Growth curves of Leishmania braziliensis braziliensis Promastigotes and surface antigen expression before and after adaptation to Schneider's drosophila medium as assessed by anti-Leishmania human sera

Leishmania braziliensis braziliensis(MHOM/BR/75/M2903) was grown in Schneider's Drosophila medium. In one set of experiments promastigotes were already adapted to the medium by means of serial passages whereas in the second cells were grown in a biphasic medium and transfered to the liquid. Growth was more abundant for culture medium adapted cells; degenerate cells in small numbers as well as dead ones were present from day 5 for promastigotes adapted to liquid medium and from day 3 for newly adapted cells. Synthesis of surface antigens differed according to length of cell culture as assessed by the titer of five mucocutaneous leishmaniasis sera on subsequent days. Five days of culture for cells already adapted to the culture medium and 3 days for newly adapted ones were judged to be the best for the preparation of immunofluorescence antigens.

Induction of iron regulated proteins during normal growth of Neisseria meningitidis in a chemically defined medium

The expression of iron regulated proteins (IRPs) in vitro has been obtained in the past by adding iron chelators to the culture after bacterial growth, in the presence of an organic iron source. We have investigated aspects concerning full expression of the meningococcal IRPs during normal growth, in defined conditions using Catlin medium, Mueller Hinton and Tryptic Soy Broth (TSB). The expression of IRPs varied between different strains with respect to Ethylenediamine Di-ortho-Hidroxy-phenyl-acetic acid (EDDA) concentrations, and according to culture medium, and also between different lots of TSB. For each strain, a specific set of IRPs were expressed and higher EDDA concentrations, or addition of glucose, or use of different culture media did not resulted in a differential expression of IRPs. We were not able to grow N. meningitidis under normal growth conditions using Desferal. We looked for a good yield of outer membrane vesicles (OMVs) expressing IRPs in iron-deficient Catlin medium containing EDDA and Hemin. Culture for 32 h at 30ºC after growing for 16 h at 37ºC supported good bacterial growth. Bacterial lysis was noted after additional 24 h at 30ºC. Approximately 4 times more OMVs was recoverable from a culture supernatant after 24 h at 30ºC than from the cells after 16 h at 37ºC. The IRP were as well expressed in OMVs from culture supernatant obtained after 24 h at 30ºC as from the cells after 16 h at 37ºC.

Reproduction of Leishmania (Leishmania ) infantum chagasi in conditioned cell culture growth medium

Leishmanias can be produced by inoculation in conditioned McCoy cell culture growth medium (CGM). Leishmania (Leishmania) infantum chagasi (100 parasites) grown in NNN medium was inoculated in 2.5 mL CGM, kept in plates (24 wells) and its multiplication was observed for five days (120 hours). After day 5, the medium was saturated with the flagellate forms of the parasite (promastigotes). The reproduction of the leishmanias was observed every 24 hours and the number of parasites was calculated by counting the parasites in a drop of 10 µ L and photomicrographied. So the number of Leishmanias was adjusted to 1 mL volume. The advantage of the technique by isolation of Leishmania in CGM demonstrated in this study is its low cost and high efficacy even with a small quantity of parasites (10² promastigotes) used as inoculum. Additionally, isolation of the leishmania can be obtained together with an increase in their density (180 times) as observed by growth kinetics, within a shorter time. These results justify the use of this low-cost technique for the isolation and investigation of the behavior and multiplication of Leishmania both in vertebrates and invertebrates, besides offering means of obtaining antigens, whether whole antigens (leishmanias) or the soluble antigens produced by the parasites which may be useful for the production of new diagnostic kits.

CO-INFECTION OF DENGUE VIRUS BY SEROTYPES 1 AND 4 IN PATIENT FROM MEDIUM SIZED CITY FROM BRAZIL

SUMMARY The natural co-infection with dengue virus can occur in highly endemic areas where different serotypes have been observed for many years. We report one case of DENV-1/DENV-4 co-infection in human serum detected by molecular tests. Phylogenetic analysis of the sequences obtained indicated the presence of genotype V and II for DENV-1 and DENV-4, respectively.

