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1.
Int J Mol Sci ; 22(6)2021 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-33803717

RESUMO

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


Assuntos
Antifúngicos/farmacologia , Durapatita/química , Fluconazol/farmacologia , Galactanos/química , Nanopartículas/química , Cicatrização/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Doença Crônica , Fungos/efeitos dos fármacos , Hidrogéis/química , Cinética , Testes de Sensibilidade Microbiana , Muramidase/metabolismo , Nanopartículas/ultraestrutura , Plâncton/efeitos dos fármacos , Porosidade , Espectroscopia de Infravermelho com Transformada de Fourier , Fatores de Tempo , Difração de Raios X
2.
Int J Mol Sci ; 22(4)2021 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-33669913

RESUMO

Candida albicans is a pathogenic fungus that is increasingly developing multidrug resistance (MDR), including resistance to azole drugs such as fluconazole (FLC). This is partially a result of the increased synthesis of membrane efflux transporters Cdr1p, Cdr2p, and Mdr1p. Although all these proteins can export FLC, only Cdr1p is expressed constitutively. In this study, the effect of elevated fructose, as a carbon source, on the MDR was evaluated. It was shown that fructose, elevated in the serum of diabetics, promotes FLC resistance. Using C. albicans strains with green fluorescent protein (GFP) tagged MDR transporters, it was determined that the FLC-resistance phenotype occurs as a result of Mdr1p activation and via the increased induction of higher Cdr1p levels. It was observed that fructose-grown C. albicans cells displayed a high efflux activity of both transporters as opposed to glucose-grown cells, which synthesize Cdr1p but not Mdr1p. Additionally, it was concluded that elevated fructose serum levels induce the de novo production of Mdr1p after 60 min. In combination with glucose, however, fructose induces Mdr1p production as soon as after 30 min. It is proposed that fructose may be one of the biochemical factors responsible for Mdr1p production in C. albicans cells.


Assuntos
Candida albicans/efeitos dos fármacos , Farmacorresistência Fúngica/efeitos dos fármacos , Fluconazol/farmacologia , Frutose/farmacologia , Proteínas Fúngicas/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Candida albicans/citologia , Carbono/farmacologia , Proliferação de Células/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Frutose/sangue , Proteínas de Fluorescência Verde/metabolismo , Frações Subcelulares/metabolismo
3.
J Med Microbiol ; 70(3)2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33688802

RESUMO

Introduction. Trichosporon asahii has been recognized as an opportunistic agent having a limited sensitivity to antifungal treatment.Hypothesis/Gap Statement. Molecular mechanisms of azole resistance have been rarely reported for Trichosproron asahii. Similar to other fungi, we hypothesized that both ERG11 gene mutation and efflux pumps genes hyper-expression were implicated.Aim. The current work aimed to study the sensitivity of clinical T. asahii isolates to different antifungal agents and to explore their resistance mechanisms by molecular methods including real-time PCR and gene sequencing.Methods. The sensitivity of T. asahii isolates to fluconazole, amphotericin B and voriconazole was estimated by the Etest method. Real-time PCR was used to measure the relative expression of Pdr11, Mdr and ERG11 genes via the ACT1 housekeeping gene. Three pairs of primers were also chosen to sequence the ERG11 gene. This exploration was followed by statistical study including the receiver operating characteristic (ROC) curve analysis to identify a relationship between gene mean expression and the sensitivity of isolates.Results. In 31 clinical isolates, the resistance frequencies were 87, 16.1 and 3.2 %, respectively, for amphotericin B, fluconazole and voriconazole. Quantitative real-time PCR demonstrated that only Mdr over-expression was significantly associated with FCZ resistance confirmed by univariate statistical study and the ROC curve analysis (P <0.05). The ERG11 sequencing revealed two mutations H380G and S381A in TN325U11 (MIC FCZ=8 µg ml-1) and H437R in TN114U09 (MIC FCZ=256 µg ml-1) in highly conserved regions (close to the haem-binding domain) but their involvement in the resistance mechanism has not yet been assigned.Conclusion. T. asahii FCZ resistance mechanisms are proven to be much more complex and gene alteration sequence and/or expression can be involved. Only Mdr gene over-expression was significantly associated with FCZ resistance and no good correlation was observed between FCZ and VCZ MIC values and relative gene expression. ERG11 sequence alteration seems to play a major role in T. asahii FCZ resistance mechanism but their involvement needs further confirmation.


