Chromatographic and Biological Screening of Chosen Species of Schisandraceae Family: Schisandra chinensis, S. rubriflora, S. sphenanthera, S. henryi and Kadsura japonica.

HPLC and TLC profiling was carried out for leaf and fruit extracts of five Schisandraceae species: Schisandra chinensis, S. rubriflora, S. spehenanthera, S. henryi and Kadsura japonica. HPLC measurements confirmed presence of lignans and phenolic compounds in fruits and leaves of all tested species. The most abundant in lignans was S. chinensis fruit extract in which 15 compounds were detected (e. g.: schisandrol A, schisanhenol, γ-schisandrin, gomisin N). The effect-directed detection, i. e., TLC-direct bioautography against Bacillus subtilis, showed exceptionally high activity for S. chinensis and S. rubriflora fruit extracts. On the other hand, TLC-DB enzyme tests (α-glucosidase, lipase, tyrosinase and acetylcholinesterase (AChE) inhibition assays) showed that all fruit and leaf extracts have ability to inhibit the above-mentioned enzymes (except for the K. japonica fruit). The leaf extracts showed much stronger antioxidant activity than the fruit ones, which were assessed and compared using both TLC-direct bioautography and spectrophotometric measurements based on ABTS, DPPH and FRAP tests.

Convergence of residential property prices in the Szczecin agglomeration in the context of the COVID-19 pandemic.

The outbreak of the COVID-19 pandemic has left a great mark on all of humanity. In addition to the immediate threat to health and life, the virus in a way also infected other spheres of human life. The lockdowns, which lasted for many weeks, had a very big impact on various sectors of the economy. Most of them experienced a severe slowdown or recession. However, there were areas that benefited from the fact that the people in many countries stayed at home and worked remotely. Examples of such areas include the computer and tablets market, online stores, and delivery companies. This proves that even in difficult circumstances like a pandemic, we are not facing a total global recession. In such a context, it is worth looking at the impact of the COVID-19 pandemic on the real estate market. This area of the economy can experience the impact of the virus in different ways. On the one hand, a general increase in global uncertainty can cause certain investment projects to freeze or be abandoned. Lockdowns cause difficulties in accessing workers at construction sites. Populations losing their jobs during a pandemic have difficulty paying rent or mortgage payments. On the other hand, rising inflation and low interest rates on bonds and bank deposits may encourage people to protect their savings by buying property. Remote working increases the demand for larger apartments that need to serve as offices in addition to residential functions. Remote work also means not having to commute to workplaces, which may encourage people to move to cheaper locations without having to give up a well-paying job in a larger but also more expensive city. These two groups of impulses are contradictory and may have a completely different impact on the shape of the real estate market understood as the structure of prices and the number of transactions concluded. The study is part of a broader research effort to assess the impact of the COVID-19 pandemic on various areas of the economy. Previous research conducted in the Szczecin agglomeration proved the existence of the price divergence phenomenon. The main city of the agglomeration showed a tendency to pull in the economic potential from the region, which translated into a faster increase in real estate prices in the capital of the region in comparison with the surrounding cities. The aim of this study is to verify whether the pandemic and the socio-economic processes triggered by it, such as the development of remote working, changed the process of divergence into a process of convergence, but whether the faster growth of residential property prices in the main city of the agglomeration, which started years ago, has not been stopped. For this purpose, α-convergence and σ-convergence models were used. The study covers the period 2015-2021. The study area is the city of Szczecin and the four largest cities adjacent to the capital of the West Pomeranian Voivodeship in Poland.

In Vitro Activity of Ebselen and Diphenyl Diselenide Alone and in Combination with Drugs against Trichophyton mentagrophytes Strains.