Medium-term cryopreservation of rabies virus samples

Introduction The cryopreservation of rabies virus has been described in detail in the literature. To date, little information is available on the use of cryoprotective agents for cold preservation of this virus, and the available data focus only on short-term virus preservation. In this study, we investigated the medium-term cryopreservation of samples of rabies virus using different cryopreservation protocols. Methods The cryopreservation protocols for the rabies virus samples were performed at -20°C and were divided according to the variables of time and cryoprotectant type used. The laboratory tests (intracerebral inoculation of mice, viral titration and direct immunofluorescence) were performed at regular intervals (360 and 720 days) to assess the viability of the viral samples according to the different preservation techniques used. Results After 1 year of cryopreservation, the fluorescence intensity of intracellular corpuscles of the rabies virus and the median survival time of the mice differed between the positive controls and the treatments with the cryoprotectants. After 2 years, most of the samples subjected to the cryopreservation protocols (including the controls) did not produce fluorescence. However, the virus samples exposed to the cryoprotectant sucrose (68% solution) responded positively in the direct immunofluorescence assay and in the intracerebral inoculation of the mice. Conclusions Medium-term cryopreservation of the rabies virus inactivates the viral sample. However, the cryoprotectant agent sucrose (68%) produces a preservative effect in cryopreserved rabies virus samples.

Multi-temporal analysis of radiometric changes in satellite images of forest edges to infer selective-logging areas in the Amazon forest

Radiometric changes observed in multi-temporal optical satellite images have an important role in efforts to characterize selective-logging areas. The aim of this study was to analyze the multi-temporal behavior of spectral-mixture responses in satellite images in simulated selective-logging areas in the Amazon forest, considering red/near-infrared spectral relationships. Forest edges were used to infer the selective-logging infrastructure using differently oriented edges in the transition between forest and deforested areas in satellite images. TM/Landsat-5 images acquired at three dates with different solar-illumination geometries were used in this analysis. The method assumed that the radiometric responses between forest with selective-logging effects and forest edges in contact with recent clear-cuts are related. The spatial frequency attributes of red/near infrared bands for edge areas were analyzed. Analysis of dispersion diagrams showed two groups of pixels that represent selective-logging areas. The attributes for size and radiometric distance representing these two groups were related to solar-elevation angle. The results suggest that detection of timber exploitation areas is limited because of the complexity of the selective-logging radiometric response. Thus, the accuracy of detecting selective logging can be influenced by the solar-elevation angle at the time of image acquisition. We conclude that images with lower solar-elevation angles are less reliable for delineation of selecting logging.

Selective serotonin-reuptake inhibitor and norepinephrine dopamine reuptake inhibitor antidepressants do not affect natural killer cell activity in vitro

OBJECTIVE: This study aims to evaluate the citotoxic activity of two commonly used anti-depressants: paroxetine and bupropion. We also evaluated the in vitro natural killer activity (NKA) after incubating the blood samples with the antidepressants. METHODS: Peripheral blood samples from 15 healthy volunteers were collected and the mononuclear cells (PBMCs) were isolated and incubated for 24h with (or without = control cells) paroxetine and bupropion, in concentrations of 30, 100 and 1000 ng/ml. After the incubation period in both groups, the amount of dead cells was calculated using trypam blue technique. NKA was evaluated using the classic51Cr release assay. CONCLUSIONS: PBMCs dead cells occurred in both groups and in proportion to all pharmacological concentrations. Nevertheless, the NKA was not affected, even with the reduction in the number of effective cells.

Propriedades psicométricas da Cocaine Selective Severity Assessment (CSSA) em mulheres usuárias de crack

Objetivo Este estudo teve como objetivo adaptar a Cocaine Selective Severity Assessment (CSSA) para o português do Brasil e verificar as propriedades psicométricas do instrumento em uma amostra de usuárias de crack. Métodos Após as etapas de tradução e adaptação, 125 mulheres usuárias de crack, internadas em uma unidade pública de desintoxicação, foram avaliadas. Para caracterização da amostra e análise das validades concorrente, de construto e preditiva, foram utilizados os seguintes instrumentos: SCID-I, ASI-6, BDI-II e CCQ-B. Resultados A análise fatorial exploratória identificou cinco fatores, com níveis adequados de consistência interna tanto para os fatores quanto para o escore geral da CSSA. Quanto à validade concorrente, a CSSA vai ao encontro de instrumentos já utilizados na clínica e em pesquisas. Em relação à validade de construto e preditiva, a CSSA pode ser sensível ao declínio dos sintomas de abstinência durante o processo de desintoxicação do crack. Conclusões Nossos achados foram além da tradução e adaptação da CSSA, proporcionando testes de validade e sugerindo que a CSSA é um instrumento confiável na avaliação dos sintomas de abstinência do crack.

