[Investigation of Cryptococcus Colonization and Mating Genotype in Environmental Samples]. / Çevresel Örneklerde Cryptococcus Kolonizasyonunun ve Esey Genotipinin Arastirilmasi.

Cryptococcus species are fungal pathogens that pose a serious threat to human life and can cause meningoencephalitis in immunocompromised and healthy individuals. It was estimated that approximately 112000 people die every year due to cryptococcal-related infections all over the world, especially in immunocompromised individuals. Cryptococcus species can be found in soil, bat dung, pigeon droppings, and various tree species in addition to humans. Despite the majority of Cryptococcus species being haploid opportunistic human pathogens, it is known that the ability to undergo sexual reproduction plays a significant role in the expansion of species distribution and the increase in virulence. In Cryptococcus species, sexual reproduction is governed by the mating genotype gene region called the MAT locus. Pathogenic Cryptococcus species have two mating types (MATa and MATα), defined by the presence of one of two alternative alleles at a single MAT locus. In this study, various tree species (eucalyptus, olive and carob) in a total of seven regions in Mersin (Gülnar, Göksu, Narlikuyu, Ayas, Kizkalesi, and Tarsus) and Hatay provinces were examined to detect Cryptococcus species. The aim of this study was to determine the environmental distribution and sexual genotypes of Cryptococcus species in these regions. In the present study, samples were collected from a total of 750 trees, including olive, eucalyptus, and carob trees. The samples were incubated on Staib agar medium containing 0.1% biphenyl and 0.5% chloramphenicol. Colonies that formed brown pigment were identified as C.neoformans using conventional and molecular methods. The sexual genotypes were determined by comparing the lengths of the STE20 gene from the isolates compared with those of reference C.neoformans strains. Growth was observed in 97 (12.9%) of 750 samples collected from eucalyptus (n= 236), olive (n= 303) and carob (n= 211) trees. All 97 isolates were determined to be C.neoformans var. grubii. The highest positivity was found in Narlikuyu (78.2%), and from carob (9.4%) and olive (3.5%) trees. Cryptococcus species was not detected in any of the samples derived from eucalyptus trees. Based on the lengths of the STE20 gene, it was determined that all C.neoformans var. grubii isolates were in the MAT Aα genotype. The data obtained regarding the environmental distribution of Cryptococcus species and the distribution of genes involved in sexual reproduction are believed to provide valuable guidance in terms of the potential clinical implications of environmental Cryptococcus hotspots and regional species characteristics in our country.

Synthesis of Graphene Quantum Dot Magnesium Hydroxide Nanocomposites and Investigation of Their Antioxidant and Antimicrobial Activities.

In this paper, we synthesized graphene quantum dots magnesium hydroxide nanocomposites (GQDs/Mg(OH)2). The synthesized nanocomposites were characterized by UV-Vis spectroscopy, X-ray diffraction (XRD), Scanning electron microscopy (SEM), Transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), and Malvern Zetasizer. The antimicrobial and antioxidant properties of the obtained GQDs/Mg(OH)2 nanocomposites were investigated. GQDs/Mg(OH)2 nanocomposites have MIC values of 15.625 µg/mL against fungi (C. metapsilosis and C. parapsilosis) and 62.5 µg/mL against Gram (+) (S. pneumonia and E. faecalis) and Gram (-) (E. coli). The synthesized GQDs/Mg(OH)2 nanocomposites showed moderate antioxidant activity. The results showed that at 100-µg/mL GQDs/Mg(OH)2 nanocomposite concentration, the H2O2 scavenging activity was 62.18%.

Investigation of the Chemical Composition, Antimicrobial, and Antioxidant Activity of Endemic Onosma halophila Boiss. & Heldr.

The Onosma halophila Boiss. & Heldr. belongs to the Boraginaceae family and it is endemic species from Turkey that distributes in Salt Lake (Tuz Gölü) and the surrounding salty steppes. In this study, the chemical content, antimicrobial, and antioxidant activity of endemic O. halophila were determined for the first time. Thirty-one components were identified by GC-MS analysis in O. halophila. Antimicrobial activity was tested against a total of eight microorganisms, including three Gram-positive, three Gram-negative bacterial strains, and two fungal strains, using the Micro dilution technique. The obtained extracts showed strong antifungal and antibacterial activity. The MIC value of extracts samples against the tested strains ranged from 15.625 to 125 µg/mL. In addition, it was determined that the extracts had different levels of antioxidant activity. The IC50 values were determined 45.20-1760 µg/mL for DPPH radical scavenging assay, 3.125-1016 µg/mL for H2O2 radical scavenging assay, and 147.12-1837 µg/mL for superoxide radical scavenging assay, respectively. As a result, it has been determined that O. halophila has the potential to be used in complementary medicine and various ethnobotanical fields in future due to the important components it contains.

