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1.
J Infect Chemother ; 29(7): 713-717, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37001753

RESUMO

Candida auris is a health hazard because of its antifungal resistance and the potential to cause healthcare-associated outbreaks. To our knowledge, no previous cases of candidemia caused by C. auris have been reported in Japan. Herein, we report the first known case of clade I C. auris candidemia in a Japanese man with coronavirus disease 2019 (COVID-19) infection who was medically evacuated from the Philippines. A 71-year-old Japanese man traveled to Cebu Island in the Philippines 5 months before admission to our hospital. He contracted severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the Philippines and was admitted to the intensive care unit (ICU) in a local hospital. During his medical evacuation, we implemented precautions given his history of COVID-19 and pneumonia caused by multi-drug-resistant Acinetobacter baumannii complex. His blood culture revealed that C. auris infection was treated with antifungal agents but he did not survive. No evidence of nosocomial transmission was found among other patients in the ICU. This case study determines that accurate detection of C. auris, appropriate antifungal agent selection, precautions, and patient isolation are crucial to prevent nosocomial outbreaks, especially in patients with a history of multidrug-resistant organism (MDRO) colonization or international hospitalization. Medical professionals should recognize the risk of MDROs in international medical evacuation settings, considering the recent resumption of cross-border travel after the COVID-19 pandemic.


Assuntos
COVID-19 , Candidemia , Infecção Hospitalar , Masculino , Humanos , Idoso , Candidemia/microbiologia , Candida auris , Candida , COVID-19/epidemiologia , Pandemias , Japão , SARS-CoV-2 , Testes de Sensibilidade Microbiana , Filipinas , Antifúngicos/farmacologia , Antifúngicos/uso terapêutico , Infecção Hospitalar/microbiologia
2.
Antimicrob Agents Chemother ; 66(6): e0005922, 2022 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-35546111

RESUMO

Trichophyton indotineae causes dermatophytosis that is resistant to terbinafine and azole compounds. The aim of this study was to determine the mechanisms of resistance to itraconazole (ITC) and voriconazole (VRC) in strains of T. indotineae. Two azole-sensitive strains (ITC MIC < 0.125 µg/mL; VRC MIC < 0.06 µg/mL) and four azole-resistant strains (ITC MIC ≥ 0.5 µg/mL; VRC MIC ≥ 0.5 µg/mL) were used for the investigation. The expression of MDR genes encoding multidrug transporters of the ABC family for which orthologs have been identified in Trichophyton rubrum and those of CYP51A and CYP51B encoding the targets of azole antifungal compounds were compared between susceptible and resistant strains. TinMDR3 and TinCYP51B were overexpressed in T. indotineae resistant strains. Only small differences in susceptibility were observed between TinMDR3 disruptants and parental strains overexpressing TinMDR3. Whole-genome sequencing of resistant strains revealed the creation of a variable number of TinCYP51B tandem repeats at the specific position of their genomes in three resistant strains. Downregulation of TinCYP51B by RNA interference (RNAi) restored the susceptibility of azole-resistant strains. In contrast, overexpression of TinCYP51B cDNA conferred resistance to a susceptible strain of T. indotineae. In conclusion, the reduced sensitivity of T. indotineae strains to azoles is mainly due to the overexpression of TinCYP51B resulting from additional copies of this gene.


Assuntos
Azóis , Esterol 14-Desmetilase/genética , Trichophyton , Antifúngicos/farmacologia , Azóis/farmacologia , Farmacorresistência Fúngica/genética , Amplificação de Genes , Itraconazol/farmacologia , Testes de Sensibilidade Microbiana , Trichophyton/genética , Voriconazol
3.
Microbiol Immunol ; 66(6): 292-298, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35229341

RESUMO

Skin colonization by the emerging pathogen Candida auris is common in outbreaks within medical settings. Culture-based screening of patients is an effective management strategy to control the pathogen, and the newly developed CHROMagar™ Candida Plus medium is claimed to enable the presumptive identification of C. auris. Here, we evaluated the use of this medium with 63 C. auris strains comprising its four well-established clades, as well as genetically related comparators, including species from the Metschnikowia clade. The colors and halos of both confluent growth and discrete colonies of all the tested strains were compared. It was found that on CHROMagar™ Candida Plus, C. auris formed characteristic white colonies with blue-green halos that were more evident after 72 hr of incubation at 35°C than after 48 hr. However, distinguishing between closely related species such as Candida haemulonii, Candida pseudohaemulonii, and Candida duobushaemulonii required the consideration of parameters other than color, including colony size and growth ability at 35°C. In conclusion, the novel chromogenic medium CHROMagar™ Candida Plus constitutes an easy screening tool for C. auris.


