B virus diagnosis presents a challenge largely complicated by the asymptomatic infection of rhesus macaques, and extremely pathogenic fatal infections in humans. Humoral detection of antibodies is generally performed using whole virus antigen for which preparation requires strict biosafety measures and specialized BSL-4 facilities. As an alternative to utilizing B virus antigen, we describe the production of a truncated form of B virus envelope glycoprotein D, gD 287, in a baculovirus expression system, and evaluate its diagnostic potential as an antigen in recombinant ELISA. After purification and characterization, gD 287 was tested using 22 negative and 72 positive macaque sera samples previously classified using the traditional method. We find that sensitivity and specificity of the recombinant ELISA are dependent on antibody titer of tested serum and gD 287 shows good to excellent predictive potential for identification of positive sera with titers higher than 500.

Hemos realizado en la Región de Murcia una investigación epidemiológica sobre los coronavirus porcinos- virus de la Gastroenteritis Transmisible (TGEV) y/o Coronavirus Respiratorio Porcino (PCRV)- mediante una técnica inmuno-enzimática (ELISA indirecta). La encuesta serológica se ha llevado a cabo en 1987 en la población de cerdos reproductores y la muestra investigada (6.000 reproductores de 480 granjas) es representativa respecto a los reproductores y a las explotaciones – y dentro de ellas según cuatro categorías de tamaños: familiares (1-9), pequeñas (10-40), medianas (50-99) y grandes (≥100)- censadas en cuatro zonas geográficas (Cartagena, Valle del Guadalentín, Huerta de Murcia y resto de la Región). En Murcia se han detectado anticuerpos de coronavirus porcinos (TGEV y/o PCRV) en 937 reproductores (prevalencia 15,61±0.91%) de 107 granjas de reproducción (prevalencia 22,29±3.72%). Los coronavirus porcinos están muy difundidos en toda la Región de Murcia, con mayor prevalencia en Cartagena y Valle del Guadalentín, y en explotaciones con más de 50 reproductores.

A 20 weeks project at Karolinska Institutet (KI), Huddinge, Sweden is in this master thesis summarized. Alzheimer’s disease is the most common form of dementia in the world. One of the pathological hallmarks seen in AD patients consists of amyloid plaques assembled of beta amyloid (Aβ) peptide aggregates. A lot of research has been done on Aβ40 and Aβ42 but not on the longer variant with 43 residues. An earlier study by Welander et al, quantified the Aβ43 peptide from amyloid plaque cores with high-performance liquid chromatography coupled to mass-spectrometry (HPLC-MS/MS)1. Here, I present the initial development of an enzyme-linked immunosorbent assay (ELISA) with the goal to quantify Aβ43 peptides in soluble fractions of human brain tissue. An ELISA method with the possibility to quantify Aβ43 peptides from cerebral spinal fluid might have the prospect to serve as a diagnostic tool for AD in the future. Commercial ELISA kits coated with antibodies against all Aβ species was not suitable for detecting Aβ43 in soluble brain tissue from human AD patients. This is due to the high amount of Aβ40 (and in some extent Aβ42) in the samples, which will bind to the same epitope as Aβ43 on the capturing antibody. These shorter Aβ species will be in excess and bind to the capturing antibody thereby ousting Aβ43 from binding in. A better way for quantifying Aβ43 with ELISA might instead be to coat a polystyrene plate with α-Aβ43 antibodies, which are c-terminal specific to Aβ43. This will abolish the competition between the different Aβ species and function as an immunoprecipitation of unwanted species. This yielded adequate quantification of Aβ43 (2.64 pM) from tris-buffer saline (TBS) fractions from a human brain sample from AD.