Effectiveness of FastFung agar in the isolation of Malassezia furfur from skin samples.

BACKGROUND: Lipophilic basidiomycetous yeasts of the Malassezia genus can cause various skin diseases, such as seborrheic dermatitis, pityriasis versicolor, folliculitis and atopic dermatitis, and even life-threatening fungemia in newborns and immunocompromised individuals. Routine mycological media used in clinical practice do not contain sufficient lipid ingredients required for the growth of Malassezia species. A recently developed medium, FastFung agar, is promising for culturing fastidious fungal species. METHODS: In this study, we compared FastFung agar and mDixon agar for culturing Malassezia species from nasolabial fold and retroauricular specimens of 83 healthy individuals and 187 and 57 patients with acne vulgaris and seborrheic dermatitis, respectively. RESULTS: Malassezia species were identified using conventional tests and matrix-assisted laser desorption/ionisation mass spectrometry. In total, 96 of 654 samples (14.6%) contained Malassezia species. The total isolation rate was significantly higher in patients with seborrheic dermatitis (40.4%) than in healthy volunteers (21.7%; p < .05), and the rate of M. furfur isolation was significantly higher for patients with acne vulgaris (13.9%) and seborrheic dermatitis (24.6%) than for healthy individuals (1.5%; p < .05). FastFung agar was superior to mDixon agar in M. furfur isolation (p = .004) but showed similar performance in the case of non-M. furfur species (p > .05). Among cultured Malassezia species, perfect agreement between mDixon agar and FastFung agar was found only for M. globosa (κ = 0.90). CONCLUSION: Our results indicate that FastFung agar favours the growth of Malassezia species and should be useful in clinical mycology laboratories.

Development and Analysis of qPCR for the Identification of Arthroconidial Yeasts of the Genus Magnusiomyces.

The arthroconidial yeasts Magnusiomyces capitatus and M. clavatus are emerging opportunistic pulmonary pathogens. They are closely related and difficult to distinguish based on morphological and physiological traits. We applied an SYBR® green-based quantitative PCR (qPCR) assay to identify the species. We analyzed 30 reference strains originating from clinical and environmental sources by targeting the Rpb2 gene encoding the second largest subunit of RNA polymerase II. The qPCR assays were tested by direct identification of M. capitatus and M. clavatus in spiked sputum and household dishwasher swabs, respectively, as models for clinical and environmental samples. The assays were proved to be reliable for species-level identification of both species, with 100% sensitivity and 100% specificity, lowest inter-assay deviations (RSDr ≤ 1.65%, R2 values >0.99), detection limit of 10 theoretical copy number of target DNA, and detection cell limit of ≥5000 yeast cells from spiked sputum samples. The developed qPCR assay is a practical molecular approach for the detection of M. capitatus and M. clavatus that can be used as a stand-alone assay or in conjunction with culture-dependent approaches.

Genes Encoding Proteolytic Enzymes Fungalysin and Subtilisin in Dermatophytes of Human and Animal Origin: A Comparative Study.

Dermatophytes are among the most successful fungal pathogens in humans, but their virulence mechanisms have not yet been fully characterized. Dermatophytic fungi secrete proteases in vivo, which are responsible for fungal colonization and degradation of the keratinized tissue during infection. In the present study, we used PCR to investigate the presence of genes encoding fungalysins (MEP) and subtilisins (SUB) in three dermatophyte species whose incidence is increasing in Europe: the anthropophilic Trichophyton rubrum (n = 58), zoophilic Microsporum canis (n = 33), and Trichophyton benhamiae (n = 6). MEP2 and SUB4 genes were significantly correlated with T. rubrum; MEP3 and SUB1 were mostly frequently harbored by M. canis; and MEP1, 2, and 4 and SUB3-7 were most frequently harbored by T. benhamiae isolates (p < 0.05). Furthermore, MEP1-5 and SUB1-3 genes were significantly more prevalent among human clinical isolates of M. canis (n = 17) than among asymptomatic cat isolates of M. canis (n = 16; p < 0.05). Unidentified MEP and/or SUB genes in some isolates in the current study may suggest that other gene repertoires may be involved in the degradation of keratin. The presented analysis of the incidence of MEP and SUB virulence genes in three dermatophyte species of diverse origins provides an insight into the host-fungus interaction and dermatophyte pathogenesis.