Assuntos
Candida , Candidíase , Ágar , Antifúngicos , Candida auris , Candidíase/diagnóstico , Humanos
4.
Mycopathologia ; 187(2-3): 225-233, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35347533

RESUMO

INTRODUCTION: Otomycosis is a superficial infection of the external ear caused by fungal pathogens. The genera Aspergillus and Candida are considered the main fungal causative agents, with the predominance of Aspergillus section Nigri. The present study aimed to evaluate the clinical symptoms of patients with otomycosis and predisposing factors and to identify fungal etiological agents using molecular approaches. We also present an overview of published papers on tympanic membrane perforation (TMP) secondary to otomycosis. MATERIALS AND METHODS: An otorhinolaryngologist collected specimens from external ear canals of patients with suspected otomycosis based on the patient's history and clinical examinations. The specimens were collected using sterile swabs. Fungal isolates were confirmed in clinical specimens by direct microscopy and culture methods. Fungal isolates were identified based on molecular approaches. RESULTS: In total, specimens from 211 patients with suspected otomycosis were examined. The presence of fungi was confirmed in about 51% of patients based on fungal elements in direct microscopy and culture-positive fungi. Aspergillus tubingensis was the most commonly isolated species (52.77%), followed by Aspergillus niger (25.92%). Otomycosis due to infection with Candida species was observed in 16% of cases. Of note, in 36.11% of cases, otomycosis was associated with TMP. CONCLUSION: A mycological examination is indispensable for a correct diagnosis in patients with otitis extern. TMP should be considered in patients with otomycosis, as it appears to be relatively common in this population.


Assuntos
Otomicose , Perfuração da Membrana Timpânica , Antifúngicos/uso terapêutico , Candida , Hospitais , Humanos , Irã (Geográfico)/epidemiologia , Otomicose/epidemiologia , Otomicose/microbiologia , Prevalência , Perfuração da Membrana Timpânica/tratamento farmacológico , Perfuração da Membrana Timpânica/epidemiologia
5.
Microbiol Immunol ; 65(11): 463-471, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34251696

RESUMO

The International Space Station (ISS) is a closed facility that orbits the earth carrying not only its crew but also microorganisms. We have participated in microbiota analysis projects for the Japanese Experiment Module KIBO (ISS; operations nomenclature: Microbe-I, II, III, and IV) and were in charge of fungal screening. The interior of KIBO was sampled using swabs and microbe detection sheets (MDSs) for fungal detection. The dominant genera obtained by culture were Aspergillus and Penicillium. DNA analyses of the fungal biota using a clone library showed that KIBO was dominated by Malassezia, a fungal inhabitant of human skin. Three fungal species, Aspergillus sydowii, Penicillium palitans, and Rhodotorula mucilaginosa, which grew under microgravity in KIBO were observed under a field emission-scanning electron microscope on the ground. No novel phenotypic characteristics were noted. The results of antifungal susceptibility testing of all isolates did not differ significantly from previous reports of corresponding fungi. In Microbe-I (August 2009), MDSs were culture negative, while in the next stages the CFU of MDSs were 10 for Microbe-II (February 2011), 24 for Microbe-III (October 2012), and 151 for Microbe-IV (February 2015). These results indicated that fungi inside KIBO are increasing and expanding over time, and therefore continuous surveillance is crucial.


