Cytokine inflammatory response in dairy cows with mastitis caused by Streptococcus agalactiae.

Introduction: The aim of the study was evaluation of the concentrations of interleukin (IL)-1ß, IL-8, IL-12ß and tumour necrosis factor alpha (TNF-α) in the serum and milk of cows with mastitis caused by Streptococcus agalactiae. Material and Methods: A total of 60 milk samples from diseased cows and 30 milk samples from healthy cows were included in the study. Blood and milk samples were taken from Holstein-Friesian cows from three herds (two in tie-stall and one in a free-stall housing system) in Lublin Province in Poland. The concentrations of cytokines in blood serum and quarter milk samples were determined by ELISA. Results: The levels of IL-1ß, IL-8, IL-12ß and TNF-α were significantly higher in the milk of cows suffering from mastitis caused by S. agalactiae compared to the milk of healthy cows (263.03 vs 55.36 pg/mL, 298.34 vs 131.82 pg/mL, 604.10 vs 139.17 pg/mL and 460.86 vs 78.82 pg/mL, respectively). In the group of sick cows, cytokine levels were significantly higher in milk than in serum (263.03 vs 55.25 pg/mL for IL-1ß, 298.34 vs 164.22 pg/mL for IL-8, 604.10 vs 70.34 pg/mL for IL-12ß and 460.86 vs 104.78 pg/mL for TNF-α). Conclusion: The results confirm the involvement of the entire bovine immune system to protect against the bacteria first locally in the udder. The response of the mammary gland to infection caused by S. agalactiae is rapid and already very strong at the beginning of the infection.

Multi-Host Pathogen Staphylococcus aureus-Epidemiology, Drug Resistance and Occurrence in Humans and Animals in Poland.

Staphylococcus aureus is a drug resistant pathogen with zoonotic potential commonly isolated from humans and animals. The aim of this study was to compare the occurrence of drug resistance, resistance genes, sequence types (STs), and genotypes of S. aureus isolated from humans, livestock, and wildlife in eastern Poland. A high percentage of isolates resistant to penicillin (63%), erythromycin (39%), clindamycin (37%), tetracycline (31%), and methicillin (MRSA-19%), an intermediate resistant to vancomycin (VISA-13%), and a multidrug resistant (MDR-39%) was obtained. Multilocus sequence typing analysis showed the presence of 35 different STs (with dominance ST 15, ST 45, ST 7, and ST 582 in human, and ST 398 and ST 8139 in porcine and cattle isolates, respectively), including 9 new ones that had never been reported before (ST 8133-8141). Identical genotypic patterns were detected among porcine and cattle isolates as well as from humans and cattle. A high percentage of MDR, MRSA, and VISA in humans and livestock combined with the presence of the same genotypes among S. aureus isolated from human and cattle indicates the circulation of strains common in the region and their zoonotic potential. There is a need to develop new strategies to counteract this phenomenon according to the One Health policy.

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.

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.

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.

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.

Complementary effect of mechanism of multidrug resistance in Trichophyton mentagrophytes isolated from human dermatophytoses of animal origin.

BACKGROUND: Dermatophytoses have gained interest worldwide due to the increased resistance to terbinafine and azoles and difficulty in management of these refractory diseases. OBJECTIVES: In this study, we identified and analysed Trichophyton mentagrophytes clinical isolates obtained from humans with infections of animal origin. METHODS: We used quantitative real-time PCR (qRT-PCR) to examine the transcriptional modulation of three MDR genes (PDR1, MDR2 and MDR4) and analysed squalene epoxidase (SQLE) gene sequences from multidrug-resistant Trichophyton mentagrophytes isolates. RESULTS: The expression profile revealed a 2- to 12-fold increase in mRNA accumulation in the presence of any of the antifungals, compared to cells incubated without drugs. A statistically significant relationship between the isolates exposed to itraconazole and increased expression of the tested genes was revealed. Substantially lower transcription levels were noted for cells exposed to luliconazole, that is, a third-generation azole. Additionally, in the case of 50% of terbinafine-resistant strains, Leu397Phe substitution in the SQLE gene was detected. Furthermore, the reduced susceptibility to itraconazole and voriconazole was overcome by milbemycin oxime. CONCLUSIONS: In conclusion, our study shed more light on the role of the ABC transporter family in T. mentagrophytes, which, if overexpressed, can confer resistance to single azole drugs and even cross-resistance. Finally, milbemycin oxime could be an interesting compound supporting treatment with azole drugs in the case of refractory dermatomycoses.

