Complete coding sequence, sequence analysis and transmembrane topology modelling of Trypanosoma brucei rhodesiense putative oligosaccharyl transferase (TbOST II).

The partial nucleotide sequence of putative Trypanosoma brucei rhodesiense oligosaccharyl transferase gene was previously reported. Here, we describe the determination of its full-length nucleotide sequence by Inverse PCR (IPCR), subsequent biological sequence analysis and transmembrane topology modelling. The full-length DNA sequence has an Open Reading Frame (ORF) of 2406 bp and encodes a polypeptide of 801 amino acid residues.

Roles of CD122+ cells in resistance against Neospora caninum infection in a murine model.

Innate cells, such as natural killer (NK) cells and NKT cells, play essential roles as primary effector cells at the interface between the host and parasite until establishment of adaptive immunity. However, the roles of NK and NKT cells in defense against Neospora caninum have not been well clarified. NK and NKT cells were depleted by the treatment with an anti-CD122 (interleukin-2 receptor beta chain) monoclonal antibody (mAb, TM-β1) in vivo.

Observations on Cryptosporidium life cycle stages during excystation.

Cryptosporidium parvum (HNJ-1 strain, genotype 2) merozoites were released from oocysts directly during an incubation and excystation procedure without bleach treatment. They were polymorphic, mostly spindle-shaped; others were bean shaped, actively motile, and underwent division. Merozoites survived for short time-period in an in vitro culture system, but could not be established in a subsequent cultivation effort in RPMI medium.

Molecular characterization of Cryptosporidium from animal sources in Qinghai province of China.

The presence of Cryptosporidium oocysts in 20 zoo animals of the Xining Zoo, 16 farm yaks and 42 farm goats in Qinghai province, China was investigated by an immunofluorescence test (IFT). The species and/or genotypes were determined by nested polymerase chain reaction (PCR) and sequence analysis of a fragment of the small subunit (SSU) rRNA gene. Cryptosporidium oocysts were found in 16 zoo animals, 2 yaks, and 15 goats by IFT.

Effects of exposure to a 50 Hz electric field on plasma levels of lactate, glucose, free Fatty acids, triglycerides and creatine phosphokinase activity in hind-limb ischemic rats.

We previously reported that extremely low frequency electric fields (ELF-EFs) affect energy metabolism in stressed conditions. To further confirm this, the effect of exposure to ELF-EFs on the experimental ischemic rat was examined. The test was based on a comparison of rats treated with EF alone, ischemic surgery alone, the combination of EF with ischemic surgery, or no treatment (double sham).

Effects of a 50 Hz electric field on plasma lipid peroxide level and antioxidant activity in rats.

The effects of exposure to extremely low frequency electric fields (ELF EFs) on plasma lipid peroxide levels and antioxidant activity (AOA) in Sprague-Dawley rats were studied. The test was based on comparisons among rats treated with a combination of the oxidizing agent, 2,2'-azobis(2-aminopropane) dihydrochloride (AAPH) and 50 Hz EF of 17.5 kV/m intensity for 15 min per day for 7 days, AAPH alone, EF alone or no treatment.

Expression of P23 of Cryptosporidium parvum in Toxoplasma gondii and evaluation of its protective effects.

In this study, P23 of Cryptosporidium parvum sporozoites, an immunodominant surface protein, was stably expressed in Toxoplasma gondii (Tg/P23) and its protective effects were evaluated in a mouse model. The molecular weight and antigenic property of P23 expressed by Tg/P23 were similar to those of the native P23. Mice immunized with lysed Tg/P23 tachyzoites produced specific neutralizing antibodies against C.

Epidemiological survey of Babesia gibsoni infection in dogs in eastern Japan.

To determine the distribution of Babesia gibsoni infection in dogs in the eastern part of Japan, an epidemiological survey of dogs suspected of having B. gibsoni infection was attempted using the polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). Thirty-five of 115 such dogs (30.

Serodiagnosis of Babesia gibsoni infection in dogs by an improved enzyme-linked immunosorbent assay with recombinant truncated P50.

The surface antigen P50 of Babesia gibsoni is an important candidate for the development of a diagnostic reagent for canine piroplasmosis. In order to establish an effective diagnostic method for practical use, the gene encoding truncated P50 (P50t) lacking a signal peptide and C-terminal hydrophobic regions were cloned and expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST). More than 90% portion of the GST-P50t was expressed as a soluble form, in contrast with GST-P50f (full-length), which was completely expressed as an insoluble form.

The detection of bovine lactoferrin binding protein on Trypanosoma brucei.

Trypanosoma brucei, the causative agent of sleeping sickness in humans, requires transferrin (TF) for growth. Therefore, T. brucei has a TF receptor that allows it to obtain iron from TF.

Serodiagnosis of canine Babesia gibsoni infection by enzyme-linked immunosorbent assay with recombinant P50 expressed in Escherichia coli.

The entire P50 gene encoding a surface protein of Babesia gibsoni was cloned into the bacteria expression vector pGEX-4T-3 and subsequently expressed in Escherichia coli as a glutathione S-transferase fusion protein. The purified recombinant P50 was evaluated in an enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of B. gibsoni infection in dogs.

60 Hz electric field upregulates cytosolic Ca2+ level in mouse splenocytes stimulated by lectin.

The effect of a 60 Hz electric field (EF) on alteration of cytosolic free Ca2+ level ([Ca2+]c) was examined in mouse splenocytes stimulated by lectins, namely concanavalin A (ConA) or phytohemagglutinin. In order to understand the role of EF on alterations in [Ca2+]c and to determine whether EF exposure increased cell mortality the splenocytes were cultured under the 60 Hz EFs producing current densities of 6 or 60 microA/cm2 for 30 min or 24 h. Cell mortality was less than 2% in experimental all conditions.

