Effect of pressure on refolding of recombinant pentameric cholera toxin B.

The production of recombinant proteins is an essential tool for the expansion of modern biological research and biotechnology. The expression of heterologous proteins in Escherichia coli often results in an incomplete folding process that leads to the accumulation of inclusion bodies (IB), aggregates that hold a certain degree of native-like secondary structure. High hydrostatic pressure (HHP) impairs intermolecular hydrophobic and electrostatic interactions, leading to dissociation of aggregates under non-denaturing conditions and is therefore a useful tool to solubilize proteins for posterior refolding. Cholera toxin (CT) is composed of a non-toxic pentamer of B subunits (CTB), a useful adjuvant in vaccines, and a toxic subunit A (CTA). We studied the process of refolding of CTB using HHP. HHP was shown to be effective for dissociation of CTB monomers from IB. Posterior incubation at atmospheric pressure of concentrated CTB (1mg/ml) is necessary for the association of the monomers. Pentameric CTB was obtained when suspensions of CTB IB were compressed at 2.4kbar for 16h in the presence of Tween 20 and incubated at 1bar for 120h. Soluble and biologically active pentameric CTB was obtained, with a yield of 213mg CTB/liter of culture. The experience gained in this study can be important to improve the refolding of proteins with quaternary structure.

Neospora caninum: evaluation of vertical transmission in slaughtered beef cows (Bos indicus).

Neospora caninum is a protozoan parasite that causes the most important reproductive problems in cattle worldwide. The objective of this study was to evaluate the possibility of vertical transmission of N. caninum in zebus breed beef cows (Bos indicus) submitted for slaughter at an abattoir in the northern region of the State of Paraná, southern Brazil. One hundred and fifty-nine cows were evaluated: 83 pregnant (in different stages of gestation) and 76 non-pregnant. Serum determination of N. caninum was evaluated by indirect ELISA (Idexx). Blood (with EDTA) from pregnant cows and tissue samples (brain and heart) from their fetuses were collected and used for PCR analyses. Antibodies against N. caninum were observed in 14.6% (12/83) of pregnant and in 15.8% (12/76) of non-pregnant cows. Antibodies against the parasites were detected in one fetus (1.4%). The PCR analyses revealed that 6.0% (5/83) of cows and 4.8% (4/83) of fetuses evaluated were positive to specific N. caninum primers. These positive fetuses were between 4 and 6 months of age. Thus, considering PCR and serology as an indicative of vertical transmission in fetuses, 4.8% of fetuses were infected by N. caninum during gestation.

[Mechanisms of secondary degeneration in the central nervous system during acute neural disorders and white matter damage]. / Mecanismos de degeneración secundaria en el sistema nervioso central durante los trastornos neuronales agudos y el daño en la sustancia blanca.

INTRODUCTION: Acute neurodegenerative diseases, including stroke and traumatic brain and spinal cord injury, possess an elevated worldwide incidence. Two distinct lesive patterns can be identified after these destructive events: primary damage, an early consequence of the primary pathological event, and secondary neural degeneration (SND), a group of pathological events inducing late degeneration in cells not or even only partially affected by the primary damage. This pathological mechanism is an important contributing factor for functional deficits and target for therapeutic approaches. Several factors are involved on the SND etiology, including excitotoxicity, inflammation, and oxidative stress. AIM: To review the main mechanisms underlying the SND occurring after acute neural disorders. DEVELOPMENT: The more recent findings about the eliciting processes of SND degeneration are discussed, as well as their significance to degeneration of white matter tracts. CONCLUSIONS: The characterization of the events underlying SND is of fundamental importance for the development of new therapeutic approaches effective enough to decrease the functional deficits, contributing to the improvement of the quality of life of people suffering neurological diseases. These therapeutic approaches must be validated in experimental models of both brain and spinal cord diseases, which effectively simulate human neural disorders protecting both gray and white matters for a better neuroprotective efficacy.

Neurotropism and neuropathological effects of selected rhabdoviruses on intranasally-infected newborn mice.