Diagnostic and epidemiological analysis of Trichophyton benhamiae infection on an alpaca (Vicugna pacos) farm in Poland.

Alpacas (Vicugna pacos) are growing in popularity and are increasingly being presented for veterinary care. Literature reports indicate that dermatophytosis occurring in alpacas accounted for about 3% of dermatological diagnoses. However, there are no reports regarding species of dermatophytes associated with alpacas and reservoirs of infection. In this study, we investigate the diagnosis and epidemiological origin procedure and the virulence enzymes activities of Trichophyton benhamiae isolates obtained from alpacas from a breeding farm. Identification was carried out traditionally by correlating clinical manifestations with micro- and macroscopic examination, and molecular differentiation methods based on Internal Transcribed Spacer (ITS) sequences. Epidemiological analysis was carried out on the basis of Melting Point PCR (MP -PCR) and Amplified Fragment Lenght Polymorphism (AFLP) genotyping. The production of virulence factors was evaluated phenotypically using specific test media. The results obtained from diagnostic tests indicated that the etiological factor of dermatophytosis is T. benhamiae. The same species was also isolated from cowsheds and insects. The MP-PCR and AFLP analyses indicated high invariability of the genomes of the strains isolated from the animals, cowsheds, and insects. In conclusion, animal husbandry outside the natural ecological niche may increase predisposition to dermatophytosis. The treatment of animals alone is insufficient, one should be aware that only elimination of all fungal sources is a long-term success and the key point of therapy.

Analysis of the occurrence and molecular characteristics of drug-resistant strains of Enterococcus faecalis isolated from the gastrointestinal tract of insectivorous bat species in Poland: A possible essential impact on the spread of drug resistance?

Bats are poorly understood as a reservoir of multidrug-resistant strains; therefore, the aim of this study was to determine molecular characterization of multidrug-resistant Enterococcus strains isolated from bat species from Poland. A multi-stage analysis based on targeted isolation of drug-resistant strains (selective media with tetracycline, chloramphenicol, gentamicin, streptomycin, and vancomycin), determination of the phenotypic profile of drug-susceptibility using the disc diffusion method, and amplification of DNA fragments surrounding rare restriction sites (ADSRRS-fingerprinting) was used for the isolation and differentiation of strains. The applied strategy finally allowed identification of E. faecalis resistant to at least one antimicrobial in 47.2% of the single-animal group and in 46.9% of the pooled samples of bat's guano. Out of the 36 distinct isolates, 69% met the criteria of multi-drug resistance, with a dominant combination of resistance to tetracycline, erythromycin, and rifampicin. Simultaneously, 41.6% of the strains were high-level aminoglycoside resistant (HLAR). In most strains, phenotypic resistance was reflected in the presence of at least one gene encoding resistance to a given drug. Moreover, our research results show that some genes were detected simultaneously in the same strain statistically significantly more frequently. This may confirm that the spread of some genes (tetM and ermB or aph (3')-IIIa as well as gelE and aac (6')-Ie-aph (2″)-Ia or ant (6)-Ia) is associated with their common occurrence on the same mobile genetic element. To our knowledge, this is the first analysis of multidrug-resistance among E. faecalis isolated from bats. Our research demonstrates that the One Health concept is not associated exclusively with food-producing animals and humans, but other species of wildlife animals should be covered by monitoring programs as well. We confirmed for the first time that bats are an important reservoir of multi-resistant E. faecalis strains and could have a great impact on environmental resistance.

Wildlife omnivores and herbivores as a significant vehicle of multidrug-resistant and pathogenic Escherichia coli strains in environment.

The phenomenon of resistance of Escherichia coli strains in free-living animals has been constantly expanding in recent years. However, the data are still fragmented and, due to the growing threat to public health, there is a constant need to search for and analyse new reservoirs and indicate their role and importance in the circulation of resistance genes in the environment. Therefore, the target group in this study were free-living non-predatory animals as reservoirs of drug-resistant and potentially virulent E. coli strains. We obtained 70 different isolates, including 71.4% of multidrug-resistant strains. In strains isolated from all species of animals, we determined high resistance to ampicillin (95.7%), tetracycline (64.3%), streptomycin (51.4%) and chloramphenicol (38.6%). Every third of the E. coli-positive individual was a carrier of more than one resistant clone. Moreover, 11.4% of isolates among the resistant strains had the ExPEC, ETEC, or EHEC pathotype. Our study confirmed that not only free-living predatory animals are reservoirs of resistance but also many synanthropic species of herbivores and omnivores contribute substantially to the spread of resistant and virulent E. coli strains.

