Toxoplasma gondii: Comparison of a rhoptry-ELISA with IFAT and MAT for antibody detection in sera of experimentally infected pigs

Indirect ELISA and IFAT have been reported to be more sensitive and specific than agglutination tests. However, MAT is cheaper, easier than the others and does not need special equipment. The purpose of this study was to compare an enzyme linked immunosorbent assay using crude rhoptries of Toxoplasma gondii as coating wells (r-ELISA) with indirect fluorescence antibody test (IFAT) and modified agglutination test (MAT) to detect anti-T. gondii antibodies in sera of experimentally infected pigs. Ten mixed breed pigs between 6.5 and 7.5 weeks old were used. All pigs were negative for the presence of T. gondii antibodies by IFAT (titre < 16), r-ELISA (OD < 0.295) and MAT (titre < 16). Animals received 7 × 107 viable tachyzoites of the RH strain by intramuscular (IM) route at day 0. Serum samples were collected at days -6, 0, 7, 14, 21, 28, 35, 42, 50, and 57. IFAT detected anti-T. gondii antibodies earlier than r-ELISA and MAT. The average of antibody levels was higher at day 35 in IFAT (Log10 = 2.9) and in MAT (Log10 = 3.5), and at day 42 in r-ELISA (OD = 0.797). The antibody levels remained high through the 57th day after inoculation in MAT, and there was a decrease tendency in r-ELISA and IFAT. IFAT was used as gold standard and r-ELISA demonstrated a higher prevalence (73.3%), sensitivity (94.3%), negative predictive value (83.3%), and accuracy (95.6%) than MAT. Kappa agreements among tests were calculated, and the best results were shown by r-ELISA × IFAT (κ = 0.88, p < 0.001). Cross-reaction with Sarcocystis miescheriana was investigated in r-ELISA and OD mean was 0.163 ± 0.035 (n = 65). Additionally, none of the animals inoculated with Sarcocystis reacted positively in r-ELISA. Our results indicate that r-ELISA could be a good method for serological detection of T. gondii infection in pigs. © 2005 Elsevier Inc. All rights reserved.

Avaliação de diferentes híbridos suínos submetidos à insensibilização elétrica e gasosa (CO 2). Parte 1 - Mensuração de indicadores sanguíneos de estresse

Pigs of three genetics lineages A, B and C marketed in Brazil, with alive weight from 100 to 120 kg were submitted to the manual electric stunning (Karl Schermer 220-230/250 volts, 45-60 Hz and 1.4 -1.5 A) and to the collective gaseous system (COMBI-BUTINA 90% CO 2). Blood samples, for levels determination of creatine phosphokinase (CPK), lactate and cortisol, as well as samples of the semimembranosus muscle (10 g) for the determination of the gene halothane, were collected. Being compared the electric and gaseous stunning systems, the electric stunning did demonstrate to be more stressful providing larger plasmatic concentrations of cortisol (p ≤ 0.001) and lactate (p ≤ 0.001) for the genetic lineages A and C, in the studied conditions. However it didn't observe significant differences beween the sanguine indicators and stunning systems in subject when the lineage B was considered. Significant differences among the genetic lineages A, B and C were obtained being compared the plasmatic values of creatine phosphokinase (p ≤ 0.001), lactate (p ≤ 0.001) and cortisol (p ≤ 0.001) when stunned with the gaseous system, however when the electric system was used only the cortisol values presented significant differences (p ≤ 0.001). The presence of the gene halothane (Nn) was only observed in the lineage B.

