RESUMO
Fungi of the genus Candida are important etiological agents of superficial and life-threatening infections in individuals with a compromised immune system. One of the main characteristics of Candida is its ability to form highly drug tolerance biofilms in the human host. Biofilms are a dynamic community of multiple cell types whose formation over time is orchestrated by a network of transcription regulators. In this brief review, we provide an update of the processes involved in biofilm formation by Candida spp. (formation, treatment, and control), as well as the transcriptional circuitry that regulates its development and interactions with other microorganisms. Candida albicans is known to build mixed species biofilms with other Candida species and with various other bacterial species in different host niches. Taken together, these properties play a key role in Candida pathogenesis. In addition, this review gathers recent studies with new insights and perspectives for the treatment and control of Candida biofilms.
Assuntos
Biofilmes/crescimento & desenvolvimento , Candida/fisiologia , Biofilmes/efeitos dos fármacos , Candida/efeitos dos fármacos , Candida/genética , Candida/ultraestrutura , Adesão Celular/genética , Adesão Celular/fisiologia , Estudo de Associação Genômica Ampla , Humanos , Microscopia Eletrônica de Varredura , Nanotecnologia/tendências , Elementos Reguladores de Transcrição/genética , Elementos Reguladores de Transcrição/fisiologiaRESUMO
The pomegranate (Punica granatum) sarcotesta contains a chitin-binding lectin (PgTeL) with antibacterial activity against human pathogenic species. In this work, the structural stability of PgTeL was evaluated by fluorimetric analysis and the lectin was evaluated for cytotoxicity to human peripheral blood mononuclear cells (PBMCs) and antifungal activity against Candida albicans and Candida krusei. PgTeL folding was impaired when lectin was incubated at pH≥6.0. On the other hand, the lectin did not undergo unfolding even when heated at 100°C. PgTeL (1, 10, and 100µg/mL) was not cytotoxic to PBMCs. Antifungal activity was detected for C. albicans (MIC: 25µg/mL; MFC: 50µg/mL) and C. krusei (MIC and MFC of 12.5µg/mL). Treatment of yeast cells with PgTeL resulted in decrease of intracellular ATP content even at sub-inhibitory concentrations (½MIC and »MIC) and induced lipid peroxidation. In addition, PgTeL damaged the integrity of fungal cell wall of both species, with more pronounced effects in C. krusei. The lectin showed significant antibiofilm activity on C. albicans at sub-inhibitory concentrations (0.195 and 0.39µg/mL). In conclusion, PgTeL is an anti-Candida agent whose action mechanism involves oxidative stress, energetic collapse, damage to the cell wall and rupture of yeast cells.
Assuntos
Candida albicans/efeitos dos fármacos , Candida/efeitos dos fármacos , Lectinas/farmacologia , Lythraceae/química , Antifúngicos/química , Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Candida/metabolismo , Candida/ultraestrutura , Candida albicans/metabolismo , Candida albicans/ultraestrutura , Parede Celular/efeitos dos fármacos , Humanos , Concentração de Íons de Hidrogênio , Lectinas/química , Lectinas/isolamento & purificação , Peroxidação de Lipídeos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , TemperaturaRESUMO
OBJECTIVES: This study aimed to assess the biofilm-forming ability of Candida spp. from the ocular conjunctiva of horses and to investigate the antifungal susceptibility of these biofilms. PROCEDURES: Initially, the biofilm-forming ability of 15 strains was assessed by crystal violet staining, which reveals the fungal biomass adhered to the polystyrene plates, and scanning electron microscopy. Then, the minimum inhibitory concentrations (MICs) of amphotericin B, fluconazole, itraconazole, and caspofungin were initially determined against strains in planktonic form. Afterward, antifungal susceptibility of mature biofilms was evaluated by exposing them to 10 × MIC and 50 × MIC of the tested drugs, followed by the assessment of their metabolic activity, using the oxidoreduction indicator XTT. Results were analyzed through ANOVA and Tukey's post-test, and P-values below 5% led to significant conclusions. RESULTS: Eight strains produced biofilms and were classified as strong (1/15), moderate (3/15) and weak (4/15) producers, according to the amount of crystal violet retained by the adhered fungal biomass. Biofilm metabolic activity of one C. tropicalis did not decrease after exposure to the tested antifungals, while biofilm metabolic activity of five strains was reduced by amphotericin B, but not the other drugs. One C. parapsilosis sensu stricto and one C. glabrata showed significant reduction in biofilm metabolic activity after exposure to fluconazole, itraconazole, and caspofungin, but not amphotericin B. CONCLUSIONS: The results demonstrate that Candida from the ocular conjunctiva of horses can pose as a risk to animal health as they are capable of forming biofilms, which are commonly involved in fungal keratitis.
