Characterization of monoclonal antibodies directed against swine leukocytes

Hybridomas were produced from fusions of the SP2/0 mouse myeloma with splenic cells from: 1) an outbred Sprague Dawley rat immunized with swine peripheral blood mononuclear (PBM) cells; 2) a (CBA/NDub X BALB/c Dub) F1 mouse immunized with concanavalin A (Con A) activated swine PBM cells and 3) a (BALB/c Dub X C3H/He Dub) F1 mouse immunized with swine thymocytes. The resulting supernatants were screened by a microcytotoxicity assay for activity against swine PBM cells. Four hybridomas (MSA1, MSA2, MSA3 and MSA4) were selected, cloned and characterized by their cell reactivity and effect on mitogenic assays. MSA1 and MSA2 belong to the rat IgG2b subclass. MSA3 and MSA4 are of the mouse IgG2a subclass. These monoclonal antibodies reacted in the following manner: MSA1 with monocytes, granulocytes, red blood cells and bone marrow cells; MSA2 with subset of T cells; MSA3 with B cells and subsets of T cells and monocytes (class II molecule) and MSA4, a pan-T cell reagent (E-rosette receptor). The involvement of the various cell types reactive to the different monoclonal antibodies in the mitogenic response of swine PBM cells to Con A, phytohemagglutinin (PHA) or pokeweed mitogen (PWM) was investigated by cellular depletion with monoclonal antibody plus complement. Cellular depletion of PBM cells with the following monoclonal antibodies plus complement treatment resulted in: MSA1, almost total reduction in the mitogenic response to low doses of Con A or PWM; MSA2, partial reduction in the proliferative responses to any concentration of Con A, PHA or PWM; MSA3, partial reduction in proliferative responses to low concentrations of Con A or PWM and 4) MSA4, total elimination of any proliferative response to Con A, PHA or PWM.     

Lack of influence of nursery temperature on the response of weanling pigs to supplemental vitamins C and E

Three trials using 240 weanling pigs were conducted during winter months to determine the influence of nursery temperature ("comfortable" vs "cold") on the response of weanling pigs to added vitamin C (700 ppm) or E (55 IU/kg) to a corn-soybean meal diet. A "comfortable" temperature schedule (27 C initially with a weekly 2 C drop) was maintained in one nursery, with the temperature schedule in the "cold" nursery about 8 C lower. Plasma concentrations of vitamin C and vitamin E were elevated when the respective vitamins were added to the diet, but were not consistently influenced by nursery temperatures. Daily gain, daily feed intake and feed efficiency were not improved with the added C or E. Daily feed intake was increased and feed-to-gain ratios were larger for pigs housed in the "cold" nursery compared with pigs housed in the "comfortable" nursery; however, daily gains were similar in the two environments. Pigs housed in the "cold" nursery were slightly stressed, as indicated by heavier adrenal glands, but the antibody response and serum glucocorticoid concentrations were not significantly affected by either diet or temperature.     

Estimation of receiver-operating characteristic curves to determine accuracy of a competitive enzyme-linked immunosorbent assay for the serodiagnosis of Brucella infection in domestic water buffalo (Bubalus bubalis) and cattle

OBJECTIVE: To estimate receiver-operating characteristic (ROC) curves for a competitive ELISA (c-ELISA) that is used in serodiagnosis of brucellosis in water buffalo and cattle, to determine the most appropriate positive cutoff value for the c-ELISA in confirmation of infection, and to evaluate species differences in c-ELISA function. SAMPLE POPULATION: Sera from 4 herds of cattle (n = 391) and 4 herds of water buffalo (381). PROCEDURE: Serum samples were evaluated for Brucella-specific antibodies by use of a c-ELISA. On the basis of previous serologic test results, iterative simulation modeling was used to classify animals as positive or negative for Brucella infection without the use of a gold standard. Accuracy of c-ELISA for diagnosis of infection was compared between cattle and water buffalo by comparison of areas under ROC curves. RESULTS: A positive cutoff value of 30% inhibition for c-ELISA yielded sensitivity and specificity estimates, respectively, of 83.9 and 92.6% for cattle and 91.4 and 95.4% for water buffalo. A positive cutoff value of 35% inhibition yielded sensitivity and specificity estimates, respectively, of 83.9 and 96.2% for cattle and 88.0 and 974% for water buffalo. Areas under ROC curves were 0.94 and 0.98 for cattle and water buffalo, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: ROC curves can be estimated by use of iterative simulation methods to determine optimal cutoff values for diagnostic tests with quantitative outcomes. A cutoff value of 35% inhibition for the c-ELISA was found to be most appropriate for confirmation of Brucella infection in cattle and water buffalo.     