Mating type (MAT) locus and possible sexuality of the opportunistic pathogen Exophiala dermatitidis.

Sexual reproduction among the black yeasts is generally limited to environmental saprobic species and is rarely observed among opportunists in humans. To date, a complete sexual cycle has not been observed in Exophiala dermatitidis. In this study, we aimed to gain insight into the reproductive mode of E. dermatitidis by characterizing its mating type (MAT) locus, conducting MAT screening of environmental and clinical isolates, examining the expression of the MAT genes and analyzing the virulence of the isolates of different mating types. Similar to other members of the Pezizomycotina, the E. dermatitidis genome harbors a high mobility group (HMG) domain gene (MAT1-2-1) in the vicinity of the SLA2 and APN2 genes. The MAT loci of 74 E. dermatitidis isolates (11 clinical and 63 environmental) were screened by PCR, and the surrounding region was amplified using long-range PCR. Sequencing of the ∼ 12-kb PCR product of a MAT1-1 isolate revealed an α-box gene (MAT1-1-1). The MAT1-1 idiomorph was 3544-bp long and harbored the MAT1-1-1 and MAT1-1-4 genes. The MAT1-2 idiomorph was longer, 3771-bp, and harbored only the MAT1-2-1 gene. This structure suggests a heterothallic reproduction mode. The distribution of MAT among 74 isolates was ∼ 1:1 with a MAT1-1:MAT1-2 ratio of 35:39. RT-PCR analysis indicated that the MAT genes are transcribed. No significant difference was detected in the virulence of isolates representing different mating types using a Galleria mellonella model (P > 0.05). Collectively, E. dermatitidis is the first opportunistic black yeast in which both MAT idiomorphs have been characterized. The occurrence of isolates bearing both idiomorphs, their approximately equal distribution, and the expression of the MAT genes suggest that E. dermatitidis might reproduce sexually.

Mating genotypes and susceptibility profiles of clinical isolates of Candida glabrata from Turkey.

The sexual cycle of Candida glabrata is not known; however, genomic evidence is indicative of recombination among subpopulations and the genome harbours genes necessary for undergoing mating and meiosis, which may increase fitness. The relationship between specific mating type-like (MTL) loci and antifungal susceptibility is not well understood in C. glabrata. We investigated different combinations of clinical C. glabrata isolate mating types and their antifungal susceptibility profiles. Allele profiles of the mating genes of 103 clinical C. glabrata isolates were identified, and their antifungal susceptibility to azoles, echinocandins and amphotericin B were compared. The majority (88.3%) of screened isolates harboured the a allele in the locus. The MTL1, MTL2 and MTL3 loci harboured a (88.3%), a (95.1%), and α (71.8%) alleles, respectively. The C. glabrata isolates were susceptible to echinocandins but displayed high minimal inhibitory concentrations (MICs) for azoles. The MIC ranges and MIC90 values of all isolates were 1.0 to ≥64 and 8.0 µg mL-1 for fluconazole, 0.06 to ≥16.0 and 0.5 µg mL-1 for voriconazole, 0.06 to ≥16.0 and 1.0 µg mL-1 for posaconazole, ≤0.015 to 0.06, and 0.03 µg mL-1 for caspofungin, ≤0.015 to 0.06 and 0.015 µg mL-1 for anidulafungin and 0.5-2 and 2.0 µg mL-1 for amphotericin B, respectively. The mating gene alleles of the clinical C. glabrata isolates were not associated with differences in the MICs of the tested antifungals, except for the MTL3 α-allele and echinocandins. The mating genotypes of the clinical C. glabrata isolates had no recognisable common effect on antifungal susceptibility.

[Prediction of the species distribution of Cryptococcus neoformans throughout Turkey]. / Türkiye genelinde Cryptococcus neoformans'in tür dagiliminin tahmini.