Assuntos
Antifúngicos/farmacologia , Azóis/farmacologia , Biofilmes/efeitos dos fármacos , Candida/efeitos dos fármacos , Túnica Conjuntiva/microbiologia , Anfotericina B/farmacologia , Animais , Candida/classificação , Candida/fisiologia , Candida/ultraestrutura , Caspofungina , Equinocandinas/farmacologia , Infecções Oculares Fúngicas/microbiologia , Infecções Oculares Fúngicas/veterinária , Fluconazol/farmacologia , Doenças dos Cavalos/microbiologia , Cavalos , Itraconazol/farmacologia , Ceratite/microbiologia , Ceratite/veterinária , Lipopeptídeos/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
Candida infection is an important cause of morbidity and mortality on immunosuppressed patients. This growing trend has been associated with resistance to the antimicrobial therapy and the ability of microorganism to form biofilms. TTO oil is used as antimicrobial which shows antibiofilm activity against Candida species. However, it presents problems due to its poor solubility and high volatility. The present study aimed to evaluate in vitro antibiofilm activity of TTO nanoparticles against many Candida species. It was performed the characterization of the oil and nanoparticles. The levels of exopolysaccharides, proteins, and the biomass of biofilms were measured. The chromatographic profile demonstrated that the TTO oil is in accordance with ISO 4730 with major constituents of 41.9% Terpinen-4-ol, 20.1% of γ-Terpinene, 9,8% of α-Terpinene, and 6,0% of 1,8-Cineole. The TTO nanoparticles showed pH of 6.3, mean diameter of 158.2 ± 2 nm, polydispersion index of 0.213 ± 0.017, and zeta potential of -8.69 ± 0.80 mV. The addition of TTO and its nanoparticles represented a significant reduction of biofilm formed by all Candida species, as well as a reduction of proteins and exopolysaccharides levels. It was possible to visualize the reduction of biofilm in presence of TTO nanoparticles by Calcofluor White method.
Assuntos
Antifúngicos/administração & dosagem , Biofilmes/efeitos dos fármacos , Candida/efeitos dos fármacos , Candida/fisiologia , Melaleuca/química , Nanopartículas/administração & dosagem , Extratos Vegetais/administração & dosagem , Antifúngicos/química , Proteínas de Bactérias/metabolismo , Biomassa , Candida/ultraestrutura , Testes de Sensibilidade Microbiana , Nanopartículas/química , Extratos Vegetais/química , Óleos de Plantas/administração & dosagem , Óleos de Plantas/químicaRESUMO
To clarify the interactions between a common food spoilage yeast and two pathogenic bacteria involved in outbreaks associated with fruit juices, the present paper studies the effect of the interplay of Candida krusei, collected from UF membranes, with Escherichia coli O157:H7 and Salmonella enterica in the overall process of adhesion and colonization of abiotic surfaces. Two different cases were tested: a) co-adhesion by pathogenic bacteria and yeasts, and b) incorporation of bacteria to pre-adhered C. krusei cells. Cultures were made on stainless steel at 25°C using apple juice as culture medium. After 24 h of co-adhesion with C. krusei, both E. coli O157:H7 and S. enterica increased their counts 1.05 and 1.11 log CFU cm2, respectively. Similar increases were obtained when incorporating bacteria to pre-adhered cells of Candida. Nevertheless C. krusei counts decreased in both experimental conditions, in a) 0.40 log CFU cm2 and 0.55 log CFU cm2 when exposed to E. coli O157:H7 and S. enterica and in b) 0.18 and 0.68 log CFU cm2, respectively. This suggests that C. krusei, E. coli O157:H7, and S. enterica have a complex relationship involving physical and chemical interactions on food contact surfaces. This study supports the possibility that pathogen interactions with members of spoilage microbiota, such as C. krusei, might play an important role for the survival and dissemination of E. coli O157:H7 and Salmonella enterica in food-processing environments. Based on the data obtained from the present study, much more attention should be given to prevent the contamination of these pathogens in acidic drinks.