Immunocytological confirmation of bone marrow metastases in a dog with cholangiocarcinoma

The case history of a four-year-old, male Bernese mountain dog is presented. Carcinoma cells were detected in the liver by ultrasound-guided fine-needle aspiration. Bone marrow aspirated from the iliac crest and the left femur showed a distinct infiltration by carcinoma cells. Immunocytological examination of the liver and bone marrow metastases showed a negative staining result for large spectrum cytokeratin (CK) KL1, a strong positive result for CK7 and a focal weak positive result for CK20. The dog was euthanased due to the grave prognosis. Histopathological examination revealed metastatic cholangiocarcinoma. The authors conclude that cytological and immunocytological examination of bone marrow aspirates should be used more frequently for the detection of distant metastases of carcinomas in small animal medicine.     

A recombinant vaccine expressing a mammalian Mycobacterium sp. antigen is immunostimulatory but not protective in striped bass

A recombinant vaccine was constructed for piscine mycobacteriosis utilizing a Brucella abortus strain RB51 vector expressing a mammalian Mycobacterium sp. 85A antigen. Juvenile striped bass were inoculated with the resulting construct at doses equivalent to 10(6), 10(7), 10(8), 10(9), and 10(10) colony-forming units/fish. Blood and tissue samples from these fish demonstrated significant specific humoral and cell-mediated immune responses towards the 85A antigen in a dose-dependent manner. However, survival studies determined that inoculated fish failed to demonstrate cross-protective responses after live Mycobacterium marinum challenge 70 days post-inoculation.     

Comparison of serologic tests for detection of Brucella infections in cattle and water buffalo (Bubalus bubalis)

OBJECTIVE: To estimate sensitivity and specificity of 4 commonly used brucellosis screening tests in cattle and domestic water buffalo of Trinidad, and to compare test parameter estimates between cattle and water buffalo. ANIMALS: 391 cattle and 381 water buffalo. PROCEDURE: 4 Brucella-infected herds (2 cattle and 2 water buffalo) and 4 herds (2 of each species) considered to be brucellosis-free were selected. A minimum of 100 animals, or all animals > 1 year of age, were tested from each herd. Serum samples were evaluated for Brucella-specific antibodies by use of standard plate agglutination test (SPAT), card test (CT), buffered plate agglutination test (BPAT), and standard tube agglutination test (STAT). A Bayesian approach was used to estimate sensitivity and specificity of diagnostic tests without the use of a gold standard, assuming conditional independence of tests. RESULTS: Sensitivity and specificity estimates in cattle, respectively, were SPAT, 66.7 and 98.9; CT, 72.7 and 99.6; BPAT, 88.1 and 98.1; and STAT, 80.2 and 99.3. Corresponding test estimates in water buffalo, respectively, were SPAT, 51.4 and 99.3; CT, 90.4 and 99.4; BPAT, 96.3 and 90.7; and STAT, 75.0 and 98.8. Sensitivity of the CT and specificity of the BPAT were different between cattle and water buffalo with at least 95% probability. CONCLUSIONS AND CLINICAL RELEVANCE: Brucellosis serologic test performance varied by species tested, but BPAT had the highest sensitivity for screening cattle and water buffalo. Sensitivity and specificity of more than 2 screening tests can be estimated simultaneously without a gold standard by use of Bayesian techniques.     

