Flexible camphor diamond-like carbon coating on polyurethane to prevent Candida albicans biofilm growth.

Camphor was incorporated in diamond-like carbon (DLC) films to prevent the Candida albicans yeasts fouling on polyurethane substrates, which is a material commonly used for catheter manufacturing. The camphor:DLC and DLC film for this investigation was produced by plasma enhanced chemical vapor deposition (PECVD), using an apparatus based on the flash evaporation of organic liquid (hexane) containing diluted camphor for camphor:DLC and hexane/methane, mixture for DLC films. The film was deposited at a low temperature of less than 25°C. We obtained very adherent camphor:DLC and DLC films that accompanied the substrate flexibility without delamination. The adherence of camphor:DLC and DLC films on polyurethane segments were evaluated by scratching test and bending polyurethane segments at 180°. The polyurethane samples, with and without camphor:DLC and DLC films were characterized by Raman spectroscopy, scanning electron microscopy, atomic force microscopy, and optical profilometry. Candida albicans biofilm formation on polyurethane, with and without camphor:DLC and DLC, was assessed. The camphor:DLC and DLC films reduced the biofilm growth by 99.0% and 91.0% of Candida albicans, respectively, compared to bare polyurethane. These results open the doors to studies of functionalized DLC coatings with biofilm inhibition properties used in the production of catheters or other biomedical applications.

Application of post-discharge region of atmospheric pressure argon and air plasma jet in the contamination control of Candida albicans biofilms

Introduction:Candida species are responsible for about 80% of hospital fungal infections. Non-thermal plasmas operated at atmospheric pressure are increasingly used as an alternative to existing antimicrobial strategy. This work investigates the action of post-discharge region of a non-thermal atmospheric plasma jet, generated by a gliding arc reactor, on biofilms of standard strain of Candida albicans grown on polyurethane substrate. Methods Samples were divided into three groups: (i) non-treated; (ii) treated with argon plasma, and (iii) treated with argon plus air plasma. Subsequently to plasma treatment, counting of colony-forming units (CFU/ml) and cell viability tests were performed. In addition, the surface morphology of the samples was evaluated by scanning electron microscopy (SEM) and optical profilometry (OP). Results Reduction in CFU/ml of 85% and 88.1% were observed in groups ii and iii, respectively. Cell viability after treatment also showed reduction of 33% in group ii and 8% in group iii, in comparison with group i (100%). The SEM images allow observation of the effect of plasma chemistry on biofilm structure, and OP images showed a reduction of its surface roughness, which suggests a possible loss of biofilm mass. Conclusion The treatment in post-discharge region and the chemistries of plasma jet tested in this work were effective in controlling Candida albicans biofilm contamination. Finally, it was evidenced that argon plus air plasma was the most efficient to reduce cell viability.

Antifungal activity of extracts and isolated compounds from Buchenavia tomentosa on Candida albicans and non-albicans.

AIM: This study aimed to evaluate the antifungal activity of Buchenavia tomentosa extract and bioactive compounds on six Candida species. MATERIALS & METHODS: The antimicrobial activity of extract was evaluated using standard strains and clinical isolates. Cytotoxicity was tested in order to evaluate cell damage caused by the extract. Extract was chemically characterized and the antifungal activity of its compounds was evaluated. RESULTS: Extract showed antifungal activity on Candida species. Candida non-albicans were more susceptible than Candida albicans. Low cytotoxicity for extract was observed. The isolated compounds presented antifungal activity at least against one Candida spp. and all compounds presented antifungal effect on Candida glabrata. CONCLUSION: Extracts from Buchenavia tomentosa showed promising antifungal activity on Candida species with low cytotoxicity. Gallic acid, corilagin and ellagic acid showed promising inhibitory activity on Candida glabrata.

Study of photodynamic therapy in the control of isolated microorganisms from infected wounds--an in vitro study.