Assuntos
Fungos , Astronave , Aspergillus , Fungos/genética , Humanos , Japão , Penicillium , Rhodotorula
6.
Mycopathologia ; 186(6): 871-876, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34410567

RESUMO

The prevalence of fungal otitis externa, or otomycosis, has been increasing in recent decades. Fungi may act as primary pathogens in this condition, or they may occur as secondary infections after prolonged ototopical treatment with antibiotics, which alters the flora of the external auditory canal (EAC) and enables overgrowth of its fungal inhabitants. We report here a case of otomycosis by Candida parapsilosis, Malassezia obtusa, and Malassezia furfur as a secondary infection following prolonged otic ofloxacin treatment. To the best of our knowledge, although isolation of C. parapsilosis and M. furfur from the EAC is not uncommon, the recovery of M. obtusa has not yet been reported. We also conducted a literature review of the searchable data on PubMed concerning the isolation of Malassezia species from the human EAC.


Assuntos
Malassezia , Otite Externa , Otomicose , Fungos , Humanos , Ofloxacino , Otite Externa/diagnóstico , Otite Externa/tratamento farmacológico
7.
Mycopathologia ; 186(2): 189-198, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33475901

RESUMO

BACKGROUND: Candida auris is an emerging pathogen associated with outbreaks in clinical settings. Isolates of the pathogen have been geographically clustered into four clades with high intra-clade clonality. Pathogenicity varies among the clades, highlighting the importance of understanding these differences. OBJECTIVES: To examine the physiological and biochemical properties of each clade of C. auris to improve our understanding of the fungus. METHODS: Optimal growth temperatures of four strains from three clades, East Asia, South Asia and South Africa, were explored. Moreover, assimilation and antifungal susceptibility properties of 22 C. auris strains from the three clades were studied. RESULTS: The optimal growth temperatures of all strains were 35-37 °C. Assimilation testing demonstrated that the commercial API ID 32 C system can be used to reliably identify C. auris based on the biochemical properties of the yeast. Notably, C. auris can be uniquely differentiated from commonly clinical fungi by its ability to assimilate raffinose and inability to utilize D-xylose, suggesting a useful simple screening tool. The antifungal susceptibility results revealed that all strains are resistant against fluconazole (minimal inhibitory concentration (MIC) 4 to > 64 µg/mL) and miconazole (MIC 8 to > 16 µg/mL), with strains from the Japanese lineage showing relatively lower MIC values (1-4 µg/mL). Conversely, itraconazole, voriconazole, amphotericin B, micafungin and caspofungin were active against most of the tested strains. On the clade level, East Asian strains generally showed lower MICs against azoles comparing to the other clades, while they displayed MICs against flucytosine higher than those of strains from South Africa and South Asia clades. CONCLUSION: Our data suggest a simple identification approach of C. auris based on its physiological and biochemical properties and highlight aspects of C. auris population from various clades.


Assuntos
Antifúngicos , Candida , Anfotericina B , Antifúngicos/uso terapêutico , Ásia , Fluconazol , Testes de Sensibilidade Microbiana
8.
Mycoses ; 63(7): 694-703, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32277529

RESUMO

BACKGROUND: Dermatophytes are a group of keratinophilic fungi of medical importance. Despite a relatively long history of molecular taxonomic studies, there is still a need for information on genetic polymorphism in wider variety of genomic loci. OBJECTIVES: Our goal was to study partial DNA topoisomerase 2 gene (TOP2) polymorphism in dermatophytes. METHODS: We performed DNA sequencing of TOP2 in 26 dermatophyte species along with ribosomal internal transcribed spacer (ITS) sequencing. RESULTS: The number of polymorphic sites in TOP2 data set was similar to that one in ITS data set. Nannizzia species formed paraphyletic group in TOP2 tree. Trichophyton simii was paraphyletic in concatenated TOP2-ITS tree, one of its two clades contained solely Iranian isolates. CONCLUSIONS: Our results revealed several unresolved problems in the taxonomy of dermatophytes, including probable polyphyly of the genus Nannizzia and the species T simii.


