Adhesion and biofilm formation by two clinical isolates of Trichosporon Cutaneum in various environmental conditions.

Trichosporon spp. is an emerging opportunistic pathogen and a common cause of both superficial and invasive infections. Although Trichosporon asahii is the most frequently isolated species, Trichosporon cutaneum is also widely observed, as it is the predominant agent in cases of white Piedra and onychomycosis. Trichosporon spp. is a known to produce biofilms, which serve as one of its virulence mechanisms, however, there is limited data available on biofilms formed by T. cutaneum. Thus, the aim of this study was to assess the adhesion and biofilm formation of two clinical isolates of T. cutaneum under various environmental conditions (including temperature, nutrient availability, and carbon source), as well as their tolerance to fluconazole. Adhesion was tested on common abiotic substrates (such as silicone, glass, and stainless steel), revealing that T. cutaneum readily adhered to all surfaces tested. CV staining was applied for the evaluation of the environment influence on biofilm efficiency and it was proved that the nutrient availability has a major impact. Additionaly, fluorescent staining was employed to visualize the morphology of T. cutaneum biofilm and its survival in the presence of fluconazole. Hyphae production was shown to play a role in elevated biofilm production in minimal medium and increased tolerance to fluconazole.

New silicate-substituted hydroxyapatite materials doped with silver ions as potential antifungal agents.

BACKGROUND: Hydroxyapatites (HAp) are widely used as medical preparations for e.g., bone replacement or teeth implants. Incorporation of various substrates into HAp structures could enhance its biological properties, like biocompatibility or antimicrobial effects. Silver ions possess high antibacterial and antifungal activity and its application as HAp dopant might increase its clinical value. RESULTS: New silicate-substituted hydroxyapatites (HAp) doped with silver ions were synthesized via hydrothermal methods. The crystal structure of HAp was investigated by using the X-ray powder diffraction. Antifungal activity of silver ion-doped HAp (with 0.7 mol%, 1 mol% and 2 mol% of dopants) was tested against the yeast-like reference and clinical strains of Candida albicans, C. glabrata, C. tropicalis, Rhodotorula rubra, R. mucilaginosa, Cryptococcus neoformans and C. gattii. Spectrophotometric method was used to evaluate antifungal effect of HAp in SD medium. It was shown that already the lowest dopant (0.7 mol% of Ag+ ions) significantly reduced fungal growth at the concentration of 100 µg/mL. Increase in the dopant content and the concentration of HAp did not cause further growth inhibition. Moreover, there were some differences at the tolerance level to Ag+ ion-doped HAp among tested strains, suggesting strain-specific activity. CONCLUSIONS: Preformed studies confirm antimicrobial potential of hydroxyapatite doped with silver. New Ag+ ion-HAp material could be, after further studies, considered as medical agent with antifungal properties which lower the risk of a surgical-related infections.

The First Comprehensive Biodiversity Study of Culturable Fungal Communities Inhabiting Cryoconite Holes in the Werenskiold Glacier on Spitsbergen (Svalbard Archipelago, Arctic).

Cryoconite holes on glacier surfaces are a source of cold-adapted microorganisms, but little is known about their fungal inhabitants. Here, we provide the first report of distinctive fungal communities in cryoconite holes in the Werenskiold Glacier on Spitsbergen (Svalbard Archipelago, Arctic). Due to a combination of two incubation temperatures (7 °C and 24 ± 0.5 °C) and two media during isolation (PDA, YPG), as well as classical and molecular identification approaches, we were able to identify 23 different fungi (21 species and 2 unassigned species). Most of the fungi cultured from cryoconite sediment were ascomycetous filamentous micromycetes. However, four representatives of macromycetes were also identified (Bjerkandera adusta, Holwaya mucida, Orbiliaceae sp., and Trametes versicolor). Some of the described fungi possess biotechnological potential (Aspergillus pseudoglaucus, A. sydowii, Penicillium expansum, P. velutinum, B. adusta, and T. versicolor), thus, we propose the Arctic region as a source of new strains for industrial applications. In addition, two phytopathogenic representatives were present (P. sumatraense, Botrytis cinerea), as well as one potentially harmful to humans (Cladosporium cladosporioides). To the best of our knowledge, we are the first to report the occurrence of A. pseudoglaucus, C. allicinum, C. ramotenellum, P. sumatraense, P. velutinum, P. cumulodentata, B. adusta, and T. versicolor in polar regions. In all likelihood, two unassigned fungus species (Orbiliaceae and Dothideomycetes spp.) might also be newly described in such environments. Additionally, due to experimenting with 10 sampling sites located at different latitudes, we were able to conclude that the number of fungal spores decreases as one moves down the glacier. Considering the prevalence and endangerment of glacial environments worldwide, such findings suggest their potential as reservoirs of fungal diversity, which should not be overlooked.

