Antifungal activity from Ocimum gratissimum L. towards Cryptococcus neoformans

Cryptococcal infection had an increased incidence in last years due to the explosion of acquired immune deficiency syndrome epidemic and by using new and effective immunosuppressive agents. The currently antifungal therapies used such as amphotericin B, fluconazole, and itraconazole have certain limitations due to side effects and emergence of resistant strains. So, a permanent search to find new drugs for cryptococcosis treatment is essential. Ocimum gratissimum, plant known as alfavaca (Labiatae family), has been reported earlier with in vitro activity against some bacteria and dermatophytes. In our work, we study the in vitro activity of the ethanolic crude extract, ethyl acetate, hexane, and chloroformic fractions, essential oil, and eugenol of O. gratissimum using an agar dilution susceptibility method towards 25 isolates of Cryptococcus neoformans. All the extracts of O. gratissimum studied showed activity in vitro towards C. neoformans. Based on the minimal inhibitory concentration values the most significant results were obtained with chloroformic fraction and eugenol. It was observed that chloroformic fraction inhibited 23 isolates (92%) of C. neoformans at a concentration of 62.5 microg/ml and eugenol inhibited 4 isolates (16%) at a concentration of 0.9 microg/ml. This screening may be the basis for the study of O. gratissimum as a possible antifungal agent.     

Detection of hepatitis C virus RNA by in situ hybridization in paraformaldehyde fixed biopsies

Fourteen hepatitis C virus (HCV) chronically infected patients were submitted to routine liver biopsy for histological evaluation. Liver samples were assayed to HCV-RNA by in situ hybridization, using digoxigenin labeled probe. HCV genotypes were found to be predominantly type 1 (71.4%), followed by genotype 3 (21.4%), and genotype 2 (7.2%). Alanine-aminotransferase levels were raised in 10 patients. The histopathological scores were minimal (21.4%), mild (57.2%), and moderate (21.4%). Viral RNA was detected in liver cells from nine patients (64.3%). ISH method provides localization and poor confirmation of HCV RNA in the liver tissue of HCV chronic patients.     

Peridomiciliary colonies of Triatoma vitticeps (Stal, 1859) (Hemiptera, Reduviidae, Triatominae) infected with Trypanosoma cruzi in rural areas of the state of Espírito Santo, Brazil

In Brazil, the colonization of human dwellings by triatomines occurs in areas with native vegetation of the caatinga or cerrado types. In areas of Atlantic forest such as in the Brazilian state of Espírito Santo, there are no species adapted to live in human habitations. The few autochthonous cases of Chagas disease encountered in Espírito Santo have been attributed to adult specimens of Triatoma vitticeps that invade houses from forest remnants. In recent years, the entomology unit of the Espírito Santo State Health Secretariat has recorded nymphs infected with flagellates similar to Trypanosoma cruzi in rural localities. Entomological surveys were carried out in the residences and outbuildings in which the insects were found, and serological examinations for Chagas disease performed on the inhabitants. Four colonies were found, all associated with nests of opossums (Didelphis aurita), 111 specimens of T. vitticeps, and 159 eggs being collected. All the triatomines presented flagellates in their frass. Mice inoculated with the faeces presented trypomastigotes in the circulating blood and groups of amastigotes in the cardiac muscle fibres. Serological tests performed on the inhabitants were negative for T. cruzi. Even with the intense devastation of the forest in Espírito Santo, there are no indications of change in the sylvatic habits of T. vitticeps. Colonies of this insect associated with opossum nests would indicate an expansion of the sylvatic environment into the peridomicile.     

Panleucogating as an accurate and affordable flow cytometric protocol to analyse lymphocyte subsets among HIV-positive patients on HAART treatment in Mozambique

In Africa tens of millions of people are HIV+. Prevention alone is not effective, and needs to be coupled with anti-retroviral treatment (HAART). Laboratory tests as CD4+ T cell count are fundamental tools in HIV disease monitoring, but they require costly equipment, reagents and specialised manpower. The goal of this study was to minimise and optimise the reagents needed for a reliable routine CD4+ cell count in a resource-poor setting (Mozambique). Panleucogating protocol (PLG), requires two antibodies only, CD45 and CD4, or three if CD8 is requested for special clinical reasons. PLG was compared with the current protocol used in two Mozambique hospitals, based on FSC/SSC gating and CD3/CD4/CD8 staining. 189 samples from HIV+ patients, included in the Community of Sant'Egidio's DREAM program and on HAART were processed with both protocols. The overall correlation of the lymphocyte subsets measurements was satisfactory, with r2 always >0.96. The Bland-Altman analysis of CD4+ cell count showed a negative bias when CD4+ cells were <15%, due to the imprecise FSC/SSC gating used previously. When CD4+ cells were >15% the negative bias tended to zero, further confirming the better quality of the PLG gating strategy. Two- or three color PLG protocol, in double platform, currently seems the most accurate and affordable method to monitor CD4+ lymphocytes and CD4/CD8 ratio by flow cytometry in resource-poor medical settings.     

