Atividade antifúngica de Caryocar brasiliensis (Caryocaraceae) sobre Cryptococcus neoformans

A grande incidência de criptococose em decorrência do aumento crescente de indivíduos imunodeprimidos e os efeitos colaterais aos fármacos utilizados para o tratamento desta infecção, tem incentivado a pesquisa de novos agentes antifúngicos. Através da técnica de diluição em ágar, foi verificada a atividade antifúngica (in vitro) de diferentes constituintes de Caryocar brasiliensis sobre Cryptococcus neoformans. Verificou-se que a cera epicuticular retirada da folha, coletada em período de baixo índice pluviométrico (170,8mm de água), foi a parte mais ativa da planta, inibindo o crescimento de 91,3% (21/23) dos isolados de Cryptococcus neoformans em concentração < a 250µg/mL.

Human serum antibody reactivity towards Paracoccidioides brasiliensis antigens treated with sodium metaperiodate

In this study, we evaluated the profile of anti-Paracoccidioides brasiliensis immunoglobulin isotypes in serum from patients with the acute and chronic forms of paracoccidioidomycosis, using the whole Paracoccidioides brasiliensis antigen and the antigen treated with sodium metaperiodate. All the immunoglobulin isotypes present in the serum from patients with the acute and chronic forms of paracoccidioidomycosis presented higher reactivity towards the whole antigen than to the antigen treated with metaperiodate (P < 0.05). The reactivity of IgG and IgM to the antigen treated with metaperiodate was greater in serum from patients with the acute form of the disease (P < 0.05), while IgA was more reactive in serum from patients with the chronic form (P < 0.05). There was greater reactivity of IgG1 and IgG2 to the whole antigen and the antigen treated with metaperiodate in the serum from patients with paracoccidioidomycosis than there was in serum from patients with other parasitic infections (P < 0.05). Furthermore, IgG1 from patients with the acute form recognized the 19kDa, 27kDa and 31kDa antigens in the western blot test. Thus, the results suggest that modifications to the epitopes of Paracoccidioides brasiliensis antigens may help to improve the immunodiagnosis of paracoccidioidomycosis.

The Triplaria tree (Triplaris spp) and Pseudomyrmex ants: a symbiotic relationship with risks of attack for humans

The authors report a massive attack by Pseudomyrmex ants on a human who touched a Triplaria - novice tree (Triplaris spp). The ants naturally live in these trees and their stings cause intense pain and discrete to moderate local inflammation. The problem is common in some Brazilian regions and can be prevented by identifying the trees.

High molecular mass fraction in clinical isolates of Paracoccidioides brasiliensis

INTRODUCTION: Different serum levels of the IgG/IgE for Paracoccidioides brasiliensis high mass molecular (hMM) fraction (~366kDa) in the acute and chronic forms of the disease have been reported. Considering the nonexistence of hMM fraction investigation involving clinical isolates of P. brasiliensis, the present study aimed to investigate the presence of the hMM fraction (~366kDa) in cell free antigens (CFA) from P. brasiliensis clinical isolates. METHODS: CFA from 10 clinical isolates and a reference strain (Pb18) were submitted to SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by gel image capturing and densitometer analysis. Additionally, CFA from 20 isolates and Pb18 were analyzed by capture ELISA (cELISA) using polyclonal (polAb) or monoclonal (mAb) antibodies to the hMM fraction. RESULTS: The presence of the hMM component was observed in CFA of all samples analyzed by SDS-PAGE/densitometry and by cELISA. In addition, Pearson's correlation test demonstrated stronger coefficients between hMM fraction levels using pAb and mAb (R = 0.853) in cELISA. CONCLUSIONS: The soluble hMM fraction was present in all the P. brasiliensis clinical isolates analyzed and the reference strain Pb18, which could be used as a source of this antigen. The work also introduces for first time, the cELISA method for P. brasiliensis hMM fraction detection. Analysis also suggests that detection is viable using polAb or mAb and this methodology may be useful for future investigation of the soluble hMM fraction (~366kDa) in sera from PCM patients.

A semi-nested PCR assay for molecular detection of Paracoccidioides brasiliensis in tissue samples

INTRODUCTION: Paracoccidioidomycosis is a systemic infection caused by Paracoccidioides brasiliensis. METHODS: In this study, a semi-nested PCR for paracoccidioidomycosis diagnosis was developed. The primers ITS1 and ITS4 were used in the first reaction, while the primers MJ03 and ITS1 primer were used in the second reaction. The semi-nested PCR was used to investigate biopsies of five patients with oral lesions that resembled paracoccidioidomycosis. RESULTS: The semi-nested PCR was positive for four samples and negative for a sample from a patient later diagnosed with leishmaniasis. CONCLUSIONS: The new semi-nested PCR describe is useful for paracoccidioidomycosis diagnosis.