Assuntos
Arthrodermataceae/enzimologia , Arthrodermataceae/genética , DNA Topoisomerases Tipo II/genética , DNA Fúngico/genética , Polimorfismo Genético , Arthrodermataceae/classificação , DNA Espaçador Ribossômico/genética , Irã (Geográfico) , Filogenia , Análise de Sequência de DNA
9.
Mycopathologia ; 185(4): 691-698, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32710393

RESUMO

BACKGROUND: Candida albicans is the most frequent pathogenic fungus in oral cavities. It adheres to dental tissues as part of dental plaques and contributes to caries formation. OBJECTIVES: To evaluate the effect of silver diamine fluoride (SDF) on reducing C. albicans adhesion on dentine surfaces. METHODS: Flat dentine surfaces were prepared from bovine dental disks, and samples were divided into three groups. The first and second groups were pretreated for 3 min with 299 mM or 2.99 M SDF, respectively, and the third group (control) did not undergo any SDF pretreatment. All samples were washed, inoculated with C. albicans suspension onto their dentine surface, incubated at 30 °C for 6 h, and washed again to remove any nonadherent cells. The abundance of adherent cells was investigated using colorimetric and real-time polymerase chain reaction approaches. Subsequently, the morphological changes in C. albicans by pretreatment with SDF were observed under a scanning electron microscope (SEM). RESULTS: SDF inhibited candidal growth at concentrations as low as 2.99 µM. Dentine disks pretreated with 299 mM or 2.99 M SDF displayed significantly fewer adhered cells as compared with the control group. Upon pretreatment with SDF, SEM images showed severe morphological changes in the cellular walls, in a dose-dependent manner, suggesting a fungicidal effect of SDF against the yeast. CONCLUSION: SDF should be considered for clinical applications aimed at inhibiting dental plaque caused by C. albicans, particularly in children and elderly individuals.


Assuntos
Candida albicans , Dentina/microbiologia , Compostos de Amônio Quaternário/farmacologia , Compostos de Prata/farmacologia , Animais , Candida albicans/efeitos dos fármacos , Bovinos , Fluoretos Tópicos/farmacologia
10.
Artigo em Inglês | MEDLINE | ID: mdl-31501141

RESUMO

The mechanisms of terbinafine resistance in a set of clinical isolates of Trichophyton rubrum have been studied recently. Of these isolates, TIMM20092 also showed reduced sensitivity to azoles. The azole resistance of TIMM20092 could be inhibited by milbemycin oxime, prompting us to examine the potential of T. rubrum to develop resistance through multidrug efflux transporters. The introduction of a T. rubrum cDNA library into Saccharomyces cerevisiae allowed the isolation of one transporter of the major facilitator superfamily (MFS) conferring resistance to azoles (TruMFS1). To identify more azole efflux pumps among 39 ABC and 170 MFS transporters present within the T. rubrum genome, we performed a BLASTp analysis of Aspergillus fumigatus, Candida albicans, and Candida glabrata on transporters that were previously shown to confer azole resistance. The identified candidates were further tested by heterologous gene expression in S. cerevisiae Four ABC transporters (TruMDR1, TruMDR2, TruMDR3, and TruMDR5) and a second MFS transporter (TruMFS2) proved to be able to operate as azole efflux pumps. Milbemycin oxime inhibited only TruMDR3. Expression analysis showed that both TruMDR3 and TruMDR2 were significantly upregulated in TIMM20092. TruMDR3 transports voriconazole (VRC) and itraconazole (ITC), while TruMDR2 transports only ITC. Disruption of TruMDR3 in TIMM20092 abolished its resistance to VRC and reduced its resistance to ITC. Our study highlights TruMDR3, a newly identified transporter of the ABC family in T. rubrum, which can confer azole resistance if overexpressed. Finally, inhibition of TruMDR3 by milbemycin suggests that milbemycin analogs could be interesting compounds to treat dermatophyte infections in cases of azole resistance.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Antifúngicos/farmacologia , Azóis/farmacologia , Trichophyton/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Antifúngicos/metabolismo , Azóis/metabolismo , Farmacorresistência Fúngica , Humanos , Macrolídeos/metabolismo , Macrolídeos/farmacologia , Testes de Sensibilidade Microbiana , Terbinafina/metabolismo , Terbinafina/farmacologia , Tinha/tratamento farmacológico , Tinha/microbiologia , Trichophyton/metabolismo
11.
Med Mycol ; 57(6): 703-709, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-30649423