Toxoplasma gondii: Detection by mouse bioassay, histopathology, and polymerase chain reaction in tissues from experimentally infected pigs

In the present study, we evaluated three techniques, mouse bioassay, histopathology, and polymerase chain reaction (PCR) to detect Toxoplasma gondii infection in tissues from experimentally infected pigs. Twelve mixed breed pigs, seronegative for T. gondii using an indirect immunofluorescent antibody test (IFAT), were used. Ten pigs were infected with 4 × 104 VEG strain oocysts, and two were maintained as uninfected controls. Animals were killed 60 days pos infection. Muscle (heart, tongue, diaphragm, and masseter) and brain samples were collected to investigate the presence of T. gondii tissue cysts by the different assay methods. For the bioassay, samples of brain (50 g) and pool of muscle samples (12.5 g of tongue, masseter, diaphragm, and heart) were used. PCR was performed using Tox4 and Tox5 primers which amplified a 529 bp fragment. The DNA extraction and PCR were performed three times, and all tissue samples were tested individually (brain, tongue, masseter, diaphragm, and heart). For histopathology, fragments of tissues were fixed in 10% of buffered formal saline and stained with HE. Histopathological results were all negative. PCR showed 25/150 (16.6%) positive samples, being 17/120 (14.1%) and 8/30 (26.6%) from muscle, and brain tissues, respectively. Tissue cysts of T. gondii were identified by mouse bioassay in 54/98 (55.1%) samples, being 31/48 (64.6%) from muscle samples, and 23/50 (46.0%) from brain samples. Toxoplasma gondii isolation in muscle samples by mouse bioassay was higher than in PCR (P < 0.01). Results indicate that DNA from pig tissues interfered with 529-bp-PCR sensitivity, and mouse bioassay was better than PCR in detecting T. gondii in tissues from pigs. © 2006 Elsevier Inc. All rights reserved.

Production of monoclonal antibodies against Streptococcus mutans antigens

Several studies have been conducted in the last decades aiming to obtain an anti-canies vaccine, however some studies have demonstrated cross reactivity between Streptococcus mutans surface antigens and the human cardiac tissue. In this work, the reactivity of five anti-Streptococcus mutans monoclonal antibodies (MoAb) (24A, 56G, G8, E8 and F6) was tested against oral streptococci, cardiac antigens and skeletal and cardiac myosins, aiming to evaluate the specificity of these MoAb. The hybrid producers of immunoglobulins of the IgG 2b class were cloned by limit dilution and expanded in vivo. MoAb were tested by ELISA. The hybrid 24A reacted with S. mutans CCT 1910, S. salivarius CCT 0365 and S. pyogenes T23. No reactivity difference was observed among the tested species. Cross reactivity with heart and cardiac myosin was not confirmed and only reaction with myosin of skeletal muscle was observed (p = 0.0381). The hybrid 56G reacted with all the tested microorganisms and there was statistically significant difference between S. mutans and S. pyogenes T23 (p < 0.001). This hybrid also reacted with myosin of skeletal muscle (p = 0.0095). C8, E8 and F6 presented low reactivity against oral streptococci strains and no reactivity against cardiac antigens. The data of this study showed that the 24A and 56G anti-S. mutans MoAb presented reactivity with S. pyogenes and S. salivarius, reinforcing the occurrence of common antigens between these species. The tested MoAb presented low cross-reactivity with myosin of skeletal muscle, but anti-heart activity could not be confirmed.

Efeito da adição de probiótico e/ou prebiótico em dietas de leitões desmamados sobre o desempenho, incidência de diarréia e contagem de coliformes totais

In the Experiment 1, 50 pigs weaned at the 21 days were fed with either of five diets: T1 - basal diet; T2 - basal diet + antibiotic; T3 - basal diet + probiotic; T4 - basal diet + prebiotic; T5 - basal diet + simbiotic. The variables studied were body weight, feed intake and feed conversion in the Phase 1 (21 to 43 days), Phase 2 (44 to 57 days), Phase 3 (58 to 70 days) and Total Phase (21 to 70 days). During the performance experiment, a fecal survey score was conducted to verify diarrhea incidence. In the Experiment 2, 44 pigs weaned at 21 days were fed with the same diets of Experiment 1. Pigs were slaughtered at three differents ages (at weaning, at seven and 14 days after weaning). A segment of the small intestine was collected for analisys of total coliformes. The results showed that the best performance was obtained with the utilization of prebiotic and simbiotic. There were not differences in relation to diarrhea incidence among the treatments studied. The addition of probiotic and/or prebiotic in the diet prevented increase of colonization by pathogenic bacterias from seven to 14 days after weaning.