BACKGROUND: Dermatophytoses are one of the most prevalent infectious diseases in the world for which the pace of developing new drugs has not kept pace with the observed therapeutic problems. Thus, searching for new antifungals with an alternative and novel mechanism of action is necessary. OBJECTIVE: This study aimed to evaluate the antifungal activity of ebselen and diphenyl diselenide against Trichophyton mentagrophytes clinical isolates. METHODS: In vitro antifungal susceptibility was assessed for organoselenium compounds used alone or in combination with allylamines and azoles according to the 3rd edition of the CLSI M38 protocol. RESULTS: Ebselen demonstrated high antifungal activity with MICGM equal to 0.442 µg/mL and 0.518 µg/mL in the case of human and animal origin strains, respectively. The values of MICGM of diphenyl diselenide were higher: 17.36 µg/mL and 13.45 µg/mL for the human and animal isolates, respectively. Synergistic or additive effects between terbinafine and ebselen or diphenyl diselenide were observed in the case of 12% and 20% strains, respectively. In turn, the combination of itraconazole with diphenyl diselenide showed a synergistic effect only in the case of 6% of the tested strains, whereas no synergism was shown in the combination with ebselen. CONCLUSIONS: The results highlight the promising activity of organoselenium compounds against Trichophyton mentagrophytes. However, their use in combinational therapy with antifungal drugs seems to be unjustified due to the weak synergistic effect observed.

Laboratory Diagnosis and In Vitro Antifungal Susceptibility of Trichophyton quinckeanum from Human Zoonoses and Cats.

The "One Health" concept increasingly demonstrates the global spread of pathogenic (also eukaryotic) microorganisms and their zoonotic potential. Dermatophytes can cause superficial mycoses in humans and animals. Furthermore, the number of transmissions from asymptomatic carriers to humans has been on the rise over the last few years. This study was focused on the detailed characterisation of clinical isolates of Trichophyton quinckeanum with epidemiological analyses and characterisation of their in vitro antifungal susceptibility patterns. The isolated dermatophytes were identified with a combination of conventional and molecular methods. In turn, their susceptibility in vitro was tested according to the Clinical and Laboratory Standards Institute (CLSI) M38 ed.3 protocol. A total of 36 strains were isolated, with 21 cases of T. quinckeanum zoonoses resulting from direct contact with symptomatic cats (58.3%). The other 15 strains (41.7%) were isolated simultaneously from healthy cats and their owners. All strains showed high susceptibility to allylamine, pyridinone, and phenyl morpholine derivatives but were resistant to fluconazole and ketoconazole. In conclusion, our study shows the frequency of zoonoses contracted from asymptomatic cats. Moreover, the antifungal susceptibility profiles indicate the serious risk posed to animal owners by resistant strains of T. quinckeanum, which are often responsible for recalcitrant-to-treatment cases.

Airborne dermatophyte propagules concentration in cowsheds as an underestimated reservoir of potential zoonoses.

AIMS: Indoor air quality in stables, cowsheds or henhouses has recently become in interest due to the potential risks of zoonotic infections. Cowsheds are commonly known to have high fungal loads, particularly if insufficient attention is paid to the monitoring and control of the indoor microclimate around three elements, that is heating, ventilation and lighting. The aim of this study was to determine the concentrations and spectrum of dermatophyte propagules in the indoor air of cowsheds. METHODS AND RESULTS: Air samples were collected on five farms, and the dermatophyte species were identified using MALDI-TOF MS analysis. The quantitative analysis of the fungal pollutants showed an average of 0.084 dermatophyte propagules (CFU) per m3 of flowing air in spring and 0.0239 CFU/m3 in the summer. Dermatophyte species were identified in case of 64.6% of the obtained colonies. Trichophyton verrucosum as dominant species was isolated on all five farms. In turn, Nannizzia gypsea was isolated on four farms and Trichophyton mentagrophytes as well as Paraphyton cookei were isolated on two farms. CONCLUSIONS: This study demonstrated that indoor aerosol appears to be one of the underestimated risks of dermatophyte infections. Moreover, the risk of zoonotic infections is posed by airborne zoophilic dermatophytes, especially T. verrucosum, whose prevalence of infections has been increasing in recent years. SIGNIFICANCE AND IMPACT OF THE STUDY: The ability of dermatophytes to infect animals and humans is thought to be a consequence of not only their adaptation to new ecological niches but also occurring as an aerosol component, which we demonstrate for the first time in this study. The microclimate of the cowshed may be an underestimated reservoir of zoophilic dermatophytes, which pose a zoonotic threat to farmers, animal breeders and veterinarians.