RESUMO

Chronic pulmonary aspergillosis (CPA) is a common subtype of pulmonary aspergillosis and a life-threatening disease. However, its diagnosis remains difficult due to the lack of specific clinical features and radiologic findings, as well as the difficulty of isolating Aspergillus spp. We developed a novel species-specific detection method of medically important aspergilli using a loop-mediated isothermal amplification (LAMP) for CPA. Specific LAMP primer sets for Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, Aspergillus terreus, and Aspergillus nidulans were designed. The use of the LAMP assay was validated using respiratory specimens (CPA cases, n = 21; nonaspergillosis cases, n = 23). A total of 15 cases were positive in the CPA group (A. fumigatus, n = 5; A. flavus, n = 1; A. niger, n = 1; A. terreus, n = 7; A. nidulans, n = 1), but only three in the non-CPA group (A. niger, n = 2; A. terreus n = 1). The sensitivity and specificity of the diagnosis of CPA by the LAMP system were 71.4% and 87.0%, respectively. In conclusion, we developed a species-specific detection approach for five medically important aspergilli using the LAMP method. The system showed high sensitivity and specificity for diagnosis of CPA.


Assuntos
Aspergillus/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico , Aspergilose Pulmonar/diagnóstico , Aspergilose Pulmonar/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Aspergillus/classificação , Doença Crônica , Primers do DNA/genética , Feminino , Humanos , Pulmão/microbiologia , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Especificidade da Espécie
12.
Respiration ; 97(1): 84-91, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30343287

RESUMO

BACKGROUND: Filamentous basidiomycetes (f-BMs) are involved in some unexplained chronic cough (UCC) cases that can be improved by the administration of antifungal agents. The disease concept was termed fungus-associated chronic cough (FACC). The current diagnostic criteria warrant environmental fungi isolation from respiratory specimens, which is hardly conceivable for such fungi. OBJECTIVES: This study aimed to detect the f-BMs Bjerkandera adusta, the most common pathogen in FACC, from respiratory specimens of patients with UCC using real-time polymerase chain reaction (PCR). It also evaluated the applicability of the PCR system to detect antifungal agent responders among patients with unexplained cough. METHODS: The PCR system specific to B. adusta was developed and its utility was evaluated using sputum samples from 23 patients with chronic cough. RESULTS: B. adusta was detected in 10 out of 14 patients with UCC (71.4%), in contrast to only 2 out of 9 patients with non-UCC (22.2%; p < 0.05 with the Fisher's exact test). The copy number of the samples correlated with the therapeutic impact score for cough symptoms following the oral administration of itraconazole. CONCLUSION: Development of the real-time PCR system enabled us to demonstrate that many patients with UCC might be influenced by B. adusta, a fact evidenced by the improvement of symptoms with itraconazole administration in most PCR-positive patients. This method would help in detecting itraconazole responders among patients with UCC when the isolation of f-BMs is not achievable.


Assuntos
Coriolaceae/genética , Tosse/diagnóstico , DNA Fúngico/análise , Itraconazol/uso terapêutico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Escarro/microbiologia , Idoso , Idoso de 80 Anos ou mais , Antifúngicos/uso terapêutico , Doença Crônica , Coriolaceae/isolamento & purificação , Tosse/tratamento farmacológico , Tosse/microbiologia , Estudos de Viabilidade , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos Testes , Estudos Retrospectivos
13.
Mycoses ; 61(3): 213-217, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29125646

RESUMO

Central nervous system (CNS) infections due to filamentous basidiomycetes are extremely rare. We encountered a case of epidural abscess due to Schizophyllum commune that extended from sinusitis. A 53-year-old Japanese man presented at our hospital with a headache. Computed tomography (CT) of the cranium and sinuses showed ethmoid and sphenoid sinusitis with no intracranial abnormalities. The patient was diagnosed with acute sinusitis and underwent antibiotic treatment. However, the symptoms deteriorated, and the patient came to our hospital again with consciousness disturbance. CT scan of the cranium and sinuses showed no improvement of sinusitis after antibiotic therapy and an epidural abscess emerged in the middle cranial fossa. Therefore, emergency craniotomy and endoscopic sinus fenestration were performed. Filamentous fungal elements were observed in both rhinorrhoea and epidural abscess. The symptoms improved after the operation and administration of liposomal amphotericin B. The clinical isolate was identified as S. commune by a molecular-based method. To our knowledge, this is the first report of epidural abscess due to this fungus. Although rare, clinicians should be aware that S. commune could be a causative agent of CNS infections.