Avaliação histológica da pele após exposição a gel acrescido de hialuronidase associado ou não a ultra-som

The objective of this work was to verify the effect in the skin of male swines gel (G) containing hyaluronidase (H) associated or not to ultrasound (US). In different areas was applied G; G+US; G+H; G+H+US and mesotherapy (M). Skin fragment was processed in paraffin. To evidence hyaluronic acid (HA) coloration with Alcian Blue (AB) was used and coloration with Hematoxilin/Eosin for morphometry. It was observed that G+H and G+H+US did not reduce coloration for the AB nor presented significant differences for the morphometry. When H was applied mesoteraphycally coloration for the AB diminished. Then, the use of H associated or with US did not seem efficient in the HA reduction.

Taenia solium and Taenia saginata: Identification of sequence characterized amplified region (SCAR) markers

Cysticercosis is one of the most important zoonosis, not only because of the effects on animal health and its economic consequences, but also due to the serious danger it poses to humans. The two main parasites involved in the taeniasis-cysticercosis complex in Brazil are Taenia saginata and Taenia solium. Differentiating between these two parasites is important both for disease control and for epidemiological studies. The purpose of this work was to identify genetic markers that could be used to differentiate these parasites. Out of 120 oligonucleotide decamers tested in random amplified polymorphic DNA (RAPD) assays, 107 were shown to discriminate between the two species of Taenia. Twenty-one DNA fragments that were specific for each species of Taenia were chosen for DNA cloning and sequencing. Seven RAPD markers were converted into sequence characterized amplified region (SCAR) markers with two specific for T. saginata and five specific for T. solium as shown by agarose gel electrophoresis. These markers were developed as potential tools to differentiate T. solium from T. saginata in epidemiological studies. © 2007 Elsevier Inc. All rights reserved.

Produção de pimentão em substratos e fertirrigação com efluente de biodigestor

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Evaluation of IFA, MAT, ELISAs and immunoblotting for the detection of anti-Toxoplasma gondii antibodies in paired serum and aqueous humour samples from experimentally infected pigs

The study evaluated the efficiency of diagnostic laboratory methods to detect anti-Toxoplasma gondii antibodies in paired serum and aqueous humour samples from experimentally infected pigs. 18-mixed breed pigs were used during the experiment; these were divided into two groups, G1 (infected group, n = 10) and G2 (uninfected group, n = 8). Infection was performed with 4 × 10 4 VEG strain oocysts at day 0 by the oral route in G1 animals. All pigs were euthanized at day 60, when retina, aqueous humour, and blood samples were collected. Anti-T. gondii antibody levels were assessed in serum (s) and aqueous humour (ah) by indirect immunofluorescence assay (IFA), modified agglutination test (MAT), m-ELISA (using crude membranes from T. gondii tachyzoites as antigen) and r-ELISA (using rhoptries from T. gondii tachyzoites as antigen). Polymerase chain reactions (PCR) of samples from the retina were performed by using Tox4 and Tox5 primers. Antibody titers of G1 animals ranged from 128 to 1024 and from 16 to 256 in serum and aqueous humour, respectively. There were differences in the correlation coefficients between IFA(s) × IFA (ah) (r = 0.62, P = 0.05), MAT(s) × MAT (ah) (r = 0.97, P < 0.0001); however, there was no significant difference between r-ELISA(s) × r-ELISA (ah) (r = 0.14, P = 0.7). Antibodies present in serum and aqueous humour recognized similar antigens. Samples of retina were positive by PCR in 30% (3/10) of infected pigs. G2 animals remained without antibody levels and were PCR negative throughout the experiment. These results suggest that the use of a combination of tests and immunoblotting for paired aqueous humour and serum samples could improve the sensitivity and specificity for the diagnosis of ocular toxoplasmosis. © 2007 Elsevier Ltd. All rights reserved.