Viral neurotropism is the ability of viruses to infect neuronal cells. This is well studied for herpesviruses, rabies-related viruses, and a few others, but it is poorly investigated among almost all arboviruses. In this study, we describe both the neurotropism and the neuropathological effects of Amazonian rhabdoviruses on the brains of experimentally infected-newborn mice. Suckling mice were intranasally infected with 10(-4) to 10(-8) LD50 of viruses. Animals were anaesthetized and perfused after they had become sick. Immunohistochemistry using specific anti-virus and anti-active caspase three antibodies was performed. All infected animals developed fatal encephalitis. Survival time ranged from 18 h to 15 days. Viruses presented distinct species-dependent neurotropism for CNS regions. Histopathological analysis revealed variable degrees of necrosis and apoptosis in different brain regions. These results showed that viruses belonging to the Rhabdoviridae family possess distinct tropism for CNS structures and induce different pattern of cell death depending on the CNS region.

Prevalence of Neospora caninum antibodies and factors associated with their presence in dairy cattle of the north of Paraná state, Brazil.

To determine the prevalence of Neospora caninum antibodies and associated factors, blood sera from 623 female dairy cattle from 23 farms in the north of the state of Paraná, Brazil, were analyzed by means of the indirect immunofluorescent-antibody test (IFAT > or = 25). Serum samples from 134 dogs living on the same farms also were tested for N. caninum antibodies (IFAT > or = 50), and the presence of dogs was associated with the prevalence of N. caninum antibodies in cattle. The overall seroprevalence in cattle was 14.3%, mainly in animals over 24 months of age. Seroprevalence in Holsteins (15.1% of 558) was greater than in mixed-breed cattle (7.7% of 65). Age (> or =24 months) of cattle, feeding silage and/or concentrate produced on the farm were associated; antibodies were found in 21.6% of dogs; and the presence of dogs was associated with the prevalence of N. caninum antibodies in cattle.

Prevalence of Neospora caninum antibodies in dogs from dairy cattle farms in Parana, Brazil.

Serum samples from 134 dogs from 22 cattle dairy farms in the northern region of Parana State, Brazil, were tested for antibodies to Neospora caninum in an indirect fluorescent antibody test. Antibodies (> or = 1:50) to N. caninum were found in 29 (21.6%) of the 134 dogs, and seropositive dogs were found on 14 (63.6%) of the 22 dairy cattle farrms. The antibody titers of dogs were 1:50 (3 dogs), 1:100 (7 dogs), 1:200 (7 dogs), 1:400 (6 dogs), and > or = 1:800 (6 dogs). The low prevalence (9%) in < 1-yr-old dogs compared with the 2- to 3-fold higher prevalence in older dogs (17-29%) suggests postnatal exposure to N. caninum infection.

Craniopharyngioma in the Mongolian gerbil (Meriones unguiculatus): a case report.

A case of spontaneously occurring infiltrative craniopharyngioma in a laboratory-reared gerbil is reported. The histopathological and ultrastructural features are described.

A anti-sepsia das maos no preparo da equipe cirurgica. / Hand asepsis in the preparation of the surgical team

Foi estudada a anti-sepsia pre-operatoria das maos em 30 estudantes preparados em condicoes semelhantes as dos componentes da equipe cirurgica. Para o experimento os estudantes foram divididos em tres grupos: grupo testemunha em que a ecovacao das maos e antebracos era feita exclusivamente om agua e sabao comum; grupo alcool iodado com a anti-sepsia completada com a aplicacao de alcool-iodado e grupo PVPI, em que a escovacao era feita com o uso de solucao de polivinil pirrolidona-iodo (PVPI). Terminada a anti-sepsia vestiam aventais e luvas esterilizados, durante periodos estabelecidos de 30, 60 e 120 minutos, simulando o trabalho operatorio. Antes do inicio da escovacao ao final da anti-sepsia das maos e ao termino dos prazos estabelecidos com o uso da vestimenta cirurgica,eram colhidas amostras na superficie das maos e dedos, para cultura e exame bacteriologicos. Pelos resultados obtidos concluiu-se que a escovacao seguida da aplicacao de alcool-iodado tem o melhor poder anti-septico imediato e a feita com PVPI tem acao mais duradoura, impedindo o retorno dos microrganismos a superficie cutanea. Recomenda-se a anti-sepsia com o uso de povidone principalmente nas cirurgias mais demoradas