Assuntos
Antifúngicos/farmacologia , Basidiomycota , Sistema Enzimático do Citocromo P-450/genética , Farmacorresistência Fúngica/genética , Proteínas Fúngicas/genética , Tricosporonose/microbiologia , Anfotericina B/farmacologia , Basidiomycota/efeitos dos fármacos , Basidiomycota/genética , Fluconazol/farmacologia , Humanos , Voriconazol/farmacologia
4.
J Med Microbiol ; 70(3)2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33560202

RESUMO

The genus Candida spp. has been highlighted as one of the main etiological agents causing fungal infections, with Candida albicans being the most prominent, responsible for most cases of candidemia. Due to its capacity for invasion and tissue adhesion, it is associated with the formation of biofilms, mainly in the environment and hospital devices, decreasing the effectiveness of available treatments. The repositioning of drugs, which is characterized by the use of drugs already on the market for other purposes, together with molecular-docking methods can be used aiming at the faster development of new antifungals to combat micro-organisms. This study aimed to evaluate the antifungal effect of diazepam on mature C. albicans biofilms in vitro and its action on biofilm in formation, as well as its mechanism of action and interaction with structures related to the adhesion of C. albicans, ALS3 and SAP5. To determine the MIC, the broth microdilution test was used according to protocol M27-A3 (CLSI, 2008). In vitro biofilm formation tests were performed using 96-well plates, followed by molecular-docking protocols to analyse the binding agent interaction with ALS3 and SAP5 targets. The results indicate that diazepam has antimicrobial activity against planktonic cells of Candida spp. and C. albicans biofilms, interacting with important virulence factors related to biofilm formation (ALS3 and SAP5). In addition, treatment with diazepam triggered a series of events in C. albicans cells, such as loss of membrane integrity, mitochondrial depolarization and increased production of EROs, causing DNA damage and consequent cell apoptosis.


Assuntos
Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Candida/efeitos dos fármacos , Diazepam/farmacologia , Farmacorresistência Fúngica/efeitos dos fármacos , Ácido Aspártico Endopeptidases/metabolismo , Candida/patogenicidade , Fluconazol/farmacologia , Proteínas Fúngicas/metabolismo
5.
ABCS health sci ; 46: e021203, 09 fev. 2021. tab
Artigo em Inglês | LILACS | ID: biblio-1147180

RESUMO

INTRODUCTION: The resistance of fungal species to drugs usually used in clinics is of great interest in the medical field. OBJECTIVE: To evaluate susceptibility and in vitro response of species of Trichophyton spp. to antifungal drugs of interest in clinical medicine. METHODS: 12 samples of clinical isolates from humans were used, nine of T. mentagrophytes and three of T. tonsurans. Susceptibility tests were performed according to the agar diffusion (AD) and broth microdilution (BM) methods. RESULTS: In the AD method, the species T. tonsurans presented a percentage of sensitivity of 33% in relation to amphotericin B and 66% to itraconazole, with 100% resistance to ketoconazole and fluconazole. T. mentagrophytes also showed 100% resistance to ketoconazole in this technique, with 11% sensitivity to ketoconazole, 22% to itraconazole and 22% of samples classified as sensitive dose dependent. In the MC method, the species T. tonsurans presented a sensitivity percentage of 66%, 55% and 33% in relation to ketoconazole, fluconazole and itraconazole, respectively. The T. mentagrophytes species presented sensitivity percentages of 11%, 11%, 33% and 55% for amphotericin B, itraconazole, ketoconazole and fluconazole, respectively. CONCLUSION: There was resistance in vitro of the species of T. mentagrophytes and T. tonsurans against the antifungal fluconazole and relative resistance against ketoconazole in the AD method. In BM, however, important percentages of sensitivity were observed for the two species analyzed in relation to the antifungals fluconazole and ketoconazole when compared to itraconazole and amphotericin B.


INTRODUÇÃO: A resistência de espécies fúngicas às drogas usualmente empregadas no meio clínico é motivo de grande interesse na área médica. OBJETIVO: Avaliar susceptibilidade e resposta in vitro de espécies de Trichophyton spp. a drogas antifúngicas de interesse em clínica médica. MÉTODOS: Foram utilizadas 12 amostras de isolados clínicos de humanos, sendo nove de T. mentagrophytes e três de T. tonsurans. Foram realizados testes de susceptibilidade segundo os métodos de difusão em ágar (DA) e microdiluição em caldo (MC). RESULTADOS: No método de DA, a espécie T. tonsurans apresentou percentual de sensibilidade de 33% em relação à anfotericina B e de 66% ao itraconazol, com 100% de resistência frente ao cetoconazol e ao fluconazol. A espécie T. mentagrophytes também apresentou 100% de resistência frente ao cetoconazol nesta técnica, com 11% de sensibilidade ao cetoconazol, 22% ao itraconazol e 22% das amostras classificadas como sensível dose dependente. No método de MC, a espécie T. tonsurans apresentou percentual de sensibilidade de 66%, 55% e 33% em relação ao cetoconazol, fluconazol e itraconazol, respectivamente. A espécie T. mentagrophytes apresentou percentuais de sensibilidade de 11%, 11%, 33% e 55% para anfotericina B, itraconazol, cetoconazol e fluconazol, respectivamente. CONCLUSÃO: Houve resistência in vitro das espécies do T. mentagrophytes e T. tonsurans frente ao antifúngico fluconazol e resistência relativa frente ao cetoconazol no método de DA. Na MC, no entanto, foram observados importantes percentuais de sensibilidade das duas espécies analisadas frente aos antifúngicos fluconazol e cetoconazol quando comparadas ao itraconazol e à anfotericina B.


