Aspidosperma (Apocynaceae) plant cytotoxicity and activity towards malaria parasites. Part II: experimental studies withAspidosperma ramiflorum in vivo and in vitro

Several species of Aspidosperma plants are used to treat diseases in the tropics, including Aspidosperma ramiflorum, which acts against leishmaniasis, an activity that is experimentally confirmed. The species, known as guatambu-yellow, yellowperoba, coffee-peroba andmatiambu, grows in the Atlantic Forest of Brazil in the South to the Southeast regions. Through a guided biofractionation of A. ramiflorum extracts, the plant activity against Plasmodium falciparum was evaluated in vitro for toxicity towards human hepatoma G2 cells, normal monkey kidney cells and nonimmortalised human monocytes isolated from peripheral blood. Six of the seven extracts tested were active at low doses (half-maximal drug inhibitory concentration < 3.8 µg/mL); the aqueous extract was inactive. Overall, the plant extracts and the purified compounds displayed low toxicity in vitro. A nonsoluble extract fraction and one purified alkaloid isositsirikine (compound 5) displayed high selectivity indexes (SI) (= 56 and 113, respectively), whereas compounds 2 and 3 were toxic (SI < 10). The structure, activity and low toxicity of isositsirikine in vitro are described here for the first time in A. ramiflorum, but only the neutral and precipitate plant fractions were tested for activity, which caused up to 53% parasitaemia inhibition of Plasmodium bergheiin mice with blood-induced malaria. This plant species is likely to be useful in the further development of an antimalarial drug, but its pharmacological evaluation is still required.

Experimental studies on Trypanosoma (Schizotrypanum) cruzi strains isolated from man, from animals and from triatomine bugs in Brazil

A study on nine strains of T. cruzi isolated from man, from animais and from triatomine bugs in Brazil are described. The parasites were slightly viscerotropic in white mice in six of the strains, highly viscerotropic and cardiotropic in two strains, and asymptomatic on one strain. Mechanical and cyclical passage from infected to healthy mice, and treatment of the infected mice with immunosuppressant drug, did not increase the blood parasitaemia or strain virulence. The results of biometric studies on the blood trypanosomes from each strain are also described. The various aspects on the importance of T. cruzi strain Identification are emphasized and discussed.

Generalization of temporal order detection skill learning: two experimental studies of children with dyslexia

The objective of this study was to investigate the phenomenon of learning generalization of a specific skill of auditory temporal processing (temporal order detection) in children with dyslexia. The frequency order discrimination task was applied to children with dyslexia and its effect after training was analyzed in the same trained task and in a different task (duration order discrimination) involving the temporal order discrimination too. During study 1, one group of subjects with dyslexia (N = 12; mean age = 10.9 ± 1.4 years) was trained and compared to a group of untrained dyslexic children (N = 28; mean age = 10.4 ± 2.1 years). In study 2, the performance of a trained dyslexic group (N = 18; mean age = 10.1 ± 2.1 years) was compared at three different times: 2 months before training, at the beginning of training, and at the end of training. Training was carried out for 2 months using a computer program responsible for training frequency ordering skill. In study 1, the trained group showed significant improvement after training only for frequency ordering task compared to the untrained group (P < 0.001). In study 2, the children showed improvement in the last interval in both frequency ordering (P < 0.001) and duration ordering (P = 0.01) tasks. These results showed differences regarding the presence of learning generalization of temporal order detection, since there was generalization of learning in only one of the studies. The presence of methodological differences between the studies, as well as the relationship between trained task and evaluated tasks, are discussed.

Recent studies on Schistosoma intercalatum: taxonomic status, puzzling distribution and transmission foci revisited