The effective treatment of infected wounds continues to be a serious challenge, mainly due to the rise of antibiotic-resistant bacteria. Photodynamic therapy (PDT) refers to the topical or systemic administration of a non-toxic, photosensitizing agent (PS), followed by irradiation with visible light of a suitable wavelength. The possibility of applying the PDT locally is what makes it so favorable to the treatment of infected wounds. The goal of this study was to evaluate the action of the PDT in the inactivation in vitro of microorganisms coming from infected wounds, using methylene blue (MB) and photodithazine (PDZ) as the PS and comparing the efficacy of these two compounds for PDT on bacteria. For the application of PDT, isolated microorganisms identified from material collected from wounds were suspended in a saline solution containing 10(6) viable cells/ml. Each isolated microorganism was submitted to PDT with MB and with PDZ in accordance with the following treatment groups: N/T--no treatment; T1--PDT with PDZ; T2--PDT with MB; T3--irradiation without PS; T4--treatment with PDZ without light; and T5--treatment with MB without light. As a light source, an LED-based device was used (Biopdi/Irrad-Lead 660), composed of 54 LEDs, each with 70 mW of power in the 660 nm region of the electromagnetic spectrum. Each tray of 96 wells was irradiated with an intensity of 25 mW/cm(2) and a dose of light of 50 J/cm(3) for 33 min. All the tests were made in duplicate. It was then concluded that the PDT with PDZ was capable of inhibiting the growth of gram-positive bacteria samples, however it did not have the same effect on gram-negative bacteria, which showed growth greater than 100,000 CFU; the PDT with MB showed an effectiveness on gram-positive as well as gram-negative bacteria, for it was able to inhibit bacterial growth in both cases.

Photodynamic inactivation of yeast and bacteria by extracts of Alternanthera brasiliana.

This study was undertaken to evaluate the effect of Alternathera brasiliana (Amaranthaceae) extracts as photosensitizing agents in photodynamic antimicrobial therapies (PACT) against Staphylococcus aureus, Staphylococcus epidermidis and Candida dubliniensis. The crude hexane and ethanol extracts were obtained from A. brasiliana whole plant and showed absortion from 650 to 700 nm. Also, singlet molecular oxygen (1O2) production (type II photosensitization reaction) was examined, and the results show that 1,3-diphenylisobenzofuran photodegradation was greatly enhanced in the presence of the A. brasiliana extracts. One plate in each assay was irradiated while the other was not irradiated, the number of colony-forming units per milliliter (CFU/mL) was obtained, and data analyzed by the Tukey test. The chemical composition of the extracts was determined by chromatographic and spectrometric techniques; steroids, triterpenes, and flavonoids were identified. Laser irradiation alone at 685 nm using diode laser, output power of 35 mW, and energy of 28 J/cm2, or non-irradiated crude extracts in sub-inhibitory concentration did not reduce the number of CFU/mL significantly, whereas irradiated hexane and ethanol extracts, in sub-inhibitory concentrations, inhibited the growth of these microorganisms. The photoactivation of hexane and ethanol extracts of A. brasiliana, in sub-inhibitory concentrations, using red laser radiation at 685 nm had an antimicrobial effect.

Candidemia by species of the Candida parapsilosis complex in children's hospital: prevalence, biofilm production and antifungal susceptibility.

Opportunistic infections are an increasingly common problem in hospitals, and the yeast Candida parapsilosis has emerged as an important nosocomial pathogen. The aims of this study were to determine and compare (i) the prevalence rate among C. parapsilosis complex organisms isolated from blood in a public children's hospital in São Paulo state, (ii) the ability of the complex C. parapsilosis species identified to produce biofilm and (iii) the antifungal susceptibility profiles. Forty-nine (49) specimens of isolated blood yeast were analyzed, previously identified as C. parapsilosis by conventional methods. After the molecular analysis, the isolates were characterized as C. parapsilosis sensu stricto (83.7 %), C. orthopsilosis (10.2 %) and C. metapsilosis (6.1 %). All species were able to form biofilm. The species with the highest biofilm production was C. parapsilosis sensu stricto, followed by C. orthopsilosis and further by C. metapsilosis. All of the strains have demonstrated similar susceptibility to fluconazole, caspofungin, voriconazole, cetoconazole and 5-flucytosine. Only one strain of C. parapsilosis was resistant to amphotericin B. Regarding itraconazole, 66.6 and 43.9 % isolates of C. metapsilosis and C. parapsilosis, respectively, have demonstrated to be susceptible dose-dependent, with one isolate of the latter species resistant to the drug. Candida parapsilosis sensu stricto has demonstrated to be the less susceptible, mainly to amphotericin B, caspofungin and "azoles" such as fluconazole. Therefore, C. metapsilosis and C. orthopsilosis are still involved in a restricted number of infections, but these data have become essential for there are very few studies of these species in Latin America.