Assuntos
Trichophyton/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Farmacorresistência Fúngica , Suscetibilidade a Doenças/microbiologia , Antifúngicos/farmacologia , Tinha/microbiologia , Tinha/tratamento farmacológico , Contagem de Colônia Microbiana , Fluconazol/farmacologia , Anfotericina B/farmacologia , Itraconazol/farmacologia , Cetoconazol/farmacologia
6.
Drug Dev Ind Pharm ; 47(2): 246-258, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33416006

RESUMO

The aim of this work was to prepare and optimize mucoadhesive nanostructured lipid carrier (NLC) impregnated with fluconazole for better management of oral candidiasis. The NLCs were fabricated using an emulsification/sonication technique. The nanoparticles consisted of stearic acid, oleic acid, Pluronic F127, and lecithin. Box-Behnken design, artificial neural networking, and variable weight desirability were employed to optimize the joint effect of drug concentration in the drug/lipid mixture, solid lipid concentration in the solid/liquid lipid mixture, and surfactant concentration in the total mixture on size and entrapment. The optimized NLCs were coated with chitosan. The nanoparticles were characterized by surface charge, spectroscopic, thermal, morphological, mucoadhesion, release, histopathological, and antifungal properties. The nanoparticles are characterized by a particle size of 335 ± 13.5 nm, entrapment efficiency of 73.1 ± 4.9%, sustained release, minor histopathological effects on rabbit oral mucosa, and higher fungal inhibition efficiency for an extended period of time compared with fluconazole solution. Coating the nanoparticles with chitosan increased its adhesion to rabbit oral buccal mucosa and improved its anti-candidiasis activity. It is concluded that mucoadhesive lipid-based nanoparticles amplify the effect of fluconazole on Candida albicans in vitro. This finding warrants pre-clinical and clinical studies in oral candidiasis disease models to corroborate in vitro findings.


Assuntos
Candidíase Bucal , Fluconazol/farmacologia , Lipídeos/química , Nanopartículas , Nanoestruturas , Animais , Candidíase Bucal/tratamento farmacológico , Portadores de Fármacos , Fluconazol/administração & dosagem , Fluconazol/química , Aprendizado de Máquina , Tamanho da Partícula , Coelhos
7.
Artigo em Inglês | MEDLINE | ID: mdl-32942047

RESUMO

Independent studies from our group and others have provided evidence that sphingolipids (SLs) influence the antimycotic susceptibility of Candida species. We analyzed the molecular SL signatures of drug-resistant clinical isolates of Candida auris, which have emerged as a global threat over the last decade. This included Indian hospital isolates of C. auris, which were either resistant to fluconazole (FLCR) or amphotericin B (AmBR) or both drugs. Relative to Candida glabrata and Candida albicans strains, these C. auris isolates were susceptible to SL pathway inhibitors such as myriocin and aureobasidin A, suggesting that SL content may influence azole and AmB susceptibilities. Our analysis of SLs confirmed the presence of 140 SL species within nine major SL classes, namely the sphingoid bases, Cer, αOH-Cer, dhCer, PCer, αOH-PCer, αOH-GlcCer, GlcCer, and IPC. Other than for αOH-GlcCer, most of the SLs were found at higher concentrations in FLCR isolates as compared to the AmBR isolates. SLs were at intermediate levels in FLCR + AmBR isolates. The observed diversity of molecular species of SL classes based on fatty acyl composition was further reflected in their distinct specific imprint, suggesting their influence in drug resistance. Together, the presented data improves our understanding of the dynamics of SL structures, their synthesis, and link to the drug resistance in C. auris.