Schistosoma intercalatum, which causes human rectal schistosomiasis in Africa, still presents a great interest for its imprecise taxonomic status and its puzzling distribution in Africa. Two geographically isolated strains of S. intercalatum are recognized, the Lower Guinea strain and the Congo strain, which differ from each other in a number of morphological, biological and biochemical characteristics. Recent molecular data using RAPD markers indicate high divergence between the two strains, with values of Nei and Li's similarity indice allowing recognition of two genetically distinct taxa: experiments on pre- and post-isolating mechanisms are in progress in order to re-evaluate the taxonomic status of this polytypic species. With regard to its geographical distribution, S. intercalatum is characterized by the existence of two stable endemic areas (localized in Lower Guinea and North East of Democratic Republic of Congo) which correspond to the historical areas of species discovery, and the emergence during the last 15 years of new foci of the Lower Guinea strain outside previously known endemic areas. The absence of local adaptation of the Lower Guinea strain to its intermediate host, supported by experimental studies, may help to facilitate the spread of this strain. Nevertheless, the present restricted distribution of this species remains puzzling, because its potential snail hosts (bulinids) are widely distributed throughout much of Africa. Recent experimental and epidemiological studies suggest that interspecific sexual interactions between human schistosomes could have a role in limiting the distribution of S. intercalatum: the competitive sexual processes acting among human schistosomes show that S. haematobium and S. mansoni are always competitively dominant over S. intercalatum. These epidemiological observations lead the authors to distinguish three kinds of transmission foci for S. intercalatum.

The use of non-human primates as animal models for the study of hepatitis viruses

Hepatitis viruses belong to different families and have in common a striking hepatotropism and restrictions for propagation in cell culture. The transmissibility of hepatitis is in great part limited to non-human primates. Enterically transmitted hepatitis viruses (hepatitis A virus and hepatitis E virus) can induce hepatitis in a number of Old World and New World monkey species, while the host range of non-human primates susceptible to hepatitis viruses transmitted by the parenteral route (hepatitis B virus, hepatitis C virus and hepatitis delta virus) is restricted to few species of Old World monkeys, especially the chimpanzee. Experimental studies on non-human primates have provided an invaluable source of information regarding the biology and pathogenesis of these viruses, and represent a still indispensable tool for vaccine and drug testing.

Seroprevalence of human parvovirus B19 in a suburban population in São Paulo, Brazil

OBJECTIVE: To analyze the prevalence of IgG antibodies to human parvovirus B19. METHODS: Cross-sectional study in a suburban community in São Paulo, Southeastern Brazil, between November 1990 and January 1991. Randomly selected (N=435) representative samples of sera were collected from healthy children older than 15 days old and adults up to 40 years old. IgG antibodies were detected using ELISA. RESULTS: High prevalence of IgG antibodies to B19 parvovirus was found in 87% of newborns. The prevalence of maternally derived IgG antibodies exponentially plunged up to the 19th month of age. Low prevalence of antibodies was found in the first 4 years of life, increasing up to 72% in those aged 31-40 years. It was estimated that the average age of first infection in this population is 21 ± 7 years old and the optimal age for vaccination with a hypothetical vaccine would be 1 year of age. CONCLUSIONS: Parvovirus B19 IgG antibody prevalence was high in newborns and those aged 31-40 years. The analysis by age groups showed a pattern similar to that found in previous studies, i.e., low prevalence of infection in children that increases with age.

The effect of Zymomonas mobilis culture on experimental Schistosoma mansoni infection

C57Bl/10 male mice infected with Schistosoma mansoni were distributed into mixed, prophylactic and curative groups. A culture of Zymomonas mobilis was orally administered to mice. A 61% protection from the infection was observed in the curative group (p <0.05). Histopathological study of the livers and intestines showed similar results.

FOOD CONCENTRATION AND TEMPERATURE EFFECTS ON LIFE CYCLE CHARACTERISTICS OF TROPICAL CLADOCERA (Daphnia gessneri Herbst, Diaphanosoma sarsi Richard, Moina reticulata (Daday)): I. DEVELOPMENT TIME

