Road-killed wild animals: a preservation problem useful for eco-epidemiological studies of pathogens

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Detection of Toxoplasma gondii in the milk of experimentally infected Wistar female rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Genotypic characterization of Toxoplasma gondii in sheep from Brazilian slaughterhouses: New atypical genotypes and the clonal type II strain identified

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morphological changes of the intestinal mucosa of broilers and layers as affected by fasting before sample collection

The present study aimed at evaluating the histo-morphological changes resulting from different fasting periods before the collection of tissue samples in different segments of the small intestine (duodenum, jejunum and ileum) of 7-d-old male chicks of a broiler and a layer strain. A completely randomized experimental design in in a 2x7 factorial arrangement, being two strains with different growth rates (Ross 308 and HyLine® W36) and seven fasting periods (0, 2, 4, 6, 8, 10 and 12 hours ), with six replicates, totaling 84 birds. The comparison of the morphometrics of the duodenum, jejunum and ileum of broiler and layer chicks demonstrated faster digestive tract development in broilers relative to layers. The fasting period caused morphological changes in the liver and small and large intestines in both strains. Therefore, it must be highlighted that in studies involving organ weights and intestinal morphometrics, birds must not be submitted to fasting before tissue collection.

Histopathology of the reproductive system of male sheep experimentally infected with Toxoplasma gondii

The aim of this study was to investigate the histopathological changes in reproductive system (testicles, epididymis, seminal vesicles, and prostate) of small male ruminants after Toxoplasma gondii infection. Eight sheep were inoculated with T. gondii: group I, four sheep (2.0 x 10(5) P-strain oocysts); group II, four sheep (1.0 x 10(6) RH-strain tachyzoites); and group III, two uninfected sheep maintained as control. Infection with T. gondii was confirmed by seroconversion (indirect fluorescent antibody test-IgG) in all the infected animals beginning on post-inoculation day (PID) 7. on PID 70, all the animals were euthanized and tissue samples (testicles, epididymis, seminal vesicles, and prostate) were collected and processed for histological analysis. The main changes detected were a focal mononuclear interstitial inflammatory infiltrate in the prostate and seminal vesicles; diffuse testicular degeneration associated with calcification foci and a multifocal mononuclear interstitial inflammatory infiltrate; and a mononuclear interstitial infiltrate and focal necrotic areas of the muscle fibers surrounding the seminal vesicles. The histopathological findings of this work, along with the detection of T. gondii in the examined parenchyma tissues (immunohistochemistry) and the results obtained by other authors examining different tissues, suggest that histological changes diagnosed in the reproductive system of rams infected with T. gondii are strongly suggestive of toxoplasmatic infection.

Histometric analysis of ligature-induced periodontitis in rats: a comparison of histological section planes

The purpose of this study was to analyze the histometry of ligature-induced periodontitis in rats at different histological section depths. Sixteen male adult Wistar rats were randomly assigned to two groups: ligature and control. In the ligature group, rats received a sterile 4/0 silk ligature around the maxillary right 2nd molar. Thirty serial sections containing the 1st and 2nd molars, in which the coronal and root pulp, cementoenamel junction (CEJ) in the mesial side of the 2nd molar, interproximal alveolar bone and connective fiber attachment were clearly visible, were selected for histometric analysis. The histological sections were clustered in groups of 10 sections corresponding the buccal (B), central (C) and lingual (L) regions of the of periodontal tissue samples. The distance between the CEJ in the mesial side of the 2nd molar and the attached periodontal ligament fibers (CEJ-PL) as well as the distance between the CEJ and the alveolar bone crest (CEJ-BC) were determined. From CEJ-PL and CEJ-BC distances measured for each specimen, the measurements obtained in the B, L and C regions were recorded individually and together. Data were submitted to statistical analysis. Significant differences (p<0.001) were observed between the control and ligature groups regarding CEJ-PL (0.05 mm and 0.26 mm, respectively) and CEJ-BC (0.47 mm and 0.77 mm, respectively) measurements. Regarding the depth of the buccal, central and lingual planes, the means of CEJ-PL and CEJ-BC of both groups showed no statistically significant differences (p>0.05). In conclusion, the selection of 10 serial sections of the central region of periodontal tissue samples at any depth can be considered as representative for the evaluation of periodontal ligament fiber attachment and bone loss in ligature-induced periodontitis in rats.