Update on Stachybotrys chartarum-Black Mold Perceived as Toxigenic and Potentially Pathogenic to Humans.

In nature, there are many species of fungi known to produce various mycotoxins, allergens and volatile organic compounds (VOCs), as well as the commonly known etiological agents of various types of mycoses. So far, none of them have provoked so much emotion among homeowners, builders, conservators, mycologists and clinicians as Stachybotrys chartarum. This species compared to fungi of the genera Fusarium and Aspergillus is not as frequently described to be a micromycete that is toxigenic and hazardous to human and animal health, but interest in it has been growing consistently for three decades. Depending on the authors of any given review article, attention is focused either on the clinical aspects alongside the role of this fungus in deterioration of biomaterials, or aspects related to its biology, ecology and taxonomic position. On the one hand, it is well established that inhalation of conidia, containing the highest concentrations of toxic metabolites, may cause serious damage to the mammalian lung, particularly with repeated exposure. On the other hand, we can find articles in which authors demonstrate that S. chartarum conidia can germinate and form hyphae in lungs but are not able to establish an effective infection. Finally, we can find case reports that suggest that S. chartarum infection is linked with acute pulmonary hemorrhage, based on fungal structures recovered from patient lung tissue. New scientific reports have verified the current state of knowledge and note that clinical significance of this fungus is exceedingly controversial. For these reasons, understanding S. chartarum requires reviewing the well-known toxigenic features and harmful factors associated with this fungus, by gathering the newest ones into a coherent whole. The research problem related to this fungus seems to be not overly publicized, and there is still a demand to truthfully define the real threats of S. chartarum and phylogenetically related species. The most important problem, which should be fully elucidated as soon as possible, remains the clarification of the pathogenicity of S. chartarum and related species. Maybe it is urgent time to ask a critical question, namely what exactly do we know 28 years after the outbreak of pulmonary hemorrhage in infants in Cleveland, Ohio, USA most likely caused by S. chartarum?

Comparative characteristics of sequence types, genotypes and virulence of multidrug-resistant Enterococcus faecium isolated from various hosts in eastern Poland. Spread of clonal complex 17 in humans and animals.

The aim of the study was to carry out a comparative analysis of the occurrence of sequence types, genotypes, and virulence genes of multidrug-resistant Enterococcus faecium isolated from humans, domestic animals, and wildlife from eastern Poland. The genetic correlation of strains was determined using multilocus sequence typing, and the method of Amplification of DNA fragments Surrounding Rare Restriction Sites. The presence of 49 different STs (including 12 not reported previously) was shown. All human isolates, 56% of E. faecium isolated from poultry and 40% isolated from wildlife, belonged to the hospital-adapted CC17. E. faecium CC17 were characterized by statistically higher resistance to aminoglycosides and penicillin, the presence of the aac(6')-Ie-aph(2″)-Ia and hyl gene and mutations in gyrA and parC, compared to other strains (p > 0.05). E. faecium were classified into 73 genotypes grouped into clusters including isolates from the same host species. The present study showed that the virulence, resistance, and genotype profiles of E. faecium were correlated with the individual host but not with the sequence type. The occurrence of E. faecium CC17 in both humans and animals proves the large circulation of the complex among various hosts.

European Hedgehogs (Erinaceus europaeus L.) as a Reservoir of Dermatophytes in Poland.