Constructed wetland in wastewater treatment

Physical and chemical variables of soil and water were measured to determine the effectiveness of a constructed wetland for wastewater treatment. Eight different macrophyte species, namely Eichhornia crassipes, Alternanthera philoxerodos, Heteranthera reniformis, Hydrocotyle umbeliferae, Lidwigia elegan, Ludwigia sericea, Myriophyllum aquaticum and Thypha domingensis, were transplanted. Inlet water and outlet water were the two sampling sites evaluated. There were significant differences (p < 0,05) when limnological characteristics between inlet and outlet water from the constructed wetland were compared. In general, dissolved oxygen was over 4 mg L-1, and conductivity was high, above 80 μS cm-1. Chlorophyll-a levels generally tended to decrease at the wetland outlet and were higher during the rainy period (fish growth period). Results show that ammonia, total phosphorus, BOD5, phosphorus and organic mattel in the sediment removals in the constructed wetland were higher, indicating that macrophytes played an important role in removing these variables. The use of constructed wetland is a viable technology for the biological treatment in aquaculture and swine wastewater.

O uso da silagem de subprodutos da filetagem de peixe na alimentação de suínos em crescimento - parâmetros séricos

The study was carried out to evaluate the inclusion of increasing levels (0, 3,0 and 6.0%) of fish silage (SSFP), in the ration. The parameters studied were urea, uric acid, total protein, triglycerides, total cholesterol and lipoproteins (LDL, HDL and VLDL). Eighteen female piglets Moura weighting and crossbred Duroc x Moura, were used with 24,0± 3,0 kg in average weight and 90 days of age and eighteen male pigs Moura and crossbred Duroc x Moura, with 31,5±5,0 kg in average weight and 70 days of age in a randomized design with 3 treatments. The results showed no effect to the addition up to 6,0% of SSPF in the swine diet neither for in growth, nor to the serum parameters, even if had been found alteratum on the urea plasmatic concentrations experiments. All the parameters was closed to the normal value recommended in the literature.

Morfologia do intestino delgado de capivara - Hydrochoerus hydrochaeris (Linnaeus, 1766)

The high potential for the exploitation of capybara requires information about its digestory morphophysiology, to improve nutritional handling. In the present study, gross anatomy, light microscopy and body and intestine lengths of 25 capybaras were evaluated. The minimum and maximum small intestine lengths for females and males were, respectively, 441 cm and 1734 cm, and 355 cm and 1123 cm. These values position the capybara between canine and swine intestinal lengths. The ratio between small intestine and body length was 12:1, without differences between sexes. There were no statistically significant differences between sexes for each part of small intestine. Correlation between length of each small intestine segment and body length was positive, and statistically significant only for the duodenum. The small intestine wall was formed by mucosa, submucosa, muscular and serosa. The mucosa presented intestinal and duodenal glands, of mucosal and serosal types, respectively. The mucosa muscular layer consisted of two distinct layers in the jejunum and ileum, and a thin and single layer in the duodenum. The submucosa, formed by moderate dense connective tissue, didn't show glands. The fiber bundles of the internal layer of muscular tunic were helicoidally arranged. The gross anatomy of the capybara small intestine was similar to canine and swine intestines. Microscopically, however, subtle differences can be identified in the submucosa and internal muscular tunics.