Assuntos
Abscesso Epidural/microbiologia , Micoses/microbiologia , Seios Paranasais/microbiologia , Schizophyllum/isolamento & purificação , Anfotericina B/administração & dosagem , Anfotericina B/uso terapêutico , Antibacterianos/uso terapêutico , Abscesso Epidural/complicações , Humanos , Masculino , Pessoa de Meia-Idade , Micoses/complicações , Seios Paranasais/diagnóstico por imagem , Schizophyllum/efeitos dos fármacos , Schizophyllum/genética , Sinusite/diagnóstico , Sinusite/tratamento farmacológico , Sinusite/microbiologia , Crânio/diagnóstico por imagem , Crânio/microbiologia , Tomógrafos Computadorizados , Resultado do Tratamento
14.
Artigo em Inglês | MEDLINE | ID: mdl-28416557

RESUMO

Terbinafine is one of the allylamine antifungal agents whose target is squalene epoxidase (SQLE). This agent has been extensively used in the therapy of dermatophyte infections. The incidence of patients with tinea pedis or unguium tolerant to terbinafine treatment prompted us to screen the terbinafine resistance of all Trichophyton clinical isolates from the laboratory of the Centre Hospitalier Universitaire Vaudois collected over a 3-year period and to identify their mechanism of resistance. Among 2,056 tested isolates, 17 (≈1%) showed reduced terbinafine susceptibility, and all of these were found to harbor SQLE gene alleles with different single point mutations, leading to single amino acid substitutions at one of four positions (Leu393, Phe397, Phe415, and His440) of the SQLE protein. Point mutations leading to the corresponding amino acid substitutions were introduced into the endogenous SQLE gene of a terbinafine-sensitive Arthroderma vanbreuseghemii (formerly Trichophyton mentagrophytes) strain. All of the generated A. vanbreuseghemii transformants expressing mutated SQLE proteins exhibited obvious terbinafine-resistant phenotypes compared to the phenotypes of the parent strain and of transformants expressing wild-type SQLE proteins. Nearly identical phenotypes were also observed in A. vanbreuseghemii transformants expressing mutant forms of Trichophyton rubrum SQLE proteins. Considering that the genome size of dermatophytes is about 22 Mb, the frequency of terbinafine-resistant clinical isolates was strikingly high. Increased exposure to antifungal drugs could favor the generation of resistant strains.


Assuntos
Antifúngicos/farmacologia , Naftalenos/farmacologia , Mutação Puntual/genética , Esqualeno Mono-Oxigenase/genética , Trichophyton/efeitos dos fármacos , Trichophyton/genética , Arthrodermataceae/efeitos dos fármacos , Arthrodermataceae/enzimologia , Arthrodermataceae/genética , Farmacorresistência Fúngica/genética , Testes de Sensibilidade Microbiana , Terbinafina , Trichophyton/enzimologia
15.
Artigo em Inglês | MEDLINE | ID: mdl-28848012

RESUMO

The in vitro activities of novel azoles compared to those of five antifungal drugs against clinical (n = 28) and environmental (n = 102) isolates of black mold and melanized yeast were determined. Luliconazole and lanoconazole had the lowest geometric mean MICs, followed by efinaconazole, against tested isolates compared to the other drugs. Therefore, it appears that these new imidazole and triazole drugs are promising candidates for the treatment of infections due to melanized fungi and their relatives.