Cryptococcus neoformans is a human pathogenic yeast that causes life-threatening infections especially in immunosuppressed patients. The environmental isolation of C.neoformans from Turkey was reported as early as 2004, although this was mostly from Eucalyptus camaldulensis colonization. Successful isolations were also reported from pomegranate (Punica granatum), oriental plane (Platanus orientalis), pine tree (Pinaceae), chestnut (Castanea sativa) and salt cedar (Tamarix hispida). The investigation of the relationship between the bioclimatic factors affecting the environmental isolation sites and the colonization of pathogens is a frequently used method. With this method, detailed risk maps can be generated in which environmental colonization can be estimated. The aim of this study was to use the high-resolution bioclimatic and previously-isolated yeasts' coordinates to create a valid model for the occurrence of C.neoformans in Turkey and provide insight into ecological processes. A machine learning approach using presence-only data software, maximum entropy (MaxEnt), was used to for the prediction of C.neoformans distribution. Climatic data and environmental bioclimatic variables from WorldClim were downloaded as 30 seconds spatial resolutions. The correlation between different Turkey bioclimatic layers were analyzed with ENMTools and similar layers were discarded. Forty-one different coordinates representing C.neoformans isolation points were used to generate a predictive map. The area under the curve and the omission rate were used to validate the model. Meanwhile, Jackknife tests were applied to enumerate the contribution of different environmental variables, and then to predict the final model. Maps were created using QGIS mapping software. In this study, we have shown that the coastal region of Anatolia, which is geographically located in the Northeastern Mediterranean Basin, as well as the entire Aegean region, carry an extremely high risk for the colonization of C.neoformans. Other areas which have not previously been reported for the isolation of C.neoformans were predicted to be potential colonization hotspots, including the western part of Ataturk Dam, the Amik Plain and the Bakirçay and Gediz valleys. The maximum temperature of the warmest month, the mean temperature of the warmest quarter and the precipitation of the coldest quarter were the most important factors influencing the model's predictions. It was determined that the humidity in the environment affected the colonization especially in November. In conclusion, we produced a C.neoformans colonization risk map of Turkey for the first time. Obtaining more regional data will facilitate the identification of the regions having similar risk. This approach is useful for the clinical prediagnosis of cryptococcosis cases, which may be more common in places with environmental niches.

[Isolation of Cryptococcus neoformans from a chesnut tree (Castanea sativa), Denizli, Turkey]. / Denizli bölgesinde kestane (Castanea sativa) agacindan Cryptococcus neoformans izolasyonu.

Cryptococcus neoformans is a basidiomycetous encapsulated yeast that can cause life-threatening infections in immunosuppressed humans and animals. C.neoformans/Cryptococcus gattii infections are considered to be acquired via inhalation of aerosolized particles from the environment. Avian guano, decaying tree hollows and soil are known as environmental niches. In recent years, colonization of the woody structures of different trees such as Eucalyptus camaldulensis, Tamarix hispida, Platanus orientalis and Punica granatum has been reported in the environmental study of the western Anatolian region. Based on the results of previous studies, our country may have intensive Cryptococcus colonization niches in the western regions. The aim of this study was to investigate the presence of the colonization of C.neoformans niche in chestnut (Castanea spp.) trees on higher altitudes. In the study, the colonization of C.neoformans was screened on chestnut trees (Castanea spp.) in Aydin-Ödemis-Denizli geographical area. This area consists of mountainous terrain between the fertile plain formed by two major rivers.This region is one of the widespreading areas of chestnut farming in Anatolia. Two hundred and fourteen chestnut trees that had deep fissures or trunk hollows were screened during mid-summer 2017. A swabbing technique was used, and all samples were cultured on Staib agar medium containing biphenyl and antibiotics. Cultures were checked for ten days for suspicious brown colonies. Suspicious yeast colonies were tested for the identification of pathogenic Cryptococcus by conventional methods and canavanine-glycine-bromothymol agar reactions. ITS 1-4 primers were used for strain PCR tests. We determined the mating type and serotypes by PCR analysis of the STE20 genes using STE20 (Aa), STE20 (Aα), STE20 (Da), and STE20 (Dα) primers. V8 agar medium was used for mating cultivation. Only one (0.47%) strain of C.neoformans was isolated from 214 screened trees. This strain was confirmed by ITS 1-4 sequencing. The serotype A MATα mating type was observed. Basidium, basidiospores and clamp connections in hyphal structure were noted with MATα mating on V8 agar medium. In this study, the first C.neoformans isolate from a chestnut tree (Castanea sativa) was determined from Denizli region. Further studies of distribution of human pathogenic Cryptococcus will be helpful to determine the risk areas for the living organisms in our region.