Wildlife Carnivorous Mammals As a Specific Mirror of Environmental Contamination with Multidrug-Resistant Escherichia coli Strains in Poland.

In recent decades, the number of studies on the occurrence of resistant strains in wildlife animals has increased significantly, but data are still fragmentary. The aim of this study was to evaluate drug resistance of Escherichia coli strains isolated from wild carnivorous mammals, common in Poland. Selective media with antimicrobials (tetracycline, kanamycin, chloramphenicol, and cefotaxime) were used for isolation. Of 53 isolates shown to be distinct by the amplification of DNA fragments surrounding rare restriction site-fingerprinting method, 77.8% were multidrug-resistant (multidrug-resistant). All strains were resistant to ampicillin and many of them also exhibited resistance to tetracycline (76.2%), sulfamethoxazole (57.1%), streptomycin and kanamycin (49.2%), chloramphenicol (30.1%), and nalidixic acid (46%). In most cases, the phenotypic resistance profile was confirmed by detection of relevant genes mostly occurring in strains isolated from livestock animals and humans. Extended-spectrum ß-lactamase-producing strains were detected in one mink and three martens. The strains were carriers of blaTEM-1, blaTEM-135, and blaCTX-M-15 genes. Our research confirmed a high carrier rate of MDR E. coli, even more than one MDR strain in a single individual; therefore, wider monitoring in this group of animals should be considered.

Dermatophytosis with concurrent Trichophyton verrucosum and T. benhamiae in calves after long-term transport.

BACKGROUND: Dermatophytosis is a common problem in cattle. The aetiological factors associated with this disease are filamentous fungi with the ability to digest and grow on keratinized substrates. In cattle, and less frequently in other domestic animals and people, the dermatophyte Trichophyton verrucosum is most commonly isolated from skin lesions. The dermatophyte Trichophyton benhamiae is an important zoonotic pathogen, and the main sources of transmission are guinea pigs and other small rodents. OBJECTIVES: In this report, we show multispecies infection in calves (Bos taurus) after long-term transport and vaccination against trichophytosis. ANIMALS: Sixty animals were imported of which 32 were observed to be affected with superficial infection nine to 12 days after vaccination for dermatophytosis. METHODS AND MATERIALS: Diagnosis was made correlating the clinical signs with a micro- and macroscopic examination of cultured fungi. Molecular differentiation was used to confirm the species affiliation. RESULTS: Eight of the calves were infected with T. verrucosum alone, and 24 calves with both T. verrucosum and T. benhamiae. We suggest that the cause of this large outbreak was immunosuppression of the animals resulting from the stress of transport and administration of vaccine. CONCLUSION: Both T. verrucosum and T. benhamiae can be seen concurrently in cattle.

Intrinsic resistance to terbinafine among human and animal isolates of Trichophyton mentagrophytes related to amino acid substitution in the squalene epoxidase.

BACKGROUND: Dermatomycoses are the most common fungal infections in the world affecting a significant part of the human and animal population. The majority of zoophilic infections in humans are caused by Trichophyton mentagrophytes. Currently, the first-line drug for both oral and topical therapy is terbinafine. However, an increasing number of cases that are difficult to be cured with this drug have been noted in Europe and Asia. Resistance to terbinafine and other allylamines is very rare and usually correlated with point mutations in the squalene epoxidase gene resulting in single amino acid substitutions in the enzyme, which is crucial in the ergosterol synthesis pathway. PURPOSE: Here, we report terbinafine-resistant T. mentagrophytes isolates among which one was an etiological factor of tinea capitis in a man and three were obtained from asymptomatic foxes in Poland. METHODS: We used the CLSI protocol to determine antifungal susceptibility profiles of naftifine, amphotericin B, griseofulvin, ketoconazole, miconazole, itraconazole, voriconazole, and ciclopirox. Moreover, the squalene epoxidase gene of the terbinafine-resistant strains was sequenced and analysed. RESULTS: In the genomes of all four resistant strains exhibiting elevated MICs to terbinafine (16 to 32 µg/ml), single-point mutations leading to Leu393Phe substitution in the squalene epoxidase enzyme were revealed. Among the other tested substances, a MIC50 value of 1 µg/ml was shown only for griseofulvin. CONCLUSION: Finally, our study revealed that the terbinafine resistance phenomenon might not be acquired by exposure to the drug but can be intrinsic. This is evidenced by the description of the terbinafine-resistant strains isolated from the asymptomatic animals.

Population differentiation, antifungal susceptibility, and host range of Trichophyton mentagrophytes isolates causing recalcitrant infections in humans and animals.