The Africanized honey bee dispersal: A mathematical zoom

A general mathematical model for population dispersal featuring long range taxis is presented and exemplified by the dispersal episode of the Africanized honey bees (Apis mellifera adansonii) throughout the American Continent. The mathematical model is a discrete-time and nonlocal model represented by an integrodifference recursion. A newtaxis concept is defined and introduced into the mathematical model by an appropriate modification of the redistribution kernel. The model is capable of predicting the natural barrier for the expansion of the Africanized honey bees in the southern part of the Continent due to low winter temperatures. It also describes a sensitive expansion velocity with respect to the quality of resources, which can explain the AHB’s astounding spread rate, by using two different kinds of population dynamics strategies, one for a resourceful environment and the other for poor regions. An erratum to this article is available at .     

Drosophila wing-spot test for genotoxic assessment of pollutants in water samples from urban and industrial origin

The Caí River (Rio Grande do Sul, Brazil) is an important watercourse that receives large amounts of industrial and untreated municipal discharges in its lower course. We employed the SMART in Drosophila melanogaster to evaluate the genotoxicity of surface waters collected from Caí sites receiving direct sewage discharge: from Montenegro (Km 52) and from S?o Sebasti?o do Caí (Km 78 and 80), and from two sites under the industrial influence (Km 13.6 and 18.6). The genotoxic analysis included three collections: March, June and September 1999, which were tested at crude sample and at 50 and 25% concentrations. Considering the industrial samples from Km 18.6 and 13.6, collected in March, June and September 1999, they were characterized as not having genetic toxicity. The urban samples collected in March--Km 52, 78 and 80--showed a significant increment in the frequencies of total spots. In Km 52 and 78 the genotoxic effect was associated to both mutational and recombinational events, although for Km 80 the increases observed were mainly related to the occurrence of homologous recombination. Moreover, the Km 80 crude sample from June and all the concentrations analyzed for Km 52 in September were also able to induce mitotic recombination. These effects were only observed in the ST cross, demonstrating the genotoxins present in the urban discharges act by direct interaction with the DNA of the somatic cells. The SMART in D. melanogaster was shown to be highly sensitive to detect genotoxic agents present in the aquatic environment, and must be better exploited for monitoring areas under anthropogenic discharges.     

Filaments of the Ure2p prion protein have a cross-beta core structure

Formation of filaments by the Ure2 protein constitutes the molecular mechanism of the [URE3] prion in yeast. According to the "amyloid backbone" model, the N-terminal asparagine-rich domains of Ure2p polymerize to form an amyloid core fibril that is surrounded by C-terminal domains in their native conformation. Protease resistance and Congo Red binding as well as beta-sheet content detected by spectroscopy-all markers for amyloid-have supported this model, as has the close resemblance between 40 A N-domain fibrils and the fibrillar core of intact Ure2p filaments visualized by cryo-electron microscopy and scanning transmission electron microscopy. Here, we present electron diffraction and X-ray diffraction data from filaments of Ure2p, of N-domains alone, of fragments thereof, and of an N-domain-containing fusion protein that demonstrate in each case the 4.7A reflection that is typical for cross-beta structure and highly indicative of amyloid. This reflection was observed for specimens prepared by air-drying with and without sucrose embedding. To confirm that the corresponding structure is not an artifact of air-drying, the reflection was also demonstrated for specimens preserved in vitreous ice. Local area electron diffraction and X-ray diffraction from partially aligned specimens showed that the 4.7A reflection is meridional and therefore the underlying structure is cross-beta.     

In silco mapping of ESTs from the turkey (Meleagris gallopavo)

Sequence similarity was used to predict the position of expressed sequence tags (ESTs) in the genome of the turkey (Meleagris gallopavo). Turkey EST sequences were compared with the draft assembly of the chicken whole-genome sequence and the chicken EST database by BLASTN. Among the 877 ESTs examined, 788 had significant matches in the chicken genome sequence. Position of orthologous sequences in the chicken genome and the predicted position of the EST loci in the turkey genome are presented Genetic assignments suggest a high level of accuracy for the COMPASS predictions.     

P-type proton ATPases are involved in intracellular calcium and proton uptake in the plant parasite Phytomonas francai

The use of digitonin to permeabilize the plasma membrane of promastigotes of Phytomonas francai allowed the identification of two non-mitochondrial Ca(2+) compartments; one sensitive to ionomycin and vanadate (neutral or alkaline), possibly the endoplasmic reticulum, and another sensitive to the combination of nigericin plus ionomycin (acidic), possibly the acidocalcisomes. A P-type (phospho-intermediate form) Ca(2+)-ATPase activity was found to be responsible for intracellular Ca(2+) transport in these cells, with no evidence of a mitochondrial Ca(2+) transport activity. ATP-driven acidification of internal compartments in cell lysates and cells mechanically permeabilized was assayed spectrophotometrically with acridine orange. This activity was inhibited by low concentrations of vanadate and digitonin, was insensitive to bafilomycin A(1), and stimulated by Na(+) ions. Taken together, our results indicate that P-type ATPases are involved in intracellular Ca(2+) and H(+) transport in promastigotes of P. francai.