Overview of selected virulence attributes in Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans, Trichophyton rubrum, and Exophiala dermatitidis.

The incidence of fungal diseases has been increasing since 1980, and is associated with excessive morbidity and mortality, particularly among immunosuppressed patients. Of the known 625 pathogenic fungal species, infections caused by the genera Aspergillus, Candida, Cryptococcus, and Trichophyton are responsible for more than 300 million estimated episodes of acute or chronic infections worldwide. In addition, a rather neglected group of opportunistic fungi known as black yeasts and their filamentous relatives cause a wide variety of recalcitrant infections in both immunocompetent and immunosuppressed hosts. This article provides an overview of selected virulence factors that are known to suppress host immunity and enhance the infectivity of these fungi.

In vitro activities of antifungal drugs against environmental Exophiala isolates and review of the literature.

Exophiala is a genus of black fungi isolated worldwide from environmental and clinical specimens. Data on antifungal susceptibility of Exophiala isolates are limited and the methodology on susceptibility testing is not yet standardised. In this study, we investigated in vitro antifungal susceptibilities of environmental Exophiala isolates. A total of 87 Exophiala isolated from dishwashers or railway ties were included. A CLSI M38-A2 microdilution method with modifications was used to determine antifungal susceptibility for fluconazole, voriconazole, posaconazole, itraconazole, amphotericin B and terbinafine. Minimum inhibitory concentration (MIC) values were determined visually at 48 hours, 72 hours and 96 hours. MIC-0 endpoint (complete inhibition of growth) was used for amphotericin B and azoles, except fluconazole, for which MIC-2 endpoint (~50% inhibition compared to growth control) was used. Both MIC-0 and MIC-1 (~80% inhibition compared to growth control) results were analysed for terbinafine to enable comparison with previous studies. Fungal growth was sufficient for determination of MICs at 48 hours for all isolates except two Exophiala dermatitidis strains. At 72 hours, most active antifungal agents according to GM MIC were voriconazole and terbinafine, followed by posaconazole, itraconazole and amphotericin B in rank order of decreasing activity. While amphotericin B displayed adequate in vitro activity despite relatively high MICs, fluconazole showed no meaningful antifungal activity against Exophiala.

Genetic Diversity and Antifungal Susceptibility of Candida parapsilosis Sensu Stricto Isolated from Bloodstream Infections in Turkish Patients.

Candida parapsilosis sensu stricto is an emerging cause of hospital-acquired Candida infections, predominantly in southern Europe, South America, and Asia. We investigated the genetic diversity and antifungal susceptibility profile of 170 independent C. parapsilosis sensu stricto strains obtained from patients with candidemia who were treated at the Ege University Hospital in Izmir, Turkey, between 2006 and 2014. The identity of each strain was confirmed via PCR amplification and digestion of the secondary alcohol dehydrogenase-encoding gene. The 24-h geometric mean minimum inhibitory concentrations of the antifungal agents, in increasing order, were as follows: posaconazole, 0.10 µg/mL; voriconazole, 0.21 µg/mL; caspofungin, 0.38 µg/mL; amphotericin B, 0.61 µg/mL; anidulafungin, 0.68 µg/mL; and fluconazole, 2.95 µg/mL. Microsatellite genotyping of the isolates (using fluorescently labeled primers and a panel of four different short-nucleotide repeat fragments) identified 25, 17, 17, and 8 different allelic genotypes at the CP6, B5, CP4, and CP1 locus, respectively. Posaconazole, caspofungin, and amphotericin B showed the greatest in vitro activity of the tested systemic azole, echinocandin, and polyene agents, respectively, and the observed antifungal susceptibility of the isolates was shown to be independent of their isolation source. We obtained a combined discriminatory power of 0.99 with a total of 130 genotypes for 170 isolates tested. Finally, microsatellite profiling analysis confirmed the presence of identical genotype between separate isolates, supporting that effective surveillance and infection-prevention programs are essential to limit the impact of C. parapsilosis sensu stricto on hospitalized patients' health.

Candida parapsilosis in domestic laundry machines.

