Diversity of Trypanosoma cruzi stocks and clones derived from Chagas disease patients: I-Behavioral characterization in vitro

In this study, we isolated Trypanosoma cruzi from chronic Chagas heart disease and from megaesophagus patients. The parasite stock hSLU239 (heart disease) yielded clones h1 and h2, whereas stock mSLU142 (megaesophagus) yielded clones m1, m2, m3 and m4. The parasite growth kinetics, doubling time and differentiation in axenic liquid medium showed broad behavioral diversity. It was shown that a particular pattern of behavior for a parental stock could not necessarily be assigned for subsequent clones. This study indicates that i) each Chagas disease patient is infected with several T. cruzi populations; ii) clonal lines derived from patient samples may have different biological characteristics from the original isolate; and that iii) additional behavioral and/or molecular markers are required for further characterization of Trypanosoma cruzi stocks and clones derived from Chagas disease patients in order to identify correlations with pathology.

Local diversity of tropical and temperature ant faunas (Hymenoptera, Formicidae)

Qualitative and quantitative collections of ants made in the region of Manaus, Brazil (evergreen tropical humid forest), and in western North Carolina, USA (deciduous temperate/wet forest), were undertaken to investigate. latitudinal patterns of ant diversity and community organization on regional and local scales. We have found to date 307 ant species in the Municipality of Manaus. Totals ranging from 134 to 270+ species have been reported in the literature for other tropical regions of less than 10,000km2. In contrast, temperate ant surveys generally report only SO to 150 species in similar or larger areas. Sampling at sardine baits set 10m apart on square grids, we found forest ecosystems near Manaus to be much richer and more diverse in ants than those sampled in North Carolina: 28 species vs. 5-10 species in 50 collections and 16 vs. 3 previously unrecorded species discovered with each doubling of sample size. Room's (1975a) results from climatically simllar Papua New Guinea forest agree closely with those from Manaus. We suggest that one important factor contributing to the increased diversity of tropical, omnivorous ants may be greater variety of nest sites available for specialization.

Sequencing and identification of expressed Schistosoma mansoni genes by random selection of cDNA clones from a directional library

We have initiated a gene discovery program in Schistosoma mansoni based on the technique of Expressed Sequence Tags (ESTs), i.e. partial sequences of cDNAs obtained from single passes in automatic DNA sequencers. ESTs can be used to identify genese onf the basis of their homology whith sequences from other species deposited in DNA or protein databases. Trasncripts with sequences without matches in teh databases may represent novel parasite-specific genes. This approach has shown to be very efficient and in less than two years a broad range of novel genes has already been ascertained, more than doubling the number of known S. mansoni genes.

Biological Parameters and Molecular Markers of Clone CL Brener - The Reference Organism of the Trypanosoma cruzi Genome Project

Clone CL Brener is the reference organism used in the Trypanosoma cruzi Genome Project. Some biological parameters of CL Brener were determined: (a) the doubling time of epimastigote forms cultured in liver infusion-tryptose (LIT) medium at 28oC is 58±13 hr; (b) differentiation of epimastigotes to metacyclic trypomastigotes is obtained by incubation in LIT-20% Grace´s medium; (c) trypomastigotes infect mammalian cultured cells and perform the complete intracellular cycle at 33 and 37oC; (d) blood forms are highly infective to mice; (e) blood forms are susceptible to nifurtimox and benznidazole. The molecular typing of CL Brener has been determined: (a) isoenzymatic profiles are characteristic of zymodeme ZB; (b) PCR amplification of a 24Sa ribosomal RNA sequence indicates it belongs to T. cruzi lineage 1; (c) schizodeme, randomly amplified polymorphic DNA (RAPD) and DNA fingerprinting analyses were performed

Establishment and characterization of a new continuous cell line from Lutzomyia longipalpis (Diptera: Psychodidae) and its susceptibility to infections with arboviruses and Leishmania chagasi