The European hedgehog (Erinaceus europaeus Linnaeus) frequently colonises areas located close to human life in cities, as these are more suitable nest sites offering an abundance of food and allowing avoidance of predators. However, urbanisation has a significant impact on the epidemiology of infectious diseases, including dermatophytoses, the primary source of which are wild animals. In this study, we determined the spectrum of dermatophytes isolated from the European hedgehog and assessed their susceptibility profile to antifungal drugs. Symptomatic and asymptomatic dermatophyte infections were observed in 7.7% and 8% of the 182 examined free-living hedgehogs, respectively. In the pool of the isolated dermatophyte strains, Trichophyton erinacei was dominant (29.9%), followed by Trichophyton mentagrophytes (17.9%), Trichophyton benhamiae (13.4%), Nannizzia gypsea (11.9%), Microsporum canis (10.4%), Nannizzia nana (7.5%), Paraphyton cookei (6.0%), and Nannizzia fulva (3.0%). Susceptibility tests revealed the highest activity of luliconazole and the lowest of activity fluconazole among the azole drugs applied. Although terbinafine generally exhibited high efficacy, two Trichophyton mentagrophytes isolates showed resistance to this drug (MIC = 2 µg/ml) resulting from missense mutations in the SQLE gene corresponding to the amino acid substitution Leu393Phe. Summarising, our study has also revealed that such wildlife animals as hedgehogs can be a reservoir of pathogenic human dermatophytes, including harmful strains resistant to commonly used antifungal drugs.

A rich mosaic of resistance in extended-spectrum ß-lactamase-producing Escherichia coli isolated from red foxes (Vulpes vulpes) in Poland as a potential effect of increasing synanthropization.

In our research, we analyzed the resistance of cephalosporin-resistant E. coli strains to antimicrobial agents. The strains were collected during five years from wild animal species commonly inhabiting Poland. We have identified the type of ß-lactamases produced and the multidrug-resistance profile. Most strains (73.8%) had genes encoding ESBL enzymes, mainly CTX-M-1 and TEM. Almost all AmpC-ß-lactamase-producing isolates had the blaCMY-2 gene. Almost 70% of the strains tested showed a multi-drug resistance profile. The dominant phenotype was resistance to tetracycline (69.05%), and/or sulfamethoxazole (57.1%). We also found high resistance to quinolones: ciprofloxacin 35.7% and nalidixic acid 52.4%. The phenotypic resistance of the strains was in most cases confirmed by the presence of corresponding genes. Among strains, 26.2% were carriers of plasmid-mediated quinolone resistance genes (PMQR). MLST analysis revealed a large clonal variation of the strains, which was reflected in 28 different sequence types. More than half of the strains (54.7%) were classified into the following sequence complexes: 10, 23, 69, 101, 155, 156, 168, 354, 398, 446, and 648. Only one strain in the studied group was assigned to the ExPEC pathotype and represented sequence type 117. The results of our research have confirmed that isolates obtained from wild animals possess many resistance determinants and sequence types, which are also found in food-producing animals and humans. This reflects the doctrine of "One health", which clearly indicates that human health is inextricably linked with animal health as well as degree of environmental contamination. We conclude that the resistance and virulence profiles of strains isolated from wildlife animals may be a resultant of various sources encountered by animals, creating a rich and varied mosaic of genes, which is very often unpredictable and not reflected in the correlation between the sequence type and the gene profile of resistance or virulence observed in epidemic clones.

New Reference Genes for qRT-PCR Analysis as a Potential Target for Identification of Trichophyton verrucosum in Different Culture Conditions.

Dermatophytes are a group of filamentous fungi infecting skin, hair, and nails that raise great diagnostic difficulties. qRT-PCR is a reliable technique for quantifying gene expression with increasingly frequent use in mycological diagnostics. Knowledge of genes and molecular markers with potential to be used in the identification of dermatophytes is of great importance for the development of this branch of diagnostics. In this article, the suitability of six candidate reference genes (TUBB, ACTB, ADPRF, RPL2, SDHA, and EEF1A1) was investigated for gene expression analysis in the dermatophyte Trichophyton verrucosum, which was cultured in various mycological media that are commonly used in a diagnostic laboratory, i.e., Sabouraud, potato dextrose, and keratin-supplemented MM-Cove. The different culture conditions are extremely important factors for the growth and physiology of dermatophytes. Gene expression stability was evaluated using geNorm, NormFinder, BestKeeper, and RefFinder algorithms. Regarding the stability of expression, SDHA was the most stable housekeeping gene; hence, this gene is recommended for future qRT-PCR studies on T. verrucosum strains. These results allow us to conclude that the SDHA gene can be an additional good candidate as an identification target in the qRT-PCR technique.