Agriculture is a valuable economic resource in western Canada, but for decades farmers have focused on intensive production practices while ignoring the long-term health and maintenance of the land. In recent years, the use of conservation agricultural techniques has been encouraged in an effort to conserve prairie landscape while sustaining cropland productivity. Sustainable agricultural practices that promote soil and water conservation and benefit wildlife and prairie biodiversity include conservation tillage and planting of winter cereal crops. Many species of wild birds nest in the ground cover provided by minimum tillage and fall seeded cropland in the spring. Although habitat quality in conservation areas is superior for birds, there is potential for eggs of ground nesting birds to be exposed to herbicides during spring weed control operations. Herbicides commonly used on the prairies to control weed growth in conservational systems include 2,4-D and Buctril-M®. Since the subtlethal effects of exposure to these herbicides may include DNA damage and immunomodulation, the overall goal of this study was to assess whether <i>in ovo</i> exposure to the herbicides 2,4-D and Buctril-M® adversely affects genetic material and/or immune system function in newly hatched domestic chickens (<i>Gallus gallus</i>) and ducks (<i>Anas platyrhynchos</i>), as surrogates for wild bird species.<p>Study design attempted to reproduce actual field exposures by use of an agricultural field spray simulator to apply formulated herbicides (as opposed to pure active ingredients) at recommended crop application rates. In three separate experiments, fertile chicken eggs were sprayed with 2,4-D ester formulation or with Buctril-M® formulation, and fertile duck eggs were sprayed with 2,4-D ester formulation, during either an early (embryonic day 6) or late (embryonic day 15 for chickens or embryonic day 21 for ducks) stage of incubation. Genotoxicity and immune system function were evaluated in the hatchlings as the main toxicological endpoints to assess potential subtle effects from herbicide exposure, but additional measures of general health and development were also evaluated. Two endpoints were used to assess subtle changes to genetic integrity. The comet assay was used to detect structural damage (strand breaks) in avian lymphocyte DNA, as an index of acute genotoxic effects. Flow cytometry was used to examine potential clastogenic effects of the herbicides, by determining if chromosomal changes resulted in variability in the DNA content of avian erythrocytes. Several endpoints were examined to evaluate potential exposure-induced effects on the immune system. Immunopathological assessment of chicks and ducklings included differential lymphocyte counts, as well as immune organ weights and histopathology. The cell-mediated and humoral immune responses in hatchlings were assessed using the delayed-type hypersensitivity test and measurement of systemic antibody production in response to immunization, respectively. Exposure of fertile chicken and duck eggs to Buctril-M® or 2,4-D had no effects on the biomarkers of genetic integrity in this study. Differences in herbicide treatment (high and low concentrations) and times of exposure (early and late incubation stages) did not translate into noticeable factor effects in final model analyses for any of the genotoxicity assay variables evaluated in newly hatched chickens exposed in ovo to 2,4-D. Similarly, comet assay outcomes in chicks exposed to Buctril-M® were not significantly associated with either herbicide treatment or time of exposure as fixed effect factors. Results of the comet assay using peripheral lymphocytes from ducklings provided evidence of potential primary genetic damage associated with the time of spray exposure in ovo. Comet tail DNA content was significantly associated (P = 0.0269) with exposure times, suggesting that ducks may be increasingly sensitive to spray exposure conditions at an early stage of embryological development. Effects of exposure timing were not attributable to herbicide treatment. Although 2,4-D exposure time was associated with DNA strand breakage in ducklings, there was no evidence of chromosomal damage. However, an association between the HPCV values (a measure of DNA content variability) and time of spray exposure was observed in the experiment where 21-day-old chickens were treated in ovo with Buctril-M®. The mean HPCV value for the early exposure group (E6) was significantly greater (P = 0.0210) than that of the group treated later in incubation (E15). However, Buctril-M® the concentration of herbicide did not have any influence on this outcome, and the reason for the difference between exposure times is uncertain, but may be attributed to stress associated with manipulations during spraying. An increase in HPCV, reflecting greater intercellular DNA variability, is indicative of increased incidence of chromosomal damage, which may be an effect of disturbance during early periods of incubation as a result of exposure conditions.