Pheromone-Related Inhibitors of Ustilago hordei Mating and Tilletia tritici Teliospore Germination

ABSTRACT Ustilago hordei, the causal agent of barley covered smut, produces mating pheromones that break down to smaller peptide compounds that act as potent inhibitors of mating and germination in several fungi. The pheromones are members of the farnesylated family of proteins. Synthetic peptide analogs of the pheromone derivatives, ranging in size from 4 mers to full length pheromones, were farnesylated, methyl esterified, or both and tested for mating or teliospore germination inhibition with U. hordei or Tilletia tritici, respectively. N-Acetyl-S-farnesylcysteine, which inhibits processing of Ras, and other sulfur-containing compounds such as homocysteine or methionine, were likewise modified and tested. The most potent inhibitors were methionine methyl ester and modified 4-mer peptides from both pheromones. Alanine scanning of the inhibitory 4 mers determined that the native amino acid sequence was specific for a high level of activity. The sulfur amino acids appear to be required for inhibition. Glasshouse studies using selected antagonists of mating and teliospore germination as seed treatments inhibited covered smut of barley and common bunt of wheat, although the level of control was inconsistent. The use of pheromone-related antagonists to mating or teliospore germination is a promising, novel strategy for control of smut and bunt diseases.     

Elektronenmikroskopische Untersuchungen intrazellulärer Stäbe (Trabeculae) im Holz vonLarix undJuniperus

Zusammenfassung Die Trabeculae von Lärche und Zwergwacholder weisen einen Bau auf, der mit Ausnahme des Zentralfadens dem einer normalen Zellwand entspricht. Auf die Mittellamelle folgt zunächst die sehr dünne Primär-, danach eine dreischichtige Sekundärwand; von der Sekundärwand ist besonders die mittlere Schicht (S₂) stark verdickt. Die innere Schicht (S₃) bildet die Tertiärlamelle, die beiJuniperus mit Warzen versehen ist. Bei Trabeculae-Reihen reicht der Zentralfaden durch die tangentialen Wände der Tracheiden von einer Zelle zur nächsten.

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Electron microscopal investigation of intracellular bars (Trabeculae) in wood ofLarix andJuniperus

Summary The Trabeculae ofSanio fromLarix andJuniperus sibirica showed with exception of central or middle core a structure like a normal cell wall. Upon the middle lamella and the central core respectively follows first of all the very thin primary wall, then the three layered secondary wall; of the secondary wall above all the middle layer (S₂) is strongly thickened. The inner layer (S₃) corresponds with the tertiary lamella which inJuniperus is warty. The rows of Trabeculae only the central core reaches through the tangential walls of tracheids from one cell to the other.

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- Larix Juniperus

, , . — . .V (S₂). (S₃) , Juniperus . , .

     

Immunodeficiency in young pigs

Immunodeficiency in neonatal and young pigs was studied in terms of T-cell function. Generalized T-cell deficiency did not exist in young pigs on the basis of the in vitro response of blood mononuclear cells to a polyclonal T-cell mitogen, phytohemagglutinin. However, immunodeficiency that extended from birth up to 4 weeks, was observed in serum antibody concentration and in vitro proliferative responses of blood mononuclear cells from young pigs exposed to a low antigen dose of a T-cell dependent antigen, egg white lysozyme. The low in vitro proliferative response to lysozyme was not attributable simply to a lack of interleukin-2 production, because supplementation with human interleukin-2 did not enhance the in vitro cellular response. Also, pokeweed mitogen-stimulated B cells from young pigs up to the age of 5 to 6 weeks produced immunoglobulin concentration, which also was not affected by the addition of human interleukin-2 to the in vitro cultures. The blood mononuclear cells obtained from pigs within the first 5 to 6 weeks after birth and incubated with monoclonal antibodies reactive to all T cells (MSA4), helper T cells (74-12-4) or suppressor/cytotoxic T cells (76-2-11) did not yield consistent excess of suppressor/cytotoxic T cells.(ABSTRACT TRUNCATED AT 250 WORDS)