Embryonic tissue explants of the sand fly Lutzomyia longipalpis (Lutz & Neiva 1912) the main vector of Leishmania chagasi (Cunha and Chagas), were used to obtain a continuous cell line (Lulo). The tissues were seeded in MM/VP12 medium and these were incubated at 28ºC. The first subculture was obtained 45 days after explanting and 96 passages have been made to date. Lulo is composed of epithelioid cells, showed a 0.04 generations/hour exponential growth rate and population doubling time at 24.7 h. The cell line isoenzymatic profiles were determined by using PGI, PGM, MPI and 6-PGDH systems, coinciding with patterns obtained from the same species and colony's pupae and adults. The species karyotype characteristics were recognized (2n = 8), in which pair 1 is subtelocentric and pairs 2, 3 and 4 are metacentric. Lulo was free from bacterial, fungal, mycoplasmic and viral infection. Susceptibility to five arbovirus was determined, the same as Lulo interaction with Leishmania promastigotes.

Reproduction and fertility life table of three aphid species (Macrosiphini) at different temperatures

Temperature is a key abiotic factor influencing the development and reproduction of aphids. The effect of temperature on the reproduction of three aphid species Aulacorthum solani (Kaltenbach), Macrosiphum euphorbiae (Thomas) and Uroleucon ambrosiae (Thomas) (Aphididae, Macrosiphini) has been investigated and fertility life tables were determined. Nymphs were reared in climatic chambers at temperatures of 16, 19, 22, 25, and 28 ± 1ºC, RH 70 ± 10% and 12 h photophase. Female adult aphids developed at these temperatures were then used in experiments in which pre-reproductive and reproductive periods were evaluated every 24 h. In addition, the number of nymphs produced and longevity were determined at each temperature. The reproduction period of A. solani and M. euphorbiae decreased with increasing temperature, whereas that of U. ambrosiae was maintained between 19 and 25ºC. The total number of nymphs produced by the aphids decreased as the temperature increased. The longevities of A. solani and M. euphorbiae decreased with increasing temperature but remained stable for U. ambrosiae between 19 and 25ºC. The largest survival rate (l x) and specific fertility (m x) values were found at 16 and 22ºC for all three species. The most favourable temperature for reproduction of A. solani, M. euphorbiae and U. ambrosiae was 22ºC, as demonstrated by the l x and m x profiles, the high values of net reproductive rates and intrinsic rates of increase, and the short intervals between generation and doubling times.

Produtividade simulada de tubérculos de batata em cenários de mudanças climáticas

The objective of this study was to simulate potato (Solanum tuberosum) tuber yield in different climate change scenarios of increased carbon dioxide concentration [CO2] and air temperature, considering symmetric and asymmetric increases in minimum and maximum daily air temperatures. Spitters model was used to simulate Asterix cultivar tuber yield considering two growing seasons (spring and fall) recommended for Santa Maria, state of Rio Grande do Sul, Brazil. In each growing season, five planting dates were evaluated in climate scenarios of a hundred years with no increase in [CO2] and temperature (current scenario), and in scenarios with doubling [CO2] and temperature increases of 0, 1, 2, 3, 4, 5 and 6ºC. A symmetric increase of 4ºC and an asymmetric increase of 5ºC in air temperature offset the yield beneficial effect of increasing [CO2] during spring, whereas increase in air temperature does not affect potato tuber yield during fall. Anticipating planting date in spring and delaying it in fall decrease the negative impact of the increasing air temperature on potato tuber yield.

Íon Be H+ em campo elétrico uniforme intenso

The influence of a strong external electric field in chemical bonding is not extensively studied. In this work, the behavior of the potential energy curves of the BeH+ ion in an external electric field is investigated by means of variational calculations using molecular orbitals. For simplicity we consider the case where the external field is parallel to the molecular axis, in the direction of the dipole moment, which raises the polarization of the ion. The effects on dissociation are investigated.