"Salt mummification" - atypical method of embalming a corpse.

It is extremely rare for table salt to be used to preserve a dead body in criminal cases. In the case presented here, after the death of his 85-year-old mother, a son kept her body preserved in table salt for about 2 years to extort social benefits (pension). Before her death, the woman had been hospitalised twice due to chronic diseases. The case has been examined by the multi-disciplinary team. The unusual conditions in which the corpse was stored influenced its good condition (close to mummification), with limited colonisation of the corpse by necrophagous insects and insects involved in soft tissue biolysis (i.e. selected Diptera or Coleoptera). The use of table salt inhibited the growth of most fungi which would normally be present on a corpse stored in ambient conditions, and the corpse's surface was colonised by halophilic fungus (Scopulariopsis brevicaulis).

A Culture-Based Study of Micromycetes Isolated from the Urban Nests of Grey Heron (Ardea cinerea) in SW Poland.

There are many positive relationships between micromycetes and birds: They can spread fungal spores, and fungi facilitate cavity woodpecker excavation by preparing and modifying excavation sites. In turn, bird nests are mainly a source of potentially zoopathogenic fungi. The Wroclaw city centre hosts the biggest grey heron breeding colony in Poland with at least 240 breeding birds pairs. To assess the possible public health risks associated with bird nests, the goal of the present study was to identify cultivable fungi present in the nests of grey herons (Ardea cinerea) in Wroclaw. Additionally, attempts were made to determine whether the obtained species of fungi may pose a potential threat to animal health. Fungi were cultured at 23 and 37 ± 0.5 °C, and identified based on phenotypic and genotypic traits. Moreover, during routine inspection, visible fungal growth in some of the nests was found. Overall, 10 different fungal species were obtained in the study (Alternaria alternata, Aspergillus fumigatus, Botryotrichum piluliferum, Cladosporium cladosporioides, Epicoccum layuense, Mucor circinelloides, M.hiemalis, Penicillium atramentosum, P.coprophilum, and P.griseofulvum). They are both cosmopolitan species and a source of potential threat to humans, homoiothermous animals and plants. The greatest number of fungal species was obtained from the nest fragments with visible fungal growth incubated at 23 °C, and the least from western conifer seed bugs (Leptoglossus occidentalis) inhabiting the nests. The species such as A. fumigatus, P. coprophilum, and P.griseofulvum can be directly related to the occurrence of visible fungal growth on plant fragments of grey heron's nests.

Nanoapatites Doped and Co-Doped with Noble Metal Ions as Modern Antibiofilm Materials for Biomedical Applications against Drug-Resistant Clinical Strains of Enterococcus faecalis VRE and Staphylococcus aureus MRSA.