Virulence and Antimicrobial Resistance Pattern of Aeromonas spp. Colonizing European Pond Turtles Emys orbicularis and Their Natural Environment. First Study from Poland.

The aim of the study was to isolate and identify species belonging to the Aeromonas genus and evaluate the antimicrobial resistance and virulence patterns of isolates colonizing European pond turtles (Emys orbicularis) from natural environment of Eastern Poland. In total, 74 turtles and 15 samples of water from their natural environment were examined. More than 40 strains were isolated and identified: A. bestiarum (n = 1), A. hydrophila (n = 13), A. allosaccharophila (n = 2), A. salmonicida (n = 3), and A. veronii (n = 23). The highest incidence of resistance was noted for ampicillin (100%) and sulfamethoxazole (62.0%), followed by erythromycin and colistin (both 40.5%). Moreover, eight strains were intermediately resistant to meropenem (19%). Most Aeromonas isolates were found to possess more than one virulence gene among fla, aer, hlyA, act, ela, alt, and ast. We showed that the population of free-living European pond turtles was highly colonized by Aeromonas spp. Such strains may be an infectious agent not only for the population of turtles but also for other species of animals inhabiting their natural environment. Moreover, the undesirable properties of water quality caused by the presence of drug-resistant aeromonads could have a negative impact on human health.

Cold atmospheric pressure plasma (CAPP) as a new alternative treatment method for onychomycosis caused by Trichophyton verrucosum: in vitro studies.

PURPOSE: Anthropophilic dermatophytes as etiological factors of onychomycoses are more common than zoophilic fungi. In the case of the latter, reverse zoonoses are possible, which poses a threat to the persistence of dermatophytes in the environment. Nevertheless, without treatment, both types of tinea unguium may lead to complete nail plate destruction and secondary mixed infections with fungi and bacteria. One of the zoophilic dermatophytes that cause onychomycosis is Trichophyton verrucosum, whose prevalence has been increasing in recent years. Such infections are usually treated with allylamines and/or azoles, but such a conventional treatment of infections caused by T. verrucosum often fails or is discontinued by patients. METHODS: Herein, we reveal the results of our in vitro studies related to direct application of cold atmospheric pressure plasma (CAPP) on Trichophyton verrucosum growth, germination and adherence to nail as a new alternative treatment method of such types of dermatomycoses. RESULTS: Our in vitro studies showed that, while exposure to CAPP for 10 min delays germination of conidia and clearly impairs the fitness of the fungal structures, 15 min is enough to kill all fungal elements exposed to plasma. Moreover, the SEM images revealed that T. verrucosum cultures exposed to CAPP for 10 and 15 min were not able to invade the nail fragments. CONCLUSION: The results revealed that single exposure to CAPP was able to inhibit T. verrucosum growth and infection capacity. Hence, cold atmospheric pressure plasma should be considered as a promising alternative treatment of onychomycoses.

Modulation of ERG gene expression in fluconazole-resistant human and animal isolates of Trichophyton verrucosum.

