Disseminated Amphotericin-Resistant Fusariosis in Acute Leukemia Patients: Report of Two Cases

Disseminated fusariosis has emerged as a significant, usually fatal infection in immunocompromised hosts despite antifungal treatment. We describe here two patients with acute leukemia who developed disseminated amphotericin-resistant fusariosis, and review of six studies of cases series in the literature. Two Fusarium solani strains were isolated from blood and skin cultures of one patient, and one strain from the blood culture of the second patient. Both patients died despite antifungal treatment. Strains were identified by sequencing of ITS1 and ITS4 regions. Random amplified polymorphic DNA analysis of the three F. solani isolates showed a low degree of similarity. Screening for Fusarium spp. contaminants within our facility was negative. Using the CLSI M-38-A2 broth dilution method and E tests®, we found that the MICs were low for voriconazole (0. 12 and 0. 5 mg/L, respectively), unexpectedly high for amphotericin B (≥8 and ≥32 μg/mL, respectively) and itraconazole (≥16 mg/ml). Patients with leukemia or persistent neutropenia should be assessed for disseminated fungal infections, including biopsy and skin cultures. Antifungal susceptibility tests are important due to the possibility of the strains being amphotericin resistant. Treatments must be aggressive, with high doses of antifungals or combined therapy. © 2012 Springer Science+Business Media Dordrecht.

Photodynamic inactivation of planktonic cultures and biofilms of Candida albicans mediated by aluminum-chloride-phthalocyanine entrapped in nanoemulsions

New drug delivery systems, such as nanoemulsions (NE), have been developed to allow the use of hydrophobic drugs on the antimicrobial photodynamic therapy. This study evaluated the photodynamic potential of aluminum-chloride- phthalocyanine (ClAlPc) entrapped in cationic and anionic NE to inactivate Candida albicans planktonic cultures and biofilm compared with free ClAlPc. Fungal suspensions were treated with different delivery systems containing ClAlPc and light emitting diode. For planktonic suspensions, colonies were counted and cell metabolism was evaluated by XTT assay. Flow cytometry evaluated cell membrane damage. For biofilms, the metabolic activity was evaluated by XTT and ClAlPc distribution through biofilms was analyzed by confocal laser scanning microscopy (CLSM). Fungal viability was dependent on the delivery system, superficial charge and light dose. Free ClAlPc caused photokilling of the yeast when combined with 100 J cm-2. Cationic NE-ClAlPc reduced significantly both colony counts and cell metabolism (P < 0.05). In addition, cationic NE-ClAlPc and free ClAlPc caused significant damage to the cell membrane (P < 0.05). For the biofilms, cationic NE-ClAlPc reduced cell metabolism by 70%. Anionic NE-ClAlPc did not present antifungal activity. CLSM showed different accumulation on biofilms between the delivery systems. Although NE system showed a lower activity for planktonic culture, cationic NE-ClAlPc showed better results for Candida biofilms. Candida albicans biofilm overview after 30 min of contact with free ClAlPc. This study presents the photodynamic potential of aluminum-chloride-phthalocyanine (ClAlPc) entrapped in cationic and anionic nanoemulsions (NE) to inactivate C. albicans planktonic cultures and biofilm comparing with free ClAlPc. The photodynamic effect was dependent on the delivery system, superficial charge and light dose. Cationic NE-ClAlPc and free ClAlPc caused significant reduction in colony counts, cell metabolism and damage to the cell membrane (P < 0.05). However, only the free ClAlPc was able to cause photokilling of the yeast. The anionic NE-ClAlPc did not present antifungal activity. Although NE system showed a lower activity for planktonic culture, cationic NE-ClAlPc showed better results for Candida biofilms. © 2012 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2012 The American Society of Photobiology.

In vitro assessment of the cytotoxic, apoptotic, and mutagenic potentials of Isatin