Assuntos
Anfotericina B/farmacologia , Antifúngicos/farmacologia , Candida/metabolismo , Farmacorresistência Fúngica Múltipla/fisiologia , Fluconazol/farmacologia , Glucosilceramidas/metabolismo , Candida/efeitos dos fármacos , Candida/isolamento & purificação , Candida albicans/efeitos dos fármacos , Candida albicans/isolamento & purificação , Candida albicans/metabolismo , Candida glabrata/efeitos dos fármacos , Candida glabrata/isolamento & purificação , Candida glabrata/metabolismo , Candidíase/microbiologia , Cromatografia Líquida , Depsipeptídeos/farmacologia , Farmacorresistência Fúngica Múltipla/efeitos dos fármacos , Ácidos Graxos Monoinsaturados/farmacologia , Glucosilceramidas/classificação , Glucosilceramidas/isolamento & purificação , Humanos , Lipidômica/métodos , Espectrometria de Massas em Tandem
8.
J Med Chem ; 64(2): 1116-1126, 2021 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-33356256

RESUMO

Due to the evolution and development of antifungal drug resistance, limited efficacy of existing drugs has led to high mortality in patients with serious fungal infections. To develop novel antifungal therapeutic strategies, herein a series of carboline fungal histone deacetylase (HDAC) inhibitors were designed and synthesized, which had potent synergistic effects with fluconazole against resistant Candida albicans infection. In particular, compound D12 showed excellent in vitro and in vivo synergistic antifungal efficacy with fluconazole to treat azole-resistant candidiasis. It cooperated with fluconazole in reducing the virulence of C. albicans by blocking morphological mutual transformation and inhibiting biofilm formation. Mechanism studies revealed that the reversion of drug resistance was due to downregulation of the expression of the azole target gene ERG11 and efflux gene CDR1. Taken together, fungal HDAC inhibitor D12 offered a promising lead compound for combinational treatment of azole-resistant candidiasis.


Assuntos
Azóis/uso terapêutico , Candida albicans/efeitos dos fármacos , Candidíase/tratamento farmacológico , Carbolinas/síntese química , Carbolinas/uso terapêutico , Farmacorresistência Fúngica/efeitos dos fármacos , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/uso terapêutico , Animais , Biofilmes/efeitos dos fármacos , Candida albicans/enzimologia , Candidíase/microbiologia , Carbolinas/toxicidade , Quimioterapia Combinada , Feminino , Fluconazol/farmacologia , Proteínas Fúngicas/efeitos dos fármacos , Fungos/efeitos dos fármacos , Fungos/enzimologia , Inibidores de Histona Desacetilases/toxicidade , Humanos , Fígado/patologia , Proteínas de Membrana Transportadoras/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos ICR , Testes de Sensibilidade Microbiana
9.
Nat Commun ; 11(1): 6429, 2020 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-33353950

RESUMO

Candida auris is an emerging fungal pathogen that exhibits resistance to multiple drugs, including the most commonly prescribed antifungal, fluconazole. Here, we use a combinatorial screening approach to identify a bis-benzodioxolylindolinone (azoffluxin) that synergizes with fluconazole against C. auris. Azoffluxin enhances fluconazole activity through the inhibition of efflux pump Cdr1, thus increasing intracellular fluconazole levels. This activity is conserved across most C. auris clades, with the exception of clade III. Azoffluxin also inhibits efflux in highly azole-resistant strains of Candida albicans, another human fungal pathogen, increasing their susceptibility to fluconazole. Furthermore, azoffluxin enhances fluconazole activity in mice infected with C. auris, reducing fungal burden. Our findings suggest that pharmacologically targeting Cdr1 in combination with azoles may be an effective strategy to control infection caused by azole-resistant isolates of C. auris.


Assuntos
Azóis/farmacologia , Candida/patogenicidade , Oxindois/farmacologia , Animais , Antifúngicos/análise , Antifúngicos/química , Antifúngicos/farmacologia , Azóis/análise , Azóis/química , Candida/efeitos dos fármacos , Candida/isolamento & purificação , Avaliação Pré-Clínica de Medicamentos , Sinergismo Farmacológico , Fluconazol/farmacologia , Proteínas Fúngicas/metabolismo , Deleção de Genes , Humanos , Camundongos , Oxindois/química , Virulência/efeitos dos fármacos
10.
J Med Microbiol ; 69(10): 1221-1227, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32894212

RESUMO

This study evaluated the effect of etomidate against biofilms of Candida spp. and analysed through molecular docking the interaction of this drug with ALS3, an important protein for fungal adhesion. Three fluconazole-resistant fungi were used: Candida albicans, Candida parapsilosis and Candida tropicalis. Growing biofilms were exposed to etomidate at 31.25-500 µg ml-1. Then, an ALS3 adhesive protein from C. albicans was analysed through a molecular mapping technique, composed of a sequence of algorithms to perform molecular mapping simulation based on classic force field theory. Etomidate showed antifungal activity against growing biofilms of resistant C. albicans, C. parapsilosis and C. tropicalis at all concentrations used in the study. The etomidate coupling analysis revealed three interactions with the residues of interest compared to hepta-threonine, which remained at the ALS3 site. In addition, etomidate decreased the expression of mannoproteins on the surface of C. albicans. These results revealed that etomidate inhibited the growth of biofilms.


