22 software. Results: In the pregnant women, 17 (68%) and 14 (56%) samples from 25 IgM+/ IgG+ cases and, 7 (25%) and 6 (21.4%) samples from 28 IgG+/IgM- cases were positive by RE and B1 real time PCR, respectively. Likewise, in immunocompromised group, 20 (66.6%) and 17 (56.6%) samples from 30 IgM+/ IgG+ cases and 2 (7.4%) and 2 (7.4%) samples from 27 IgG+/ IgM- cases were positive by RE and B1 real time PCR, respectively. Conclusion: Probe-based real time PCR assay is a quantitative approach for early diagnosis of T. gondii infection in clinical samples. Moreover, this method can be more appropriate in diagnosis of acute and reactivated toxoplasmosis. In addition our results indicated that RE gene is more sensitive than B1 gene.

Background and Purpose: By using advanced detection/identification methods, the list of emerging uncommon opportunistic yeast infections is rapidly expanding worldwide. Our aim in the present study was sequence-based species delineation of previously unidentified yeasts obtained from a clinically yeast collection. Materials and Methods: A total of twenty three out of the 855 (5.7%) yeast isolates which formerly remained unidentified by PCR-RFLP method, were subjected to sequence analysis of the entire internal transcribed spacers (ITS) regions of rDNA. The precise species recognition was performed by the comparison of the sequences with the reliable GenBank database. Results: Sequencing analysis of the ITS region of the strains revealed several uncommon yeasts that were not reported previously in Iran. The species include Hanseniaspora uvarum, Saccharomyces cerevisiae, Sporidiobolus salmonicolor, Pichia fabianii, Pichia fermentans, Candida famata, Candida inconspicua, Candida maqnoliae, Candida guilliermondii, Candida kefyr, Candida rugosa, Candida lusitaniae, Candida orthopsilosis, and Candida viswanathii. Conclusion: We identified several rare clinical isolates selected from a big collection at the species level by ITS-sequencing. As the list of yeast species as opportunistic human fungal infections is increasing dramatically, and many isolates remain unidentified using conventional methods, more sensitive and specific advanced approaches help us to clarify the aspects of microbial epidemiology of the yeast infections.

Background: Rhipicephalus sanguineus is the most widely distributed tick in the world, which is partly due to its biological flexibility and the global distribution of its major host, the domestic dog. In Mediterranean region it could be principal reservoir host for Leishmania infantum, usually transmitted by the phlebotomine sand flies. In this study, we evaluated the vector potential of R. sanguineus in transmitting L. infantum to uninfected dogs. Methods: During 2014, five dogs with clinical manifestations of canine visceral leishmaniasis (CVL), high anti-Leishmania antibody titers and tick infestation, were selected from CVL endemic areas (Tehran and Alborz provinces). At least, twenty live ticks were removed from each infected dog. After morphological identification, the ticks were divided into two groups; ticks belonging to the first group were dissected for parasitological examinations and semi-nested PCR assay, and those of the second group were selected for the transmission of CVL caused by L. infantum to uninfected dogs. Following tick infestation, all uninfected dogs were kept for 9 months and examined monthly for clinical and serological tests. Results: Nearly, 67% of ticks were infected by L. infantum using the semi-nested PCR. All other parasitological tests of ticks were negative. Clinical examinations and serological tests of the investigated dogs revealed negative results. Nested-PCR test results performed on splenic biopsy samples of dogs were also negative. Conclusion: L. infantum-positive R. sanguineus ticks were unable to transfer L. infantum from infected dogs to healthy ones. The detection of L. infantum DNA in ticks collected from naturally infected dogs by semi-nested PCR does not prove their vectorial competence.