The effects of food concentration and temperature on embryonic and postem-bryonic duration of three tropical species, Daphnia gessneri(1.5mm), Diaphanosoma sarsi(1.2mm) and Moina reticulata(0.8mm), were investigated as part of life cycle studies which included growth, body size and reproduction. These are the very first experimental studies undertaken on these species. The long-term growth experiments were performed under controlled laboratory conditions at all combinations of temperature (22"C, 27"C and 32"C) and constant food concentration (0.03, 0.05, 0.10, 0.25, 0.50 and 1.00 mgC/L) of the unicellular green alga Scenedesmus acutus.Animals were examined twice daily throughout their life cycle from the neonate to third adult instar. In all three species, temperature exerted the most powerful influence on embryonic duration but there was also a smaller food effect. In D. gessneri,postembry-onic durations remained more or less the same at food levels 0.25 mgC/L but were influenced by temperature. At food concentrations of 0.1 mgC/L or lower, postembryonic durations became increasingly prolonged, particularly at high temperatures. This threshold concentration is affected by temperature: in D. gessneri,it was 0.1 mgC/L at 22oC and 27oC but higher at 32oC (between 0.25 and 0.50 mgC/L). At the same temperature of 27oC, the food threshold level varied between species: it was higher (0.25 mgC/L) for D. sarsiand lower (0.05 mgC/L) for M. reticulatacompared with D. gessneri(0.1 mgC/L). In both embryonic and postembryonic durations there is a body size effect as the absolute durations were longest in the largest species and shortest in the smallest species In all three species, prolongation of postembryonic duration at combinations of high temperature and lowered food levels was accompanied by increased number of juvenile instars.

Multivariate Analysis for Animal Selection in Experimental Research

Background: Several researchers seek methods for the selection of homogeneous groups of animals in experimental studies, a fact justified because homogeneity is an indispensable prerequisite for casualization of treatments. The lack of robust methods that comply with statistical and biological principles is the reason why researchers use empirical or subjective methods, influencing their results. Objective: To develop a multivariate statistical model for the selection of a homogeneous group of animals for experimental research and to elaborate a computational package to use it. Methods: The set of echocardiographic data of 115 male Wistar rats with supravalvular aortic stenosis (AoS) was used as an example of model development. Initially, the data were standardized, and became dimensionless. Then, the variance matrix of the set was submitted to principal components analysis (PCA), aiming at reducing the parametric space and at retaining the relevant variability. That technique established a new Cartesian system into which the animals were allocated, and finally the confidence region (ellipsoid) was built for the profile of the animals’ homogeneous responses. The animals located inside the ellipsoid were considered as belonging to the homogeneous batch; those outside the ellipsoid were considered spurious. Results: The PCA established eight descriptive axes that represented the accumulated variance of the data set in 88.71%. The allocation of the animals in the new system and the construction of the confidence region revealed six spurious animals as compared to the homogeneous batch of 109 animals. Conclusion: The biometric criterion presented proved to be effective, because it considers the animal as a whole, analyzing jointly all parameters measured, in addition to having a small discard rate.

High-fat Diet Promotes Cardiac Remodeling in an Experimental Model of Obesity

AbstractBackground:Although nutritional, metabolic and cardiovascular abnormalities are commonly seen in experimental studies of obesity, it is uncertain whether these effects result from the treatment or from body adiposity.Objective:To evaluate the influence of treatment and body composition on metabolic and cardiovascular aspects in rats receiving high saturated fat diet.Methods:Sixteen Wistar rats were used, distributed into two groups, the control (C) group, treated with isocaloric diet (2.93 kcal/g) and an obese (OB) group, treated with high-fat diet (3.64 kcal/g). The study period was 20 weeks. Analyses of nutritional behavior, body composition, glycemia, cholesterolemia, lipemia, systolic arterial pressure, echocardiography, and cardiac histology were performed.Results:High-fat diet associates with manifestations of obesity, accompanied by changes in glycemia, cardiomyocyte hypertrophy, and myocardial interstitial fibrosis. After adjusting for adiposity, the metabolic effects were normalized, whereas differences in morphometric changes between groups were maintained.Conclusion:It was concluded that adiposity body composition has a stronger association with metabolic disturbances in obese rodents, whereas the high-fat dietary intervention is found to be more related to cardiac morphological changes in experimental models of diet-induced obesity.

Host genetic and epigenetic factors in toxoplasmosis

Analysing human genetic variation provides a powerful tool in understanding risk factors for disease. Toxoplasma gondii acquired by the mother can be transmitted to the fetus. Infants with the most severe clinical signs in brain and eye are those infected early in pregnancy when fetal immunity is least well developed. Genetic analysis could provide unique insight into events in utero that are otherwise difficult to determine. We tested the hypothesis that propensity for T. gondii to cause eye disease is associated with genes previously implicated in congenital or juvenile onset ocular disease. Using mother-child pairs from Europe (EMSCOT) and child/parent trios from North America (NCCCTS), we demonstrated that ocular and brain disease in congenital toxoplasmosis associate with polymorphisms in ABCA4 encoding ATP-binding cassette transporter, subfamily A, member 4 previously associated with juvenile onset retinal dystrophies including Stargardt's disease. Polymorphisms at COL2A1 encoding type II collagen, previously associated with Stickler syndrome, associated only with ocular disease in congenital toxoplasmosis. Experimental studies showed that both ABCA4 and COL2A1 show isoform-specific epigenetic modifications consistent with imprinting, which provided an explanation for the patterns of inheritance observed. These genetic and epigenetic risk factors provide unique insight into molecular pathways in the pathogenesis of disease.