In vitro antifungal activity of fatty acid methyl esters of the seeds of Annona cornifolia A.St.-Hil. (Annonaceae) against pathogenic fungus Paracoccidioides brasiliensis

INTRODUCTION: Fatty acids are abundant in vegetable oils. They are known to have antibacterial and antifungal properties. METHODS: Antifungal susceptibility was evaluated by broth microdilution assay following CLSI (formerly the NCCLS) guidelines against 16 fungal strains of clinical interest. RESULTS: In this work, fatty acid methyl esters (FAME) was able to inhibit 12 clinical strains of the pathogenic fungus Paracoccidioides brasiliensis and were also active in the bioautographic assay against Cladosporium sphaerospermum. CONCLUSIONS: FAME was a more potent antifungal than trimethoprim-sulfamethoxazole against P. brasiliensis under the experimental conditions tested.

The influence of carbohydrates in the interaction of Paracoccidioides brasiliensis with CCL-6 cells in vitro

INTRODUCTION: Little is known about the early events in the interaction between Paracoccidioides brasiliensis and its host. To understand the effect of carbohydrates in the interaction between the fungus and epithelial cell in culture, we analyzed the influence of different carbohydrate solutions on the adhesion of P. brasiliensis yeast cells to CCL-6 cells in culture. METHODS: Fungal cells were cultivated with the epithelial cell line, and different concentrations of D-fucose, N-acetyl-glucosamine, D-mannose, D-glucosamine, D-galactosamine, sorbitol and fructose were added at the beginning of the experiment. Six hours after the treatment, the cells were fixed and observed by light microscopy. The number of P. brasiliensis cells that were adhered to the CCL-6 monolayer was estimated. RESULTS: The number of adhesion events was diminished following treatments with D-fucose, N-acetyl-glucosamine, D-mannose, D-glucosamine and D-galactosamine as compared to the untreated controls. Sorbitol and fructose-treated cells had the same adhesion behavior as the observed in the control. P. brasiliensis propagules were treated with fluorescent lectins. The FITC-labeled lectins WGA and Con-A bound to P. brasiliensis yeast cells, while SBA and PNA did not. CONCLUSIONS: The perceptual of adhesion between P. brasiliensis and CCL-6 cells decreased with the use of D-mannose, N-acetyl-glucosamine and D-glucosamine. The assay using FITC-labeled lectins suggests the presence of N-acetyl-glucosamine, α-mannose and α-glucose on the P. brasiliensis cell surface. An enhanced knowledge of the mediators of adhesion on P. brasiliensis could be useful in the future for the development of more efficient and less harmful methods for disease treatment and control.

An optimized one-tube, semi-nested PCR assay for Paracoccidioides brasiliensis detection

Introduction Herein, we report a one-tube, semi-nested-polymerase chain reaction (OTsn-PCR) assay for the detection of Paracoccidioides brasiliensis. Methods We developed the OTsn-PCR assay for the detection of P. brasiliensis in clinical specimens and compared it with other PCR methods. Results The OTsn-PCR assay was positive for all clinical samples, and the detection limit was better or equivalent to the other nested or semi-nested PCR methods for P. brasiliensis detection. Conclusions The OTsn-PCR assay described in this paper has a detection limit similar to other reactions for the molecular detection of P. brasiliensis, but this approach is faster and less prone to contamination than other conventional nested or semi-nested PCR assays.

Toxicological profile of deltamethrin in Triatoma brasiliensis (Hemiptera: Reduviidae) in State of Ceará, Northeastern Brazil

INTRODUCTION: Triatoma brasiliensis is the species of greatest epidemiological relevance in the semi-arid region of Brazil. This species is predominantly found in domestic environments, and it has the ability to build large colonies with high levels of natural infection via Trypanosoma cruzi. Thus, T. brasiliensis is one of the most efficient transmitters of Chagas disease (CD) to humans. Despite household spraying with residual insecticides, many areas report persistent reinfestations for reasons that remain poorly understood. Therefore, this study sought to characterize the toxicological profile of deltamethrin in T. brasiliensis from areas with persistent reinfestation in State of Ceará, Brazil. METHODS: The susceptibility reference lineage (SRL) was derived from Umari. Serial dilutions of deltamethrin were prepared and applied to the dorsal abdomen of first instar nymphs. The control group received only pure acetone. Mortality was evaluated after 72h. Qualitative tests assessed mortality in response to a diagnostic dose of 1xLD99 (0.851 nanograms of active ingredient per treated nymph) of the SRL. RESULTS: The susceptibility profile characterization of the T. brasiliensis populations revealed 50% resistance ratios (RR50) that ranged from 0.32 to 1.21. The percentage of mortality in response to the diagnostic dose was 100%. CONCLUSIONS: We demonstrated that T. brasiliensis was highly susceptible to deltamethrin. The control difficulties found might be related to the recolonization of the triatomines originating from neighboring environments and the possible operational failures related to the process of spraying that enabled specimens less susceptible to deltamethrin to survive.