Assuntos
Candida/efeitos dos fármacos , Farmacorresistência Fúngica/efeitos dos fármacos , Etomidato/farmacologia , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Etomidato/metabolismo , Fluconazol/farmacologia , Proteínas Fúngicas/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular/métodos
11.
PLoS One ; 15(9): e0238428, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32941438

RESUMO

OBJECTIVES: Recurrent vulvovaginal candidiasis (RVVC) causes significant morbidity. Candida albicans is the main pathogen associated with both sporadic and recurrent candidiasis. Due to unsatisfactory treatment effect, the impact of chlorhexidine digluconate and fluconazole alone or in combination on C. albicans and biofilm was investigated. METHODS: Vaginal C. albicans isolates from 18 patients with recurrent candidiasis and commensals from 19 asymptomatic women were isolated by culture. Crystal violet, XTT and colony forming unit assay were used to analyze the effect of chlorhexidine digluconate and fluconazole on growth of C. albicans, formation of new and already established, mature, biofilm. RESULTS: Fluconazole reduced the growth of planktonic C. albicans. However, in established biofilm, fluconazole had no effect on the candida cells and was not able to disperse and reduce the biofilm. By contrast, chlorhexidine digluconate had a direct killing effect on C. albicans grown both planktonically and in biofilm. Chlorhexidine digluconate also dispersed mature biofilm and inhibited formation of new biofilm. No major differences were observed between commensal isolates and candida causing recurrent vulvovaginitis with respect to biofilm or growth after chlorhexidine digluconate treatment. CONCLUSION: Biofilm is a problem in patients with recurrent vulvovaginal candidiasis reducing the effect of antifungal treatment. Development of new treatment strategies are urgently needed to decrease the recurrences. In already established biofilm, chlorhexidine digluconate dispersed the biofilm and was more effective in eradicating candida compared to fluconazole. Future treatment strategy may thus be a combination of chlorhexidine digluconate and fluconazole and prophylactic use of chlorhexidine digluconate to prevent biofilm formation and restrict infections.


Assuntos
Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Clorexidina/análogos & derivados , Adulto , Antifúngicos/farmacologia , Biofilmes/crescimento & desenvolvimento , Candida/efeitos dos fármacos , Candida albicans/isolamento & purificação , Candidíase/tratamento farmacológico , Candidíase Vulvovaginal/tratamento farmacológico , Candidíase Vulvovaginal/microbiologia , Clorexidina/metabolismo , Clorexidina/farmacologia , Feminino , Fluconazol/farmacologia , Humanos , Testes de Sensibilidade Microbiana/métodos , Vagina/microbiologia
12.
Int J Nanomedicine ; 15: 3681-3693, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32547022

RESUMO

Background: Fungal infections are becoming more prevalent and threatening because of the continuous emergence of azole-resistant fungal infections. The present study was aimed to assess the activity of free Methylglyoxal (MG) or MG-conjugated chitosan nanoparticles (MGCN) against fluconazole-resistant Candida albicans. Materials and Methods: A novel formulation of MGCN was prepared and characterized to determine their size, shape and polydispersity index. Moreover, the efficacy of fluconazole or MG or MGCN was determined against intracellular C. albicans in macrophages and the systematic candidiasis in a murine model. The safety of MG or MGCN was tested in mice by analyzing the levels of hepatic and renal toxicity parameters. Results: Candida albicans did not respond to fluconazole, even at the highest dose of 20 mg/kg, whereas MG and MGCN effectively eliminated C. albicans from the macrophages and infected mice. Mice in the group treated with MGCN at a dose of 10 mg/kg exhibited a 90% survival rate and showed the lowest fungal load in the kidney, whereas the mice treated with free MG at the same dose exhibited 50% survival rate. Moreover, the administration of MG or MGCN did not induce any liver and kidney toxicity in the treated mice. Conclusion: The findings of the present work suggest that MGCN may be proved a promising therapeutic formulation to treat azole-resistant C. albicans infections.


Assuntos
Candidíase/tratamento farmacológico , Quitosana/química , Farmacorresistência Fúngica , Fluconazol/uso terapêutico , Nanopartículas/química , Aldeído Pirúvico/uso terapêutico , Animais , Antifúngicos/farmacologia , Antifúngicos/uso terapêutico , Candida albicans/efeitos dos fármacos , Candida albicans/crescimento & desenvolvimento , Candidíase/microbiologia , Modelos Animais de Doenças , Farmacorresistência Fúngica/efeitos dos fármacos , Feminino , Fluconazol/farmacologia , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Nanopartículas/ultraestrutura , Tamanho da Partícula , Aldeído Pirúvico/farmacologia
13.
Sci Rep ; 10(1): 7769, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385378