<p>Among the panel of immunotoxicity tests conducted to evaluate the effects of <i>in ovo</i> exposure to 2,4-D and Buctril-M® on the developing avian immune system, only heterophil/ lymphocyte (H/L) ratios and relative immune organ weights were significantly associated with either herbicide treatment or time of spray exposure in all three experiments. In 21-day-old chicks exposed in ovo to 2,4-D, relative bursa weight was associated with the different herbicide treatments (P = 0.0006). Relative bursa weights were significantly lower in chicks in the low dose group, while the opposite effect was observed in the high dose chicks, compared with the controls. It is unlikely that the observed decrease in bursa weight in the low dose group is causally related to herbicide exposure because a consistent dose-response effect was not observed, but this outcome may be explained by a compensatory immune response. The relative spleen weights of newly hatched chickens exposed in ovo to Buctril-M® exhibited a significant association with herbicide treatment (P = 0.0137). Relative spleen weights for birds in the low dose treatment groups were significantly different than both the control (P = 0.0179) and high dose groups (P = 0.0125). However, there was no significant difference between high dose and control groups, and this outcome reduces the likelihood of a causal relationship between spleen weight and herbicide exposure. In the parallel experiment involving in ovo exposure to 2,4-D to ducklings, relative bursa weight was associated with time of spray exposure (P = 0.0434). Ducklings that hatched from eggs exposed to spray on day 6 of incubation exhibited greater mean relative bursa weights than the birds exposed to spray at a later incubation stage (E21). This result implies that spray exposure during earlier stages of development may result in conditions which affect the humoral immune response, if increased bursal weight is associated with increased B lymphocyte and antibody production. In the same experiment, mean H/L ratios in peripheral blood samples from 21-day-old ducklings were significantly different between the groups treated with the high concentration of 2,4-D and water (control) (P = 0.0395). Although ratios from the birds in the low dose groups were not significantly different from the control groups, changes in H/L ratio values demonstrate a dose dependent relationship with increasing herbicide exposure.<p>Residue analysis of chicken and duck eggs in this study measured transfer of herbicide through the shell and into the embryo 24 hours and up to 5 days (chickens only) after spraying. Mean 2,4-D residue concentrations were higher in both chicken and duck eggs from the high dose (10X) groups than in eggs exposed to the recommended field rate of herbicide application (1X). Embryo residue concentrations in the chicken eggs increased from the day following exposure to 5 days after spraying, in both low and high dose groups. This observation indicates that the risk of contaminant-induced adverse effects may continue to increase for at least several days after exposure, thereby influencing the concentration of herbicide to which the developing embryo is exposed.<p>On the Canadian prairies, wild bird eggs are potentially to be exposed to 2,4-D and Buctril-M® during various stages of embryonic development. The present study examined effects of herbicide exposure at two distinct times during incubation, and demonstrated the potential for subtle impacts on genetic integrity and the immune system. Results indicate that spray exposure during earlier stages of organogenesis may cause more significant adverse effects. Given the possible harmful consequences of the observed changes on the long-term health of wild birds, further research is needed in order to better characterize the risks of in ovo agrochemical exposure in prairie ecosystems.

