Selection of Neospora caninum antigens stimulating bovine CD4+ve T cell responses through immuno-potency screening and proteomic approaches

ABSTRACT: Neospora caninum is recognised worldwide as a major cause of bovine infectious abortion. There is a real need to develop effective strategies to control infection during pregnancy which may lead to either abortion or congenital transmission. Due to the intracellular nature of the parasite, cell-mediated immune (CMI) responses involving CD4+ve, CD8+ve, γ/δ TCR+ve T cells and NK cells, as well as production of IFN-γ, are thought to be important for protective immunity. In this study we applied a combination of proteomic and immunological approaches to identify antigens of N. caninum that are recognized by CD4+ve T cell lines derived from infected cattle. Initially, N. caninum tachyzoite Water Soluble Antigens (NcWSA) were fractionated by size-exclusion HPLC and then screened for immune-potency using CD4+ve T cell lines. LC-ESI-MS/MS (liquid chromatography electrospray ionisation tandem mass spectrometry) was employed to catalogue and identify the proteins comprising three immunologically selected fractions and led to the identification of six N. caninum target proteins as well as sixteen functional orthologues of Toxoplasma gondii. This approach allows the screening of biologically reactive antigenic fractions by the immune cells responsible for protection (such as bovine CD4+ve cells) and the subsequent identification of the stimulating components using tandem mass spectrometry.     

Use of key feature questions in summative assessment of veterinary medicine students

Purpose

To prove the hypothesis that procedural knowledge might be tested using Key Feature (KF) questions in written exams, the University of Veterinary Medicine Hannover Foundation (TiHo) pioneered this format in summative assessment of veterinary medicine students. Exams in veterinary medicine are either tested orally, practically, in written form or digitally in written form. The only question formats which were previously used in the written e-exams were Type A Single-choice Questions, Image Analysis and Short Answer Questions. E-exams are held at the TiHo using the electronic exam system Q [kju:] by CODIPLAN GmbH.

Methods

In order to examine less factual knowledge and more procedural knowledge and thus the decision-making skills of the students, a new question format was integrated into the exam regulations by the TiHo and some examiner used this for the first time in the computer based assessment. Following a successful pilot phase in formative e-exams for students, KF questions were also introduced in summative exams. A number of multiple choice questions were replaced by KF questions in four computer based assessment in veterinary medicine. The subjects were internal medicine, surgery, reproductive medicine and dairy science.

Results

The integration and linking of KF questions into the computer based assessment system Q [kju:] went without any complications. The new question format was well received both by the students and the teaching staff who formulated the questions.

Conclusion

The hypothesis could be proven that Key Feature questions represent a practicable addition to the existing e-exam question formats for testing procedural knowledge. The number of KF questions will be therefore further increased in examinations in veterinary medicine at the TiHo.     

THE CANINE TROCHANTERIC FOSSA: A RADIOGRAPHIC AND ANATOMIC STUDY

Radiographic features of the region of the trochanteric fossa were evaluated bilaterally on 175 ventrodorsal radiographs of canine hip joints and findings were compared to anatomic specimens. The fossa showed a variable radiographic appearance, which was affected by positioning and anatomic diversity. On a small number of radiographs (n = 12), the trochanteric fossa was clearly defined as a circular area of radiopacity surrounded by a radiolucent halo. This appearance was significantly influenced by medial rotation of the femur (P < 0.001). On some anatomic specimens an irregular protuberance representing the site of attachment of the gemelli, internal and external obturator muscles was seen within the fossa, surrounded by a variable number of small openings, which were thought to be nutrient foramina. Dissection identified a small number of minor branches of the medial circumflex femoral artery entering these.     

Effect of Fish Density During Transportation on Stress and Mortality of Juvenile Tambaqui Colossoma macropomum

The increased demand for juvenile tambaqui Colossoma macropomum for grow-out ponds and stocking programs in the Amazon state of Brazil has increased the transportation of this species. This study was designed to determine the optimum density of juvenile tambaqui during transportation in closed containers. Fish (51.9 ± 3.3 g and 14.9 ± 0.4 cm) were packed in sealed plastic bags and transported for 10 h at four densities: 78, 156, 234, and 312 kg/m³. After transportation, fish from each density were kept in separate 500-L tanks for 96 h. Mortality, 96-h cumulative mortality, water quality, and blood parameters (hematocrit, plasma cortisol, and glucose) were monitored. Fish mortality after transportation was significantly lower at densities of 78 and 156 kg/m³ than at 234 and 312 kg/m³. Cumulative mortality was significantly lower at a density of 78 kg/m³. Dissolved oxygen after 10 h of transportation remained high at a density of 78 kg/m³, but reached critically low values at all other densities. Ammonia concentration was highest at the lowest density and was lower at higher densities. Carbon dioxide concentration was lowest at the density of 78 kg/m³ but higher in the other treatments. Plasma glucose and cortisol increased significantly immediately after transportation at densities of 156, 234, and 312 kg/m³, returning to control values by 24 h. The best density for juvenile tambaqui during a 10-h transportation haul in a closed container was 78 kg/m³. At this density there was no fish mortality, water quality was kept within acceptable values, and fish were not stressed.     