Abundance of Conepatus chinga (Carnivora, Mephitidae) and other medium-sized mammals in grasslands of southern Brazil

Between January 2007 and December 2010, the abundance of medium-sized mammals was studied, with special focus on the Molina's hog-nosed skunk, Conepatus chinga (Molina, 1782), at four locations in southern Brazil. In this study, transect line methodology was used to obtain data for Distance Analyses. Transects were traveled by car at night, searching with spotlights along the edges of secondary roads in agricultural landscapes. Along 1,811 km, we obtained 620 observations of 20 mammal species. The most common species was the exotic European hare, Lepus europaeus (Pallas, 1778); the highest abundance estimated for South America was observed in one of the study areas, where its density was estimated as 32 individuals/km². Carnivores were the most commonly recorded mammals, represented by 10 species and comprising 51% of all observations. Molina's hog-nosed skunk occurred in all study areas, but occurred in sufficient numbers to obtain density estimates in only two of the areas. We estimated 1.4 to 3.8 individuals/km², in the first density estimate made by the transect method for a member of Conepatus in the Neotropics. These values are similar to those estimated for North American species of Mephitidae. In Brazil, C. chinga is apparently more abundant in the Pampa biome than in the grasslands of the Atlantic Forest. For two other carnivores, Lycalopex gymnocercus (Fisher, 1814) and Cerdocyon thous (Linnaeus, 1766), we estimated preliminary densities that were similar to those previously cited for different regions.

Diversity and habitat preference of medium and large-sized mammals in an urban forest fragment of southwestern Amazon

We assessed the species composition and abundance of medium and large-sized mammals in an urban forest fragment in the Brazilian Amazon, and recorded the preference of some species for particular phytophysiognomies. We placed nine transects with 20 sand plots each in three phytophysiognomies: open rainforest with a dominance of bamboos (OFB), open rainforest with palm trees (OFP), and dense rainforest (DF). We calculated species abundance as the number of records/plot.day, in a total of 2,700 plots.day. We recorded twelve mammal species; Sylvilagus brasiliensis (Linnaeus, 1758) and Dasyprocta fuliginosa (Wagler, 1831) were the most abundant. The results differed among phytophysiognomies: DF presented the highest mammal diversity, whereas the species composition of OFP was less similar than that of other phytophysiognomies. Rodents showed higher preference for OFP and Sylvilagus brasiliensis was more abundant in OFB. The study area showed high species richness, with the occurrence of mesopredators, but there was a predominance of common species adaptable to disturbed environments, which reflects the severe isolation degree of the forest fragment and the hunting pressure that is still present.

A vegetação do município de Ilhéus, Estado da Bahia: III - caracterização da vegetação pelo valôr dos índices das espécies

The indices found are analysed as a whole and general conclusions are drawn from them which may be of use in understanding many of the problems offered by the local flora (Ilhéus). The first column of the tables presented indicates the biological form of the species, showing the nature of the flora and the constitution of the climax. A total of 200 species of phanerophyta were found; 69 macrophanerophyta (trees), 54 are mesophanerophyta (treelets) and 77 are nanophanerophyta (shrubs). The macrophanerophyta are consequently considered as dominants and the meso-and nanophanerophyta as codominants (the biological forms: chamaephyta, hemicriptophyta, criptophyta, geophyta, therophyta, epiphyta and hydrophyta are subdominants), the more so as the first cover 80% and the others more or less 50%. This points to a climax of trees and a local vegetation mainly composed of trees also. The smaller forms are left out as they are beyond the present scope of this sort of wort in Brazil. The third column of ecological formulae indicates the reaction of the constituent species to light (C = sciophilous, F = photophilous and I = indifferents), the biological types of vegetation (H = hygrophytes, X = xerophytes and M = mesophytes) and the fidelity of the species to the climax. Of the species studied: 25 are pioneers (P. Table I), 63 are accidentals (A. Table II), 35 are companion species (O. Table III), 19 show preferences (E. Table with vitality Vn), 44 are selective (S. Table V) and 13 exclusive species (L. Table VI). This leads to the conclusion that the vegetation of the region is in full reconstitution. As to the ecological characteristics of the 200 species studied, 89 are either pioneers (a class separated by the author) or accidentals; this means that the devastated zones are being reconstituted in the subsere both with members of the prisere and alien species. Of the remaining species, 54 are companion, or accompanying species, which appear in most subclimax, serclímax and quasiclimax associations, and 57 are real constituents of the local climax. As all the species except the pioneers, selectives and exclusives (xerophytes and mesophytes) may be considered as hygrophytes this type evidently predominantes in the region and may constitute a hygrophilous serclimax and quasiclímax. In regard to light 101 are sciophilous, 32 indiferents and 67 photophilous. This leads to the conclusion that the vegetation comprises mainly tolerant species, showing the hygrophilous and mesophilous character of the region with a vegetation composed mostly of trees. The presence a large number of sciophilous species is easy to understand as the hygrophilous and mesophilous habitats and the dominance of trees favour the germination and growth of tolerant species. The last two columns analyse the percentage of individuals present and the occurrent classes to which they belong: 92 species vary between 1 and 9%; 50 between to 10 and 19%; 36 between 20 and 29%; 14 between 30 and 39%; and 8 between 40 and 49%. Only 8 species belong to occurrence class V; 14 to classe IV; 36 to class III; 50 to class II; and 92 to class I. This leads to the conclusion that the local formation is very unsociable and very complex, though the median coverture is 80% and the number of species is very large. The analysis of the data also shows that the climax is being reconstituted in the subsere with elements drawn from the prisere and alien species introduced either by man (following desvastation) or by other consequent factors (such as brusque changes of microclimates due to total or partial destruction). This modifies the subclimax appreciably and apparently also the climax of the local regional subsere. As a final conclusion it is suggested that as in the subsere the pioneer formation is xerophilous, the prisere also beging as a xerosere; but as there are and probably always were hydrophilous formation evolving in the same climate, the local climax is composed of species with medium exactions, that is of relative mesophites.