The main aim of our research was to investigate antiadhesive and antibiofilm properties of nanocrystalline apatites doped and co-doped with noble metal ions (Ag+, Au+, and Pd2+) against selected drug-resistant strains of Enterococcus faecalis and Staphylococcus aureus. The materials with the structure of apatite (hydroxyapatite, nHAp; hydroxy-chlor-apatites, OH-Cl-Ap) containing 1 mol% and 2 mol% of dopants and co-dopants were successfully obtained by the wet chemistry method. The majority of them contained an additional phase of metallic nanoparticles, in particular, AuNPs and PdNPs, which was confirmed by the XRPD, FTIR, UV-Vis, and SEM-EDS techniques. Extensive microbiological tests of the nanoapatites were carried out determining their MIC, MBC value, and FICI. The antiadhesive and antibiofilm properties of the tested nanoapatites were determined in detail with the use of fluorescence microscopy and computer image analysis. The results showed that almost all tested nanoapatites strongly inhibit adhesion and biofilm production of the tested bacterial strains. Biomaterials have not shown any significant cytotoxic effect on fibroblasts and even increased their survival when co-incubated with bacterial biofilms. Performed analyses confirmed that the nanoapatites doped and co-doped with noble metal ions are safe and excellent antiadhesive and antibiofilm biomaterials with potential use in the future in medical sectors.

Microbial traces and their role in forensic science.

Forensic microbiology, also known as the microbiology of death, is an emerging branch of science that is still underused in criminal investigations. Some of the cases might be difficult to solve with commonly used forensic methods, and then they become an operational field for microbiological and mycological analyses. The aim of our review is to present significant achievements of selected studies on the thanatomicrobiome (micro-organisms found in the body, organs and fluids after death) and epinecrotic community (micro-organisms found on decaying corpses) that can be used in forensic sciences. Research carried out as a part of the forensic microbiology deals with the thanatomicrobiome and the necrobiome-communities of micro-organisms that live inside and outside of a putrefying corpse. Change of species composition observed in each community is a valuable feature that gives a lot of information related to the crime. It is mainly used in the estimation of post-mortem interval (PMI). In some criminal investigations, such noticeable changes in the microbiome and mycobiome can determine the cause or the actual place of death. The microbial traces found at the crime scene can also provide clear evidence of guilt. Nowadays, identification of micro-organisms isolated from the body or environment is based on metagenome analysis and 16S rRNA gene amplicon-based sequencing for bacteria and ITS rRNA gene amplicon-based sequencing for fungi. Cultivation methods are still in use and seem to be more accurate; however, they require much more time to achieve a final result, which is an unwanted feature in any criminal investigation.

First Speleomycological Study on the Occurrence of Psychrophilic and Psychrotolerant Aeromycota in the Brestovská Cave (Western Tatras Mts., Slovakia) and First Reports for Some Species at Underground Sites.

Most underground ecosystems are heterotrophic, fungi in these objects are dispersed in the air in the form of spores, and they may be potentially hazardous to mammals. Research in underground sites has focused on mesophilic airborne fungi and only a few concerned cold-adapted species. Therefore, the goal of our research was the first report of psychrophilic and psychrotolerant aeromycota in the Brestovská Cave using culture-based techniques with genetic and phenotypic identification. Plates with PDA medium containing sampled biological material were incubated at 8 ± 0.5 °C. The density of mycobiota inside the cave ranged from 37.4 to 71 CFU 1 m-3 of air and 63.3 CFU 1 m-3 of air outside the cave. Thus, the level of fungal spores did not exceed the standards for the mycological quality of the air. A total of 18 species were isolated during the study, and some species may be potentially dangerous to people with weakened immune system. All fungal species were present inside the cave and only seven of them were outside. Cladosporium cladosporioides dominated in the external air samples and Mortierella parvispora was cultured most frequently from internal air samples. To our knowledge, this is the first discovery of the fungal species such as Coniothyrium pyrinum, Cystobasidium laryngis, Filobasidium wieringae, Leucosporidium drummii, M. parvispora, Mrakia blollopis, Nakazawaea holstii, and Vishniacozyma victoriae in the air inside the underground sites. Moreover, C. pyrinum, C. laryngis, L. drummii, M. blollopis, and N. holstii have never been detected in any component of the underground ecosystems. There are possible reasons explaining the detection of those species, but global warming is the most likely.

Mitochondrial Function Are Disturbed in the Presence of the Anticancer Drug, 3-Bromopyruvate.