Assuntos
Aderência Bacteriana , Candida/fisiologia , Escherichia coli O157/fisiologia , Sucos de Frutas e Vegetais/microbiologia , Salmonella enterica/fisiologia , Aço Inoxidável , Candida/isolamento & purificação , Candida/ultraestrutura , Meios de Cultura/química , Escherichia coli O157/crescimento & desenvolvimento , Escherichia coli O157/ultraestrutura , Manipulação de Alimentos , Microbiologia de Alimentos , Conservação de Alimentos , Malus/microbiologia , Salmonella enterica/crescimento & desenvolvimento , Salmonella enterica/ultraestrutura , UltrafiltraçãoRESUMO
CONTEXT: Uncaria tomentosa D.C. (Rubiaceae) has several biological activities, including activity against resistant Candida strains. The synergistic interaction with terbinafine or fluconazole can be an important alternative to overcome this resistance. OBJECTIVES: The potential synergy between a water insoluble fraction (WIF) from Uncaria tomentosa bark and the antifungals terbinafine (TRB) and fluconazole (FLZ) against non-Candida albicans resistant strains was investigated. MATERIALS AND METHODS: TRB and FLZ, alone and combined with WIF, were tested by the checkerboard procedure using the micro-dilution technique against seven isolates of Candida glabrata and C. krusei. The molecular interactions occurring outside the cell wall were evaluated by scanning electron microscopy, Fourier transform infrared (FT-IR) and differential scanning calorimetry (DSC) analysis. RESULTS: The checkerboard inhibitory assay demonstrated synergy for WIF:TRB and WIF:FLZ combinations, respectively. The best synergistic cell damage was demonstrated unequivocally for the associations of WIF and TRB (1.95:4.0 µg/mL) and WIF and FLZ (1.95:8.0 µg/mL). The comparison of the FT-IR spectra of the antifungal alone, and in combination with WIF, allows recognizing clear differences in 3000, 1600, 1400, and 700-800 cm-1 bands. Additionally, modifications on TRB and FLZ thermograms were clearly noticed after their combination with WIF. CONCLUSIONS: DSC and infrared analysis demonstrated intermolecular interactions between WIF and either TRB or FLZ. Hence, quite likely the synergistic effect is related to interaction events occurring outside the cell wall between antifungal and cat's claw proanthocyanidins. A direct action on the cell wall is suggested, without connection with the ABC efflux pump mechanism.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Unha-de-Gato/química , Farmacorresistência Fúngica/efeitos dos fármacos , Fluconazol/farmacologia , Naftalenos/farmacologia , Extratos Vegetais/farmacologia , Antifúngicos/isolamento & purificação , Varredura Diferencial de Calorimetria , Candida/crescimento & desenvolvimento , Candida/ultraestrutura , Parede Celular/efeitos dos fármacos , Parede Celular/ultraestrutura , Sinergismo Farmacológico , Microscopia Eletrônica de Varredura , Fitoterapia , Casca de Planta , Extratos Vegetais/isolamento & purificação , Plantas Medicinais , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier , Terbinafina , Água/químicaRESUMO
The microbial synthesis of nanoparticles is a green chemistry approach that combines nanotechnology and microbial biotechnology. The aim of this study was to obtain silver nanoparticles (SNPs) using aqueous extract from the filamentous fungus Fusarium oxysporum as an alternative to chemical procedures and to evaluate its antifungal activity. SNPs production increased in a concentration-dependent way up to 1 mM silver nitrate until 30 days of reaction. Monodispersed and spherical SNPs were predominantly produced. After 60 days, it was possible to observe degenerated SNPs with in additional needle morphology. The SNPs showed a high antifungal activity against Candida and Cryptococcus , with minimum inhibitory concentration values ≤ 1.68 µg/mL for both genera. Morphological alterations of Cryptococcus neoformans treated with SNPs were observed such as disruption of the cell wall and cytoplasmic membrane and lost of the cytoplasm content. This work revealed that SNPs can be easily produced by F. oxysporum aqueous extracts and may be a feasible, low-cost, environmentally friendly method for generating stable and uniformly sized SNPs. Finally, we have demonstrated that these SNPs are active against pathogenic fungi, such as Candida and Cryptococcus.