Presença de substâncias fisològicamente ativas em culturas de E. Coli isolados das fezes de pacientes alérgicos pela técnica de Heist-Cohen

Four patients with several allergic symptoms have been studied. Heist-Cohen pathogen selective method was positive in feces of all these patients for E. coli. Other causes that could be responsible for the allergic symptoms were discarded. Samplets after seeding in a medium physiologically inactive have been tested in guinea pig illea following Cohnheim-Magnus technique. It was demonstrated that all samples isolated by the Heist-Cohen technique were physiologically active.(Fig.1). Ten other strains of E. coli taken at random from the collection of I.O.C. and tried according similar technique, brought evidence that only three samples possessed such activity. This confirms the works of KOESSLER-HANKE (1922). In order to obtain a quick relief from allergic symptoms, one of us had employed in some others, identical cases an antibiotic (Teramycin, Chas Pfeizer) before the specific disesitization was done (Figs. 2, 3, 4, 5). Such method were choice since LABORDE, PARROT, and URQUIA (1953) have demonstrated the importance of production of histamine, from components of the bowel's flora in the production of allergic symptoms.

Chagas' disease: selective affinity and cytotoxicity of Trypanosoma cruzi-immune lymphocytes to parasympathetic ganglion cells

The megaesophagus and megacolon endemic in South America are related , to Chagas' disease. These mega conditions are found in patients with chronic Chagas's infection, when the parasite is not demonstrable in the lesions. These are characterized by depopulation of parasympathetic ganglion cells, dilation and hypertrophy of the viscera. In the experiments described here we deminstrate a selective affinity and adherence of Trypanosoma cruzi-immune lymphocytes to myenteric, parasympathetic ganglion cells, leading to neuronolysis. None of these features are observed when non-immune lymphocytes from control rabbits are used, or when the immune lymphocytes are allowed to react with CNS neurons. This demonstration is an indication of the high degree of specificity of the destruction of parasympathetic neurons in Chagas' disease. We postulate that the T. cruzi-immune lymphocyte rejection of parasympathetic neurons, but not of CNS neurons, might be related to recognition of a cross-reacting antigenic determinant secreted only by the target neurons. In favor of this interpretation is the observation of lymphocytic infiltrates and parasympathetic ganglion cell destruction in chronic Chagas' infection in the absence of encephalitis.

Liquid nitrogen cryopreservation of Bracoccidioides brasiliensis in Fava's Netto medium

The aplicability of Fava's Netto medium in the liquid nitrogen cryopreservation technique of Paracoccidioides brasiliensis cells was demonstrated.

The change of behavior of two strains of Leishmania after cultivation in a defined medium

Attempts have been made to characterize two strains of Leishmania that became infective to golden hamsters only after they had been maintained for several years in a chemically defined culture medium. Observations were made on the growth rates of promastigotes in vitro, course of infection in hamsters, morphology of amastigotes, and electrophoretic mobility patterns of eight isoenzymes. Information was obtained about the buoyant densities of n-DNA and k-DNA, and one strain was tested against monoclonal antibodies. The identity of both strains remains obscure.