Objective(s): The nosocomial infections by Aspergillus species are associated with constructions and increased dust loads in hospital indoors. Our main object was to find the environmental sources of Aspergillus species causing hospital acquired infections. Materials and Methods: The clinical and environmental samplings were performed during 18 months from spring 2010 to summer 2011 in Imam educational hospital, Urmia, Iran. A morphological diagnosis was performed including microscopic characterization of isolated aspergillus from cultured specimens and polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) for the identification in the level of species. Random amplified polymorphic DNA – PCR RAPD-PCR using random primers for rDNA gene was performed to compare Aspergillus isolates of clinical cases with the relevant environmental sources. Results: Use of RAPD method resulted various differential patterns, so that some Aspergillus isolates from the clinical and hospital indoor were completely matched (matched pairs) and some other Aspergillus isolates were not matched. In the case of matched pairs, Aspergillus niger and A. flavus isolated from broncoalveolar lavage and sinus discharge were relevant to those of air conditioner and walls surfaces, respectively. Conclusion: The hospital sources for the Aspergillus clinical isolates included air condition and walls. RAPD-PCR analysis can play a trivial role to find the hospital sources of Aspergillus clinical isolates.

Background and Aim: Aspergillus fumigatus is the cause of more than 90% of invasive pulmonary aspergillosis. The aim of this study is identification and detection of A.fumigatus in bronchoalveolar lavage (BAL) samples patients suspected to have tuberculosis by using of Nested-PCR. Materials and Methods: In this assay, a set of 100 BAL samples which were collected in 9 month, between the years 2012-2013, from patients with suspected tuberculosis in Tehran hospital, were examined by direct, culture and molecular tests. Manual phenol-chloroform method was used in order to extract DNA. Specific primers of Aspergillus genus and beta-actin primers were used for duplex-PCR. A pairs of internal primers were utilized in order to identification of A.fumigatus (Nested-PCR method). Finally, some of the PCR-products were sequenced for confirming results. Results: Overall, 12 samples (12%) were aspergillus-positive by molecular test while 11% and 10% samples were positive by direct and culture test, respectively. The sensitivities and specificities of PCR method compare with direct examination was 100% and 98.8%, respectively. The study also showed that sensitivities and specificities of PCR method compare with culture was 100% and 97.7%, respectively. After sequencing PCR product sample, isolated species were recognized as A. fumigatus. Conclusions: This study showed that PCR method in compare with the direct method and the culture has a higher sensitivity in the detection of Aspergillus species, also Present study displayed that can be consider Aspergillus species and A. fumigatus in suspicious to tuberculosis BAL samples

Background: Surveillance of dermatophytosis is essential to determine the likely changes in etiological trends and distribution profile of this infection. In this study beta tubulin gene (BT2), was used as the first time in a PCR-RFLP format to clarify the distribution of dermatophytosis agents in some parts of Iran.Methods: A total of 603 clinical isolates was obtained from 500 patients in Tehran, Isfahan, Mazandaran and Guilan provinces. The isolates were identified using macro/micro-morphological criteria and electrophoretic patterns of PCR amplicons of BT2after digestion with each of the restriction enzymes FatI, HpyCH4V, MwoI and Alw21I.Results: Among the patients, 59.2% were male and 40.8% female. The most prevalent clinical form was tinea pedis (42.4%), followed by tinea cruris (24.2%), tinea unguium (12.3%), tinea corporis (10.8%), tinea faciei (4%), tinea manuum (3.14%), tinea capitis (3%) and tinea barbae (0.16%), respectively. Trichophyton interdigitale ranked the first, followed by T. rubrum, Epidermophyton floccosum, Microsporum canis, T. tonsurans, T. erinacei and T. violaceum (each 0.49%) and the less frequent species were T. schoenleinii, M. gypseum and T. anamorph of Arthroderma benhamiae (each 0.16%). A case of scalp infection by E. floccosum was an exceptional event in the study. No case of T. verrucosum was found.Conclusion: Trichophyton species and E. floccosum are yet the predominant agents of infection in Iran, while Microsporum species are decreasing. T. interdigitale and Tinea pedis remain as the most causal agent and clinical form of dermatophytosis, respectively. It seems that BT2 can be a useful genetic marker for epidemiological survey of common pathogenic dermatophytes.

