Experimentally induced intravaginal Tritrichomonas foetus infection in a mouse model

The interest to develop research on the host-parasite relationship in bovine tritrichomonosis has accomplished the use of experimental models alternative to cattle. The BALB/c mouse became the most appropriate species susceptible to vaginal Tritrichomonas foetus infection requiring previous estrogenization. For the need of an experimental model without persistent estrogenization and with normal estrous cycles, the establishment and persistence of vaginal infection on BALB/c mouse with different concentrations of T. foetus in two experimental groups was evaluated. Group A was treated with 5mg of b-estradiol 3-benzoate to synchronize the estrous, 48 hours before the T. foetus vaginal inoculation, and Group B was inoculated in natural estrus. At 5-7 days after treatment, estrogenic effect decreased allowing all animals to cycle regularly during the experiment. From the first week post-infection, samples of vaginal mucus were taken from all animals during 34 weeks, in order to evaluate the course of infection and the stage of the estrus cycle. Group A showed 93.6% of infected animals, and Group B showed 38%. Different doses of T. foetus were assayed to establish the vaginal infection, with a persistence of 34 weeks. Although different behavior was observed in each subgroup belonging to either Group A or Group B, there were no significant differences among the infecting doses used. The b-estradiol 3-benzoate treatment had a favorable effect on the establishment of the infection (P<0.0001), but it did not influence its persistence (P=0.1097). According to the results, an experimental mouse model is presented, appropriate for further studies on mechanisms of pathogenicity, immune response, protective evaluation of immunogen and therapeutic effect of drugs.

Evaluation of tear production with modified Schirmer tear test-1 during the neonatal period in cats

The study aimed to evaluate tear production by means of modified Schirmer tear tes-1 (mSTT-1) in neonate cats. Likewise, correlation between mSTT-1 and STT-1 was assessed in vitro. Standard SST strips were cut in half and after eye lid opening, tear production of neonates (n=15) was daily measured in both eyes (mSTT-1), until the 7th day, and at day 14, 21, and 28. Animals were daily weighted until 28 days of age. Results were compared statistically (P<0.05). During the first 7 days, the overall mSTT-1 mean was 0.76 wetting/minute. Significant differences between right and left eyes were not observed at any time point (P=1.00). Tear secretion increased significantly, from the 14th to 28th day, in comparison with 7 first days (P<0.05). Positive correlation between maturity parameters and tear secretion was observed (P<0.0001). Distance between slopes of each strip changed significantly (P<0.0001). It was concluded that tear secretion in the neonatal period of cats is very below the reference values described for young and adults of the same species. It is not possible to extrapolate results obtained with mSTT-1 to standard STT-1.

Matched case-control study evaluating the frequency of the main agents associated with neonatal diarrhea in piglets

A case-control study was carried out in litters of 1 to 7-day-old piglets to identify the main infectious agents involved with neonatal diarrhea in pigs. Fecal samples (n=276) from piglets were collected on pig farms in the State of Rio Grande do Sul, Brazil, from May to September 2007. Litters with diarrhea were considered cases (n=129) and normal litters (n=147) controls. The samples were examined by latex agglutination test, PAGE, conventional isolating techniques, ELISA, PCR, and microscopic methods in order to detect rotavirus, bacterial pathogens (Escherichia coli, Clostridium perfringens type A and C, and Clostridium difficile), and parasites (Coccidian and Cryptosporidium spp.). Outbreaks of diarrhea were not observed during sampling. At least one agent was detected in fecal samples on 25 out of 28 farms (89.3%) and in 16 farms (57.1%) more than one agent was found. The main agents diagnosed were Coccidia (42.86%) and rotavirus (39.29%). The main agents identified in litters with diarrhea were Clostridium difficile (10.6%), Clostridium perfringens type A (8.8%) and rotavirus (7.5%); in control litters, Clostridium difficile (16.6%) and Coccidian (8.5%). Beta hemolytic Escherichia coli and Clostridium perfringens type C were not detected. When compared with controls, no agent was significantly associated with diarrhea in case litters. These findings stress the need for caution in the interpretation of laboratorial diagnosis of mild diarrhea in neonatal pigs, as the sole detection of an agent does not necessarily indicate that it is the cause of the problem.