Utilização da microespectroscopia infravermelha (FT-IR) para teste de algoritmos estatísticos na diferenciação dos micro-organismos Candida albicans, Candida dubliniensis e Candida parapsilosis / Using FT-IR microspectroscopy for testing statistical algorithms for differentiation of Candida albicans, Candida dubliniensis and Candida parapsilosis

As espécies do gênero Candida são causadoras de diversas infecções fúngicas e, nos últimos anos, tem sido desenvolvidas novas tecnologias para auxiliar nos diagnósticos microbiológicos. Dentre as técnicas está a espectroscopia infravermelha junto com a análise estatística multivariada. O objetivo deste trabalho é comparar dois métodos: estatístico (análise multivariada) e não-estatístico (ajuste de curva), utilizando os espectros infravermelhos de Candida albicans, Candida dubliniensis e Candida parapsilosis para testar o potencial do uso de Análise Estatística Multivariada para discriminação de espectros de micro-organismos. Para isso foram obtidos, utilizando o Spectrum Spotlight 400 da PerkinElmer, 54 espectros infravermelhos, sendo 18 de cada espécie, na faixa de 4000 a 1000 cm-1, com resolução de 4 cm-1, no modo de transmissão, a 20 ºC. A análise dos espectros foi realizada através de três métodos: (1) inspeção visual direta dos espectros; (2) análise estatística multivariada; (3) ajuste de curva para a determinação de estruturas secundárias de proteínas. Na região de 1200 a 1000 cm-1, os espectros apresentam diferenças que podem ser percebidas numa inspeção visual direta. Uma banda próxima de 1070 cm-1 e outra próxima de 1045 cm-1 apresentam intensidades relativas diferentes para os três espectros. Por outro lado, as bandas da amida I, na região de 1710 a 1590 cm-1, apresentam aspectos visuais semelhantes com máximo em 1651 cm-1 para os espectros dos três micro-organismos. Esse fato torna possível submeter a análise estatística multivariada a um teste de sua capacidade de diferenciar três espectros de Candida. A análise estatística multivariada foi aplicada aos 54 espectros para investigar as regiões de 4000 a 1000 cm-1 com exceção da região de 2600 a 2300 cm-1 e de 1710 a 1590 cm-1 que corresponde a das bandas da amida I. A técnica selecionada foi a análise por componentes principais (PCA, Principal Componente Analysis), utilizando os primeiros quatro componentes principais, em conjunto com a técnica hierárquica de análise de agrupamento (HCA, Hierarchical Clustering Analysis) segundo o método de Ward. Foi utilizado para esta análise o software MINITAB 15 e o resultado mostra uma clara discriminação dos espectros dos três micro-organismos nas duas regiões consideradas. Adicionalmente foi obtido o espectro médio de cada micro-organismo nas bandas da amida I na região de 1710 a 1590 cm-1. Os três espectros médios assim obtidos foram analisados pelo método de ajuste de curva que não é estatístico para determinar as estruturas secundárias de proteínas. Para esta análise o software ORIGIN 7.5 foi utilizado e os resultados obtidos mostram estruturas conformacionais diferentes nos três micro-organismos. Esses resultados confirmam a discriminação obtida através da análise estatística multivariada e visual. Pode-se concluir que as análises estatísticas multivariadas baseadas em análise por componentes principais e análise de agrupamento com uso do algoritmo Ward é potencialmente útil para discriminar micro-organismos através de seus espectros infravermelhos. Além disso, as análises mostram que as bandas da amida I dos espectros infravermelhos de Candida albicans, Candida dubliniensis e Candida parapsilosis fornecem um conjunto de dados cuja estrutura de agrupamento é conhecida e que pode ser útil para testar e validar algoritmos estatísticos de análise de agrupamento.