Back to the future in Chagas disease: from animal models to patient cohort studies, progress in immunopathogenesis research

Despite the wealth of information generated by trans-disciplinary research in Chagas disease, knowledge about its multifaceted pathogenesis is still fragmented. Here we review the body of experimental studies in animal models supporting the concept that persistent infection by Trypanosoma cruzi is crucial for the development of chronic myocarditis. Complementing this review, we will make an effort to reconcile seemingly contradictory results concerning the immune profiles of chronic patients from Argentina and Brazil. Finally, we will review the results of molecular studies suggesting that parasite-induced inflammation and tissue damage is, at least in part, mediated by the activities of trans-sialidase, mucin-linked lipid anchors (TLR2 ligand) and cruzipain (a kinin-releasing cysteine protease). One hundred years after the discovery of Chagas disease, it is reassuring that basic and clinical research tends to converge, raising new perspectives for the treatment of chronic Chagas disease.

Influence of the polymorphisms of the α-major regulatory element HS-40 on in vitro gene expression

The α-MRE is the major regulatory element responsible for the expression of human α-like globin genes. It is genetically polymorphic, and six different haplotypes, named A to F, have been identified in some population groups from Europe, Africa and Asia and in native Indians from two Brazilian Indian tribes. Most of the mutations that constitute the α-MRE haplotypes are located in flanking sequences of binding sites for nuclear factors. To our knowledge, there are no experimental studies evaluating whether such variability may influence the α-MRE enhancer activity. We analyzed and compared the expression of luciferase of nine constructs containing different α-MRE elements as enhancers. Genomic DNA samples from controls with A (wild-type α-MRE) and B haplotypes were used to generate C-F haplotypes by site-directed mutagenesis. In addition, three other elements containing only the G→A polymorphism at positions +130, +199, and +209, separately, were also tested. The different α-MRE elements were amplified and cloned into a plasmid containing the luciferase reporter gene and the SV40 promoter and used to transiently transfect K562 cells. A noticeable reduction in luciferase expression was observed with all constructs compared with the A haplotype. The greatest reductions occurred with the F haplotype (+96, C→A) and the isolated polymorphism +209, both located near the SP1 protein-binding sites believed not to be active in vivo. These are the first analyses of α-MRE polymorphisms on gene expression and demonstrate that these single nucleotide polymorphisms, although outside the binding sites for nuclear factors, are able to influence in vitro gene expression.

Chikungunya risk for Brazil

This study aimed to show, based on the literature on the subject, the potential for dispersal and establishment of the chikungunya virus in Brazil. The chikungunya virus, a Togaviridae member of the genusAlphavirus, reached the Americas in 2013 and, the following year, more than a million cases were reported. In Brazil, indigenous transmission was registered in Amapa and Bahia States, even during the period of low rainfall, exposing the whole country to the risk of virus spreading. Brazil is historically infested by Ae. aegypti and Ae. albopictus, also dengue vectors. Chikungunya may spread, and it is important to take measures to prevent the virus from becoming endemic in the country. Adequate care for patients with chikungunya fever requires training general practitioners, rheumatologists, nurses, and experts in laboratory diagnosis. Up to November 2014, more than 1,000 cases of the virus were reported in Brazil. There is a need for experimental studies in animal models to understand the dynamics of infection and the pathogenesis as well as to identify pathophysiological mechanisms that may contribute to identifying effective drugs against the virus. Clinical trials are needed to identify the causal relationship between the virus and serious injuries observed in different organs and joints. In the absence of vaccines or effective drugs against the virus, currently the only way to prevent the disease is vector control, which will also reduce the number of cases of dengue fever.