Candida parapsilosis, although a human commensal, acts as an opportunistic pathogen associated with nosocomial infections, with a rising incidence worldwide. Its ecological characteristics are poorly understood. Human-made environments within dwellings, such as dishwashers and water distribution systems, represent major sources of fungi such as C. parapsilosis. Here, we investigated the presence of members of the C. parapsilosis complex in 99 washing machines in various dwellings in the city of Mersin, Turkey. We sampled three sites in each washing machine: (i) the washing powder drawers, (ii) fabric softener drawers, and (iii) rubber seals around the washing machine doors. Additionally, we recorded the type of cleanser used by each customer. Of note, 25.3% of sampled washing machines harbored C. parapsilosis strains, later identified as the members of the C. parapsilosis sensu stricto via internal transcribed spacer (ITS) sequencing. Out of the 29 isolates obtained, biofilm-forming ability and proteinase and esterase activities were recorded in 14, 11, and 4 of the isolates, respectively. Our results suggest that the washing machines investigated abundantly harbored C. parapsilosis sensu stricto; however, no single preferred isolation site or association with cleanser type was observed (P > .05). Furthermore, C. parapsilosis isolates grew at temperatures ranging from 10°C to 37°C, at pH values ranging from 4 to 10, and were found to tolerate 5-10% NaCl. Domestic laundry appliances as a potential source of C. parapsilosis infections are discussed.

Design of potent fluoro-substituted chalcones as antimicrobial agents.

Owing to ever-increasing bacterial and fungal drug resistance, we attempted to develop novel antitubercular and antimicrobial agents. For this purpose, we developed some new fluorine-substituted chalcone analogs (3, 4, 9-15, and 20-23) using a structure-activity relationship approach. Target compounds were evaluated for their antitubercular efficacy against Mycobacterium tuberculosis H37Rv and antimicrobial activity against five common pathogenic bacterial and three common fungal strains. Three derivatives (3, 9, and 10) displayed significant antitubercular activity with IC50 values of ≤16,760. Compounds derived from trimethoxy substituent scaffolds with monofluoro substitution on the B ring of the chalcone structure exhibited superior inhibition activity compared to corresponding hydroxy analogs. In terms of antimicrobial activity, most compounds (3, 9, 12-14, and 23) exhibited moderate to potent activity against the bacteria, and the antifungal activities of compounds 3, 13, 15, 20, and 22 were comparable to those of reference drugs ampicillin and fluconazole.

ATR-FTIR Spectroscopy Highlights the Problem of Distinguishing Between Exophiala dermatitidis and E. phaeomuriformis Using MALDI-TOF MS.

The present study compared two chemical-based methods, namely, matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, to understand the misidentification of Exophiala dermatitidis and Exophiala phaeomuriformis. The study utilized 44 E. dermatitidis and 26 E. phaeomuriformis strains, which were partially treated with strong acids and bases for further evaluation. MALDI-TOF MS and ATR-FTIR spectroscopy data of the two Exophiala species were compared. Data groupings were observed for the chromic acid- and nitric acid-treated species when the black yeast sources were categorized as creosoted-oak sleepers, concrete sleepers, or dishwasher isolates. The MALDI-TOF MS data for the metalloenzyme-containing regions were consistent with the ATR-FTIR spectroscopy data. These results indicated that environmental isolates might contain metals not found in human isolates and might interfere with chemical-based identification methods. Therefore, MALDI-TOF MS reference libraries should be created for clinical strains and should exclude petroleum-associated environmental isolates.

Virulence markers of opportunistic black yeast in Exophiala.

The black yeast genus Exophiala is known to cause a wide variety of diseases in severely ill individuals but can also affect immunocompetent individuals. Virulence markers and other physiological parameters were tested in eight clinical and 218 environmental strains, with a specific focus on human-dominated habitats for the latter. Urease and catalase were consistently present in all samples; four strains expressed proteinase and three strains expressed DNase, whereas none of the strains showed phospholipase, haemolysis, or co-haemolysis activities. Biofilm formation was identified in 30 (13.8%) of the environmental isolates, particularly in strains from dishwashers, and was noted in only two (25%) of the clinical strains. These results indicate that virulence factors are inconsistently present in the investigated Exophiala species, suggesting opportunism rather than pathogenicity.

MALDI-TOF MS-based identification of black yeasts of the genus Exophiala.