Isatin (1H-indole-2,3-dione) is a chemical found in various medicinal plant species and responsible for a broad spectrum of pharmacological and biological properties that may be beneficial to human health, as an anticonvulsant, antibacterial, antifungal, antiviral, and anticancer agent. The aim of the present study was to determine in vitro the cytotoxic, mutagenic, and apoptotic effects of isatin on CHO-K1 and HeLa cells using the MTT viability assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide), micronucleus (MN) test, apoptosis index, and nuclear division index (NDI). The 5 isatin concentrations evaluated in the mutagenicity and apoptosis tests were 0.5, 1, 5, 10, and 50 μM, selected through a preliminary MTT assay. Positive (doxorubicin, DXR) and negative (phosphate buffered saline, PBS) control groups were also included in the analysis. Isatin did not exert a mutagenic effect on CHO-K1 after 3 and 24 h of treatment or on HeLa cells after 24 h. However, 10 and 50 μM concentrations inhibited cell proliferation and promoted apoptosis in both CHO-K1 and HeLa cells. Data indicate that the cytotoxic, apoptotic, and antiproliferative effects of isatin were concentration independent and cell line independent. The authors thank Profa Dra Eiko Nakagawa Itano for the use the spectrophotometer and the Conselho Nacional para o Desenvolvimento Científico e Tecnológico for master's scholarships to P. M. Cândido-Bacani and grants to T. R. Calvo, W. Vilegas, E. A. Varanda and I. M. S. Cólus. The Conselho Nacional para o Desenvolvimento Científico e Tecnológico provided funding for this study. © 2013 Taylor & Francis Group, LLC.

Hybrid nanocomposites containing carboxymethylcellulose and silver nanoparticles

Silver nanoparticles have high temperature stability and low volatility, and at the nanoscale are known to be an effective antifungal and antimicrobial agent. The present investigation involves the synthesis of silver nanoparticle/carboxymethylcellulose nanocomposites. The nanoparticles synthesised in this study had sizes in the range of 100 and 40 nm. The nanocomposites formed by a combination of metallic nanoparticles and carboxymethylcellulose were characterised by contact angle measurements, solubility tests, thermal and mechanical analyses, and morphological images. Improvements in the hydrophobic properties were observed with inclusion of the nanoparticles in the nanocomposites, with the best results occurring after the addition of 40 nm nanoparticles in a carboxymethylcellulose matrix. The silver nanoparticles tend to occupy the empty spaces in the pores of the carboxymethylcellulose matrix, inducing the collapse of these pores and thereby improving the tensile and barrier properties of the film. Copyright © 2013 American Scientific Publishers All rights reserved.

Estudo duplo cego com Isoconazol e Terbinafina tópicos no tratamento de um paciente com Tinea nigra plantaris bilateral e sugestões para novos diagnósticos diferenciais

The authors report a case of bilateral Tinea nigra plantaris treated through a double-blind study with the topical antifungal agents Isoconazole and Terbinafine. The objective of the study was to clinically compare the efficacy of these two topical antifungal agents on days 10, 20 and 30 of the treatment. No significant clinical differences were found, as all the plantar lesions regressed completely by the end of the treatment. Our conclusion was that in the case reported, the topical antifungal agents Isoconazole and Terbinafine demonstrated identical efficacy as a clinical cure. We also suggest the inclusion of injuries caused by arthropods of the Diplopoda Class in the differential diagnosis of Tinea nigra plantaris, due to the persistent acral hyperpigmentation.

Microplate alamarblue assay for paracoccidioides susceptibility testing

CLSI method M27-A3 is not available for use with dimorphic fungi, such as those of the Paracoccidioides genus. In this study, we developed a microdilution method and added the alamarBlue reagent to test the responses of Paracoccidioides brasiliensis and Paracoccidioides lutzii against amphotericin B and itraconazole antifungals. The test proved to be sensitive, practical, and inexpensive and can be used to monitor the activity of low-growth microorganisms and their response to various drugs. © 2013, American Society for Microbiology.

Anti-Candida properties of urauchimycins from actinobacteria associated with Trachymyrmex ants

After decades of intensive searching for antimicrobial compounds derived from actinobacteria, the frequency of isolation of new molecules has decreased. To cope with this concern, studies have focused on the exploitation of actinobacteria from unexplored environments and actinobacteria symbionts of plants and animals. In this study, twenty-four actinobacteria strains isolated from workers of Trachymyrmex ants were evaluated for antifungal activity towards a variety of Candida species. Results revealed that seven strains inhibited the tested Candida species. Streptomyces sp. TD025 presented potent and broad spectrum of inhibition of Candida and was selected for the isolation of bioactive molecules. From liquid shake culture of this bacterium, we isolated the rare antimycin urauchimycins A and B. For the first time, these molecules were evaluated for antifungal activity against medically important Candida species. Both antimycins showed antifungal activity, especially urauchimycin B. This compound inhibited the growth of all Candida species tested, with minimum inhibitory concentration values equivalent to the antifungal nystatin. Our results concur with the predictions that the attine ant-microbe symbiosis may be a source of bioactive metabolites for biotechnology and medical applications. © 2013 Thais D. Mendes et al.