Assuntos
Antifúngicos/farmacologia , Ascomicetos/efeitos dos fármacos , Imidazóis/farmacologia , Triazóis/farmacologia , Leveduras/efeitos dos fármacos , Adulto , Ascomicetos/isolamento & purificação , Pré-Escolar , Farmacorresistência Fúngica/genética , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Micoses/tratamento farmacológico , Adulto Jovem
16.
Med Mycol ; 55(5): 555-562, 2017 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-27811180

RESUMO

Molecular evolution has dominated taxonomic studies for decades, replacing traditional methods for identification and classification. However, there is a need for better markers to resolve the problems that have limited their usefulness. In this report, we introduce the protein tag (Ptag) sequence, a highly polymorphic amino acid sequence within the C-terminal region of the largest subunit of RNA polymerase I, as a new systematic tag sequence for delineating the evolutionary history of medically important fungi. As Ptag sequences are highly polymorphic between species and low within species, 42 fungal species representing the main taxonomic groups in the phyla Ascomycota and Basidiomycota were tested. The phylogenetic tree inferred from the Ptag sequences showed high consistency with the accepted classification of the Assembling the Fungal Tree of Life (AFTOL) project. Moreover, it could resolve the interspecies phylogenetic relationships of the tested taxa. In contrast, the phylogeny inferred from the nucleotide tag (Ntag) sequence, encoding the Ptag peptide, displayed lesser discriminatory power in resolving the phylogenetic relationships among distantly related taxa. In the case of closely related fungal species, the phylogenetic trees for Ptag and Ntag sequences were consistent with the tree for ITS1 sequences of 11 dermatophytic species. Taken together, the Ptag sequences should contribute to inferring phylogeny among species whereas the Ntag sequences should be useful to analyze variations among closely related species to resolve taxonomic issues in fungi.


Assuntos
Ascomicetos/classificação , Ascomicetos/genética , Basidiomycota/classificação , Basidiomycota/genética , Micoses/microbiologia , Filogenia , RNA Polimerase I/genética , Sequência de Aminoácidos , Sequência de Bases , DNA Fúngico/química , DNA Fúngico/genética , Marcadores Genéticos/genética , RNA Polimerase I/química , Análise de Sequência de DNA , Especificidade da Espécie
17.
Mycopathologia ; 182(1-2): 33-43, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27435975

RESUMO

Dermatophytes are a group of closely related fungi that nourish on keratinized materials for their survival. They infect stratum corneum, nails, and hair of human and animals, accounting the largest portion of fungi causing superficial mycoses. Huge populations are suffering from dermatophytoses, though the biology of these fungi is largely unknown yet. Reasons are partially attributed to the poor amenability of dermatophytes to genetic manipulation. However, advancements in this field over the last decade made it possible to conduct genetic studies to satisfying extents. These included genetic transformation methods, indispensable molecular tools, i.e., dominant selectable markers, inducible promoter, and marker recycling system, along with improving homologous recombination frequency and gene silencing. Furthermore, annotated genome sequences of several dermatophytic species have recently been available, ensuring an optimal recruitment of the molecular tools to expand our knowledge on these fungi. In conclusion, the establishment of basic molecular tools and the availability of genomic data will open a new era that might change our understanding on the biology and pathogenicity of this fungal group.


Assuntos
Arthrodermataceae/genética , Genética Microbiana/métodos , Biologia Molecular/métodos , Arthrodermataceae/patogenicidade , Arthrodermataceae/fisiologia , Humanos
18.
Antimicrob Agents Chemother ; 60(2): 873-80, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26596944

RESUMO

Despite the existing treatment options for onychomycosis, there remains a strong demand for potent topical medications. ME1111 is a novel antifungal agent that is active against dermatophytes, has an excellent ability to penetrate human nails, and is being developed as a topical agent for onychomycosis. In the present study, we investigated its mechanism of action. Trichophyton mentagrophytes mutants with reduced susceptibility to ME1111 were selected in our laboratory, and genome sequences were determined for 3 resistant mutants. The inhibitory effect on a candidate target was evaluated by a spectrophotometric enzyme assay using mitochondrial fractions. Point mutations were introduced into candidate genes by a reverse genetics approach. Whole-genome analysis of the 3 selected mutants revealed point mutations in the structural regions of genes encoding subunits of succinate dehydrogenase (complex II). All of the laboratory-generated resistant mutants tested harbored a mutation in one of the subunits of succinate dehydrogenase (SdhB, SdhC, or SdhD). Most of the mutants showed cross-resistance to carboxin and boscalid, which are succinate dehydrogenase inhibitors. ME1111 strongly inhibited the succinate-2,6-dichlorophenolindophenol reductase reaction in Trichophyton rubrum and T. mentagrophytes (50% inhibitory concentrations [IC50s] of 0.029 and 0.025 µg/ml, respectively) but demonstrated only moderate inhibition of the same reaction in human cell lines. Furthermore, the target protein of ME1111 was confirmed by the introduction of point mutations causing the amino acid substitutions in SdhB, SdhC, and SdhD found in the laboratory-generated resistant mutants, which resulted in reduced susceptibility to ME1111. Thus, ME1111 is a novel inhibitor of the succinate dehydrogenase of Trichophyton species, and its mechanism of action indicates its selective profile.


