Genetic variability of Mahanarva sp (Hemiptera: Cercopidae) collected from different sites in Brazil.

Spittlebugs are the leading cause of damage to tall grasses. Annual losses are estimated to reach 2.1 billion dollars in sugarcane crops and grazing land throughout the world. Correct identification of these species is difficult due to similarities in color, body size and male genitalia. Molecular markers have been useful in the identification and assessment of genetic diversity of many species. We investigated the genetic diversity of the spittlebug species Mahanarva fimbriolata, M. spectabilis and M. liturata and looked for markers that could aid in their identification. DNA from 34 spittlebug specimens, collected from six different regions of Brazil (Brasília, Campo Grande, Valença, Presidente Prudente, Juiz de Fora, and Porto Alegre), was analyzed with 29 RAPD primers, generating 501 polymorphic markers. High genetic variability was found among individuals M. fimbriolata (0.37), M. spectabilis (0.18) and M. liturata (0.69). Species-specific molecular RAPD markers were identified for each of the three species; these could be used as auxiliary tools for their correct identification.

Selenium and mercury in Brazilian Cerrado soils and their relationships with physical and chemical soil characteristics.

The objective of this study was to determine the natural concentrations of Hg and Se in 45 representative soil profiles from the Cerrado biome in central Brazil, and to correlate their concentrations with soil chemical and physical characteristics. The study area was composed of three sub-regions: Goiás, Northwest of Minas Gerais, and Minas Gerais Triangle. Selenium and Hg concentrations were determined by acid digestion and atomic absorption spectroscopy. Data were subjected to analysis of variance on the means of the Hg and Se variables within each soil class at two depths, followed by multivariate statistical methods. The Hg concentrations ranged from 15 to 182 µg kg-1 and the Se concentrations ranged from 22 to 72 µg kg-1. The soil characteristics that most contributed to Hg concentrations in the soils, according to principal component analysis, were Fe2O3, FeO, TiO2, pH, P2O5, and effective CEC. In general, the soils of the Cerrado biome have deficient Se concentrations. The Humic Rhodic Acrustoxes have Hg concentrations above the prevention reference value for soils of Minas Gerais.

Two New Species of the Spittlebug Genus Ocoaxo Fennah (Hemiptera: Cercopidae) from Mexico, and Keys for the Groups, Group Three, and First Subgroup.

Insects of the family Cercopidae are known as spittlebugs or froghoppers and are represented by 62 genera in the Neotropical region. One of these genera is Ocoaxo Fennah, 1968 with 30 species. The most recent species to be accepted into this genus, Ocoaxo costaricanus, was described by Nast (Ann Zool 33:93-101, 1975). Herein, two new species of Ocoaxo from Mexico are described. One of these new species forms a complex together with Ocoaxo assimilis (Walker) and Ocoaxo varians (Stål). The complex has economic importance in the mountainous areas of the states of Puebla and Oaxaca because it attacks Pinus spp. and causes a disorder called "pine decline." Additionally, dichotomous keys were designed to identify the Ocoaxo Fennah groups and also the species of the subgroup bivittus.

Critical mercury concentration in tropical soils: Impact on plants and soil biological attributes.