3-bromopuryvate (3-BP) is a compound with unique antitumor activity. It has a selective action against tumor cells that exhibit the Warburg effect. It has been proven that the action of 3-BP is pleiotropic: it acts on proteins, glycolytic enzymes, reduces the amount of ATP, induces the formation of ROS (reactive oxygen species), and induces nuclear DNA damage. Mitochondria are important organelles for the proper functioning of the cell. The production of cellular energy (ATP), the proper functioning of the respiratory chain, or participation in the production of amino acids are one of the many functions of mitochondria. Here, for the first time, we show on the yeast model that 3-BP acts in the eukaryotic cell also by influence on mitochondria and that agents inhibiting mitochondrial function can potentially be used in cancer therapy with 3-BP. We show that cells with functional mitochondria are more resistant to 3-BP than rho0 cells. Using an MTT assay (a colorimetric assay for assessing cell metabolic activity), we demonstrated that 3-BP decreased mitochondrial activity in yeast in a dose-dependent manner. 3-BP induces mitochondrial-dependent ROS generation which results in ∆sod2, ∆por1, or ∆gpx1 mutant sensitivity to 3-BP. Probably due to ROS mtDNA lesions rise during 3-BP treatment. Our findings may have a significant impact on the therapy with 3-BP.

Greater Mouse-Eared Bats (Myotis myotis) Hibernating in the Nietoperek Bat Reserve (Poland) as a Vector of Airborne Culturable Fungi.

Bats can contribute to an increase of aeromycota in underground ecosystems and might be a vector/reservoir of microorganisms; however, there is no information about the number and species composition of fungi around hibernating bats. One of the most common species in Europe with direct human contact is the greater mouse-eared bat (Myotis myotis). The goal of our research was the first report of the airborne fungi present in the close vicinity of hibernating M. myotis in the Nietoperek bat reserve (Western Poland) by the use of culture-based techniques and genetic and phenotypic identifications. Aerobiological investigations of mycobiota under hibernating bats were performed on two culture media (PDA and YPG) and at two incubation temperatures (7 and 24 ± 0.5 °C). Overall, we detected 32 fungal species from three phyla (Ascomycota, Basidiomycota, and Zygomycota) and 12 genera. The application of YPG medium and the higher incubation temperature showed higher numbers of isolated fungal species and CFU. Penicillium spp. were dominant in the study, with spores found outside the underground hibernation site from 51.9% to 86.3% and from 56.7% to 100% inside the bat reserve. Penicillium chrysogenum was the most frequently isolated species, then Absidia glauca, Aspergillus fumigatus, A. tubingensis, Mortierella polycephala, Naganishia diffluens, and Rhodotorula mucilaginosa. Temperature, relative humidity, and the abundance of bats correlated positively with the concentration of airborne fungal propagules, between fungal species diversity, and the concentration of aeromycota, but the number of fungal species did not positively correlate with the number of bats. The air in the underground site was more contaminated by fungi than the air outside; however, the concentration of aeromycota does not pose a threat for human health. Nevertheless, hibernating bats contribute to an increase in the aeromycota and as a vector/reservoir of microscopic fungi, including those that may cause allergies and infections in mammals, and should be monitored.

New Approach to Antifungal Activity of Fluconazole Incorporated into the Porous 6-Anhydro-α-l-Galacto-ß-d-Galactan Structures Modified with Nanohydroxyapatite for Chronic-Wound Treatments-In Vitro Evaluation.

New fluconazole-loaded, 6-Anhydro-α-l-Galacto-ß-d-Galactan hydrogels incorporated with nanohydroxyapatite were prepared and their physicochemical features (XRD, X-ray Diffraction; SEM-EDS, Scanning Electron Microscopy-Energy Dispersive X-ray Spectroscopy; ATR-FTIR, Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy), fluconazole release profiles and enzymatic degradation were determined. Antifungal activity of pure fluconazole was tested using Candida species (C. albicans, C. tropicalis, C. glabarata), Cryptococcus species (C. neoformans, C. gatti) and Rhodotorula species (R. mucilaginosa, R. rubra) reference strains and clinical isolates. Standard microdilution method was applied, and fluconazole concentrations of 2-250 µg/mL were tested. Moreover, biofilm production ability of tested isolates was tested on the polystyrene surface at 28 and 37 ± 0.5 °C and measured after crystal violet staining. Strains with the highest biofilm production ability were chosen for further analysis. Confocal microscopy photographs were taken after live/dead staining of fungal suspensions incubated with tested hydrogels (with and without fluconazole). Performed analyses confirmed that polymeric hydrogels are excellent drug carriers and, when fluconazole-loaded, they may be applied as the prevention of chronic wounds fungal infection.