Assuntos
Antifúngicos/metabolismo , Candida/efeitos dos fármacos , Cryptococcus/efeitos dos fármacos , Fusarium/metabolismo , Nanopartículas Metálicas , Prata/metabolismo , Antifúngicos/uso terapêutico , Candida/classificação , Candida/ultraestrutura , Extratos Celulares , Cryptococcus/classificação , Cryptococcus/ultraestrutura , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Inibidores do Crescimento , Nanopartículas Metálicas/uso terapêutico , Testes de Sensibilidade Microbiana , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Prata/análise , Prata/uso terapêuticoRESUMO
The microbial synthesis of nanoparticles is a green chemistry approach that combines nanotechnology and microbial biotechnology. The aim of this study was to obtain silver nanoparticles (SNPs) using aqueous extract from the filamentous fungus Fusarium oxysporum as an alternative to chemical procedures and to evaluate its antifungal activity. SNPs production increased in a concentration-dependent way up to 1 mM silver nitrate until 30 days of reaction. Monodispersed and spherical SNPs were predominantly produced. After 60 days, it was possible to observe degenerated SNPs with in additional needle morphology. The SNPs showed a high antifungal activity against Candida and Cryptococcus , with minimum inhibitory concentration values ≤ 1.68 µg/mL for both genera. Morphological alterations of Cryptococcus neoformans treated with SNPs were observed such as disruption of the cell wall and cytoplasmic membrane and lost of the cytoplasm content. This work revealed that SNPs can be easily produced by F. oxysporum aqueous extracts and may be a feasible, low-cost, environmentally friendly method for generating stable and uniformly sized SNPs. Finally, we have demonstrated that these SNPs are active against pathogenic fungi, such as Candida and Cryptococcus .
Assuntos
Antifúngicos/metabolismo , Candida/efeitos dos fármacos , Cryptococcus/efeitos dos fármacos , Fusarium/metabolismo , Nanopartículas Metálicas , Prata/metabolismo , Antifúngicos/uso terapêutico , Extratos Celulares , Candida/classificação , Candida/ultraestrutura , Cryptococcus/classificação , Cryptococcus/ultraestrutura , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Inibidores do Crescimento , Testes de Sensibilidade Microbiana , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Nanopartículas Metálicas/uso terapêutico , Prata/análise , Prata/uso terapêuticoRESUMO
The production of virulence attributes in three reference strains and 11 clinical isolates primarily identified as Candida parapsilosis was evaluated. Morphological and phenotypical tests were not able to discriminate among the three species of the C. parapsilosis complex; consequently, molecular methods were applied to solve this task. After employing polymerase chain reaction-based methods, nine clinical strains were identified as C. parapsilosis sensu stricto and two as C. orthopsilosis. Protease, catalase, and hemolysin were produced by all 14 strains, while 92.9% and 78.6% of strains secreted, respectively, esterase and phytase. No phospholipase producers were detected. Mannose/glucose, N-acetylglucosamine, and sialic acid residues were detected at the surface of all strains, respectively, in high, medium, and low levels. All strains presented elevated surface hydrophobicity and similar ability to form biofilm. However, the adhesion to inert substrates and mammalian cells was extremely diverse, showing typical intrastrain variations. Overall, the strains showed (1) predilection to adhere to plastic over glass and the number of pseudohyphae was more prominent than yeasts and (2) the interaction process was slightly enhanced in macrophages than fibroblasts, with the majority of fungal cells detected inside them. Positive/negative correlations were demonstrated among the production of these virulence traits in C. parapsilosis complex.