The present study aimed detection of a human pathogen B. bugdorferi sensu lato species in suspected Lyme borreliosis (LB) patients in Turkey by PCR analysis and supportive serologic tests. The 152 clinical samples (140 serum and blood, 10 cerebrospinal fluid (CSF), 1 synovial fluid, 1 skin biopsy specimens) from 140 patients sent from 22 different cities of Turkey to The Spirochetal Diseases Diagnosis Laboratory of Central Veterinary Control and Research Institute were analysed. Serum samples were subjected to ELISA with a commercial kit and all of the blood, CSF, synovial fluid and skin biopsy samples were examined by PCR. In PCR analysis two primer sets targeting the ospA gene located on the plasmid and ribosomal 23S rRNA gene of B. burgdorferi sensu lato were used. The results indicated that 32,1% (45 of 140) seropositivity was detectable by ELISA. Our results support that there is a risk of acquiring LB in different regions of Turkey. Although considerable positive detections were recorded using serologic tests,none of the specimens were positive in PCR analysis. Further studies on PCR based methods for detection of B. burgdorferi sensu lato in patients with a high clinical probability of LB apparently may require that the specimen should be taken in the early phases and before the administration of any therapeutic agent.

No description available.

Increased reactive oxygen species (ROS) have been reported to be at the centre of various diseases. Although several reports have implicated elevated levels of ROS in the pathogenesis of diabetes mellitus, the early detection of ROS is still not attainable. This limitation causes difficulty in the early diagnosis of ROS related disorders. The presence of high levels of ROS was reported to result in differential expression of antioxidant genes involved in protecting cells from their deleterious effects. Among the antioxidant genes that are expressed, it was postulated that expression of metallothioneins (MTs) are also induced. MTs are low molecular weight, cysteine-rich proteins involved in metal homeostasis and reported to harbour antioxidant function. The aim of this investigation was to explore MTs as biomarkers for elevated levels of ROS in whole blood of type 2 diabetic (T2D) individuals. The level of ROS in diabetic, non-diabetic as well as individuals at risk of developing T2D was determined via the use of biochemical assays. Real-Time PCR was utilised to analyse the expression of MTs and the presence of MT proteins was analysed via the ELISA. In this study it was observed that diabetic individuals had elevated levels of ROS. However, no significant difference in the expression of MTs and the presence of MT proteins between the diabetic and non-diabetic individuals was observed. In vitro experimental conditions indicated that MT expression is induced by elevated levels of ROS. In pathological conditions the ROS-dependent induction of MT expression needs to be elucidated further. It therefore can be suggested that MTs can not yet be utilised as biomarkers for the detection of elevated levels of ROS in pathological conditions with ROS aetiology. This investigation also highlights the fact that blood is not an optimal medium in which this objective can be attained. / Thesis (Ph.D. (Biochemistry))--North-West University, Potchefstroom Campus, 2005.

La Asamblea Barrial de Villa Elisa se inscribe en el escenario público platense como una forma de participación colectiva que, a través de la reflexión y acción de sus integrantes, tiende a redefinir las subjetividades sociales impactando en la construcción de ciudadanía. Tal interpretación se logra a partir del estudio de los datos recabados durante reuniones, marchas y entrevistas protagonizadas por los asambleístas en 2006. Así el concepto de diálogo desarrollado por Paulo Freire, es utilizado como categoría de análisis que, junto al diseño de diferentes subcategorías permite identificar a la asamblea como un espacio de interacción y participación ciudadana. Asimismo, resulta imprescindible dar cuenta del marco histórico en el que se inscribe el desarrollo de este movimiento que surge tras la crisis político institucional cuya máxima expresión tuvo lugar el 19 y 20 de diciembre de 2001. Los vecinos se juntaron ante la necesidad de exteriorizar inquietudes vinculadas tanto a su experiencia y formación personal como a un sentimiento compartido de inconformismo social. El diálogo aparece entonces, como el espíritu común de la asamblea en el que se manifiesta el pensamiento de quienes participan del encuentro. / Programa de investigación: Comunicación, Prácticas Socioculturales y Subjetividad