Mercury is a toxic element that becomes a problem when present at high concentrations in soils. Mercury toxicity in soils varies depending on chemical species, concentration, exposure routes, and organism vulnerability. There is little information regarding the toxicity of Hg in tropical soils, especially for establishing safe levels of this pollutant. The purpose of this study was to investigate Hg concentrations in two tropical soils and their effect on oats and common beans, as well as on soil biological attributes. The experiment was carried out in a greenhouse, following ISO 11.269-2 and OECD-208 guidelines. Oat and common bean were cultivated in a Typic Hapludox (TyHpx) and Rhodic Acrudox (RhAcx) contaminated with HgCl2 at the following concentrations: 0, 2.5, 5.0, 10.0, 20.0, 40.0, and 80.0 mg of Hg kg-1 of dry soil. The biological variables analyzed were seedling emergence, vegetative growth, chlorophyll content (SPAD index), gas exchange (photosynthetic rate, internal CO2 concentration, transpiration rate, and stomatal conductance), and Hg concentration and accumulation in shoot dry matter. Microbial biomass carbon, soil basal respiration, and metabolic quotient (qCO2) were also analyzed. Due to the sorptive characteristics of TyHpx, it had higher Hg concentrations than RhAcx. Mercury showed toxic effects on both oat and common bean species. However, common bean was affected only at concentrations higher than 20 mg kg-1. The microbial community showed high sensitivity to soil Hg concentrations, but external factors, such as the plant species cultivated, influenced the sensitivity of the community. The microbiota was most sensitive in pots with common bean, and this effect was more pronounced at low clay and low organic matter contents (TyHpx). In this study, the concentration of 0.36 mg kg-1 was critical for Hg in these soils, based on its deleterious effects on oat and common bean and on biological soil attributes.

Natural variation of selenium in Brazil nuts and soils from the Amazon region.

Brazil nut tree (Bertholletia excelsa) is native of the Amazon rainforest. Brazil nuts are consumed worldwide and are known as the richest food source of selenium (Se). Yet, the reasoning for such Se contents is not well stablished. We evaluated the variation in Se concentration of Brazil nuts from Brazilian Amazon basin, as well as soil properties, including total Se concentration, of the soils sampled directly underneath the trees crown, aiming to investigate which soil properties influence Se accumulation in the nuts. The median Se concentration in Brazil nuts varied from 2.07 mg kg-1 (in Mato Grosso state) to 68.15 mg kg-1 (in Amazonas state). Therefore, depending on its origin, a single Brazil nut could provide from 11% (in the Mato Grosso state) up to 288% (in the Amazonas state) of the daily Se requirement for an adult man (70 µg). The total Se concentration in the soil also varied considerably, ranging from <65.76 to 625.91 µg kg-1, with highest Se concentrations being observed in soil samples from the state of Amazonas. Se accumulation in Brazil nuts generally increased in soils with higher total Se content, but decreased under acidic conditions in the soil. This indicates that, besides total soil Se concentration, soil acidity plays a major role in Se uptake by Brazil nut trees, possibly due to the importance of this soil property to Se retention in the soil.

Distribution patterns of Neotropical primates (Platyrrhini) based on Parsimony Analysis of Endemicity.

The Parsimony Analysis of Endemicity (PAE) is a method of historical biogeography that is used for detecting and connecting areas of endemism. Based on data on the distribution of Neotropical primates, we constructed matrices using quadrats, interfluvial regions and pre-determinated areas of endemism described for avians as Operative Geographic Units (OGUs). We codified the absence of a species from an OGU as 0 (zero) and its presence as 1 (one). A hypothetical area with a complete absence of primate species was used as outgroup to root the trees. All three analyses resulted in similar groupings of areas of endemism, which match the distribution of biomes in the Neotropical region. One area includes Central America and the extreme Northwest of South America, other the Amazon basin, and another the Atlantic Forest, Caatinga, Cerrado and Chaco.

Detection of ecto-5'-nucleotidase on rat B and T lymphocyte subpopulations using a monoclonal antibody in rosetting reactions.

