Isaac Roitman

Fine structure study of Leptomonas samueli by the freeze-fracture technique.

The ultrastructure of promastigotes ofLeptomonas samueli was studied by the freeze-fracture technique. Both the P and E faces of the plasma membrane ofL. samueli have intramembranous particles. However, the particle density was higher in the E than in the P face. The P face of the flagellar membrane had few particles whereas abundant flat membrane particles were evident on the E face. An arrangement of particles indicating membrane specializations was seen on the flagellar membrane and on the membrane lining the flagellar pocket. Cellular structures such as the nucleus, peroxisomes, the mitochondrion, and microtubules were also studied.

An electron microscopic and cytochemical study of Leptomonas samueli.

The fine structure of promastigotes ofLeptomonas samueli is described. This protozoan revealed several features in common with other trypanosomatids. A large membrane-bound cavity containing many vesicles was observed near the nucleus. Pinocytotic vesicles were seen arising from the membrane lining the flagellar pocket. They are associated with some microtubules which originate in the flagellar pocket region and extend toward the multivesicular structure. Morphometric analysis made on electron micrographs showed a mitochondrial relative volume of 0.11 and a peroxisome relative volume of 0.08. Determination of the number of sub-pellicular microtubules in different regions of the protozoon body show that the largest number is found in the region containing the Golgi complex. Carbohydrates were detected using the periodic-acid-thiosemicarbazide-silver proteinate technique. Reaction products were seen in the plasma membrane, in the membrane of the Golgi complex, and in the membranes which form the multivesicular structure. Two cytochemical methods were used to locate basic proteins. Using the ammoniacal silver method reaction products were seen only in the nucleus. With the ethanolic phosphotungstic acid method reaction products were seen in the microtubules which form the flagellum, in the peroxisome-like organelle, and at the region of adhesion of the flagellum to the cell body.

INCORPORATION OF AMMONIUM IN AMINO ACIDS BY TRYPANOSOMA CRUZI

Ammonium ions were incorporated into L-glutamate and alpha-ketoglutarate in epimastigote forms of Trypanosoma cruzi through the following enzymatic systems: NADPH and NADH-dependent glutamate dehydrogenase, NADPH-dependent glutamate synthase, L-glutamine synthetase and NADH-dependent glutamate synthase in order of decreasing specific activity (mumoles of product formed/min/mg protein). The pH optima and Kms for the glutamate dehydrogenase system were determined. Disc electrophoresis showed the presence of cathodic bands of GDH activity, which were highly dependent on NADP+.

Effect of lipopolysaccharide (LPS) on the metabolism and cell surface of the trypanosomatid Herpetomonas megaseliae

Abstract 1. 1. The effect of lipopolysaccharide (LPS) on the surface and metabolism of Herpetomonas megaseliae was studied. 2. 2. No inhibition of growth was detected even with the use of high concentrations of LPS (100 μg/ml). 3. 3. LPS stimulated the differentiation-transformation of pro- to opisthomastigote forms at low concentrations (2 × 10 −3 μ g/ml). 4. 4. LPS-treated cells showed an increased O 2 uptake when glucose and glycerol were used as substrate. In addition such cells induce O 2 consumption when proline was present. 5. 5. LPS-treated cells showed an increased content of galactose and xylose and a marked decrease of rhamnose and mannose, whereas glucose amount remained unchanged. 6. 6. LPS-treated cells showed an increased agglutination in the presence of Con A and a decrease in the affinity for anti-flagellate serum. 7. 7. The results of items 3, 4, 5 and 6 are discussed in relation to the cell differentiation of Herpetomonas .

Imunofluorescência indireta no pênfigo foliáceo endêmico. contribuição para sua padronização