Ricinus communis treatment of denture stomatitis in institutionalised elderly

This study compared the effectiveness of Ricinus communis (RC) with Nystatin (NYS) and Miconazole (MIC) in the treatment of institutionalised elderly with denture stomatitis (DS). They (n = 30) were randomly distributed into three groups: MIC, NYS or RC. Clinical and mycological evaluations were performed prior to the use of the antifungal (baseline) and repeated after 15 and 30 days of treatment. The sample was clinically examined for oral mucosal conditions. Standard photographs were taken of the palate, and the oral candidiasis was classified (Newton's criteria). Mycological investigation was performed by swabbing the palatal mucosa, and Candida spp. were quantified by counting the number of colony-forming units (cfu mL-1). The clinical and mycological data were analysed, respectively by Wilcoxon and Student's t-test (α = 0·05). Significant improvement in the clinical appearance of DS in the MIC and RC groups was observed between the 1st and 3rd collections (MIC - P = 0·018; RC - P = 0·011) as well as between the 2nd and 3rd collections (MIC - P = 0·018; RC - P = 0·011). Neither groups showed a statistically significant reduction in cfu mL-1 at any time. Although none of the treatments decreased the cfu mL-1, it was concluded that Ricinus communis can improve the clinical condition of denture stomatitis in institutionalised elderly patients, showing similar results to Miconazole. © 2013 Blackwell Publishing Ltd.

Fungos endofíticos: Uma fonte inexplorada e sustentável de novos e bioativos produtos naturais

Endophytic fungi are a rich source of new and biologically active natural products. They colonize a relatively unexplored ecological habitat and their secondary metabolism is particularly active, presumably due to metabolic interactions with their hosts. In the course of our continuing investigations for new and bioactive compounds from endophytic fungi from brazilian flora Alibertia macrophylla, Caseria sylvestris, Ocotea corymbosa, Cassia spectabilis, Piper aduncum, Cryptocaria mandioccana, Xylopia aromatica and Palicourea marcgravii were investigated. Forty two natural products were isolated and their structures were established on the basis of comprehensive spectral analysis, mainly using 1D and 2D NMR experiments. The compounds were tested in their antifungal, antioxidant, anticholinesterasic and anticancer activities.

Identification of Candida species in the clinical laboratory: A review of conventional, commercial, and molecular techniques

In healthy individuals, Candida species are considered commensal yeasts of the oral cavity. However, these microorganisms can also act as opportunist pathogens, particularly the so-called non-albicans Candida species that are increasingly recognized as important agents of human infection. Several surveys have documented increased rates of C. glabrata, C. tropicalis, C. guilliermondii, C. dubliniensis, C. parapsilosis, and C. krusei in local and systemic fungal infections. Some of these species are resistant to antifungal agents. Consequently, rapid and correct identification of species can play an important role in the management of candidiasis. Conventional methods for identification of Candida species are based on morphological and physiological attributes. However, accurate identification of all isolates from clinical samples is often complex and time-consuming. Hence, several manual and automated rapid commercial systems for identifying these organisms have been developed, some of which may have significant sensitivity issues. To overcome these limitations, newer molecular typing techniques have been developed that allow accurate and rapid identification of Candida species. This study reviewed the current state of identification methods for yeasts, particularly Candida species. © 2013 John Wiley & Sons A/S.

Evaluation of a propolis water extract using a reliable RP-HPLC methodology and in vitro and in vivo efficacy and safety characterisation

Since the beginning of propolis research, several groups have studied its antibacterial, antifungal, and antiviral properties. However, most of these studies have only employed propolis ethanolic extract (PEE) leading to little knowledge about the biological activities of propolis water extract (PWE). Based on this, in a previous study, we demonstrated the anti-inflammatory and immunomodulatory activities of PWE. In order to better understand the equilibrium between effectiveness and toxicity, which is essential for a new medicine, the characteristics of PWE were analyzed. We developed and validated an RP-HPLC method to chemically characterize PWE and PEE and evaluated the in vitro antioxidant/antimicrobial activity for both extracts and the safety of PWE via determining genotoxic potential using in vitro and in vivo mammalian micronucleus assays. We have concluded that the proposed analytical methodology was reliable, and both extracts showed similar chemical composition. The extracts presented antioxidant and antimicrobial effects, while PWE demonstrated higher antioxidant activity and more efficacious for the most of the microorganisms tested than PEE. Finally, PWE was shown to be safe using micronucleus assays. © 2013 Bruno Alves Rocha et al.