Films of Candida albicans, Candida dubliniensis and Candida parapsilosis were prepared and the infrared spectra of these films were obtained in the region 4000 to 1000 cm-1, with resolution of 4 cm-1, in the transmission mode, at 20 ºC. Fifty four spectra were obtained, 18 of each microorganism, with the PerkinElmer Spotlight 400 FT-IR, which has a microscope attached to a FT-IR spectrophotometer. The spectra were analyzed through three methods: (1) mere visual inspection; (2) multivariate statistical analysis; (3) curve-fitting for determining secondary structures of proteins. In the region 1200 to 1000 cm-1, the spectral bands show differences that can be seen by a mere visual inspection. On the other hand, the amide I bands, in the region 1710 to 1590 cm-1, have the same visual aspect for the three microorganisms. Multivariate statistical analysis was applied to analyze these amide I bands of all the 54 spectra. Principal component analysis (PCA) and techniques of hierarchical cluster analysis (HCA, Hierarchical Clustering Analysis) according to Ward's method were applied using the software MINITAB 15. The results show a clear discrimination of the three microorganisms. The average spectrum of each microorganism was obtained in the amide I band. Each average spectrum was analyzed by curve-fitting for the determination of secondary structures of proteins. The software used was the ORIGIN 7.5 and the results confirm the discrimination obtained through multivariate statistical analysis. This result shows that multivariate statistical analysis can be useful to discriminate infrared spectra of different microorganisms. Furthermore, this work shows that the amide I bands of the infrared spectra of Candida albicans, Candida dubliniensis, and Candida parapsilosis provide a set of data of known group structure that can be useful to test statistical algorithms of cluster analysis.

Atividade antifúngica do óleo essencial de melaleuca alternifolia sobre leveduras isoladas de candidíase bucal de gestantes HIV positivas / Antifungal activity of melaleuca alternifolia essential oil on yeasts isolated from HIV positive pregnant women with oral candidiasis

A candidíase bucal é a infecção fúngica mais comum em portadores de HIV e, com episódios recorrentes em pacientes com Aids. Com o objetivo de pesquisar novos e eficazes agentes antifúngicos contra cepas resistentes, foi investigada a atividade antifúngica do óleo essencial de Melaleuca alternifolia, em diferentes concentrações, em leveduras isoladas de candidíase oral. O experimento foi realizado por meio da técnica de difusão em ágar. Foram avaliadas cepas padrão de Candida albicans ATCC 10231, Candida tropicalis ATCC 157, Candida glabrata ATCC 30070, Candida krusei ATCC 6258 e Candida dubliniensis ATCC 778157e os isolados da cavidade bucal de gestantes HIV positivas, sendo sete C. albicans, um C. tropicalis, um C.glabrata e um C. krusei. O óleo essencial foi analisado nas quantidades de 20 e 50 μL, nas concentrações de 10 a 100%, variando de 10 em 10%. Todas as cepas analisadas foram suscetíveis ao óleo essencial de M.alternifolia nas concentrações de 70% e 50 %, respectivamente, nos volumes de 20 μL e 50 μL. O potencial antifúngico do óleo essencial de M. alternifolia desperta interesse para o desenvolvimento de novos fármacos.

Oral candidiasis is a fungal infection mostly common in people infected with HIV and the recurrent episodes occur in patients with AIDS. Aiming at searching, a new and efficacious drug against resistant strains, the antifungal activity of essential oil from Melaleuca alternifolia at different concentrations was assessed on yeasts isolated from oral candidiasis. The experiment was performed by using agar diffusiontechnique; and the antifungal effect was evaluated on the standard strains of Candida albicans ATCC 10231, Candida tropicalis ATCC 157, Candida glabrata ATCC 30070, Candida krusei ATCC 6258 and Candida dubliniensis ATCC 778 157, and on the isolated from the oral cavity of HIV positive pregnant women, being seven C. albicans, one C. tropicalis, one C glabrata and one C. krusei. The essential oil was analyzedin quantities of 20 and 50 μL at 10-100% concentrations, ranging from 10 to 10%. All of the tested strainswere susceptible to the M. alternifolia essential oil at concentrations of 70% and 50% in volumes of 20 μL and 50 μL, respectively. The antifungal activity of essential oil from M. alternifolia holds one´s attention in manufacturing new and effective drugs.