Temperatura ambiente e concentrações séricas de cortisol em suínos em crescimento-terminação

Thirty-six castrated males and females Landrace x Large-White pigs (74 to 149 days of age) were randomly allotted to two environmental conditions: high temperature in a climatic chamber (HT; 22.2 to 32.8 °C) and comfort temperature in a conventional shed (CT; 17.6 to 26.6 °C), with night-and-day variations. Blood samples were weekly collected from animals of both HT and CT conditions for determination of serum cortisol levels. Cortisol levels of both sexes were not different, and there was no interaction with environmental temperature. Pigs of HT showed significantly higher average cortisol level (P<0.01) than the CT ones (7.06 and 4.82 mg/dL, respectively). Increasing in serum cortisol was continuous and linear (P<0.05) during the experimental period, suggesting the cortisol as a possible indicator of the heat stress in growing-finishing pigs.

Leitões lactentes e desmamados alimentados com dietas farelada ou extrusada seca e umida

This work was conducted throughout three phases of swine's growth (7 days from birth till the weaning age; from weaning until 7 days later and from 8 to 21 days after weaning); taking as objective the evaluation of the development of suckling and weaned piglets fed with a bran or extruded, dried and wet diet. We've used, for this paper, a total number of 31 pigs breeds, weaned and in average 21 day's lifetime and 3 different rations: dried bran; dried extruded bran and wet extruded bran (2 volumes of water to one volume of ration). During the 8th and 21st day after weaning all the animals received the same bran dried ration. Water and ration were supplied ad libitum. The experimental delimitation was totally random, with 3 treatments and eleven repetitions in the two first phases and five repetitions in the third phase, the multivariate variance analysis applied in the inferences about the results and from this we could conclude that the weight daily gain and the ration daily consumption were significantly not affected, i.e. by the processing, by the dry material ration content. However, we could observe a tendency to increase ration consumption and weight gaining when the animals were fed with bran extruded wet ration.

Influence of environmental temperature, dietary energy level and sex on performance and carcass characteristics of pigs

Pigs are quite sensitive to high environmental temperatures and the thermoregulation mechanisms represent great expenses in energy for heating loss, reducing animal well-being and production performance, and altering carcass quality. The aim of this study was to assess the effects of sex and dietary energy level in growing-finishing pigs submitted to characteristic seasonal variation of temperature in subtropical humid climate, and to propose a mathematical model to predict growth performance and carcass characteristics. Twenty-eight crossbred growing-finishing pigs were randomly allotted to twelve treatments, in a 2x2x3 factorial trial (2 sex; 2 environmental conditions, and 3 energy levels). Heat stress condition (climatic chamber) showed temperatures of 31 oC at 7:00 and 22 oC at 17:00 (maximum of 33 °C) and thermal comfort condition (stall) showed temperatures of 18 °C at 7:00 and 24 °C (maximum of 27 °C). Pigs were fed ad libitum with diets containing 12.2 (low), 13.6 (medium) and 15.0 (high) MJ ME/ kg DM. Voluntary feed intake, daily weight gain, and final body weight were higher (P<0.01) at thermal comfort condition and were influenced by sex (P<0.01) in growing pigs. Feed to gain ratio decreased as the energy level increased (P<0.01), with values of 2.67, 2.59, and 2.32 (12.2, 13.6, and 15.0 MJ ME/kg DM, respectively). There was energy level and sex interaction only for daily weight gain. Regarding finishing pigs, environmental conditions also showed effects (P<0.01) on voluntary feed intake, daily weight gain, and final body weight. Performance of pigs was better at thermal comfort condition. Feed to gain ratio values were 3.55, 3.42, and 2.95 for low, medium, and high energy level, respectively. Interactions between energy level and sex were observed for voluntary feed intake, daily weight gain, and final body weight (P<0.05). Carcass yield and quality were affected by environmental condition and dietary energy level. Both hot and cold carcass weight increased as energy of ration increased. Cold carcass weight increased by 1.142 kg/MJ EM whereas backfat thickness was up to 252 mm/MJ EM. Longissimus thoracis muscle thickness was around 16 mm smaller in pigs under heat stress, but lean content was 2.68% higher in those animals. Regression equations were proposed to predict the performance values in the different situations studied.