Assuntos
Candida/classificação , Fenótipo , Biofilmes , Candida/fisiologia , Candida/ultraestrutura , Membrana Celular/química , Membrana Celular/metabolismo , Glicosilação , Humanos , Hidrólise , Interações Hidrofóbicas e Hidrofílicas , Tipagem Molecular , Filogenia , RNA Fúngico , RNA Ribossômico 28S , Virulência/genéticaRESUMO
Ethanolic crude extracts prepared from the arils and seeds, pericarp, peels and from the whole fruit of Punica granatum, known as pomegranate, had their antifungal activity tested against Candida spp. The ethanolic crude extracts were analyzed by Mass Spectrometry and yielded many compounds such as punicalagin and galladydilacton. The extracts from the pericarp and peel showed activity against Candida spp., with MICs of 125 µg/mL. The effect of pericarp and peel extracts upon the morphological and structure of C. albicans and C. krusei were examined by scanning and transmission electron microscopy, with the visualization of an irregular membrane and hyphae, formation of vacuoles and thickening of the cell wall. The data obtained revealed potential antimicrobial activity against yeasts cells of the Candida genus, and the bioactive compounds could be responsible for changes in cell morphology and structure. The data obtained open new perspectives for future research in continuation to this study, where information such as determination of the site of action of the compounds could contribute to an alternative therapy against these organisms.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Extratos Vegetais/farmacologia , Lythraceae/química , Antifúngicos/síntese química , Antifúngicos/isolamento & purificação , Candida/ultraestrutura , Membrana Celular/efeitos dos fármacos , Membrana Celular/ultraestrutura , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Organelas/efeitos dos fármacos , Organelas/ultraestrutura , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificaçãoRESUMO
Ethanolic crude extracts prepared from the arils and seeds, pericarp, peels and from the whole fruit of Punica granatum, known as pomegranate, had their antifungal activity tested against Candida spp. The ethanolic crude extracts were analyzed by Mass Spectrometry and yielded many compounds such as punicalagin and galladydilacton. The extracts from the pericarp and peel showed activity against Candida spp., with MICs of 125 µg/mL. The effect of pericarp and peel extracts upon the morphological and structure of C. albicans and C. krusei were examined by scanning and transmission electron microscopy, with the visualization of an irregular membrane and hyphae, formation of vacuoles and thickening of the cell wall. The data obtained revealed potential antimicrobial activity against yeasts cells of the Candida genus, and the bioactive compounds could be responsible for changes in cell morphology and structure. The data obtained open new perspectives for future research in continuation to this study, where information such as determination of the site of action of the compounds could contribute to an alternative therapy against these organisms.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Lythraceae/química , Extratos Vegetais/farmacologia , Antifúngicos/síntese química , Antifúngicos/isolamento & purificação , Candida/ultraestrutura , Membrana Celular/efeitos dos fármacos , Membrana Celular/ultraestrutura , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Organelas/efeitos dos fármacos , Organelas/ultraestrutura , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificaçãoRESUMO
Vulvovaginal candidiasis (VVC) is a very common cause of fungal infection that remains a significant problem worldwide, especially concerning its complex pathogenicity. Biofilm dynamics from vaginal isolates requires further investigation. Different assays, such as cell surface hydrophobicity (CSH), biofilm production, fungal metabolism by 2H-tetrazolium-5-carboxanilide (XTT) and phenazine methosulfate (PMS), scanning electron microscopy (SEM) and confocal scanning laser microscopy (CSLM) were used in order to determine the ability of five Candida species isolates from VVC patients to form in vitro biofilms and their ultrastructural characteristics. All yeasts demonstrated the ability to produce biofilm and showed viability up to 48 h after the completion of assay, confirmed by SEM and CSLM, but differences were observed between them. SEM and CSLM also revealed that all VVC isolates adhered only in blastoconidia form, except for Candida parapsilosis. Even though, only one isolate from each Candida species has been used, the results of high biofilm formation, metabolic activity and CSH showed by Candida albicans and C. parapsilosis, as well as by the ultrastructural characteristics, suggest that these species exhibit greater ability of adherence in relation to the others. Ours results support the theory that virulence potential is multifactorial and that other factors not evaluated in this study could be involved in the CVV physiopathogeny.
Assuntos
Biofilmes/crescimento & desenvolvimento , Candida/crescimento & desenvolvimento , Candida/ultraestrutura , Candidíase Vulvovaginal/microbiologia , Candida/isolamento & purificação , Candida/fisiologia , Feminino , Humanos , Interações Hidrofóbicas e Hidrofílicas , Metilfenazônio Metossulfato/metabolismo , Viabilidade Microbiana , Microscopia Confocal , Microscopia Eletrônica de Varredura , Propriedades de Superfície , Sais de Tetrazólio/metabolismoRESUMO
The research of alternative substances to treat infections caused by Candida species is a need. Aromatic plants have the ability to produce secondary metabolites, such as essential oils (EO). The antimicrobial properties of Aloysia triphylla (L'Her.) Britton (cedrón) EO has been previously described. The aims of this work were to determine the antimicrobial activity and the effect on the cell structure of the EO of A. triphylla against Candida sp isolated from human illnesses. The EO was obtained by hydrodistillation of A. triphylla leaves. The minimum inhibitory concentration (MIC) was performed with microdilution method and the minimum fungicidal concentration (MFC) was determined. A. triphylla EO's showed antifungal activity against all yeast: C. albicans, C. dubliniensis, C. glabrata, C. krusei, C. guillermondii, C. parapsilosis and C. tropicalis which were resistant to fluconazol (150 mg/mL). The range of MIC values was from: 35 to 140 microg/mL and the MFC: 1842 to 2300 microg/mL. The time of killing at the MFC against C. albicans (3 x 10(5) UFC/mL) was 140 min. The dates of OD620 and OD260 suggest lysis and loss of absorbing material, respectively. The HROM shows distortion in morphology and shape of the cell, with large vacuoles in the cytoplasm. These studies clearly show that A. triphylla EO is a promising alternative for the treatment of candidiasis.