The major problems in determining the causative factors of the high prevalence of dermatophytoses include the lack of a well-standardized antifungal susceptibility testing method, the low consistency of in vitro and clinical minimal inhibitory concentration values, the high genomic diversity of the population, and the unclear mechanism of pathogenicity. These factors are of particular importance when the disease is recalcitrant and relapses. Herein, we identified and characterized Trichophyton mentagrophytes isolates obtained from therapy-resistant cases in humans and animals. We used genomic diversity analysis of 17 human and 27 animal clinical isolates with the MP-PCR technique, determined their phenotypic enzymatic activity and host range, and performed antifungal susceptibility testing to currently available antifungal drugs from various chemical groups. Genomic diversity values of 35.3% and 33.3% were obtained for clinical isolates from humans and animals, respectively, yet without any relationship to the host species or antifungal drug to which resistance in therapy was revealed. The highest activity of keratinase enzymes was recorded for fox, guinea pig, and human hairs. These hosts can be considered as the main species in the host range of these isolates. A phenyl morpholine derivative, i.e. amorolfine, exhibited superior activity against strains obtained from both humans and animals with the lowest MIC50. Interestingly, high compliance of terbinafine in vitro resistance with clinical problems in the treatment with this substance was shown as well. The high resistance of dermatophytes to drugs is the main cause of the recalcitrance of the infection, whereas the other features of the fungus are less important.

Identification of emerging trends in the prevalence of dermatophytoses in alpacas (Vicugna pacos) farmed in Poland.

The increasing number of dermatophytoses among animals observed recently in developed countries may be connected with relocation of many exotic species outside their natural living environment. Moreover, an impact on this situation may also be exerted by relapses related to limited compliance with antifungal treatment regimes. Many exotic animals, including camelids, imported to European countries are connected with cases of tuberculosis or zoophilic dermatophytoses in humans. In the present study, we identified and comprehensively analysed dermatophyte infections in alpacas from breeding farms in Poland. As part of this study, we determined the prevalence of dermatophyte infections in alpacas. The conventional and molecular mycological diagnostic procedures applied led to unambiguous identification of the aetiological factors of symptomatic dermatophytoses and asymptomatic animals, that is Trichophyton benhamiae and T. verrucosum. Furthermore, the susceptibility tests allowed choosing the best therapeutic option and revealed superior activity of allylamine drugs against all strains. Finally, in the case of strains isolated from symptomatic dermatophytoses, our study revealed a significantly higher virulence level expressed by high activity of chosen enzymes, especially related to keratinolytic and haemolytic activity. In conclusion, this report indicates that farmed alpacas can be a reservoir or vector for contagious zoophilic dermatophyte infection. For this reason, imported animals should be subjected to meticulous monitoring to detect not only symptomatic infections but also asymptomatic animals.

A significant number of multi-drug resistant Enterococcus faecalis in wildlife animals; long-term consequences and new or known reservoirs of resistance?

As the last link in the food chain in a complex ecosystem covering at least three different environmental spheres, species of wildlife carnivorous mammals constitute a group accumulating potential pathogens and factors resulting from human activity, including the emergence of drug resistance. Therefore, the aim of this study was to evaluate the level and range of resistance in commensal E. faecalis isolated from wildlife carnivorous mammals and genetic relationships in terms of the source of these strains as well as resistance and virulence genes. Differentiation between strains was performed based on ADSRRS-fingerprinting method. The results showed that almost half of the tested animals (48%) were carriers of at least one multidrug resistant E. faecalis strain. Moreover, 44% of MDR-positive animals showed two or three strains differing in both the genotype and the resistance phenotype. A significant percentage of strains were resistant to high-level aminoglycosides (from 20% to even 57.5%). The resistance and virulence gene profiles showed a rich panel of genes closely related to isolates from nosocomial infection and from livestock animals. The presence of the same genotypes in different hosts reflects not only a possible transfer of genes between E. faecalis strains but also exchange of strains between animals. The obtained results reflect a very high level of contamination of animals that are not subjected to targeted antibiotic therapy, which may suggest the degree of pollution of the environment. Wildlife animals and their environment can be a link closing the circulation cycle of genes and even epidemiologically important strains; therefore, there is a high risk that this pool will never run out and will be maintained at a high level.

Comparison of in vitro activities of 11 antifungal agents against Trichophyton verrucosum isolates associated with a variety hosts and geographical origin.