Inhibitory effect of antiserum to surface antigen P50 of Babesia gibsoni on growth of parasites in severe combined immunodeficiency mice given canine red blood cells.

The inhibitory effect of an antiserum to surface protein P50 of Babesia gibsoni on the growth of the parasite was determined with severe combined immunodeficiency mice given canine red blood cells. The antiserum to the recombinant P50 protein significantly inhibited the parasite growth, indicating that P50 might be a useful vaccine candidate.

Immunology. Antigen-presenting cells in the gut.

It has been known for the past 85 years that mucosal responses can be stimulated by local presentation of antigen and that the gut immune system is capable of mounting both primary and secondary responses to potentially harmful antigens while avoiding the expression of damaging responses to harmless dietary proteins. How these types of responses are induced and regulated remains unclear. In the gut attention has for some time been focused on Peyer's patches (PP) due to evidence that they play a vital role in the induction of humoral and cellular responses.

Serodiagnosis of Neospora caninum infection in cattle by enzyme-linked immunosorbent assay with recombinant truncated NcSAG1.

Neospora caninum is a veterinary medically important pathogen capable of causing abortion in cattle and neuromuscular paralysis in dogs. The surface antigen 1 of N. caninum (NcSAG1) is an important candidate for the development of a diagnostic reagent for neosporosis.

The detection of bovine lactoferrin binding protein on Toxoplasma gondii.

Lactoferrin (LF), a member of the transferrin (TF) protein family, is an iron-binding protein that is known to interact with bacteria through a specific receptor. We examined the binding of bovine LF (bLF), bovine TF (bTF), and ovotransferrin (OTF) by Toxoplasma gondii using a fluorescence test and the streptavidin-biotin (SAB) method using biotin-streptavidin, and found that bLF, bTF, and OTF bound to the protein components of T. gondii.

Characterisation of Toxoplasma gondii engineered to express mouse interferon-gamma.

Recent studies have shown the feasibility of using Toxoplasma gondii as an expression system for heterologous protein. For better understanding of the mechanism of interferon-gamma (IFN-gamma) dependent immunity to T. gondii, the parasites were stably transfected with IFN-gamma gene, under control of the GRA1 promoter.

A role for balance of interferon-gamma and interleukin-4 production in protective immunity against Neospora caninum infection.

A suitable balance in the production of Th1/Th2-type cytokines has a crucial role in the control of microbial infections. We investigated cytokine production patterns and effects during Neospora caninum infection, based on two mouse models and an in vitro system. In the acute infection of N.

Molecular characterization of a gene encoding a 29-kDa cytoplasmic protein of Babesia gibsoni and evaluation of its diagnostic potentiality.

A cDNA expression library prepared from Babesia gibsoni merozoite mRNA was screened with B. gibsoni-infected dog serum. cDNA encoding 29-kDa protein was cloned and designated as the P29 gene.

High-level expression of truncated surface antigen P50 of Babesia gibsoni in insect cells by baculovirus and evaluation of its immunogenicity and antigenicity.

Previously, we identified an immunodominant antigen, P50 of Babesia gibsoni. In the present study, the gene encoding the truncated P50 (rP50t) without a C-terminal hydrophobic region (29 amino acids [aa]) was expressed in insect cells by a recombinant baculovirus. The highly hydrophobic C-terminal 20-aa regions seems to be a transmembrane region, which was evidenced by the fact that rP50t was effectively secreted into the supernatant of insect cells infected with the recombinant baculovirus.

A comparison of the phenotype of dendritic cells derived from discrete Peyer's patch macrophages of non-infected and Toxoplasma gondii infected mice.

A comparison of the expression of surface membrane antigens between dendritic cells (DC) derived from Peyer's patch macrophages (DPP-DC) of non-infected and Toxoplasma gondii (T. gondii) infected mice was performed. C57BL/6J mice aged 6-8 weeks of both sexes were infected orally with a 0.

Viability and infectivity of Cryptosporidium parvum oocysts detected in river water in Hokkaido, Japan.

The viability and infectivity of Cryptosporidium parvum (C. parvum) oocysts, detected in water samples collected from river water in Hokkaido, were investigated using Severe Combined Immunodeficient (SCID) mice. The water samples collected from September 27 through October 10, 2001 by filtration using Cuno cartridge filters were purified and concentrated by the discontinuous centrifugal flotation method.

Recombinant bovine herpesvirus-1 expressing p23 protein of Cryptosporidium parvum induces neutralizing antibodies in rabbits.

In order to develop a vaccine against cryptosporidiosis in cattle, we constructed a recombinant bovine herpesvirus-1 (BHV-1) expressing an immunodominant surface protein, p23, of Cryptosporidium parvum sporozoites. In the recombinant virus, the p23 gene under the control of a CAG promoter and a gene coding for an enhanced green fluorescent protein were integrated into the gG gene of BHV-1. Despite a low frequency of homologous recombination, cloning of the recombinants was easy because of the specific fluorescence of the plaques formed by recombinants.

Phenotype and function of murine discrete Peyer's patch macrophage derived - dendritic cells.

The phenotype and function of peritoneal cavity macrophage-derived dendritic cells (PEC-DC) was previously reported. In this study we have gone further in using our established culture system to generated discrete Peyer's patch dendritic cells (DPP-DC) from murine discrete Peyer's patch macrophages (DPP-Mø), following stimulation with granulocyte macrophage colony stimulating factor (GM-CSF) plus interleukin 4 (IL-4) for 7 days. DPP-Mø from murine small intestines were obtained by mechanical disruption of discrete Peyer's patches (DPP), followed by metrizamide density gradient centrifugation to remove Peyer's patch resident DC and debri, after which an overnight adherent step in tissue culture medium was carried out for macrophage enrichment.