Comparison of the biocompatibility of different root canal irrigants

The purpose of this study was to compare the reaction of rat subcutaneous connective tissue to 0.9% sterile saline, 2.5% sodium hypochlorite (NaOCl), 5.25% NaOCl and 2% chlorhexidine gluconate solution or gel. Six circles were demarcated on the dorsal skin of 24 male Wistar rats, leaving 2 cm between each circle. Using a syringe, 0.1 mL of each root canal irrigant was injected subcutaneously into 5 circles. In the 6th circle, the needle of an empty syringe was introduced into the skin, but no irrigant was injected (control group). Evaluations were undertaken at 2 h, 48 h, 14 days and 30 days post-procedure. Tissue samples were excised, embedded in paraffin blocks and 3-mu m-thick sections were obtained and stained with hematoxylin and eosin. The areas of inflammatory reaction were evaluated and analyzed statistically by ANOVA and Tukey's test. The control group showed few or no inflammatory reaction areas in the subcutaneous tissue. 0.9% saline solution, 2.0% chlorhexidine solution and 2.5% NaOCl showed a good biocompatibility, as very mild inflammatory reaction was detected at 14 days and tissue repair occurred at 30 days. 5.25% NaOCl was the most toxic irrigant, as the number of inflammatory cells remained elevated at 14 and 30 days. The group treated with 2.0% chlorhexidine gluconate gel presented a moderate inflammatory response at 14 days, which decreased at 30 days, being considered similar to that of the control group, 0.9% saline solution, 2.0% chlorhexidine solution and 2.5% NaOCl at this experimental period.

Expression of interferon-gamma, interferon-alpha and related genes in individuals with Down syndrome and periodontitis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Relationship of abomasal histology and parasite-specific immunoglobulin A with the resistance to Haemonchus contortus infection in three breeds of sheep

This study was carried out to evaluate the relationship of abomasal inflammatory cells and parasite-specific immunoglobulin A (IgA) in mucus, with the resistance to Haemonchus contortus infection in three breeds of sheep naturally infected with gastrointestinal nematodes. The breeds were the native Santa Ines sheep, and the European Suffolk and Ile de France breeds. Mast cells, eosinophils and globule leucocytes were enumerated in abomasal mucosa. Eosinophils within the sub-mucosa also were counted separately. Histamine concentration was estimated in abomasal tissue samples. Enzyme-linked immunosorbent assay was carried out in mucus samples to determine the level of IgA anti-H. contortus third and fifth instar. There were no significant differences among group means of these variables (P > 0.05). The correlation coefficients between fecal egg counts (FEC) x mast cells (r = -0.490; P < 0.05) and FEC x eosinophils in sub-mucosa (r = -0.714; P < 0.01) was significant in the Santa Ines sheep. In the Ile de France group, the correlation coefficients between globule leucocytes x FEC (r = -0.879; P < 0.001) and histamine x worm burden (r = -0.833; P < 0.01) were also significant. In the Santa Ines and Ile de France sheep, correlation coefficients between IgA anti-L3 x worm burden and IgA anti-L3 x FEC were negative. In general, inflammatory cells and IgA-parasite-specific in abomasum were inversely associated with H. contortus worm burden and FEC indicating that they may impair parasite development or fecundity in the three breeds of sheep. However, similar mean values of inflammatory cells and IgA were found in the resistant (Santa Ines) and in the susceptible (Suffolk and Ile de France) breeds of sheep. The enumeration of cells by histological assessment does not provide information on their functional activity, which may be different among breeds. Thus, the effect of breed on the functional activity of these and other inflammatory cells is an important area for further study. (c) 2004 Elsevier B.V. All rights reserved.

Metabolic responses to glucoprivation induced by 2-deoxy-D-glucose in Brycon cephalus (Teleostei, Characidae)

The effects of functional cytoglucopenia provoked by 2-deoxy-D-glucose (2-DG) were studied in adult Brycon cephalus, an omnivorous fish from the Amazon Basin in Brazil. Glycogen content in liver and muscle as well as plasmatic glucose, free fatty acids (FFA), insulin, and glucagon were measured. After 48 h fasting, an intraperitoneal saline injection (NaCl 0.6 g/100 ml) was administered to control fish, whereas the experimental group received 2-DG, dissolved in saline, in the dosage of 80 mg/kg (0.487 mmol/kg) or 150 mg/kg (0.914 mmol/kg) body weight; injection volume was 5 ml in all treatments. Blood and tissue samples were taken immediately before, and 2, 8, 10, and 24 h after administration of the drug or saline. Fish injected with both doses of 2-DG showed a marked increase in glycemia levels. Liver and muscle glycogen decreased after 2-DG administration and reached their lowest values 10-24 h after injection, while in control animals no significant changes were observed. Elevation in plasma glucagon was observed only in response to the maximum dosage of 2-DG administered, especially 10 h and 24 h post-injection. Plasma insulin levels were lower in animals treated with the glucose analogue but only statistically significant 24 h after drug administration. In conclusion, the administration of the non-metabolizable glucose analogue 2-DG in B. cephalus is a stimulus to generate responses towards an increase in the glucose available to tissues, which is a characteristic of a fasting situation. All the above data support the interest of 2-DG administration as a model to study carbohydrate metabolism adjustment mechanisms in fish.