Backpressure effects on the flow-pressure relation of driplines

ABSTRACT For drip irrigation design and management, it is necessary to know the relation between flow and pressure acting on emitters. In the case of subsurface drip irrigation, the backpressure phenomenon may change the hydraulic characteristics of emitters. Thus, this study aimed at determining such relationship between flow and pressure of different driplines in surface and subsurface conditions; aiming to find possible differences in hydraulic behavior. We tested four emitter types; two pressure compensating (D5000 and Hydro PCND) and two non-pressure compensating (TalDrip and Jardiline). Emitter flow rates were attained in atmospheric conditions and submerged in water, in which submergence levels represented backpressure. Assays were performed using inlet pressures of 80, 100, 120, and 150 kPa for the Hydro PCND dripline and 25, 50, 100, and 150 kPa for the other ones; the backpressures were of 0.49, 1.47, 2.45, 4.41 and 6.37 kPa with four replications. The emitters had their proportionality constants and discharge exponents changed in submerged applications, representing backpressure effect. Non-pressure compensating emitters had their discharge exponent decreased, while in pressure compensating ones, it was increased. Backpressure reduced emitter flow rates at all evaluated pressures.

A new approach to evaluate imperfection sensitivity in asymmetric bifurcation buckling analysis

A direct procedure for the evaluation of imperfection sensitivity in bifurcation problems is presented. The problems arise in the context of the general theory of elastic stability for discrete structural systems, in which the energy criterion of stability of structures and the total potential energy formulation are employed. In cases of bifurcation buckling the sensitivity of the critical load with respect to an imperfection parameter e is singular at the state given by epsilon =0, so that, a regular perturbation expansion of the solution is not possible. In this work we describe a direct procedure to obtain the relations between the critical loads, the generalized coordinates at the critical state, the eigenvector, and the amplitude of the imperfection, using singular perturbation analysis. The expansions are assumed in terms of arbitrary powers of the imperfection parameter, so that both exponents and coefficients of the expansion are unknown. The solution of the series exponents is obtained by searching the least degenerate solution. The formulation is here applied to asymmetric bifurcations, for which explicit expressions of the coefficients are obtained. The use of the method is illustrated by a simple example, which allows consideration of the main features of the formulation.

Effect of ventilation on systemic blood flow evaluated by echodopplercardiography

Systemic blood flow (Q) was measured by echodopplercardiography in 5 normal young adult males during apnea, eupnea and tachypnea. Measurements were made in a recumbent posture at 3-min intervals. Tachypnea was attained by doubling the respiratory frequency at eupnea at a constant tidal volume. An open glottis was maintained during apnea at the resting respiratory level. The Q values were positively correlated with the respiratory frequency, decreasing from eupnea to apnea and increasing from eupnea to tachypnea (P<0.05). These data demonstrate that echodopplercardiography, a better qualified tool for this purpose, confirms the positive and progressive effects of ventilation on systemic blood flow, as suggested by previous studies based on diverse technical approaches

Large-scale production and purification of recombinant protein from an insect cell/baculovirus system in Erlenmeyer flasks: application to the chicken poly(ADP-ribose) polymerase catalytic domain

A simple and inexpensive shaker/Erlenmeyer flask system for large-scale cultivation of insect cells is described and compared to a commercial spinner system. On the basis of maximum cell density, average population doubling time and overproduction of recombinant protein, a better result was obtained with a simpler and less expensive bioreactor consisting of Erlenmeyer flasks and an ordinary shaker waterbath. Routinely, about 90 mg of pure poly(ADP-ribose) polymerase catalytic domain was obtained for a total of 3 x 109 infected cells in three liters of culture