Assuntos
Candida/efeitos dos fármacos , Candidíase/microbiologia , Óleos Voláteis/química , Óleos de Plantas/química , Terpenos/isolamento & purificação , Verbenaceae/química , Argentina , Candida/citologia , Candida/ultraestrutura , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Testes de Sensibilidade Microbiana , Óleos Voláteis/farmacologia , Folhas de Planta/química , Óleos de Plantas/farmacologia , Terpenos/farmacologiaRESUMO
Paradoxical growth (PG) has been described for echinocandins and is characterized by cell growth at drug concentrations above the MIC. In this study, two isolates each of Candida albicans, C. tropicalis, C. orthopsilosis, and C. parapsilosis, all of which displaying PG in response to caspofungin, were subjected to MIC, minimal fungicidal concentration (MFC), and time-kill curve assays to evaluate the levels of PG. Cell wall components and ultrastructural modifications of the PG cells were also investigated. The results showed that when cell growth and survival were evaluated by MFC or time-kill curve assays, high concentrations of caspofungin did not show fungicidal activity against PG cells. Furthermore, for C. parapsilosis and C. orthopsilosis, time-kill curves were more discriminatory than MFCs in detecting the PG effect. The four different Candida species studied demonstrated similar alterations in cell wall components and ultrastructure associated with PG. In PG cells, ß-1,3-glucan content decreased from 2.7- to 7.8-fold, whereas chitin content increased from 4.0- to 6.6-fold. An electron microscopy study of the PG cells revealed morphological alterations, clumping of cells, enlarged cells, the absence of filamentation, abnormal septa, and accumulation of chitin in the cell wall. Also, PG cells basically exhibited a single dark high-density layer in the cell wall, indicating the loss of the ß-1,3-glucan layer. Our results present novel details about the ultrastructural alterations that occur in C. albicans, C. parapsilosis, C. orthopsilosis, and C. tropicalis during PG and show that chitin is the major component of the cell walls of PG cells. Stimulation of chitin synthesis may represent a rescue mechanism against caspofungin activity.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Candida/ultraestrutura , Parede Celular/efeitos dos fármacos , Parede Celular/ultraestrutura , Equinocandinas/farmacologia , Caspofungina , Lipopeptídeos , Testes de Sensibilidade Microbiana , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Microscopia de FluorescênciaRESUMO
The residue (vinasse) formed during the distillation of bio-ethanol and cachaça, a traditional rum-type spirit produced from sugar-cane in Brazil, is highly harmful if discharged into the environment due to high values of COD and BOD. One possibility for minimizing the impact of vinasse in soils and waters is to use the residue in the production of microbial biomass for use as an animal feed supplement that will provide high levels on nitrogen (>9% d.m.) and low content of nucleic (≤ 10% d.m.) This paper reports the production and quality of biomass produced from fermentation of Saccharomyces cerevisiae and Candida parapsilosis in culture media under 12 different culture conditions and the respective effects of each variable (glucose, yeast extract, peptone, potassium phosphate, vinasse, pH and temperature). Of the S. cerevisiae isolates tested, two (VR1 and PE2) originating from fuel alcohol-producing plants were identified as offering the best potential for the industrial production of single cell protein from vinasse due to highest biomass productivity. Our results showed a potential viable and economic use of vinasse.