A mouse monoclonal antibody to rat 5'-nucleotidase (5N 4-2 McAb) was used in the direct anti-determinant rosetting reaction (DARR) to demonstrate the ecto-5'-nucleotidase molecule in preparations of rat lymphocytes. Results indicated that 35.5 +/- 7.5% of peripheral blood lymphocytes (PBL), 37.3 +/- 4.8% of lymph node cells (LN) and 37.0 +/- 8.5% of spleen lymphocytes expressed the 5N 4-2 antigen. Depletion studies and mixed rosetting reactions (MRR) showed that the 5N 4-2 antigen was mainly expressed on rat T lymphocytes rather than on B lymphocytes: In fact 59.6 +/- 3.2% (in PBL), 76.5 +/- 0.6% (in LN) and 67.1 +/- 1.3% (in spleen) of T lymphocytes exhibited the 5N 4-2 antigen compared to only 26.5 +/- 2.6% (in PBL), 34.0 +/- 2.1% (in LN) and 46.1 +/- 12.0% (in spleen) of B lymphocytes. As expected a strong association was found between the expression of 5N 4-2 antigen and 5'-nucleotidase enzyme activity on lymphocytes. Both 5N 4-2 positive cells and enzyme activity were preferentially exhibited in the T lymphocyte subpopulation, and 92% of the enzyme activity was observed in a 5N 4-2 antigen positive subpopulation.

Iron exerts a specific inhibitory effect on CD2 expression of human PBL.

The effect of in vitro exposure to ferric citrate (Fe-citrate) on the expression of human lymphocyte surface markers was studied. The following markers were examined: E-rosette formation, CD2, CD3, CD4, CD8, CD1, CD22, CD10 and HLA-DR. Pretreatment of peripheral blood lymphocytes (PBL) with Fe-citrate at concentrations ranging from 10(-2) M to 10(-5) M resulted in the exclusive inhibition of E-rosette formation and CD2 expression. None of the other surface antigens examined appeared to be sensitive to the Fe-citrate treatment. Competition experiments further indicated that iron interacts specifically with CD2 on T lymphocytes.

In vitro biomineralization by osteoblast-like cells. I. Retardation of tissue mineralization by metal salts.

The cytocompatibility of stainless steel 316L (SS 316L) corrosion products was investigated with particular focus on the dose- and time-effect of electrochemically dissolved SS and the corresponding separate metal ions on osteogenic bone marrow derived cells. Type AISI 316L stainless steel (Fe 63.9%, Cr 18.0%, Ni 12.5%, Mo 2.8%, Si 1.2%, Mn 1.6% and C 0.025%, weight for weight) was anodically dissolved in Hank's Balanced Salt Solution (HBSS) and diluted to the following concentrations: 500 microg ml(-1) of Fe, 122 microg ml(-1) of Cr and 101 microg ml(-1) of Ni, as estimated by atomic absorption spectrometry. Similarly, salt solutions containing 50 microg ml(-1) of Fe (FeCl3 x 6H2O), 122 microg ml(-1) of Cr (CrCl3 x 6H2O) or 101 microg ml(-1) of Ni (NiNO3) were prepared. All solutions were diluted 1:10(3), 1:10(4) and 1:10(5) and their effects on cell proliferation and function of rabbit bone marrow cells were studied up to 28 days of culture. Bone marrow cells (second subculture) were cultured in alpha-Minimal Essential Medium (alpha-MEM) supplemented with 10% fetal bovine serum 10(-8) mol l(-1) dexamethasone, 2.52 x 10(-4) mol l(-1) ascorbic acid and 10(-2) mol l(-1) beta-glycerophosphate. The osteoblast response to the presence of metal ions was evaluated by biochemical assays (enzymatic reduction of MTT for evaluation of cell viability/proliferation, and estimation of alkaline phosphatase (ALP) activity) and histochemical assays (identification of ALP positive cells and calcium and phosphates deposits). Results suggest a decrease in the expression of the osteoblast phenotype in the presence of ion and alloy solutions. Stainless steel corrosion products elicited slight effects but the corresponding metal ions produced pronounced effects on the osteoblast phenotype, namely an alteration in the levels and temporal expression of ALP and lower and retarded tissue mineralization ability.

In vitro biomineralization by osteoblast-like cells. II. Characterization of cellular culture supernatants.