Streptococcus pneumonine viršutiniuose kvėpavimo takuose sutinkamos gram-teigiamos bakterijos, galinčios sukelti pneumoniją, meningitą, vidurinės ausies uždegimą, sinusitus ir kitus susirgimus. Neatspariausi šiems susirgimams yra vaikai. Vienas šio darbo tikslų buvo ištirti streptokokų paplitimą ikimokyklinio amžiaus grupės sveikų vaikų populiacijoje. Buvo ištirtas 601 nosiaryklės tepinėlis, paimtas iš 2-7 metų amžiaus vaikų, lankančių ikimokyklines ugdymo įstaigas Vilniaus mieste. Streptococcus pneumoniae išaugo 280 vaikų tepinėlių pasėliuose (47 proc.). Viso rasti 22 skirtingi serotipai; nustatyti dažniausiai šioje populiacijoje sutinkami serotipai 19F,23F, 6B, 6A, 3, 18C. Labai dažnai pneumokokai susitelkia nosiaryklės gleivinės epitelyje nesukeldami jokių ligos simptomų. Šiuo atveju ypatingą reikšmę įgauna žmogaus imuninė sistema, todėl buvo tiriamas atskirų klasių imunoglobulinų (IgG, IgA, IgM) kiekis infekuotų vaikų seilių pavyzdžiuose. Šiems tyrimams buvo panaudoti monokloniniai antikūnai, gauti hibridominės technologijos pagalba. Imunofermentiniu metodu seilių mėginiuose buvo nustatytas IgA, sIgA, IgG, IgM klasių antikūnų kiekis, o taip pat specifinių S. pneumonine antikūnų kiekis; nustatyta, kad koreliacija tarp duomenų grupių (bendras konkrečios klasės imunoglobulinų skaičius, specifinių antikūnų skaičius) yra vidutinė arba silpna. Tačiau gleivinių imuniteto vertinimas imunofermentiniu metodu yra pakankamai informatyvus ir perspektyvus, ypač dėl tos priežasties, kad... [toliau žr. visą tekstą] / Streptococcus pneumonine is a gram-positive bacterial organism that colonizes the upper respiratory tract and causes diseases such as pneumonia, meningitis, otitis media, sinusitis and others. Disease rates are particularly high in young children. One of the aims of this study was to assess the prevalence rates of streptococcal infection among the population of healthy children of preschool age. 601 nasopharyngeal samples were tested from 2-7 years age kindergardeners in Vilnius. Streptococcus pneumonine were grown from 280 samples (47 % of collected samples). 22 different serotypes were identified; the most frequent serotypes were determined 19F, 23F, 6B, 6A, 3, and 18C. Frequently S.pneumonine colonizes mucosal epithelium at nasopharynx without causing any symptoms. In that case the immunity of an organism is of great significance. The second aim of this study is to evaluate the immune response to pneumococcal infection. The amount of separate immunoglobuline classes (IgG, IgA, and IgM) were assessed in saliva samples of infected children. Monoclonal antibodies developed by hybridoma technology were applied. The amounts of classes IgA, sIgA, IgG, IgM antibodies were determined parallel to amount of specific antibodies to S.pneumonine. Correlation between data groups (total amount of actual immunoglobulin class, concentration of specific antibodies) was weak or moderate. Generally, evaluation of mucosal immunity by enzyme immunoassay is informative and perspective especially... [to full text]

Four analogues of the Lewis X trisaccharide antigen (β-D-Galp(1→4)[α-L-Fucp(1→3)]-D-GlcNAcp) in which the galactosyl residue is modified at O-4 as a methyloxy, deoxychloro, deoxyfluoro or deoxy were synthesized. The preparation of the modified 4-OMe, 4-Cl, 4-F and 4-H trichloroacetimidate galactosyl donors is described as well as their use in the glycosylation of an N-acetylglucosamine glycosyl acceptor. The resulting disaccharides were deprotected at O-3 of the glucosamine residue and fucosylated giving access to the desired protected Lewis X analogues. One step global deprotection (Na/NH3) of the protected 4”-methoxy and 4”-deoxy analogues, and two step deprotections (removal of a p-methoxybenzyl with DDQ, then Zemplén deacylation) of the 4”-deoxychloro and 4”-deoxyfluoro protected Lex analogues gave the desired compounds in good yields. The relative binding energies of the 4” manipulated Lewis X methyl glycoside analogues for anti-Lewis X mAb SH1 were determined using competitive ELISA procedures. The binding experiments revealed that substitution of the 4” site with a deoxychloro, deoxyfluoro or deoxy function had a large differential change in the free energy of binding; yet they did not completely abolish cross reactivity with native Lewis X. Substitution with a methyloxy function eliminated recognition by anti-Lewis X monoclonal antibody SH1. / NSERC