Extracts from Alternanthera maritima as natural photosensitizers in photodynamic antimicrobial chemotherapy (PACT).

This study investigated the effect of photodynamic antimicrobial chemotherapy (PACT) using extracts from Alternanthera maritima on the viability of Candida dubliniensis. Human infections constitute a great health problem. Several antifungal drugs are currently available, but their uses are limited by a number of factors, such as low potency, poor solubility, microbial resistance, and drug toxicity. Therefore, the search for new and more effective antimicrobial agents and the development of alternative therapies, such as PACT, are necessary. Crude hexane and ethanol extracts of A. maritima were produced. The prepared extracts presented absorption at 650-700 nm. For bioassays, 50 microL of culture medium, 50 microL of extract (25 mg/mL) or control, and 5 microL of a suspension of the microorganism to be tested (C. dubliniensis ATCC 778157 or ATCC 777, 10(7)CFU/mL) were placed in a sterile 96-well microtiter plate (well cross section=0.38 cm(2)). The contents of each well were irradiated with a 685-nm diode laser with an output power of 35 mW, which was distributed through the well cross section yielding an energy dosage of 28 J/cm(2). In each assay (n=6), one plate was subjected to irradiation, and one was not. For each active sample, the number of colony-forming units per milliliter (CFU/mL) was obtained, and data were analyzed by the Tukey test. The chemical compositions of the extracts were determined by chromatographic and spectroscopic techniques. The results suggest inhibition of the growth of C. dubliniensis when irradiated with a diode laser in the presence of hexane and ethanol extracts from A. maritima as photosensitizers. Laser irradiation alone or crude extracts at 25mg/mL did not significantly reduce the number of CFU/mL. Steroids, triterpenes, and flavonoids were identified in the analyzed extracts. In conclusion, the photoactivation of crude hexane and ethanol extracts of A. maritima by red laser radiation at 685 nm promoted an antimicrobial effect, showing that these natural products can be used as photosensitizers in PACT.

Atividade antifúngica do óleo essencial de Cymbopogon citratus frente a cepas de Candida albicans e Candida tropicalis isoladas de infecções nosocomiais / Antifungal activity of Cymbopogon citratus essential oil on Candida albicans and Candida tropicalis strains isolated from nosocomial infections

O óleo essencial de Cymbopogon citratus tem sido alvo de vários estudos em função do potencial antimicrobiano. Neste estudo, a atividade desse componente foi investigada em cepas do gênero Candida isoladas de infecções hospitalares. Para a condução do estudo, foram analisadas 24 isolados de Candida albicans e 15 isolados de Candida tropicalis, originados de pacientes com suspeitas de infecção hospitalar e uma cepa padrão de C. albicans ATCC 10231, por meio da técnica de difusão em ágar. O óleo essencial de C. citratus apresentou atividade antifúngica em 100% dos isolados a partir da concentração de 25% (v/v), o que indica sua ação positiva sobre as cepas hospitalares. Sugere-se a realização de estudos farmacológicos e toxicológicos desse componente para avaliar a possível aplicação clínica.

Atividade antifúngica do citral em leveduras do gênero Candida isoladas de pacientes hospitalizados / Citral antifungal activity against Candida genus yeasts isolated from hospitalized patients

O citral, componente do óleo essencial de Cymbopogon citratus, tem sido pesquisado para verificar sua ação antimicrobiana em bactérias e fungos. Com o objetivo de avaliar a atividade antifúngica do citral contra leveduras do gênero Candida, no presente trabalho foram avaliadas 32 amostras de Candida albicans, 25 de C. tropicalis, 20 de C. parapsilosis e 5 de C. glabrata, coletadas de pacientes hospitalizados. O citral foi testado nas concentrações de 10%, 15%, 25%, 35%, 50% e 60% (v/v), utilizando a técnica de difusão em ágar Sabouraud. A atividade antifúngica do citral foi constatada em todas as leveduras selecionadas nas concentrações ≥25%. Mediante os resultados obtidos, sugere-se a realização de novas pesquisas sobre citral frente às demais espécies de fungos patogênicos para conhecer as características toxicológicas e farmacológicas para que esse componente possa futuramente ser utilizado como um importante princípio ativo na produção de novos agentes antifúngicos.