The quantification of total calcium, phosphorus, iron, chromium and nickel in cell culture medium by electrochemical or spectroscopic means may require digestion of samples. Nevertheless, when pH adjustment is performed for values higher than about 6.5, the formation of two phases occurs: a white precipitate and a clear solution. Analysing both phases using microelectrodes, atomic absorption spectrometry (AAS), diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy, X-ray dispersive (XRD) analysis, scanning electron microscopy (SEM) and energy dispersive spectroscopic (EDS) analysis, it was observed that iron, chromium and nickel are not co-precipitating with the white solid phase. If quantification of calcium, phosphorus and magnesium is intended, a ten-fold dilution at least, must be performed to avoid most of these elements going into the precipitate. This knowledge is crucial if a mineralization study is going to be made.

Use of microelectrodes as electrochemical sensors of metal ions released from biomaterials.

In vitro studies to determine Fe3+ levels in mice liver samples were performed using platinum microelectrodes and atomic absorption spectrophotometry. Microelectrodes have been shown to be useful for quantitative analysis of metal ions released during the biodegradation process that occurs on implanted metallic biomaterials.

Effects of AISI 316L corrosion products in in vitro bone formation.

Rat bone marrow cells were cultured in experimental conditions that favour the proliferation and differentiation of osteoblastic cells (i.e., 2.52 x 10(-4) mol l(-1) ascorbic acid, 10(-2) mol l(-1) beta-glycerophosphate and 10(-8) mol l(-1) dexamethasone) in the absence and in the presence of stainless-steel corrosion products, for a period of 18 days. An AISI 316L stainless-steel slurry (SS) was obtained by electrochemical means and the concentrations of the major metal ions, determined by atomic absorption spectrometry, were 8.78 x 10(-3) mol l(-1) of Fe, 4.31 x 10(-3) mol l(-1) of Cr and 2.56 x 10(-3) mol l(-1) of Ni. Bone marrow cells were exposed to 0.01, 0.1 and 1% of the SS and at the end of the incubation period, control and treated cultures were evaluated by histochemical assays for the identification of the presence of alkaline phosphatase and also calcium and phosphate deposition. Cultures were further observed by scanning electron microscopy. Levels of total and ionised calcium and phosphorus in the culture media collected from control and metal exposed cell cultures were also quantified. Histochemical staining showed that control cultures presented a strong reaction for the presence of alkaline phosphatase and exhibited formation of calcium and phosphates deposits. The presence of 0.01% SS caused no detectable biological effects in these cultures, 0.1% SS impaired osteoblastic behaviour and, 1% SS resulted in cell death. In the absence of bone cells, levels of total and ionised calcium and phosphorus in the control and metal added culture medium were similar throughout the incubation period. A significant decrease in the levels of ionised calcium and phosphorus were observed in the culture medium of control cultures and also in cultures exposed to 0.01% SS after two weeks of incubation, an event related with the formation of mineral calcium phosphate deposits in these cultures. In cultures grown in the presence of 0.1 and 1% SS corrosion products, levels of calcium and phosphorus were similar to those observed in the absence of cells. Results showed that stainless-steel corrosion products above certain concentrations may disturb the normal behaviour of osteoblast-like rat bone marrow cell cultures.

Chromium determination in osteoblast-like cell culture medium by catalytic cathodic stripping voltammetry with a mercury microelectrode.

A catalytic cathodic stripping voltammetric procedure for the determination of total chromium in osteoblast-like cell culture medium using a mercury film microelectrode (MFM) was optimised. The method is based on the pre-concentration of the Cr(III)-diethylenetriaminepentaacetic acid (DTPA) complex by adsorption at the potential of-1.00 V (vs. Ag/AgC1) in the presence of 10 x 10(-3) mol/L DTPA, 0.70 mol/L sodium nitrate, 0.04 mol/L sodium acetate and 1.0 x 10(-3) mol/L potassium permanganate at pH 5.9-6.0. The limit of detection obtained for a 40 s collection time was 2.80 x 10(-10) mol/L of chromium. The results achieved by stripping voltammetry using the MFM were compared to those obtained by atomic absorption spectrometry (AAS) to ensure the reliability of the electrochemical method. This procedure proved to be an alternative to AAS and valuable in biocompatibility studies performed in vitro using osteoblast-like cells.