RESUMO

Microorganisms employ quorum sensing (QS) mechanisms to communicate with each other within microbial ecosystems. Emerging evidence suggests that intraspecies and interspecies QS plays an important role in antimicrobial resistance in microbial communities. However, the relationship between interkingdom QS and antimicrobial resistance is largely unknown. Here, we demonstrate that interkingdom QS interactions between a bacterium, Pseudomonas aeruginosa and a yeast, Candida albicans, induce the resistance of the latter to a widely used antifungal fluconazole. Phenotypic, transcriptomic, and proteomic analyses reveal that P. aeruginosa's main QS molecule, N-(3-Oxododecanoyl)-L-homoserine lactone, induces candidal resistance to fluconazole by reversing the antifungal's effect on the ergosterol biosynthesis pathway. Accessory resistance mechanisms including upregulation of C. albicans drug-efflux, regulation of oxidative stress response, and maintenance of cell membrane integrity, further confirm this phenomenon. These findings demonstrate that P. aeruginosa QS molecules may confer protection to neighboring yeasts against azoles, in turn strengthening their co-existence in hostile polymicrobial infection sites.


Assuntos
Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Candida albicans/fisiologia , Farmacorresistência Fúngica , Fluconazol/farmacologia , Pseudomonas aeruginosa/fisiologia , Percepção de Quorum , Vias Biossintéticas , Ergosterol/biossíntese , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Interações Microbianas
14.
PLoS One ; 15(5): e0233102, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32392266

RESUMO

Candida auris, a decade old Candida species, has been identified globally as a significant nosocomial multidrug resistant (MDR) pathogen responsible for causing invasive outbreaks. Biofilms and overexpression of efflux pumps such as Major Facilitator Superfamily and ATP Binding Cassette are known to cause multidrug resistance in Candida species, including C. auris. Therefore, targeting these factors may prove an effective approach to combat MDR in C. auris. In this study, 25 clinical isolates of C. auris from different hospitals of South Africa were used. All the isolates were found capable enough to form biofilms on 96-well flat bottom microtiter plate that was further confirmed by MTT reduction assay. In addition, these strains have active drug efflux mechanism which was supported by rhodamine-6-G extracellular efflux and intracellular accumulation assays. Antifungal susceptibility profile of all the isolates against commonly used drugs was determined following CLSI recommended guidelines. We further studied the role of farnesol, an endogenous quorum sensing molecule, in modulating development of biofilms and drug efflux in C. auris. The MIC for planktonic cells ranged from 62.5-125 mM, and for sessile cells was 125 mM (4h biofilm) and 500 mM (12h and 24h biofilm). Furthermore, farnesol (125 mM) also suppresses adherence and biofilm formation by C. auris. Farnesol inhibited biofilm formation, blocked efflux pumps and downregulated biofilm- and efflux pump- associated genes. Modulation of C. auris biofilm formation and efflux pump activity by farnesol represent a promising approach for controlling life threatening infections caused by this pathogen.


Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Farneseno Álcool/farmacologia , Anfotericina B/farmacologia , Biofilmes/efeitos dos fármacos , Farmacorresistência Fúngica , Fluconazol/farmacologia , Testes de Sensibilidade Microbiana , Percepção de Quorum , África do Sul
16.
J Med Microbiol ; 69(6): 830-837, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32459616

RESUMO

Introduction. Cryptococcus species are pathogens commonly associated with cases of meningoencephalitis in individuals who are immunosuppressed due to AIDS.Aim. The aim was to evaluate the effects of the antiretroviral darunavir alone or associated with fluconazole, 5-flucytosine and amphotericin B against planktonic cells and biofilms of Cryptococcus species.Methodology. Susceptibility testing of darunavir and the common antifungals against 12 members of the Cryptococcus neoformans/Cryptococcus gattii species complex was evaluated by broth microdilution. The interaction between darunavir and antifungals against planktonic cells was tested by a checkerboard assay. The effects of darunavir against biofilm metabolic activity and biomass were evaluated by the XTT reduction assay and crystal violet staining, respectively.Results. Darunavir combined with amphotericin B showed a synergistic interaction against planktonic cells. No antagonistic interaction was observed between darunavir and the antifungals used. All Cryptococcus species strains were strong biofilm producers. Darunavir alone reduced biofilm metabolic activity and biomass when added during and after biofilm formation (P<0.05). The combination of darunavir with antifungals caused a significant reduction in biofilm metabolic activity and biomass when compared to darunavir alone (P<0.05).Conclusion. Darunavir presents antifungal activity against planktonic cells of Cryptococcus species and synergism with amphotericin B. In addition, darunavir led to reduced biofilm formation and showed activity against mature biofilms of Cryptococcus species. Activity of the antifungals against mature biofilms was enhanced in the presence of darunavir.