In this study, we investigated the applicability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of Exophiala species. The analysis included a total of 110 Exophiala isolates, including 15 CBS strains representing 4 species, Exophiala dermatitidis (61), E. phaeomuriformis (36), E. crusticola (9), and E. heteromorpha (4), that had been previously identified based on internal transcribed spacer (ITS) regions. We also compared the relative efficacies of Sabouraud glucose agar (SGA) and Columbia agar (CA) for use in MALDI-TOF MS. Remarkably, we obtained a log-score value ≥2.0 by using either SGA or CA for all 15 CBS strains, indicating species-level identification. The remaining 95 Exophiala strains were identified to the genus or species levels, with identification rates of 96.8% and 90.5%, using SGA or CA, respectively. Most of the E. dermatitidis (100% and 92.9%), E. phaeomuriformis (80.6% and 83.9%), E. crusticola (50% and 100%), and E. heteromorpha (100% and 100%) isolates were correctly identified using SGA or CA, respectively. Furthermore, 58.9% and 26.3% of the strains had log-score values of ≥2.0 by using SGA and CA, respectively. Our results indicate that MALDI-TOF MS is a rapid and reliable technique with high rates of correct taxonomic identification.

Serum microRNAs; miR-30c-5p, miR-223-3p, miR-302c-3p and miR-17-5p could be used as novel non-invasive biomarkers for HCV-positive cirrhosis and hepatocellular carcinoma.

Recently, serum miRNAs have been evolved as possible biomarkers for different diseases including hepatocellular carcinoma and other types of cancers. Investigating certain serum miRNAs as novel non-invasive markers for early detection of HCV-positive cirrhosis and hepatocellular carcinoma (HCC). The expression profiles of 58 miRNA were analyzed in patient's plasma of chronic hepatitis C (CHC), HCV-positive cirrhosis and HCV-positive HCC and compared with control group samples. Totally 94 plasma samples; 64 patient plasma (26 CHC, 30 HCV-positive cirrhosis, 8 HCV-positive HCC) and 28 control group plasma, were included. The expression profiles of 58 miRNAs were detected for all patient and control group plasma samples by qRT-PCR using BioMarkTM 96.96 Dynamic Array (Fluidigm Corporation) system. In CHC group, expression profiles of miR-30a-5p, miR-30c-5p, miR-206 and miR-302c-3p were found significantly deregulated (p < 0.05) when compared versus control group. In HCV-positive cirrhosis group, expression profiles of miR-30c-5p, miR-223-3p, miR-302c-3p, miR-17-5p, miR-130a-3p, miR-93-5p, miR-302c-5p and miR-223-3p were found significantly deregulated (p < 0.05). In HCV-positive HCC group, expression profiles of miR-17-5p, miR-223-3p and miR-24-3p were found significant (p < 0.05). When all groups were compared versus control, miR-30c-5p, miR-223-3p, miR-302c-3p and miR-17-5p were found significantly deregulated for cirrhosis and HCC. These results imply that miR-30c-5p, miR-223-3p, miR-302c-3p and miR-17-5p could be used as novel non-invasive biomarkers of HCV-positive HCC in very early, even at cirrhosis stage of liver disease.

Haemolytic and co-haemolytic (CAMP-like) activity in dermatophytes.

Dermatophytes are some of the most common fungal pathogens in both humans and animals. These fungi release enzymes (e.g., keratinases) that play roles in their pathogenesis. Little is known about their haemolytic and co-haemolytic (CAMP-like) activities; however, in bacteria, these components play significant roles in pathogenesis. This study characterised these two factors in 45 dermatophyte strains (representing the genera Arthroderma, Epidermophyton, Microsporum and Trichophyton) using Columbia agar (CA) supplemented with 5% bovine, ovine and equine erythrocytes. Haemolysis was best observed on CA supplemented with ovine erythrocytes followed by equine and bovine erythrocytes, while CAMP-like reactions occurred using bovine and ovine but not equine erythrocytes. Haemolytic and CAMP-like activities were best observed using ovine and bovine erythrocytes in CA in 44 and 38 strains at 7 and 3 days respectively. Most dermatophytes recovered from both symptomatic and asymptomatic lesions had haemolytic and CAMP-like activities. We suggest that the haemolytic and CAMP-like activities are not correlated with ecological characteristics, isolation sites or clinical manifestations of dermatophytic fungi. We also believe that this study has the potential to contribute to the existing literature on dermatophytes and dermatophyte pathogenesis.