Atrophic Cardiac Remodeling Induced by Taurine Deficiency in Wistar Rats

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Molecular detection of Paracoccidioides brasiliensis in road-killed wild animals

Paracoccidioides brasiliensis infections have been little studied in wild and/or domestic animals, which may represent an important indicator of the presence of the pathogen in nature. Road-killed wild animals have been used for surveillance of vectors of zoonotic pathogens and may offer new opportunities for eco-epidemiological studies of paracoccidiodomycosis (PCM). The presence of P. brasiliensis infection was evaluated by Nested-PCR in tissue samples collected from 19 road-killed animals; 3 Cavia aperea (guinea pig), 5 Cerdocyon thous (crab-eating-fox), 1 Dasypus novemcinctus (nine-banded armadillo), 1 Dasypus septemcinctus (seven-banded armadillo), 2 Didelphis albiventris (white-eared opossum), 1 Eira barbara (tayra), 2 Gallictis vittata (grison), 2 Procyon cancrivorus (raccoon) and 2 Sphiggurus spinosus (porcupine). Specific P. brasiliensis amplicons were detected in (a) several organs of the two armadillos and one guinea pig, (b) the lung and liver of the porcupine, and (c) the lungs of raccoons and grisons. P. brasiliensis infection in wild animals from endemic areas might be more common than initially postulated. Molecular techniques can be used for detecting new hosts and mapping 'hot spot' areas of PCM.

Relationship of intestinal histology with the resistance to Trichostrongylus colubriformis infection in three breeds of sheep

Avaliaram-se a associação entre o número de células inflamatórias no intestino delgado e a resistência à infecção por Trichostrongylus colubriformis em ovinos de três raças (Santa Inês, Suffolk e Ile de France), naturalmente infectados. Mastócitos, eosinófilos e leucócitos globulares foram quantificados na mucosa intestinal. A concentração de histamina foi estimada em amostras teciduais do intestino, bem como foi determinado o comprimento de machos e fêmeas de T. colubriformis. A resposta celular foi similar na mucosa intestinal das três raças ovinas (P>0,05). Houve grande variação entre os ovinos em relação aos resultados parasitológicos e celulares, mesmo nos animais de mesma raça. em geral, os animais que apresentaram número menor de células inflamatórias tiveram cargas parasitárias maiores, contagens de ovos por grama de fezes mais altas e exemplares de T. colubriformis maiores. Os resultados indicaram que mastócitos, eosinófilos e leucócitos globulares prejudicaram o estabelecimento, o desenvolvimento e a sobrevivência dos parasitas.

Cytokine gene expression in response to Haemonchus placei infections in Nelore cattle

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immunological responses and cytokine gene expression analysis to Cooperia punctata infections in resistant and susceptible Nelore cattle

Cellular and humoral immune response, as well as cytokine gene expression, was assessed in Nelore cattle with different degrees of resistance to Cooperia punctata natural infection. One hundred cattle (male, weaned, 11-12 months old), kept together on pasture, were evaluated. Faecal and blood samples were collected for parasitological and immunological assays. Based on nematode faecal egg counts (FEC) and worm burden, the seven most resistant and the eight most susceptible animals were selected. Tissue samples of the small intestine were collected for histological quantification of inflammatory cells and analysis of cytokine gene expression (IL-2, IL-4, IL-8, IL-1 2p35, IL-13, TNF-alpha, IFN-gamma, MCP-1, MCP-2, and MUC- 1) using real-time RT-PCR. Mucus samples were also collected for IgA levels determination. Serum IgG1 mean levels against C. punctata antigens were higher in the resistant group, but significant differences between groups were only observed 14 days after the beginning of the experiment against infective larvae (1-3) and 14 and 84 days against adult antigens. The resistant group also presented higher IgA levels against C. punctata (L3 and adult) antigens with significant difference 14 days after the beginning of the trial (P < 0.05). In the small-intestine mucosa, levels of IgA anti-L3 and anti-adult C. punctata were higher in the resistant group, compared with the susceptible group (P < 0.05). Gene expression of both T(H)2 cytokines (IL-4 and IL-13) in the resistant group and T(H)1 cytokines (IL-2, IL-1 2p35, IFN-gamma and MCP-1) in the susceptible group was up-regulated. Such results suggested that immune response to C. punctata was probably mediated by TH2 cytokines in the resistant group and by T(H)1 cytokines in the susceptible group. (C) 2008 Elsevier B.V. All rights reserved.