PRELIMINARY RESULTS OF A LARGE-SCALE TREE INVENTORY OF UPLAND RAIN FOREST IN THE CENTRAL AMAZON

A large-scale inventory of trees > 10cm DBH was conducted in the upland "terra firme" rain forest of the Distrito Agropecuário da SUFRAMA (Manaus Free Zone Authority Agricultural District) approximately 65Km north of the city of Manaus (AM), Srasil. Thegeneral appearance and structure of the forest is described together with local topography and soil texture. Thepreliminary results of the Inventory provide a minimum estimate of 698 tree species in 53 families in the 40Km radius sampled, including 17 undescribed species. Themost numerically abundant families, Lecythidaceae, Leguminosae, 5apotaceae and Burseraceae as also among the most species rich families. One aspect of this diverse assemblage is the proliferation of species within certain genera, Including 26 genera In 17 families with 6 or more species or morphospecies. Most species have very low abundances of less than 1 tree per hectare. While more abundant species do exist at densities ranging up to a mean of 12 trees per ha, many have clumped distributions leading to great variation in local species abundance. The degree of similarity between hectare samples based int the Coefficient of Community similarity Index varies widely over different sample hectares for five ecologically different families. Soil texture apparently plays a significant role In determining species composition in the different one hectare plots examined while results for other variable were less consistent. Greater differences in similarity indices are found for comparisons with a one hectare sample within the same formation approximately 40Km to the south. It is concluded that homogeneity of tree community composition within this single large and diverse yet continuous upland forest formation can not be assumed.

AVALIAÇÃO DE GENÓTIPOS DE SERINGUEIRA (Hevea spp.) AO Microcyclus ulei VISANDO À IDENTIFICAÇÃO DE FONTES DE RESISTÊNCIA HORIZONTAL

Avaliou-se, sob condições controladas, a resistência de genótipos de seringueira ao mal-das-folhas. Empregaram-se dois isolados de Microcyclus ulei, pertencentes aos grupos I e II. Utilizaram-se mudas em sacos plásticos e inocularam-se folíolos nos estádios B1/B2 com uma suspensão de 2 χ 105conídios/ml. Incubaram-se as mudas em câmara úmida a 24° C por 24 horas. Determinaram-se o período latente (PL), diâmetro das lesões (DL), tipo de lesão (TL) e tipo de reação (TR). A maioria dos genótipos testados foi suscetível a pelo menos um isolado do patógeno. Os genótipos que apresentaram resistência moderada, com maior período latente e menor esporulação a um isolado, em geral, foram suscetíveis ao outro. Apenas o genotipo AM/86/271 exibiu resistência incompleta, caracterizada por maior PL, menor DL e baixa esporulação. Entretanto, o número de isolados testados não assegurou o caráter da resistência. Com base nos resultados obtidos concluiu-se que os genótipos avaliados não apresentaram perspectivas para utilização em programas de melhoramento visando resistência ao M.ulei.

Aperfeiçoamento da técnica de eletroforese para análise isoenzimática de clones de seringueira (Hevea sp.)1

Este estudo foi conduzido no sentido de otimizar as técnicas de preparo de extratos de folíolos e identificar tampões e condições de corrida que proporcionassem uma boa definição dos perfis eletroforéticos de clones de seringueira. Foram testados 25 diferentes sistemas enzimáticos utilizando-se onze sistemas tamponantes. Não foram encontradas diferenças na qualidade dos zimogramas obtidos nos diferentes estádios fenológicos analisados, em amostras submetidas a diferentes condições de centrifugação e nas diferentes plantas de um mesmo clone. Extratos de folíolos liofilizados apresentaram resolução semelhante para Adh, Pgi, 6Pgdh, Lap-1, Skdh, Acp, Mdh, ßGlu, Pgm e Idh que homogenados de folíolos não liofilizados preparados até 2 dias pós-coleta. Os padrões de Got e Est apresentaram boa nitidez somente em extratos de folíolos frescos. As demais enzimas não apresentaram resolução satisfatória. Com o uso de diferentes substratos, foi possível detectar nove regiões de atividade esterásica nas condições empregadas, embora variação genética tenha sido caracterizada para apenas três locos nos clones analisados. Os fenótipos observados para esterase-7 utilizando-se ésteres derivados de naftol foram semelhantes aos padrões encontrados utilizando L-benzoil-ßnaftilamida como substrato, indicando que esta esterase seja, provavelmente, uma en-dopeptidase. A utilização de substratos fluorogênicos permitiu ainda a detecção de uma variante eletroforética de Acp em Hevea nítida e uma variante de ßGlu em Hevea rigidifolia.