Background: Pneumocystis jirovecii causes Pneumocystis pneumonia (PCP) in immunocompromised patients with a high rate of morbidity and mortality. Colonization with this fungus may stimulate pulmonary inflammation or lead to PCP in susceptible patients. The epidemiology of this infection and routs of its transmission has poorly studied in Iran. We examined Pneumosystis colonization in patients with various lung underlying diseases.Methods: Bronchoalveolar lavage (BAL) fluids of 458 patients with different underlying diseases or pulmonary signs were collected between August 2010 and January 2012. Patients were divided into four groups: transplant recipients, malignant patients, immunosuppressive drug recipients and patients with other different lung diseases. A sensitive nested-PCR method targeted 18S ribosomal RNA gene was used for investigating P. jirovecii in the specimens.Results: P. jirovecii DNA was detected in 57 out of 458 (12.5%) BAL samples by nested-PCR. Colonization rate in malignant patients, transplant recipients, immunosuppressive therapy recipients and patients with other various lung diseases was 21.7%, 20.3%, 12.7% and 7.3%, respectively. The enzyme BanI cuts all PCR products producing fragments with the size of 228 and 104 base pair. This finding as well as sequencing of four random positive samples validated and reconfirmed the PCR results. P. jirovecii cysts were found in 5 out of 57 PCR positive samples.Conclusion: A significant number of patients with pulmonary diseases were colonized by P. jirovecii that can develop to PCP in these patients or they may transmit the fungus to other susceptible patients

Background: Although Candida albicans remains the most common fungal isolate from clinical specimens, many studies have detected a shift towards non-albicans Candida species. Despite worrying clinical pictures associated with latter species, there is little information regarding its susceptibility patterns against currently available antifungal agents, with only a small number of strains having been studied. Methods: We evaluated the in vitro antifungal susceptibilities of clinical isolates of C. orthopsilosis already identified by two-steps PCR-RFLP and reconfirmed by sequence analysis of entire ITS rDNA region, to six antifungal drugs. Results: The resulting MIC50 and MIC90 for all strains (n=18) were in increasing order, as follows: posaconazole (0.016 & 0.063 μg/ml); itraconazole (0.031 & 0.125 μg/ml); amphotericin B (0.5 & 1 μg/ml); fluconazole (0.25 & 0.5 μg/ml) and caspofungin (4 & 8 μg/ml). A uniform pattern of the MIC ranges was seen for amphotericin B, fluconazole, itraconazole, and posaconazole, while a widest range and the highest MICs were observed for caspofungin. Conclusion: Although we emphasis on the careful species designation of the clinical isolates of Candida, the antifungal susceptibility patterns of these clinically important organisms may have an application in clinical and epidemiological setting and deserve the implementation of local surveillance programs to monitor.

Background: We aimed to quicken and simplify the detection of Plasmodium in blood samples by developing and testing a pan-Plasmodium real-time PCR for accurate screening of individuals suspected of malaria. Methods: A single primer/probe set for pan-species Plasmodium-specific real time PCR targeting a conserved region of the small subunit 18S ribosomal DNA was designed and evaluated for rapid diagnosis and screening of malaria infections using dried blood smears. FTA cards were used for rapid and simple DNA extraction. Results: The primers and probes showed a positive response with the DNA extracted from bloods infected with P. falciparum and P. vivax but not with DNA extracted from various smears from uninfected blood samples. Seven positive cases positive by both microscopy and nested PCR were found among 280 blood samples taken from in South and Southeast Iran. Five samples were identified as positive for P. vivax and two as positive for P. falciparum. All positive samples were positive by real-time PCR. Furthermore, all 38-blood samples positive by microscopy were positive by real-time PCR. No microscopy-negative samples were positive by real-time PCR. Conclusion: By using a simple FTA card for DNA extraction and by application of the real-time PCR developed in this study, sensitivity similar to nested-PCR and microscopy was achieved. This format simplifies the detection of Plasmodium in large numbers of samples.