Plant-Fungal Interactions: A Case Study of Epicoccoum nigrum Link.

Epicoccum nigrum Link is a cosmopolitan species, and it has been described as both an in vitro and in vivo antagonist of many fungal pathogens of plants. However, there are no clear reports about the interactions between E. nigrum and various plant species, and about the effects of culture filtrates produced by this fungus on plants. Therefore, we assessed the interactions between E. nigrum and different plant species, such as sugar beet (Beta vulgaris L. ssp. vulgaris), spring wheat (Triticum aestivum L.), red clover (Trifolium pratense L.), and winter oilseed rape (Brassica napus L.). Additionally, we evaluated the effect of E. nigrum culture filtrates on garden cress (Lepidium sativum L.). Our study showed that the E. nigrum strains varied in terms of the color of excreted culture filtrates and showed different interactions with garden cress. Overall, fungal strains only affected adversely the sprout length in a significant way and, partially, the growth of the tested plant. In addition, we confirmed the suitability of the garden cress as a test plant in in vitro toxicological tests. Most strains of E. nigrum (61.1%) secreted enzymes expected to participate mainly in the later stages of the infection (amylases and proteases) and not those expected to operate in the early phases of host penetration (cellulases and pectinases) that were secreted by 33.3% of fungal strains. The group of pectinolytic enzymes represented the catalysts with the highest activity. Host specialization tests showed that E. nigrum was mainly re-isolated from the plant surface and the number of infected seedlings as well as the disease index depended on a studied plant species, with sugar beet and red clover being most sensitive to infection. In turn, the lowest value of the disease index caused by E. nigrum strains was recorded for spring wheat and winter oilseed rape. Overall, statistically significant differences in the growth of plant seedlings during the host specialization test were noted only for sugar beet and red clover seedlings. The seedlings of plants in the control group (without fungal inoculum) exhibited an increased length compared to those treated with E. nigrum inoculum. Our studies also showed that E. nigrum is probably a facultative saprotroph of plants and it may winter on red clover, which is presumably its main reservoirs, among the species considered.

A Culture-Based ID of Micromycetes on the Wing Membranes of Greater Mouse-Eared Bats (Myotis myotis) from the "Nietoperek" Site (Poland).

Bats play important functions in ecosystems and many of them are threatened with extinction. Thus, the monitoring of the health status and prevention of diseases seem to be important aspects of welfare and conservation of these mammals. The main goal of the study was the identification of culturable fungal species colonizing the wing membranes of female greater mouse-eared bat (Myotis myotis) during spring emergence from the "Nietoperek" underground hibernation site by the use of genetic and phenotypic analyses. The study site is situated in Western Poland (52°25' N, 15°32' E) and is ranked within the top 10 largest hibernation sites in the European Union. The number of hibernating bats in the winter exceeds 39,000 individuals of 12 species, with M. myotis being the most common one. The wing membranes of M. myotis were sampled using sterile swabs wetted in physiological saline (0.85% NaCl). Potato dextrose agar (PDA) plates were incubated in the dark at 8, 24 and 36 ± 1 °C for 3 up to 42 days. All fungi isolated from the surface of wing membranes were assigned to 17 distinct fungal isolates belonging to 17 fungal species. Penicillium chrysogenum was the most frequently isolated species. Some of these fungal species might have a pathogenic potential for bats and other mammals. However, taking into account habitat preferences and the life cycle of bats, it can be assumed that some fungi were accidentally obtained from the surface of vegetation during early spring activity. Moreover, Pseudogymnoascus destructans (Pd)-the causative agent of the White Nose Syndrome (WNS)-was not found during testing, despite it was found very often in M. myotis during previous studies in this same location.