Chronic ocular lesions associated with bi-directional microbeam radiation therapy in an experimental rat study for therapy of C6 and F98 gliomas

Objectives  To describe the chronic ocular lesions associated with microbeam radiation therapy (MRT) in an experimental rat study.
Procedures  MRT was administered bi-directionally with a skin entry dose of 350 Gy. During laterally directed irradiation, the beam entered the head on the right with the center of the beam array 3 mm posterior to the center of the right eye. During irradiation in anterior-posterior direction, the right eye was almost completely in the path of the beam array. Twelve months after MRT ophthalmic examinations were completed on 37 treated (MRT+) and 16 control (MRT–) rats. Electroretinography (ERG) was completed in two MRT+ and one MRT– rat. Histopathology was performed on eyes of 16 MRT+ and 9 MRT– rats, and retinal and choroidal thicknesses were measured.
Results  Biomicroscopic and indirect ophthalmoscopic examinations revealed fundus pallor and retinal vascular attenuation in 33 of 37 right and 2 of 37 left eyes of MRT+ rats. Cataracts were present in the right eyes of 12 of 37 MRT+ rats. ERG amplitudes were reduced in the eyes of MRT+ rats. Light microscopy revealed retinal lesions ranging in severity with loss of outer to inner retinal cell layers, in 16 of 16 right and of 8 of 16 left eyes MRT+ rats. The mean right retinal thickness of MRT+ rats was reduced.
Conclusions  Eyes within the field treated with MRT at a dose of 350 Gy develop retinal degeneration and occasionally, cataract.     

A simple way to identify non-viable cells within living plant tissue using confocal microscopy

Background  

Plant cell death is a normal process during plant development. Mutant plants may exhibit misregulation of this process, which can lead to severe growth defects. Simple ways of visualising cell death in living plant tissues can aid the study of plant development and physiology.     

Contrasting effects of puerarin and daidzin on glucose homeostasis in mice

Puerarin and daidzin are the major isoflavone glucosides found in kudzu dietary supplements. In this study, we demonstrated that puerarin significantly improves glucose tolerance in C57BL/6J-ob/ob mice, an animal model of type 2 diabetes mellitus, blunting the rise in blood glucose levels after i.p. administration of glucose. In contrast, daidzin, the O-glucoside, had a significant but opposite effect, impairing glucose tolerance as compared to saline-treated controls. When they were administered i.p. with (14)C-glucose to C57BL/6J lean mice, puerarin inhibited glucose uptake into tissues and incorporation into glycogen, while daidzin stimulated glucose uptake, showing an opposite effect to puerarin. Puerarin also antagonized the stimulatory effect of decyl-beta-D-thiomaltoside, an artificial primer of glycogen synthesis, which increases (14)C-glucose uptake and incorporation into glycogen in mouse liver and heart. A liquid chromatography-tandem mass spectrometry procedure was used to investigate the metabolism and bioavailability of puerarin and daidzin. The blood puerarin concentration-time curve by i.p. and oral administration indicated that puerarin was four times more bioavailable via i.p. injection than via the oral route of administration. This may account for the increased hypoglycemic effect seen in the i.p. glucose tolerance test vs that seen orally. Our results suggest that puerarin is rapidly absorbed from the intestine without metabolism, while daidzin is hydrolyzed to the aglycone daidzein. The opposing effects of puerarin and daidzin on glucose homeostasis may have implications for the activity of dietary supplements that contain both of these isoflavonoids.     

Ethyl hexanoate transfer modeling in carrageenan matrices for determination of diffusion and partition properties

Aroma compound properties in food matrices, such as volatility and diffusivity, have to be determined to understand the effect of composition and structure on aroma release and perception. This work illustrates the use of mass transfer modeling to identify diffusion and partition properties of ethyl hexanoate in water and in carrageenan matrices with various degrees of structure. The comparison of results obtained with a diffusive model to those obtained with a convective model highlights the importance of considering the appropriate transfer mechanism. Modeling of the preliminary experimental steps ensures correct estimation of the conditions for the main aroma release step. The obtained values of partition and diffusion coefficients are in agreement with those found in the literature (either experimentally determined or predicted by theoretical equations) and demonstrate that the structure level of carrageenan matrices has little influence on diffusion properties of ethyl hexanoate (less than 20%).