Hyperthermia increases the metastatic potential of murine melanoma

Hyperthermia, either alone or combined with radio-, immuno- or chemotherapy, can control tumor growth, but its effect on metastasis is still controversial. In the present study, we investigated the influence of hyperthermia on the metastatic potential of B16-F10 murine melanoma cells. Incubation of melanoma cells at 43ºC for 30 min led to a significant decrease in cell viability. About half of the cells survived the acute exposure to heat. These thermoresistant cells displayed a longer lag phase as compared to control unheated B16-F10 melanoma cells. Other parameters of cell growth such as doubling time and saturation density were equivalent in both control and thermoresistant cells. Both control and treated cells were adherent, but thermoresistant cells failed to spread during the first 48 h after heat exposure. B16-F10 cells colonize the lungs of C57BL/6J mice when injected intravenously; the number of lung colonies is a measure of the metastatic potential of injected cells. Median values of 22, 10.5 and 31 colonies per injected mouse were observed for control cells, cells heated to 43ºC for 30 min and thermoresistant cells, respectively, with statistically significant differences between groups (Mann-Whitney test, P<0.02). Thus, despite its cytotoxic action, heat exposure induced the acquisition of a more metastatic phenotype in a subpopulation of B16-F10 cells

Differential regulation of vitamin D receptor expression in distinct leukemic cell lines upon phorbol ester-induced growth arrest

A close correlation between vitamin D receptor (VDR) abundance and cell proliferation rate has been shown in NIH-3T3 fibroblasts, MCF-7 breast cancer and in HL-60 myeloblastic cells. We have now determined if this association occurs in other leukemic cell lines, U937 and K562, and if VDR content is related to c-myc expression, which is also linked to cell growth state. Upon phorbol myristate acetate (PMA) treatment, cells from the three lineages (HL-60, U937 and K562) differentiated and expressed specific surface antigens. All cell lines analyzed were growth inhibited by PMA and the doubling time was increased, mainly due to an increased fraction of cells in the G0/G1 phase, as determined by flow cytometry measurements of incorporated bromodeoxyuridine and cell DNA content. C-myc mRNA expression was down-regulated and closely correlated to cell growth arrest. However, VDR expression in leukemic cell lines, as determined by immunofluorescence and Northern blot assays, was not consistently changed upon inhibition of cell proliferation since VDR levels were down-regulated only in HL-60 cells. Our data suggest that VDR expression cannot be explained simply as a reflection of the leukemic cell growth state.

Fludarabine induces apoptosis in chronic lymphocytic leukemia - the role of P53, Bcl-2, Bax, Mcl-1, and Bag-1 proteins

The expression of P53, Bcl-2, Bax, Bag-1, and Mcl-1 proteins in CD5/CD20-positive B-chronic lymphocytic leukemia (B-CLL) cells from 30 typical CLL patients was evaluated before and after 48 h of incubation with 10-6 M fludarabine using multiparametric flow cytometric analysis. Protein expression was correlated with annexin V expression, Rai modified clinical staging, lymphocyte doubling time, and previous treatment. Our main goal was to determine the predictive value of these proteins in CLL cells in terms of disease evolution. Bcl-2 expression decreased from a median fluorescence index (MFI) of 331.71 ± 42.2 to 245.81 ± 52.2 (P < 0.001) after fludarabine treatment, but there was no difference between viable cells (331.57 ± 44.6 MFI) and apoptotic cells (331.71 ± 42.2 MFI) before incubation (P = 0.859). Bax expression was higher in viable cells (156.24 ± 32.2 MFI) than in apoptotic cells (133.56 ± 35.7 MFI) before incubation, probably reflecting defective apoptosis in CLL (P = 0.001). Mcl-1 expression was increased in fludarabine-resistant cells and seemed to be a remarkable protein for the inhibition of the apoptotic process in CLL (from 233.59 ± 29.8 to 252.04 ± 35.5; P = 0.033). After fludarabine treatment, Bag-1 expression was increased in fludarabine-resistant cells (from 425.55 ± 39.3 to 447.49 ± 34.5 MFI, P = 0.012), and interestingly, this higher expression occurred in patients who had a short lymphocyte doubling time (P = 0.022). Therefore, we could assume that Bag-1 expression in such situation might identify CLL patients who will need treatment earlier.