The Anticancer Drug 3-Bromopyruvate Induces DNA Damage Potentially Through Reactive Oxygen Species in Yeast and in Human Cancer Cells.

3-bromopyruvate (3-BP) is a small molecule with anticancer and antimicrobial activities. 3-BP is taken up selectively by cancer cells' mono-carboxylate transporters (MCTs), which are highly overexpressed by many cancers. When 3-BP enters cancer cells it inactivates several glycolytic and mitochondrial enzymes, leading to ATP depletion and the generation of reactive oxygen species. While mechanisms of 3-BP uptake and its influence on cell metabolism are well understood, the impact of 3-BP at certain concentrations on DNA integrity has never been investigated in detail. Here we have collected several lines of evidence suggesting that 3-BP induces DNA damage probably as a result of ROS generation, in both yeast and human cancer cells, when its concentration is sufficiently low and most cells are still viable. We also demonstrate that in yeast 3-BP treatment leads to generation of DNA double-strand breaks only in S-phase of the cell cycle, possibly as a result of oxidative DNA damage. This leads to DNA damage, checkpoint activation and focal accumulation of the DNA response proteins. Interestingly, in human cancer cells exposure to 3-BP also induces DNA breaks that trigger H2A.X phosphorylation. Our current data shed new light on the mechanisms by which a sufficiently low concentration of 3-BP can induce cytotoxicity at the DNA level, a finding that might be important for the future design of anticancer therapies.

Phenotypic and genotypic diversity of airborne fungal spores in Demänovská Ice Cave (Low Tatras, Slovakia).

This paper is the first aero-mycological report from Demänovská Ice Cave. Fungal spores were sampled from the internal and external air of the cave in June, 2014, using the impact method with a microbiological air sampler. Airborne fungi cultured on PDA medium were identified using a combination of classical phenotypic and molecular methods. Altogether, the presence of 18 different fungal spores, belonging to 3 phyla, 9 orders and 14 genera, was detected in the air of the cave. All of them were isolated from the indoor samples, and only 9 were obtained from the outdoor samples. Overall, airborne fungal spores belonging to the genus Cladosporium dominated in this study. However, the spores of Trametes hirsuta were most commonly found in the indoor air samples of the cave and the spores of C. herbarum in the outdoor air samples. On the other hand, the spores of Alternaria abundans, Arthrinium kogelbergense, Cryptococcus curvatus, Discosia sp., Fomes fomentarius, Microdochium seminicola and T. hirsuta were discovered for the first time in the air of natural and artificial underground sites. The external air of the cave contains more culturable airborne fungal spores (755 colony-forming units (CFU) per 1 m3 of air) than the internal air (from 47 to 273 CFU in 1 m3), and these levels of airborne spore concentration do not pose a threat to the health of tourists. Probably, the specific microclimate in the cave, including the constant presence of ice caps and low temperature, as well as the location and surrounding environment, contributes to the unique species composition of aeromycota and their spores in the cave. Thus, aero-mycological monitoring of underground sites seems to be very important for their ecosystems, and it may help reduce the risk of fungal infections in humans and other mammals that may arise in particular due to climate change.

Dark stains on rock surfaces in Driny Cave (Little Carpathian Mountains, Slovakia).

Mycobiota are important in underground ecology. In 2014, we discovered dark stains on clayey sediments on the walls of Driny Cave, Slovakia. Our description is based on the morphology of the fungus and the phylogenetic relationships of the internal transcribed spacer (ITS) region. In addition, data on its capacity for the production of extracellular enzymes, growth, and survival in vitro at different temperatures are reported. Our analyses revealed that this dark stains on the wall was produced by Penicillium glandicola. The fungus was able to synthesize amylases, proteases and cellulases, but not pectinases and keratinases. The vegetative structures of mycelium of this fungus are viable in vitro after storage at cool temperatures (from -72 to 5 °C), and show active growth at temperatures from 5 to 25 °C, but without spore germination, and without active growth at 30 and 37 °C. Penicillium glandicola is a psychrotolerant species and belong to var. glandicola.