Assuntos
Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Cryptococcus gattii/efeitos dos fármacos , Cryptococcus neoformans/efeitos dos fármacos , Darunavir/farmacologia , Anfotericina B/farmacologia , Células Cultivadas , Fluconazol/farmacologia , Testes de Sensibilidade Microbiana/métodos , Plâncton/microbiologia
17.
PLoS Negl Trop Dis ; 14(3): e0008137, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32231354

RESUMO

BACKGROUND: Fluconazole is used in combination with amphotericin B for induction treatment of cryptococcal meningitis and as monotherapy for consolidation and maintenance treatment. More than 90% of isolates from first episodes of cryptococcal disease had a fluconazole minimum inhibitory concentration (MIC) ≤4 µg/ml in a Gauteng population-based surveillance study of Cryptococcus neoformans in 2007-2008. We assessed whether fluconazole resistance had emerged in clinical cryptococcal isolates over a decade. METHODOLOGY AND PRINCIPAL FINDINGS: We prospectively collected C. neoformans isolates from 1 January through 31 March 2017 from persons with a first episode of culture-confirmed cryptococcal disease at 37 South African hospitals. Isolates were phenotypically confirmed to C. neoformans species-complex level. We determined fluconazole MICs (range: 0.125 µg/ml to 64 µg/ml) of 229 C. neoformans isolates using custom-made broth microdilution panels prepared, inoculated and read according to Clinical and Laboratory Standards Institute M27-A3 and M60 recommendations. These MIC values were compared to MICs of 249 isolates from earlier surveillance (2007-2008). Clinical data were collected from patients during both surveillance periods. There were more males (61% vs 39%) and more participants on combination induction antifungal treatment (92% vs 32%) in 2017 compared to 2007-2008. The fluconazole MIC50, MIC90 and geometric mean MIC was 4 µg/ml, 8 µg/ml and 4.11 µg/ml in 2017 (n = 229) compared to 1 µg/ml, 2 µg/ml and 2.08 µg/ml in 2007-2008 (n = 249) respectively. Voriconazole, itraconazole and posaconazole Etests were performed on 16 of 229 (7%) C. neoformans isolates with a fluconazole MIC value of ≥16 µg/ml; only one had MIC values of >32 µg/ml for these three antifungal agents. CONCLUSIONS AND SIGNIFICANCE: Fluconazole MIC50 and MIC90 values were two-fold higher in 2017 compared to 2007-2008. Although there are no breakpoints, higher fluconazole doses may be required to maintain efficacy of standard treatment regimens for cryptococcal meningitis.


Assuntos
Antifúngicos/farmacologia , Criptococose/microbiologia , Cryptococcus neoformans/efeitos dos fármacos , Farmacorresistência Fúngica , Fluconazol/farmacologia , Adulto , Cryptococcus neoformans/isolamento & purificação , Feminino , Hospitais , Humanos , Masculino , Testes de Sensibilidade Microbiana , Estudos Prospectivos , África do Sul
18.
Mycoses ; 63(5): 471-477, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32124485

RESUMO

INTRODUCTION: The prevalence of azole resistance in C parapsilosis is very low in most parts of the world. However, South Africa has reported an exceptionally high prevalence of azole resistance in C parapsilosis strains isolated from candidaemia cases. We aimed to determine the possible molecular mechanisms of fluconazole resistance in C parapsilosis isolates obtained through surveillance at a large neonatal unit at a South African academic hospital. METHODS: We sequenced the ERG11 and MRR1 genes of C parapsilosis isolates recovered from cases of neonatal candidemia, followed by microsatellite genotyping. A total of 73 isolates with antifungal susceptibility results were analysed. RESULTS: Of these, 57 (78%) were resistant, 11 (15%) susceptible dose-dependent and 5 (7%) susceptible. The most commonly identified amino acid substitution within the ERG11 gene was Y132F in 68% (39/57) of fluconazole-resistant isolates and none in susceptible isolates. Three amino acid substitutions (R405K, G583R and A619V) and 1 nucleotide deletion at position 1331 were identified within MRR1 gene in 19 (26%) isolates. Microsatellite genotyping grouped isolates into four clusters (50 isolates). Cluster 1 accounted for 23% (17/73) of all cases, cluster 2 for 22% (16/73), cluster 3 for 14% (10/73) and cluster 4 for 10% (7/73). We found an association between cluster type and fluconazole resistance (P-value = .004). Isolates harbouring the Y132F substitution were more likely to belong to a cluster than non-Y132F isolates. CONCLUSION: Fluconazole resistance in C parapsilosis strains from a single South African neonatal unit was associated with cluster type and predominantly driven by Y123F amino acid substitutions in the ERG11 gene.