Dermatophytes are a group of eukaryotic microorganisms characterized by high capacity to colonize keratinized structures such as the skin, hair, and nails. Over the past years, the incidence of infections caused by zoophilic species, e.g., Trichophyton verrucosum, has been increasing in some parts of the world, especially in Europe. Moreover, the emergence of recalcitrant dermatophytoses and in vitro resistant dermatophytes has become a cause of concern worldwide. Here, we analyzed the mechanisms underlying resistance to fluconazole among clinical isolates of T. verrucosum. Quantitative RT-PCR was carried out to determine the relative expression levels of mRNA transcripts of ERG3, ERG6, and ERG11 genes in the fungal samples using the housekeeping gene GAPDH as a reference. Our results showed that the upregulation of the ERG gene expression is a possible mechanism of resistance to fluconazole in this species. Furthermore, ERG11 is the most statistically significantly overexpressed gene in the pool of fluconazole-resistant T. verrucosum isolates. Additionally, we have demonstrated that exposure to fluconazole increases the levels of expression of ERG genes in fluconazole-resistant isolates of T. verrucosum. In conclusion, this study has shown one of the possible mechanisms of resistance to fluconazole among zoophilic dermatophytes, which involves the maintenance of high levels of expression of ERG genes after drug exposure.

Real-Time PCR as an Alternative Technique for Detection of Dermatophytes in Cattle Herds.

Dermatophytes are filamentous fungi with the ability to digest and grow on keratinized substrates. The ongoing improvements in fungal detection techniques give new scope for clinical implementations in laboratories and veterinary clinics, including the monitoring of the disease and carrier status. The technologically advanced methods for dermatophyte detection include molecular methods based on PCR. In this context, the aim of this study was to carry out tests on the occurrence of dermatophytes in cattle herds using qPCR methods and a comparative analysis with conventional methods. Each sample collected from ringworm cases and from asymptomatic cattle was divided into three parts and subjected to the real-time PCR technique, direct light microscopy analysis, and culture-based methods. The use of the real-time PCR technique with pan-dermatophyte primers detected the presence of dermatophytes in the sample with a 10.84% (45% vs. 34.17%) higher efficiency than direct analysis with light microscopy. Moreover, a dermatophyte culture was obtained from all samples with a positive qPCR result. In conclusion, it seems that this method can be used with success to detect dermatophytes and monitor cowsheds in ringworm cases and carriers in cattle.

In vitro evaluation of photodynamic activity of methylene blue against Trichophyton verrucosum azole-susceptible and -resistant strains.

The intense search for the "Holy Grail" of antifungal therapy can be observed today. The searches are not limited only to discovery of potential antifungal drugs, but also new therapeutic strategies involving the use of chemosensitizers to achieve synergistic effect or physicochemical factors inducing stress conditions in fungal cells. In this study was examined in vitro effectiveness of photodynamic antifungal strategy with methylene blue using a light beam with a wavelength equal to 635 nm toward the Trichophyton verrucosum susceptible and itraconazole- and/or fluconazole-resistant strains. Methylene blue used at concentration equal to 5 µg/mL and in the presence of 40 J/cm2 of light energy showed fungicidal effect toward the susceptible strains. However, for azole-resistant isolates, only the energy dose equal to 60 J/cm2 at 5 µg/mL of methylene blue allowed to kill the pathogen. This study confirms that methylene blue induced by red light has a definite inhibitory effect on zoophilic dermatophytes.

Dataset of the next-generation sequencing of variable 16S rRNA from bacteria and ITS2 regions from fungi and plants derived from honeybees kept under anthropogenic landscapes.

Forager Apis melliefera honeybees were collected from four localities located in Europe, i.e.: London, UK; Athens, Greece; Marchamalo, Spain and Lublin, Poland. Furthermore, from Asia we have collected A. mellifera as well as A. cerana foragers form Chiang Mai in Thailand We used next generation sequencing (NGS) to analyse the 16S rRNA bacterial gene amplicons based on the V3-V4 region and the ITS2 region from fungi and plants derived from honeybee samples. Amplicon libraries, were prepared using the 16S Metagenomic Sequencing Library Preparation, Preparing 16S Ribosomal RNA Gene Amplicons for the Illumina MiSeq System (Illumina®) protocol. NGS raw data are available at https://www.ncbi.nlm.nih.gov/bioproject/PRJNA686953. Furthermore, isolated DNA was used as the template for screening pathogens: Nosema apis, N. ceranae, N. bombi, tracheal mite (Acarapis woodi), any organism in the parasitic order Trypanosomatida, including Crithidia spp. (i.e., Crithidia mellificae), neogregarines including Mattesia and Apicystis spp. (i.e., Apicistis bombi). The presented data can be used to compare the metagenomic samples from different honeybee population all over the world. A higher load of fungi, and bacteria groups such as: Firmicutes (Lactobacillus); γ- proteobacteria, Neisseriaceae, and other unidentified bacteria was observed for Nosema cearana and neogregarines infected honeybees. Healthy honeybees had a higher load of plant pollens, and bacteria groups such as: Orbales, Gilliamella, Snodgrassella, and Enterobacteriaceae. More details can be found in research article [1] Ptaszynska et al. 2021.