Bats Increase the Number of Cultivable Airborne Fungi in the "Nietoperek" Bat Reserve in Western Poland.

The "Nietoperek" bat reserve located in Western Poland is one of the largest bat hibernation sites in the European Union with nearly 38,000 bats from 12 species. Nietoperek is part of a built underground fortification system from WWII. The aims of the study were (1) to determine the fungal species composition and changes during hibernation season in relation to bat number and microclimatic conditions and (2) evaluate the potential threat of fungi for bat assemblages and humans visiting the complex. Airborne fungi were collected in the beginning, middle and end of hibernation period (9 November 2013 and 17 January and 15 March 2014) in 12 study sites, one outside and 11 inside the complex. Ambient temperature (T a) and relative humidity (RH) were measured by the use of data loggers, and species composition of bats was recorded from the study sites. The collision method (Air Ideal 3P) sampler was used to detect 34 species of airborne fungi including Pseudogymnoascus destructans (Pd). The density of airborne fungi isolated from the outdoor air samples varied from 102 to 242 CFU/1 m(3) of air and from 12 to 1198 CFU in the underground air samples. There was a positive relationship between number of bats and the concentration of fungi. The concentration of airborne fungi increased with the increase of bats number. Analysis of other possible ways of spore transport to the underground indicated that the number of bats was the primary factor determining the number of fungal spores in that hibernation site. Microclimatic conditions where Pd was found (median 8.7 °C, min-max 6.1-9.9 °C and 100 %, min-max 77.5-100.0 %) were preferred by hibernating Myotis myotis and Myotis daubentonii; therefore, these species are most probably especially prone to infection by this fungi species. The spores of fungi found in the underground can be pathogenic for humans and animals, especially for immunocompromised persons, even though their concentrations did not exceed limits and norms established as dangerous for human health. In addition, we showed for the first time that the air in bats hibernation sites can be a reservoir of Pd. Therefore, further study in other underground environments and wintering bats is necessary to find out more about the potential threat of airborne fungi to bats and public health.

Mycobiota of Underground Habitats: Case Study of Harmanecká Cave in Slovakia.

Harmanecká Cave is located in the Harmanec Valley to the northwest of Banská Bystrica city, in the southern part of the Great Fatra Mountains, Slovakia. This cave is the most important underground locality of bat occurrence in Slovakia (population of 1000 to 1500 individuals). The study aimed at mycological evaluation of the air, the water, and the rock surface of Harmanecká Cave in Slovakia. The samples were taken on 24 July 2014. To examine the air, the Air Ideal 3P sampler was used. Microbiological evaluation of the rock surface was performed using swab sampling and the water by using the serial dilution technique. The authors observed a relationship between air temperature and the concentration of fungi. The concentration of airborne fungi increased with the increase in the air temperature and decreased with distance from the entrance to the cave. The density of airborne fungi isolated from the outdoor air samples was 810.5 colony-forming units (CFU) per 1 m3 of air and from 27.4 to 128.5 CFU for the indoor air samples. From the rock surface inside the cave, 45.0 to 106.6 CFU per 1 cm2 were isolated, whereas from the water, 29.9 CFU per 1 ml were isolated. Seven species of filamentous fungi were isolated from the external air samples and 12 species of filamentous fungi and 3 species of yeast-like fungi from the internal air samples. From the surface of the rocks inside the cave, 5 species of filamentous fungi and 1 species of yeast-like fungi were cultured, whereas from the water samples, 6 species of filamentous fungi were cultured. Cladosporium spp. were the fungi most frequently isolated from the external air; from the internal air, Penicillium urticae was most frequently isolated; from the rock surface, it was Gliocladium roseum; and from the water, it was P. chrysogenum. The species found in the cave can be pathogenic for humans and animals, especially for immunocompromised persons, and they can also cause biodegradation of the rocks. However, the concentration of airborne fungi inside the cave did not exceed official limits and norms stated as dangerous for the health of tourists.