Methods for obtaining reliable and reproducible results in studies of Candida biofilms formed in vitro

Biofilm formation is one of the most important attributes for virulence in Candida species and contributes to increased resistance to antifungal drugs and host immune mechanisms. These features have led to the development of several methodologies to reproduce a sessile community in vitro that can be used to study the development of a biofilm, its interaction with other microorganisms and the environment, and its susceptibility to available antifungal agents and also to search for new therapy strategies. The purpose of this review is to describe the most commonly used methods to study Candida biofilms in vitro, to discuss the benefits and limitations of the different methods to induce biofilm formation, and to analyse the architecture, viability and growth kinetics of Candida biofilms. © 2013 Blackwell Verlag GmbH.

Dimerization of aurein 1.2: Effects in structure, antimicrobial activity and aggregation of Cândida albicans cells

Antimicrobial peptides (AMPs) are a promising solution to face the antibiotic-resistant problem because they display little or no resistance effects. Dimeric analogues of select AMPs have shown pharmacotechnical advantages, making these molecules promising candidates for the development of novel antibiotic agents. Here, we evaluate the effects of dimerization on the structure and biological activity of the AMP aurein 1.2 (AU). AU and the C- and N-terminal dimers, (AU)2K and E(AU)2, respectively, were synthesized by solid-phase peptide synthesis. Circular dichroism spectra indicated that E(AU)2 has a coiled coil structure in water while (AU)2K has an α-helix structure. In contrast, AU displayed typical spectra for disordered structures. In LPC micelles, all peptides acquired a high amount of α-helix structure. Hemolytic and vesicle permeabilization assays showed that AU has a concentration dependence activity, while this effect was less pronounced for dimeric versions, suggesting that dimerization may change the mechanism of action of AU. Notably, the antimicrobial activity against bacteria and yeast decreased with dimerization. However, dimeric peptides promoted the aggregation of C. albicans. The ability to aggregate yeast cells makes dimeric versions of AU attractive candidates to inhibit the adhesion of C. albicans to biological targets and medical devices, preventing disease caused by this fungus. © 2013 Springer-Verlag Wien.

Photodynamic inactivation of biofilm: Taking a lightly colored approach to stubborn infection

Microbial biofilms are responsible for a variety of microbial infections in different parts of the body, such as urinary tract infections, catheter infections, middle-ear infections, gingivitis, caries, periodontitis, orthopedic implants, and so on. The microbial biofilm cells have properties and gene expression patterns distinct from planktonic cells, including phenotypic variations in enzymic activity, cell wall composition and surface structure, which increase the resistance to antibiotics and other antimicrobial treatments. There is consequently an urgent need for new approaches to attack biofilm-associated microorganisms, and antimicrobial photodynamic therapy (aPDT) may be a promising candidate. aPDT involves the combination of a nontoxic dye and low-intensity visible light which, in the presence of oxygen, produces cytotoxic reactive oxygen species. It has been demonstrated that many biofilms are susceptible to aPDT, particularly in dental disease. This review will focus on aspects of aPDT that are designed to increase efficiency against biofilms modalities to enhance penetration of photosensitizer into biofilm, and a combination of aPDT with biofilm-disrupting agents. © 2013 Informa UK Ltd.

Anti-Candida targets and cytotoxicity of casuarinin isolated from Plinia cauliflora leaves in a bioactivity-guided study

In addition to the bio-guided investigation of the antifungal activity of Plinia cauliflora leaves against different Candida species, the major aim of the present study was the search for targets on the fungal cell. The most active antifungal fraction was purified by chromatography and characterized by NMR and mass spectrometry. The antifungal activity was evaluated against five Candida strains according to referenced guidelines. Cytotoxicity against fibroblast cells was determined. The likely targets of Candida albicans cells were assessed through interactions with ergosterol and cell wall composition, porosity and architecture. The chemical major component within the most active antifungal fraction of P. cauliflora leaves identified was the hydrolysable tannin casuarinin. The cytotoxic concentration was higher than the antifungal one. The first indication of plant target on cellular integrity was suggested by the antifungal activity ameliorated when using an osmotic support. The most important target for the tannin fraction studied was suggested by ultrastructural analysis of yeast cell walls revealing a denser mannan outer layer and wall porosity reduced. It is possible to imply that P. cauliflora targeted the C. albicans cell wall inducing some changes in the architecture, notably the outer glycoprotein layer, affecting the cell wall porosity without alteration of the polysaccharide or protein level. © 2013 by the authors.