Assuntos
Bebidas Alcoólicas , Proteínas Alimentares/metabolismo , Etanol/metabolismo , Proteínas Fúngicas/metabolismo , Gerenciamento de Resíduos/métodos , Leveduras/metabolismo , Ração Animal , Candida/crescimento & desenvolvimento , Candida/metabolismo , Candida/ultraestrutura , Destilação , Etanol/química , Concentração de Íons de Hidrogênio , Nitrogênio/análise , Nitrogênio/metabolismo , Avaliação Nutricional , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/ultraestrutura , TemperaturaRESUMO
Although bacterial biofilms have been studied in detail, adhesion of Candida albicans and non-albicans species to an intrauterine contraceptive device (IUD) is not clear. The objective of this study was to evaluate aspects of imaging of the ultrastructure and viability of vaginal yeasts adhered to different parts of an IUD, through scanning electron microscopy (SEM) and confocal scanning laser microscopy (CSLM). We studied yeasts isolated from different patients with vulvovaginal candidiasis: C. albicans, C. glabrata, C. guillermondii, C. parapsilosis, C. tropicalis, and Saccharomyces cerevisiae. A suspension of the each yeast was prepared and incubated with IUD parts (tail, without copper, and copper-covered). SEM and CSLM showed that all the vaginal yeasts adhered to all the parts of the IUD and demonstrated viability, including 30 days after contact for C. albicans. Possibly irregularities of IUD surface contribute to the adherence process. Although all of the IUD parts contribute to retention of yeasts in the genital tract, high concentration of yeast cells on the tail may indicate the importance of this segment in maintaining the colonization by yeast cells because the tail forms a bridge between the external environment, the vagina that is colonized by yeast cells, and the upper genital tract where there is no colonization.
Assuntos
Candida/ultraestrutura , Dispositivos Intrauterinos/microbiologia , Saccharomyces cerevisiae/ultraestrutura , Biofilmes/crescimento & desenvolvimento , Candida/isolamento & purificação , Candida/fisiologia , Candidíase Vulvovaginal/microbiologia , Feminino , Humanos , Viabilidade Microbiana , Microscopia Confocal , Microscopia Eletrônica de Varredura , Saccharomyces cerevisiae/fisiologiaRESUMO
Activity-guided repeated fractionation of crude hydro alcoholic extract prepared from the fruit peel of Punica granatum on a silica-gel column yielded a compound that exhibited strong antifungal activity against Candida spp. Based on spectral analyses, the compound was identified as punicalagin. Punicalagin showed strong activity against Candida albicans and Candida parapsilosis, with MICs of 3.9 and 1.9 microg/ml, respectively. The combination of punicalagin and fluconazole showed a synergistic interaction. MIC for fluconazole decreased twofold when combined with the extract. The FIC index was 0.25. The synergism observed in disk-diffusion and checkerboard assays was confirmed in time-kill curves. The effect of punicalagin on the morphology and ultrastructure in treated yeast cells was examined by scanning and transmission electron microscopy. An irregular budding pattern and pseudohyphae were seen in treated yeasts. By transmission electron microscopy, treated cells showed a thickened cell wall, changes in the space between cell wall and the plasma membrane, vacuoles, and a reduction in cytoplasmic content. Since the punicalagin concentration effective in vitro is achievable in vivo, the combination of this agent with fluconazole represents an attractive prospect for the development of new management strategies for candidiasis, and should be investigated further in in vivo models.
Assuntos
Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Candida/efeitos dos fármacos , Fluconazol/farmacologia , Taninos Hidrolisáveis/farmacologia , Lythraceae/química , Antifúngicos/química , Antifúngicos/isolamento & purificação , Candida/ultraestrutura , Candida albicans/ultraestrutura , Cromatografia em Camada Fina , Farmacorresistência Fúngica , Sinergismo Farmacológico , Taninos Hidrolisáveis/química , Taninos Hidrolisáveis/isolamento & purificação , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Epiderme Vegetal/químicaRESUMO
Candida parapsilosis is found frequently as commensal organism on epithelial tissues, and is also an increasing cause of nosocomial infection. Scanning electron microscope (SEM) observations were used to analyse the capability of C. parapsilosis cells to adhere and grow as biofilm on human natural substrates and to compare the adherence pattern of isolates exhibiting distinct phenotypes. Cells from the crepe phenotype are predominantly elongated and form pseudohyphae whereas cells from the smooth phenotype are yeast-shaped, either in liquid cultures or on human nail and hair surfaces. The electron micrographs revealed that C. parapsilosis cells from the smooth phenotype adhered in higher number to both surfaces compared to the observed for the crepe phenotype. SEM analysis of human hair surface revealed that cells from the smooth phenotype appear as clumped blastoconidia of uniform morphology embedded in a flocculent extracellular material forming biofilm. The extracellular material and biofilm were seeing in a less extension in the crepe phenotype. A distinct adherence pattern was observed when human nail was used as substrate. Here C. parapsilosis cells seem to be linked to surface structures of human nail plate. Fibrillar extracellular material was observed connecting neighbouring cells as well as nail surface.