Phosphorus improves arsenic phytoremediation by Anadenanthera peregrina by alleviating induced oxidative stress.

Due to similarities in their chemical behaviors, studies examining interactions between arsenic (As)--in special arsenate--and phosphorus (P) are important for better understanding arsenate uptake, toxicity, and accumulation in plants. We evaluated the effects of phosphate addition on plant biomass and on arsenate and phosphate uptake by Anadenanthera peregrina, an important Brazilian savanna legume. Plants were grown for 35 days in substrates that received combinations of 0, 10, 50, and 100 mg kg(-1) arsenate and 0, 200, and 400 mg kg(-1) phosphate. The addition of P increased the arsenic-phytoremediation capacity of A. peregrina by increasing As accumulation, while also alleviating As-induced oxidative stress. Arsenate phytotoxicity in A. peregrina is due to lipid peroxidation, but not hydrogen peroxide accumulation. Added P also increased the activity of important reactive oxygen species-scavenging enzymes (catalase and ascorbate peroxidase) that help prevent lipid peroxidation in leaves. Our findings suggest that applying P represents a feasible strategy for more efficient As phytoremediation using A. peregrina.

Stimulation of human T lymphocytes via CD2 and CD3 molecules: establishment of the method and characterisation of proliferating cells.

The in vitro exposure of human lymphocytes to degradation products (Fe, Ni or Co) of metallic biomaterials causes a significant reduction of lymphocytes expressing the molecules involved in T lymphocyte activation, CD2 and CD3. A method is described which was developed for the stimulation of lymphocytes in which both CD2 and CD3 molecules were triggered simultaneously. For this purpose an anti-CD3 monoclonal antibody (mAb) was chemically bound to sheep (SE) or human (HE) erythrocytes, forming SE alpha CD3 or HE alpha CD3 conjugates, respectively, which were used for lymphocyte stimulation. Stimulation via CD2 and CD3 was compared with classical phytohaemagglutinin stimulation as well as with soluble alpha CD3 mAb stimulation. Phenotypical characterisation of DNA synthesising lymphocytes was assessed by autoradiography of 3H-thymidine pulsed cultures combined with immunocytochemical tests. Results indicate that this method of T lymphocyte stimulation via CD2 and CD3 is reliable and consistent, and it seems to be very convenient for the evaluation of immunocytotoxicity of metal ions derived from metallic biomaterials.

Mineralization in serially passaged human alveolar bone cells.

Well-characterized human bone cell cultures have been regarded as a useful tool to study bone control mechanisms and also to analyse bone/biomaterials interactions. In the present study, human alveolar bone cells were cultured in alpha-minimal essential medium (alpha-MEM) containing 10% foetal bovine serum (FBS), 50 microg/ml ascorbic acid, 10 mM sodium beta-glycerophosphate and either in the presence or in the absence of 10 nM dexamethasone (Dexa). Cultures were characterized concerning cell viability/proliferation, alkaline phosphatase (ALP), acid phosphatase (ACP) and tartraric acid resistant phosphatase (TRAP) activities, and formation of mineralized areas. Cell proliferation increased gradually for approximately 20 days. In the presence of Dexa, cells formed isolated or interconnected multilayered clusters that increased with culture time. Histochemical assays revealed strong positive reactions for ALP and calcium and phosphates deposits, mainly in relation t! o ce lls associated with the clusters. High levels of ALP activity (biochemical determination) were observed. Cells cultured in the absence of Dexa showed significantly lower ALP activity and no calcium and phosphates deposits were present. Serially passaged cells kept the proliferation rate constant but a decrease in ALP activity was observed either in the presence or in the absence of Dexa. The ability to form mineralized areas (cultures fed with Dexa) also decreased on serial subculture.

Differential effects of eight metal ions on lymphocyte differentiation antigens in vitro.