Assuntos
Antifúngicos/farmacologia , Farmacorresistência Fúngica/genética , Onicomicose/tratamento farmacológico , Fenóis/farmacologia , Pirazóis/farmacologia , Succinato Desidrogenase/genética , Trichophyton/efeitos dos fármacos , Trichophyton/genética , Administração Tópica , Substituição de Aminoácidos/genética , Arthrodermataceae/efeitos dos fármacos , Sequência de Bases , Linhagem Celular , DNA Fúngico/genética , Humanos , Dados de Sequência Molecular , Onicomicose/microbiologia , Análise de Sequência de DNA
19.
Med Mycol ; 54(2): 207-18, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26483437

RESUMO

Calcineurin is a serine/threonine protein phosphatase that consists of catalytic (calcineurin A) and regulatory (calcineurin B) subunits. The conserved protein plays important roles in various biological processes. Drug combination of fluconazole and the calcineurin inhibitor (FK506) showed synergistic effects against dermatophytes. In the current study, we identified the calcineurin A homologous gene (TmcanA) in the dermatophyte Arthroderma vanbreuseghemii (anamorph: Trichophyton mentagrophytes). Knockdown mutants were produced from A. vanbreuseghemii, resulting in a defection in growth properties in accordance with dose of the suppressing reagent. The TmcanA gene restored the ability of calcineurin A-deficient Cryptococcus neoformans strain to grow at elevated temperatures. Repression of TmcanA at 37°C resulted in severely stunted growth, suggesting that this protein plays a role in tolerance to elevated temperatures. In addition, TMCANA showed an interaction with high osmolarity glycerol (HOG) signalling pathway by governing the secretion of a secondary metabolite. Moreover, expression of the hydrophobin A gene (TmHF) decreased significantly under the TmcanA-repressive condition, suggesting that TMCANA is involved in its regulation. In conclusion, calcineurin A is a multifunctional gene that is involved in the regulation of several biological processes and therefore is worth being considered as a drug target for treatment of dermatophytoses.


Assuntos
Arthrodermataceae/enzimologia , Arthrodermataceae/genética , Calcineurina/genética , Calcineurina/metabolismo , Arthrodermataceae/crescimento & desenvolvimento , Técnicas de Silenciamento de Genes , Teste de Complementação Genética , Temperatura
20.
Microbiol Immunol ; 60(5): 295-302, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26969809

RESUMO

As a part of a series of studies regarding the microbial biota in manned space environments, fungi were isolated from six pieces of equipment recovered from the Japanese Experimental Module "KIBO" of the International Space Station and from a space shuttle. Thirty-seven strains of fungi were isolated, identified and investigated with regard to morphological phenotypes and antifungal susceptibilities. The variety of fungi isolated in this study was similar to that of several previous reports. The dominant species belonged to the genera Penicillium, Aspergillus and Cladosporium, which are potential causative agents of allergy and opportunistic infections. The morphological phenotypes and antifungal susceptibilities of the strains isolated from space environments were not significantly different from those of reference strains on Earth.


Assuntos
Equipamentos e Provisões/microbiologia , Fungos/classificação , Fungos/isolamento & purificação , Astronave , Antifúngicos/farmacologia , Fungos/citologia , Fungos/efeitos dos fármacos , Testes de Sensibilidade Microbiana
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