Assuntos
Candida/ultraestrutura , Cabelo/microbiologia , Interações Hospedeiro-Patógeno , Unhas/microbiologia , Biofilmes/crescimento & desenvolvimento , Adesão Celular , Humanos , Microscopia Eletrônica de VarreduraRESUMO
Candida biofilms are microbial communities, embedded in a polymeric matrix, growing attached to a surface, and are highly recalcitrant to antimicrobial therapy. These biofilms exhibit enhanced resistance against most antifungal agents except echinocandins and lipid formulations of amphotericin B. In this study, biofilm formation by different Candida species, particularly Candida albicans, C. tropicalis, and C. parapsilosis, was evaluated, and the effect of caspofungin (CAS) was assessed using a clinically relevant in vitro model system. CAS displayed in vitro activity against C. albicans and C. tropicalis cells within biofilms. Biofilm formation was evaluated after 48 h of antifungal drug exposure, and the effects of CAS on preformed Candida species biofilms were visualized using scanning electron microscopy (SEM). Several species-specific differences in the cellular morphologies associated with biofilms were observed. Our results confirmed the presence of paradoxical growth (PG) in C. albicans and C. tropicalis biofilms in the presence of high CAS concentrations. These findings were also confirmed by SEM analysis and were associated with the metabolic activity obtained by biofilm susceptibility testing. Importantly, these results suggest that the presence of atypical, enlarged, conical cells could be associated with PG and with tolerant cells in Candida species biofilm populations. The clinical implications of these findings are still unknown.
Assuntos
Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Candida/efeitos dos fármacos , Candida/crescimento & desenvolvimento , Equinocandinas/farmacologia , Candida/ultraestrutura , Caspofungina , Humanos , Lipopeptídeos , Microscopia Eletrônica de VarreduraRESUMO
BACKGROUND: Although Candida species are commensal microorganisms, they can cause many invasive fungal infections. In addition, antifungal resistance can contribute to failure of treatment.The purpose of this study was to evaluate the antifungal activity of inhibitors of Delta24(25)-sterol methyltransferase (24-SMTI), 20-piperidin-2-yl-5alpha-pregnan-3beta-20(R)-diol (AZA), and 24(R,S),25-epiminolanosterol (EIL), against clinical isolates of Candida spp., analysing the ultrastructural changes. RESULTS: AZA and EIL were found to be potent growth inhibitors of Candida spp. isolates. The median MIC50 was 0.5 microg.ml-1 for AZA and 2 microg.ml-1 for EIL, and the MIC90 was 2 microg.ml-1 for both compounds. All strains used in this study were susceptible to amphotericin B; however, some isolates were fluconazole- and itraconazole-resistant. Most of the azole-resistant isolates were Candida non-albicans (CNA) species, but several of them, such as C. guilliermondii, C. zeylanoides, and C. lipolytica, were susceptible to 24-SMTI, indicating a lack of cross-resistance. Reference strain C. krusei (ATCC 6258, FLC-resistant) was consistently susceptible to AZA, although not to EIL. The fungicidal activity of 24-SMTI was particularly high against CNA isolates. Treatment with sub-inhibitory concentrations of AZA and EIL induced several ultrastructural alterations, including changes in the cell-wall shape and thickness, a pronounced disconnection between the cell wall and cytoplasm with an electron-lucent zone between them, mitochondrial swelling, and the presence of electron-dense vacuoles. Fluorescence microscopy analyses indicated an accumulation of lipid bodies and alterations in the cell cycle of the yeasts. The selectivity of 24-SMTI for fungal cells versus mammalian cells was assessed by the sulforhodamine B viability assay. CONCLUSION: Taken together, these results suggest that inhibition of 24-SMT may be a novel approach to control Candida spp. infections, including those caused by azole-resistant strains.