Monitoramento de parâmetros laboratoriais em gatos sem raça definida durante o período neonatal

O objetivo do presente estudo foi à monitoração dos parâmetros laboratoriais como hemograma, enzimas hepáticas alanina aminotransferase (ALT) e gama-glutamiltransferase (GGT), glicemia e proteinograma sérico, e avaliar o efeito da idade em gatos sem raça definida durante a fase neonatal. Vinte gatos machos e fêmeas foram utilizados a partir do terceiro dia de vida até o 38º dia de idade. As amostras de sangue foram colhidas semanalmente e as análises laboratoriais (hemograma, enzimas hepáticas, glicemia e proteinograma sérico) realizadas no 3º, 10º, 17º, 24º, 31º e 38º dia de idade. Os resultados exibiram efeito significativo da idade sobre a contagem total de eritrócitos, concentração de hemoglobina, volume globular, volume corpuscular médio, concentração de hemoglobina corpuscular média, leucócitos totais, neutrófilos, eosinófilos e basófilos. Nenhum efeito foi observado em células como linfócitos, monócitos ou na concentração sérica de glicose. A análise das modificações ocorridas nos parâmetros laboratoriais durante a fase neonatal reflete o desenvolvimento fisiológico do filhote e contribui para o conhecimento do processo adaptativo em gatos neonatos durante o primeiro mês de vida, sendo útil para a avaliação clínica, diagnóstico e tratamento das doenças neonatais.

Microbiota bacteriana e citologia da região traqueobrônquica de bezerros no período neonatal

As broncopneumonias são afecções importantes na pecuária mundial, representando uma das principais causas de mortalidade de bezerros nos primeiros meses de vida. As medidas preventivas e terapêuticas adotadas geralmente são baseadas em resultados de estudos internacionais, não se conhecendo as bactérias implicadas nos quadros pneumônicos em animais criados no Brasil. Aliado a isso, no primeiro mês de vida, os bezerros demonstram imaturidade do sistema imune, o que tem sido pouco estudado em quadros pneumônicos. Desta maneira, objetivou-se estudar as broncopneumonias em bezerros neonatos, identificando bactérias do trato respiratório posterior de bezerros sadios e com pneumonias naturalmente adquiridas, bem como analisar citologicamente a resposta pulmonar frente a estes patógenos. Para isso amostras de lavado do trato respiratório foram colhidas por traqueocentese durante o primeiro mês de vida dos animais. Verificou-se que não houve diferença na microbiota traqueobrônquica de bezerros sadios em relação aos doentes, discordando dos relatos da literatura internacional, sendo constituída principalmente por: Staphylococcus sp., Bacillus sp., Streptococcus sp., Pseudomonas aeruginosa e enterobactérias, permitindo inferir que as medidas profiláticas e terapêuticas adotadas internacionalmente possam não ser tão efetivas para as criações brasileiras. Observou-se também que bezerros neonatos têm uma proporção aproximada de 1:1 de macrófagos e neutrófilos na região traqueobrônquica quando saudáveis, atingindo uma relação aproximada de 1:3 durante os quadros de broncopneumonias, sendo estes perfis provavelmente característicos da idade, período conhecido pela imaturidade do sistema imune e agravado por fatores de manejo que favoreçam uma maior inalação de agentes bacterianos.

Morphological and immunohistochemical characterization of angiogenic and apoptotic factors and the expression of thyroid receptors in the ovary of tilapia Oreochromis niloticus in captivity

Morphological and immunohistochemical characterization of angiogenic and apoptotic factors and the expression of thyroid receptors in the ovary of tilapia Oreochromis niloticus in captivity were studied. The morphological evaluation of the ovaries was performed by histological paraffin embedded and stained with HE. The immunohistochemical expressions of CDC47, VEGF, Flk-1, angiopoietin, Tie-2 and thyroid receptor (TRα) were performed by the technique of streptavidein-biotin-peroxidase. Apoptosis was assessed using the TUNEL kit. The relative expression of thyroid hormone receptors (TRα and TRβ) was assessed by RT-PCR real time. The nuclear expression of CDC47 increased with the stage of maturation of the oocyte and was observed in the follicle cells. Apoptotic bodies were observed in the follicular cells of atretic follicles and postovulatory follicles from the ovaries of 150g and 350g fish. Expression of VEGF and its receptor Flk-1 was also observed in the follicular cells, and the expression of both increased with the maturity of the oocyte, with a higher intensity observed in the full-grown follicle. The expression of angiopoietin and of its receptor (Tie 2) was discrete and moderate respectively. TRα expression was independent of follicular development. However, the 350 g tilapia exhibited higher expression of TRβ compared with the 50 g tilapia. We conclude that the proliferative activity and the expression of VEGF and its receptor increase with follicular maturation and that the TRs expression increases with ovarian maturity in tilapia (Oreochromis niloticus).