Effect-directed analysis as a method for quality and authenticity estimation of Rhodiola rosea L. preparations.

Adulterations of food and pharmaceutical preparations are the important global problem. On the one hand, fraud practices are becoming more and more sophisticated while on the other, monitoring and uncovering falsifications are insufficient. One of the most common consumer concern is the quality and authenticity of the purchased products, related to the confidence that they have composition and properties in accordance with the manufacturer's declaration on the label. This refers also to pharmaceuticals potentially delivering great health benefits such as Rhodiola rosea L. supplements. The aim of this study was defining authenticity and possible adulterations of two R. rosea preparations basing on their TLC-bioprofiles and the presence of biomarker compounds characteristic for this plant. The effect-directed analysis (EDA), i.e. TLC hyphenated with micro-chemical and biological assays performed directly on TLC plates followed by HPLC-ESI-MS was used for the bioprofiling of antioxidants, antibacterials, and inhibitors of lipase, acetylcholine, α-glucosidase and tyrosinase as well as for the identification of the biomarkers. The results pointed to the possible adulteration of one of the tested products related to the absence of two rosavins, the most important quality markers of R. rosea.

Amplicon Sequencing of Variable 16S rRNA from Bacteria and ITS2 Regions from Fungi and Plants, Reveals Honeybee Susceptibility to Diseases Results from Their Forage Availability under Anthropogenic Landscapes.

European Apis mellifera and Asian Apis cerana honeybees are essential crop pollinators. Microbiome studies can provide complex information on health and fitness of these insects in relation to environmental changes, and plant availability. Amplicon sequencing of variable regions of the 16S rRNA from bacteria and the internally transcribed spacer (ITS) regions from fungi and plants allow identification of the metabiome. These methods provide a tool for monitoring otherwise uncultured microbes isolated from the gut of the honeybees. They also help monitor the composition of the gut fungi and, intriguingly, pollen collected by the insect. Here, we present data from amplicon sequencing of the 16S rRNA from bacteria and ITS2 regions from fungi and plants derived from honeybees collected at various time points from anthropogenic landscapes such as urban areas in Poland, UK, Spain, Greece, and Thailand. We have analysed microbial content of honeybee intestine as well as fungi and pollens. Furthermore, isolated DNA was used as the template for screening pathogens: Nosema apis, N. ceranae, N. bombi, tracheal mite (Acarapis woodi), any organism in the parasitic order Trypanosomatida, including Crithidia spp. (i.e., Crithidia mellificae), neogregarines including Mattesia and Apicystis spp. (i.e., Apicistis bombi). We conclude that differences between samples were mainly influenced by the bacteria, plant pollen and fungi, respectively. Moreover, honeybees feeding on a sugar based diet were more prone to fungal pathogens (Nosema ceranae) and neogregarines. In most samples Nosema sp. and neogregarines parasitized the host bee at the same time. A higher load of fungi, and bacteria groups such as Firmicutes (Lactobacillus); γ-proteobacteria, Neisseriaceae, and other unidentified bacteria was observed for Nosema ceranae and neogregarine infected honeybees. Healthy honeybees had a higher load of plant pollen, and bacteria groups such as: Orbales, Gilliamella, Snodgrassella, and Enterobacteriaceae. Finally, the period when honeybees switch to the winter generation (longer-lived forager honeybees) is the most sensitive to diet perturbations, and hence pathogen attack, for the whole beekeeping season. It is possible that evolutionary adaptation of bees fails to benefit them in the modern anthropomorphised environment.