Assuntos
Candida parapsilosis/efeitos dos fármacos , Candida parapsilosis/genética , Candidemia/microbiologia , Farmacorresistência Fúngica/genética , Fluconazol/farmacologia , Proteínas Fúngicas/genética , Centros Médicos Acadêmicos , Substituição de Aminoácidos , Azóis/farmacologia , Genótipo , Humanos , Lactente , Recém-Nascido , Testes de Sensibilidade Microbiana , Repetições de Microssatélites , Berçários Hospitalares , África do Sul
19.
J Mycol Med ; 30(2): 100935, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32139093

RESUMO

OBJECTIVE: Dermatophytes are a group of keratinophilic fungi that invade and infect the keratinized tissues and cause dermatophytosis. We investigated effectiveness of novel triazole (luliconazole and lanaconazole) in comparison with available antifungal agents against dermatophyte species isolated from patients with tinea pedis. MATERIAL AND METHODS: A total of 60 dermatophytes species were isolated from the patients with tinea pedis. Identification of species was done by DNA sequencing of the ITS1-5.8S rDNA-ITS2 rDNA region. In vitro antifungal susceptibility testing with luliconazole and lanaconazole and available antifungal agent was done in accordance with the Clinical and Laboratory Standards Institute, M38-A2 document. RESULTS: In all investigated isolates, luliconazole had the lowest minimum inhibitory concentration (MIC) (MIC range=0.0005-0.004µg/mL), while fluconazole (MIC range=0.4-64µg/mL) had the highest MICs. Geometric mean MIC was the lowest for luliconazole (0.0008µg/mL), followed by lanoconazole (0.003µg/mL), terbinafine (0.019µg/mL), itraconazole (0.085 µg/mL), ketoconazole (0.089µg/mL), econazole (0.097µg/mL), griseofulvin (0.351 µg/mL), voriconazole (0.583µg/mL) and fluconazole (11.58µg/mL). CONCLUSION: The novel triazoles showed potent activity against dermatophytes and promising candidates for the treatment of tinea pedis caused by Trichophyton and Epidermophyton species. However, further studies are warranted to determine the clinical implications of these investigations.


Assuntos
Antifúngicos/farmacologia , Arthrodermataceae/efeitos dos fármacos , Tinha dos Pés/microbiologia , Triazóis/farmacologia , Arthrodermataceae/crescimento & desenvolvimento , Dermatomicoses/tratamento farmacológico , Dermatomicoses/microbiologia , Fluconazol/farmacologia , Griseofulvina/farmacologia , Humanos , Imidazóis/farmacologia , Itraconazol/farmacologia , Cetoconazol/farmacologia , Testes de Sensibilidade Microbiana , Terbinafina/farmacologia , Tinha/tratamento farmacológico , Tinha/microbiologia , Tinha dos Pés/tratamento farmacológico , Trichophyton/efeitos dos fármacos , Trichophyton/crescimento & desenvolvimento , Voriconazol/farmacologia
20.
Medicine (Baltimore) ; 99(11): e19494, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32176090

RESUMO

As detection rates of non-albicans Candida species are increasing, determining their pathogen profiles and antifungal susceptibilities is important for antifungal treatment selection. We identified the antifungal susceptibility patterns and predictive factors for mortality in candidemia.A multicenter retrospective analysis of patients with at least 1 blood culture positive for Candida species was conducted. Candida species were classified into 3 groups (group A, Candia albicans; group B, Candida tropicalis, and Candida parasilosis; group C, Candida glabrata and Candida krusei ) to analyze the susceptibility patterns, first-line antifungal administered, and mortality. Univariate and multivariate comparisons between outcomes were performed to identify mortality risk factors.In total, 317 patients were identified, and 136 (42.9%) had recorded mortality. Echinocandin susceptibility was higher for group A than group B (111/111 [100%] vs 77/94 [81.9%], P < .001). Moreover, group A demonstrated higher fluconazole susceptibility (144/149 [96.6%] vs 39/55 [70.9%], P < .001) and lower mortality (68 [45.3%] vs 34 [61.8%], P = .036) than those of group C. In the multivariate analysis, the sequential organ failure assessment score (odds ratio OR 1.351, 95% confidence interval 1.067-1.711, p = 0.013) and positive blood culture on day 7 of hospitalization (odds ratio 5.506, 95% confidence interval, 1.697-17.860, P = .004) were associated with a higher risk of mortality.Patients with higher sequential organ failure assessment scores and sustained positive blood cultures have an increased risk of mortality.


Assuntos
Antifúngicos/uso terapêutico , Candida/efeitos dos fármacos , Candidemia/tratamento farmacológico , Farmacorresistência Fúngica , Fluconazol/uso terapêutico , Idoso , Antifúngicos/farmacologia , Candidemia/mortalidade , Feminino , Fluconazol/farmacologia , Humanos , Masculino , Testes de Sensibilidade Microbiana , Valor Preditivo dos Testes , República da Coreia , Estudos Retrospectivos , Fatores de Risco
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