Microphotometric scanning of chromatid gaps and breaks induced by AluI and BamHI in Chinese hamster ovary cells

Chromatid gaps and breaks induced by the restriction endonucleases AluI and BamHI in the long arm of chromosome 1 of Chinese hamster ovary cells were microphotometrically scanned and mapped to a quantitative G-band map. More than 50% of chromatid breaks appeared as chromatin losses of greater than 5% of the total arm length. The majority of chromatid gaps and breaks as well as chromatin losses induced by both restriction endonucleases were non-randomly located in a region from 0.35 to 0.65 relative length units of the long arm of chromosome 1. We suggest that the access of these endonucleases to chromosomal DNA depends on the local organization of the chromatin.

Effect of taxol on chromosome aberrations induced by gamma radiation or by doxorubicin in Chinese hamster ovary cells

Combined therapy with radiation and chemotherapy has being increasingly used in cancer treatment. The effect of combinations of taxol (0.08 mug/ml) with doxorubicin (DXR, 0.5 or 1.0 mug/ml) or gamma radiation (20 or 40 cGy) was examined in two different treatment schedules (pretreatment or simultaneous treatment) using Chinese hamster ovary (CHO) cells treated at the G2 phase of the cell cycle. The results showed that taxol did not have a radiosensitizing effect on the chromosomal aberrations induced by gamma radiation nor did it have a potentiating effect on the chromosomal aberrations induced by DXR in CHO cells treated in the G2 phase of the cell cycle

Clastogenic activity of integerrimine determined in mouse micronucleus assays

The pyrrolizidine alkaloid integerrimine, obtained from Senecio brasiliensis, was tested by acute dosing at two concentrations (18.75 and 37.5 mg/kg), and at different times, to establish its ability to induce micronuclei in mouse erythrocytes. This alkaloid was able to increase the frequency of micronucleated polychromatic erythrocytes in both, bone marrow and peripheral blood erythrocytes

Synergistic effect of glucose and prolactin on GLUT2 expression in cultured neonatal rat islets

We studied the synergistic effect of glucose and prolactin (PRL) on insulin secretion and GLUT2 expression in cultured neonatal rat islets. After 7 days in culture, basal insulin secretion (2.8 mM glucose) was similar in control and PRL-treated islets (1.84 ± 0.06% and 2.08 ± 0.07% of the islet insulin content, respectively). At 5.6 and 22 mM glucose, insulin secretion was significantly higher in PRL-treated than in control islets, achieving 1.38 ± 0.15% and 3.09 ± 0.21% of the islet insulin content in control and 2.43 ± 0.16% and 4.31 ± 0.24% of the islet insulin content in PRL-treated islets, respectively. The expression of the glucose transporter GLUT2 in B-cell membranes was dose-dependently increased by exposure of the islet to increasing glucose concentrations. This effect was potentiated in islets cultured for 7 days in the presence of 2 µg/ml PRL. At 5.6 and 10 mM glucose, the increase in GLUT2 expression in PRL-treated islets was 75% and 150% higher than that registered in the respective control. The data presented here indicate that insulin secretion, induced by different concentrations of glucose, correlates well with the expression of the B-cell-specific glucose transporter GLUT2 in pancreatic islets

Morphometry and acetylcholinesterase activity of the myenteric plexus of the wild mouse Calomys callosus

The myenteric plexus of the digestive tract of the wild mouse Calomys callosus was examined using a histochemical method that selectively stains nerve cells, and the acetylcholinesterase (AChE) histochemical technique in whole-mount preparations. Neuronal density was 1,500 ± 116 neurons/cm2 (mean ± SEM) in the esophagus, 8,900 ± 1,518 in the stomach, 9,000 ± 711 in the jejunum and 13,100 ± 2,089 in the colon. The difference in neuronal density between the esophagus and other regions was statistically significant. The neuron profile area ranged from 45 to 1,100 µm2. The difference in nerve cell size between the jejunum and other regions was statistically significant. AChE-positive nerve fibers were distributed within the myenteric plexus which is formed by a primary meshwork of large nerve bundles and a secondary meshwork of finer nerve bundles. Most of the nerve cells displayed AChE activity in the cytoplasm of different reaction intensities. These results are important in order to understand the changes occurring in the myenteric plexus in experimental Chagas' disease

Rotavirus and reovirus interaction with mouse peritoneal resident phagocytic cells

Rotaviruses and reoviruses are involved in human and animal diseases. It is known that both viruses penetrate the gastrointestinal tract but their interaction with phagocytic cells is unknown. To study this interaction, peritoneal resident phagocytic cells were used and rotavirus and reovirus replication in peritoneal phagocytic cells was observed. However, rotavirus replication in these cells led to the production of defective particles since MA-104 cells inoculated with rotavirus phagocytic cell lysate did not show any evidence of virus replication. On the basis of these results, we suggest that, although reovirus dissemination may be helped by these phagocytic cells, these cells may control rotavirus infection and probably contribute to the prevention of its dissemination.