Therapeutic efficacy of topically used luliconazole vs. terbinafine 1% creams.

OBJECTIVES: Dermatomycoses of zoophilic origin, especially those caused by Trichophyton mentagrophytes, often pose considerable therapeutic problems. This is reflected in the growing number of strains of this species with resistance to terbinafine caused by a mutation in the squalene epoxidase (SQLE) gene. Therefore, it is reasonable to look for alternative therapies to the commonly used terbinafine. The aim of the present study was to assess the in vivo effectiveness of topical therapy with luliconazole or terbinafine 1% cream. METHODS: Therapeutic efficacy was assessed using direct examination in KOH with DMSO, qPCR analysis with pan-dermatophyte primers and culturing. Moreover, in vitro susceptibility tests for luliconazole and terbinafine were performed. RESULTS: The results demonstrated significantly higher antifungal activity of luliconazole than terbinafine against dermatomycoses caused by T. mentagrophytes. The geometric mean of the MIC value for luliconazole against all T. mentagrophytes strains was 0.002 µg/ml, while this value for terbinafine was 0.004 µg/ml. In all studied cases, 28-day local therapy with luliconazole contributed to complete eradication of the aetiological agent of infection. CONCLUSIONS: Given the increasingly frequent reports of difficult-to-treat dermatophytoses caused by zoophilic terbinafine-resistant strains, the 1% luliconazole cream can be alternative solution in topical therapy.

Comparative study of multidrug-resistant Enterococcus faecium obtained from different hosts.

Introduction. The possible transfer of antimicrobial resistance genes between Enterococcus faecium isolates from humans and different animal species, including those not covered by monitoring programs (e.g. pet and wildlife), poses a serious threat to public health.Hypothesis/Gap Statement. Little is known about occurrence and mechanisms of phenomenon of multidrug resistance of E. faecium isolated from various host species in Poland.Aim. The aim of the study was to characterize multidrug-resistant E. faecium isolated from humans and animals (livestock, pets and wildlife) in terms of the occurrence of genetic markers determining resistance.Methodology. Bacterial isolates were tested for phenotypic resistance and the presence of genes encoding resistance to macrolides, tetracycline, aminoglycosides, aminocyclitols and phenicols as well as efflux pump (emeA), resolvase (tndX) and integrase (Int-Tn) genes. The quinolone resistance-determining regions of gyrA and parC were sequenced.Results. Human isolates of E. faecium were characterized by high-level resistance to: ciprofloxacin, enrofloxacin, erythromycin (100 %), as well, as aminoglycosides resistance (kanamycin - 100%, streptomycin - 78 %, gentamicin - 78%). Regardless of the animal species, high level of resistance of E. faecium to tetracycline (from 88-100 %), erythromycin (from 82-94 %) and kanamycin (from 36-100 %) was observed. All E. faecium isolates from wildlife were resistant to fluoroquinolones. However, full susceptibility to vancomycin was observed in all isolates tested. Phenotypic antimicrobial resistance of E. faecium was identified in the presence of the following resistance genes: erm(B) (70%), msr(A) (50 %), tet(L) (35 %), tet(K) (34 %), tet(M) (76 %), aac(6')-Ie-aph(2″)-Ia (25%), ant(6)-Ia (31%), aph(3)-IIIa (68 %), (tndX) (23 %), and integrase gene (Int-Tn) (34 %). A correlation between an amino acid substitution at positions 83 and 87 of gyrA and position 80 of parC and the high-level fluoroquinolone resistance in E. faecium has been observed as well.Conclusion. The level and range of antimicrobial resistance and the panel of resistance determinants is comparable between E. faecium isolates, despite host species.