The high prevalence of dermatophytosis in animals is usually associated with extra expenditure on prevention, diagnosis and long-term treatment. Humans are usually infected from animals, also from asymptomatic carriers, through direct contact or indirectly via fungus-bearing hair, scales and fomites. Despite the medical importance of Trichophyton verrucosum infections, there are limited in vitro data on the fungal susceptibility to antifungal drugs, including new-generation triazoles, imidazoles and allyloamines. The aim of the current study was to evaluate comprehensively the in vitro activity of 11 antifungal drugs against a large collection of T. verrucosum isolates obtained in Poland, Latvia, Lithuania and Slovakia from humans and animals using a microdilution assay. In vitro susceptibility testing of 11 antifungal drugs was performed according to the Clinical and Laboratory Standards Institute (CLSI) document M38. The MICs of clotrimazole, ciclopirox, enilconazole, miconazole, naftifine and terbinafine against all T. verrucosum isolates were below 1 µg/mL, whereas those of fluconazole, griseofulvin, itraconazole, ketoconazole and voriconazole were above 1 µg/mL. Ciclopirox was demonstrated to have superior activity against all strains in comparison with the other drugs, whereas fluconazole exerted the weakest in vitro effect and exhibited the highest MIC values. Our study has shown that drugs of different chemical origin have satisfactory antifungal activity and can be promising candidates for the treatment of T. verrucosum dermatophytosis. Moreover, no significant disparity in drug sensitivity between isolates obtained from different hosts and geographical regions have been demonstrated.

Bats as a reservoir of resistant Escherichia coli: A methodical view. Can we fully estimate the scale of resistance in the reservoirs of free-living animals?

Bats are a poorly understood reservoir of pathogenic and multi-drug resistant microorganisms; therefore, the aim of the study was to analyze the presence of drug resistance among E. coli isolated from the species of bats occurring naturally in Poland. The strategy of isolation and identification of resistant strains from pooled and single-animal samples was based on selective media with cefotaxime, chloramphenicol, kanamycin and tetracycline, the use of the ADSRRS-fingerprinting method for genomic differentiation of isolates, and the classical methods of evaluation of phenotypic and genotypic resistance. Of the 78 isolated isolates confirmed as E. coli, there were 38 genetically distinct strains resistant at least to one antimicrobial. 71% of these strains met the multi-drug resistance criterion. Moreover, two different multidrug resistant strains were isolated from three single samples. The highest resistance was observed in the case of ampicillin (66%), kanamycin (84%), sulfamethoxazole/trimetoprim (61%/55% respectively), and streptomycin (50%), which in most cases was confirmed by the presence of an adequate gene. Two isolates from single hosts produced extended-spectrum beta-lactamases (blaCTX-M-3, blaCTX-M-15, blaTEM-1). With the exception of tetracycline resistance, which was dominant among isolates from single animals, no significant differences in the resistance of the strains from both groups of samples were observed. Bats should not be neglected as another environmental reservoir and as an unpredictable source of potential pathogenic and multidrug resistant bacteria and should be extensively studied to predict the direction of the development and range of spreading resistance.

In search of the source of dermatophytosis: Epidemiological analysis of Trichophyton verrucosum infection in llamas and the breeder (case report).

During the last few years, the number of cases of Trichophyton verrucosum isolation from humans suffering from mycoses has been constantly increasing, which is correlated with the presence of an increasing number of outdoor breeding farms. Farmers and their families as well as veterinarians and technicians involved in handling the animals are at a higher risk of infection. One of the most important aims of mycological diagnostics is epidemiological analysis. Typically, the history of the disease is not sufficient to indicate reliably and eliminate the outbreak of infection. PCR fingerprinting methods are a useful tool in this type of analysis, which is presented in this study. The main aim is to present diagnostic and epidemiological analyses of dermatophyte isolates from llamas and their breeder. In two llamas, round alopecia sites or ca. 2-cm excoriations covered with thickened scaling epidermis were noticed at the border of the head and neck with a distinct tendency towards hair loss. Tinea unguium was noticed in a nail of the breeder's right hand. Direct analysis of the material from the clinical lesions revealed the presence of arthrospores. The macro- and micromorphology of the isolates were homogeneous and characteristic for T. verrucosum. The identification analysis based on the ITS sequences confirmed the previous morphological diagnostic examination. The MP-PCR and MSP-PCR analysis indicated high invariability of the genomes of the strains isolated from the human and animals. The epidemiological research has indicated an identical source of dermatophyte infection in the breeder and the lamas. To sum up, the number of pets and farm animals is increasing and dermatologists should always be informed about possible dermatophyte transmission sources. The possibility of transmission of zoophilic dermatophytes from humans to animals is a suggestion for further analysis; therefore, this type of transmission should be considered in dermatological studies.