Aversive stimulation during the stress-hyporesponsive period does not affect the number of corticotroph cells in neonatal male rats

Immunohistochemistry was used to evaluate the effects of neonatal handling and aversive stimulation during the first 10 days of life on the number of corticotrophs in the anterior lobe of the pituitary of 11-day-old male Wistar rats. Since adult rats handled during infancy respond with reduced corticosterone secretion in response to stressors and with less behavior inhibition in novel environments, we assumed that neonatal stimulation could affect pituitary morphology during this critical period of cell differentiation. Three groups of animals were studied: intact (no manipulation, N = 5), handled (N = 5) and stimulated (submitted to 3 different aversive stimuli, N = 5). The percentage of ACTH-immunoreactive cells in the anterior lobe of the pituitary (number of ACTH-stained cells divided by total number of cells) was determined by examining three slices per pituitary in which a minimum of 200 cells were counted by two independent researchers. Although animals during the neonatal period are less reactive to stress-like stimulation in terms of ACTH and corticosterone secretion, results showed that the relative number of ACTH-stained cells of neonatal handled (0.25 ± 0.01) and aversive stimulated (0.29 ± 0.03) rats was not significantly different from intact (0.30 ± 0.03) animals. Neonatal stimulation may have a differential effect on the various subpopulations of corticotroph cells in the anterior pituitary

Relationship of cytokines and cytokine signaling to immunodeficiency disorders in the mouse

The contributions of cytokines to the development and progression of disease in a mouse model of retrovirus-induced immunodeficiency (MAIDS) are controversial. Some studies have indicated an etiologic role for type 2 cytokines, while others have emphasized the importance of type 1 cytokines. We have used mice deficient in expression of IL-4, IL-10, IL-4 and IL-10, IFN-g, or ICSBP - a transcriptional protein involved in IFN signaling - to examine their contributions to this disorder. Our results demonstrate that expression of type 2 cytokines is an epiphenomenon of infection and that IFN-g is a driving force in disease progression. In addition, exogenously administered IL-12 prevents many manifestations of disease while blocking retrovirus expression. Interruption of the IFN signaling pathways in ICSBP-/- mice blocks induction of MAIDS. Predictably, ICSBP-deficient mice exhibit impaired responses to challenge with several other viruses. This immunodeficiency is associated with impaired production of IFN-g and IL-12. Unexpectedly, however, the ICSBP-/- mice also develop a syndrome with many similarities to chronic myelogenous leukemia in humans. The chronic phase of this disease is followed by a fatal blast crisis characterized by clonal expansions of undifferentiated cells. ICSBP is thus an important determinant of hematopoietic growth and differentiation as well as a prominent signaling molecule for IFNs

Mouse inoculation for the detection of non-cultivable gastric tightly spiralled bacteria

In the present study we compared the inoculation of swine gastric mucus into the stomach of mice, the urease test and carbolfuchsin-stained smears for the diagnosis of the infection with "Gastrospirillum suis" ("Helicobacter heilmannii" type 1), an uncultivated tightly spiralled gastric bacterium. Fragments obtained from the antral and oxyntic mucosa of the stomach of 50 slaughtered pigs were used for urease test, for carbolfuchsin-stained smears and for obtaining scrapings of mucus for mouse inoculation. The mice were killed by spinal dislocation 10 days after inoculation and fragments of the antral and oxyntic mucosa were used for spiral bacterium identification (urease test and carbolfuchsin-stained smears). Among the methods employed for the diagnosis of "H. heilmannii" infection, the inoculation of gastric mucus into the stomach of mice was the most sensitive and demonstrated bacterial positivity in 31 (62.0%) swine. Direct examination showed tightly spiralled bacteria in the gastric mucosa of only 4 (8.0%) of the 50 pigs studied. Among them, 3 (6.0%) presented a positive preformed urease test. Spiral bacteria were not seen in the gastric mucosa of any control mice. These results show that the use of the mouse inoculation method improved the detection of "H. heilmannii" in swine