In vitro studies were conducted to determine the effects of metal ions known to be released from metallic implants in vivo on the expression of lymphocyte surface antigens. Normal human peripheral blood lymphocytes were exposed to various concentrations of metal ions (Fe3+, Ni2+, Co2+, Mo6+, V5+, Cr6+, Cr3+, and Ti3+) for 30 min at 37 degrees C in a 5% CO2 atmosphere, and then analyzed for their ability to form rosettes with sheep red blood cells. Following this preliminary analysis, lymphocytes were exposed to the metal ions found to inhibit the E-rosette reaction (Fe3+, Ni2+, and Co2+) in order to determine which of the following surface antigens were affected: CD2, CD3, CD4, CD8, CD1, CD22, CD10, and HLA-DR. Our results showed that the in vitro treatment of lymphocytes with Fe3+ or Co2+ caused inhibition of CD2 only, whereas Ni2+ caused inhibition of both CD2 and CD3 antigens. These findings suggest that Fe3+, Co2+, and Ni2+ ions may interfere with T cell activation since both CD2 and CD3 are involved in that process.

Toxic effects caused by stainless steel corrosion products on mouse seminiferous cells.

Stainless steel type AISI 316L is widely used in orthopaedic surgery. The toxic effects of its corrosion products on the morphology of mouse seminiferous cells were investigated. Chemical elements from stainless steel were released into a physiological medium using an electrochemical method. This metallic solution was injected subcutaneously into male Charles River mice at 72 h intervals for 10 days. Electron microscopic observations of seminiferous tubule thin sections showed that the metallic suspension caused tissue vacuolation, cell degeneration, and multinucleated cell formation. This apparent tissue toxicity induced by stainless steel corrosion products suggests that long term implantation of such biomaterials may impair spermatogenesis.

The use of rat, rabbit or human bone marrow derived cells for cytocompatibility evaluation of metallic elements.

Rat, rabbit and human bone marrow cells were cultured according to the method previously reported for cells of rat origin [1] and were exposed, or not (control), to corrosion products of a Co-Cr orthopaedic alloy as well as to metal salts containing Co2+, Cr3+ and Cr6+. Cells were cultured for 21 days and analysed for the following biochemical parameters: intracellular MTT reduction (i.e. cell viability/proliferation), alkaline phosphatase (ALP) activity and protein production. Morphological observations included both histochemistry (detection of ALP-positive cells, calcium and phosphate deposits) and scanning electron microscopy (SEM). Control cultures of rat and rabbit cells showed higher proliferation rates than human cells at the start of culture, but they all reached similar values on day 21. Protein production was parallel to cell proliferation. In contrast, ALP activity of rat cultures was much stronger than rabbit or human cultures. All cell types were able to develop the osteogenic phenotype in vitro.Co-Cr extract caused inhibitory effects on cell viability, on ALP activity and, to a lower extent, on protein production of all rat, rabbit and human cell cultures. Compared to rat and rabbit cultures, human cultures were the most sensitive to metal ions exposure.

Immune responses in cardiac transplantation. I. Detection of activated TIa+ cells in the blood during herpes virus infections.

Lymphocyte subpopulations in the blood of cardiac transplant recipients were monitored by single and double marker rosetting tests; using monoclonal antibodies to monomorphic determinants on T cells and 'Ia' antigens. Elevated absolute numbers and percentages of TIa+ cells were found in association with primary cytomegalovirus (CMV), and reactivation of Epstein-Barr virus or Herpes simplex virus. Serial tests showed that in primary CMV TIa+ cells peaked before the maximal IgM and IgG anti-viral titres measured by ELISA. Infection related antiglobulin levels increased in parallel with anti-viral IgM in primary CMV infections. Intravenous methylprednisone and blood transfusions selectively depressed TIa+ cell levels without affecting antibody titres. These results show that patients on maintenance immunosuppression of cyclosporin A and steroids can successfully mount T cell and antibody responses to herpes virus infections.