The aim of the present research was to standardize the indirect immunofluorescence reaction for Endemic Pemphigus Foliaceus (Fogo Selvagem). We found that fresh human skin was the ideal substrate and could proceed from foreskin, head, neck, or anterior abdominal wall. PBS pre-washing of the skin preceding the incubation with the serum should be avoided since the antigenicity might be diminished. TAS-calcium pre-serves the Pemphigus antigenic properties of the skin and shall be preferred as the diluent for the sera. Albumin-coated slides are useful because they increase the adherence of the skin sections. The conjugate appropriate dilution is convenientely determined by the radial immunodiffusion test (Ouchterlony method). So far as the correlation between the antibody titer and the clinical activity is concerned, we concluded that a titer of 160 or more was of bad prognosis, since it was associated with the generalized form of the disease or with cases of the localized form refractory to the usual therapy. Nevertheless, this assumption needs confirmation by further studies involving an appropriate clinical approach.O proposito da presente investigacao foi padronizar a reacao de imunofluorescencia indireta para Penfigo Foliaceo Endemico (Fogo Selvagem). Verificamos que a pele humana normal e o substrato ideal e que pode proceder de prepucio, cabeca, pescoco ou da parede abdominal anterior. A lavagem previa da pele precedendo a incubacao com o soro deve ser evitada pois a antigenicidade pode ser diminuida. O TAS-calcio preserva as propriedades antigenicas da pele e deve ser preferido como diluente para os soros. Lâminas cobertas com albumina sao uteis porque aumentam a aderencia dos cortes de pele. A diluicao apropriada do conjugado e convenientemente determinada pelo teste de imunodifusao radial (metodo de Ouch-terlony). Com referencia a correlacao entre titulo de anticorpos e atividade clinica, concluimos que um titulo igual ou maior do que 160 era de mau prognostico pois estava associado a forma generalizada da doenca ou a casos de forma localizada refratarios a terapeutica usual. Contudo, esta obervacao requer confirmacao atraves de estudos que envolvam uma abordagem clinica apropriada.The aim of the present research was to standardize the indirect immunofluorescence reaction for Endemic Pemphigus Foliaceus (Fogo Selvagem). We found that fresh human skin was the ideal substrate and could proceed from foreskin, head, neck, or anterior abdominal wall. PBS pre-washing of the skin preceding the incubation with the serum should be avoided since the antigenicity might be diminished. TAS-calcium preserves the Pemphigus antigenic properties of the skin and shall be preferred as the diluent for the sera. Albumin-coated slides are useful because they increase the adherence of the skin sections. The conjugate appropriate dilution is convenientely determined by the radial immunodiffusion test (Ouchterlony method). So far as the correlation between the antibody titer and the clinical activity is concerned, we concluded that a titer of 160 or more was of bad prognosis, since it was associated with the generalized form of the disease or with cases of the localized form refractory to the usual therapy. Nevertheless, this assumption needs confirmation by further studies involving an appropriate clinical approach.

Herpetomonas megaseliae: effect of lipopolysaccharide on cell-surface anionogenic groups

Surface anionic groups ofHerpetomonas megaseliae, untreated or treated with lipopolysaccharides (LPS), were analyzed by cell electrophoresis and cytochemistry as well as by identification of sialic acid residues with the aid of thin-layer chromatography. LPS treatment of the trypanosomatids caused a marked decrease in the negative surface charge, as well as in binding to the cell surface of cationized ferritin, colloidal iron hydroxide, and Sendai virus particles. Incubation of LPS-treated and untreated flagellates with neuraminidase resulted in decrease of their mobility to the negative pole. The foregoing findings suggest that sialic acid residues are the major anionogenic groups exposed on the surface of this protozoon. Treatment with LPS markedly alters the nature and number of these residues. OnlyN-acetylneuraminic acid can be detected on untreated flagellates, whereasN-acetyl andN-glycolylneuraminic acid derivatives are found on the organisms exposed to LPS.

Auxotrophic mutants of Crithidia fasciculata.

In pursuance of genetic studies, after exposure to ethylmethanesulfonate, 11 auxotrophic mutants of Crithidia fasciculata were cloned. Three proved uracil dependent; 3 serine dependent; and the remainder have not had their auxotrophy defined.

ESTUDO COMPARATIVO DA ATIVIDADE RESPIRATÓRIA DE: BACTÉRIAS DO GÉNERO CHROMOBACTERIUM *

Utilizando possivel nova especie de Chromobacterium, uma analise comparativa da atividade respiratoria foi realizada com a utilizacao de outras amostras de Chromobacterium, atraves do metodo polarografico. Os resultados demonstraram diferencas na atividade respiratoria de Chromobacterium violaceum (amostra Goias) em comparacao com outras Chromobacterium em presenca de varios substratos. Esses resultados sugerem que C. violaceum (amostra Goias) seja uma especie nova, ou, ao minimo, uma subespecie.