Immunohistochemical localization of bovine decorin was examined with its biological analysis in the fetal bovine rumen. By immunohistochemical staining, monoclonal antibody (mAb) 2B6, which recognizes chondroitin 4-sulfate and/or dermatan sulfate (DS), reacted specifically to the lower mesenchymal region in the developing ruminal wall. Biochemical analysis of the extract from the developing rumen revealed that the molecule detected immunohistochemically by mAb 2B6 was small DS proteoglycan, bovine decorin. These results support the view that bovine decorin is involved in organization of the fetal bovine ruminal mesenchyme as a collagenous tissue.
Protective efficacy of Actinobacillus pleuropneumoniae extracellular hemolytic toxins, RTX-toxin I (Apx I) and Apx II, was evaluated in pigs. The hemolysins were purified from culture supernatant of A. pleuropneumoniae strain HA-337, serotype 1 by immunoaffinity chromatography using a monoclonal antibody specific for Apx I or Apx II as ligand. Four pigs were vaccinated with the purified hemolysins adsorbed on aluminium phosphate gel adjuvant. Four pigs of a control group were given placebo. Hemolysin-neutralizing antibodies were detected only in vaccinated group after booster injection. One of four control pigs died following an aerosol challenge with the homologous strain, and three surviving pigs developed serious clinical signs of pneumonia and had extensive lung lesions. In contrast, there was no mortality in vaccinated group. Only transient hyperthermia was observed in two vaccinated pigs after challenge. At necropsy, two vaccinated pigs had slight localized pulmonary lesions, though the remaining two had no lung lesions at all. These results indicate that the hemolysin vaccine made of Apx I and Apx II has good protective activity against swine pleuropneumonia caused by A. pleuropneumoniae serotype 1.
The optimum pH for the measurement of serum p-phenylenediamine oxidase (Ox) activity was given (pH 6.6), and the relationship between serum Ceruloplasmin(Cp) concentration and its Ox activity was established in healthy adult horses. In adult horses, serum antigenic Cp concentrations were measured by the single radial immunodiffusion (SRID) method with the affinity-purified antibody to equine plasma Cp and compared with its Ox activity. Efficient co-relation between Cp concentration and Ox activity in the sera (r=0.93) and its Ox/Cp ratio were given. These results might contribute to the calculation of antigenic Cp concentration from its Ox activity, the analysis of holo-Cp content in serum, and the research for copper metabolism in Thoroughbred horses.
The effect of ammonia inhalation on the luminol-dependent chemiluminescence response of bronchoalveolar lavage (BAL) cells was investigated in clinically normal calves receiving eight ml of ammonia water (5,000 ppm) in the form of aerosol sprayed into the nasal cavities (four ml into each nostril) once a day for two weeks. The chemiluminescence level of the BAL cells was augmented following the sprayings of ammonia compared with prior to exposure. The level still remained high one week after the exposure was removed, but showed recovery after two weeks.
To establish an adequate experimental model for the study of immuno-neuroendocrine mechanisms underlying the behavioral changes during the acute infection, temporal relationship of various physiological responses to endotoxin administration was examined in ovariectomized goats. Immediately after intravenous injection of 200 ng/kg of lipopolysaccharide, there were an abrupt decrease of white blood cell number and a gradual increase of rectal temperature, which were followed by elevation of plasma levels of adrenocorticotropic hormone, cortisol, glucose, free fatty acids, and then later by an increase of heart rate. The results suggest that the endotoxin administration would evoke a stereotyped cascade of, febrile, neuroendocrine and metabolic as well as autonomic response to the activation of immune systems in the ruminant species.
Sarcocystis mihoensis n. sp., a heteroxenous coccidium was detected from sheep in Japan and dogs were experimentally determined as the definitive host. The cysts were 1,300-2,100 × 200-300 μm in size and had the thick cyst wall which was 10 to 12 μm thick and provided with radial striated and finger-like villar protrusions. Two 6-month old dogs fed with the cysts passed sporocysts, 15-16 × 8-9 μm in size, in the feces from day 11 to days 57-60 after ingestion. No domestic cats fed with the same cysts shed sporocysts throughout the experimental period.
Endectocidal efficacy of doramectin administered intramuscularly at a dosage of 300 μg/kg was evaluated in 464 pigs naturally infected with intestinal nematodes or mange mites on 14 commercial farms in Japan. By doramectin treatment, fecal egg counts were reduced >99% for Ascaris suum, Strongyloides ransomi, Oesophagostomum dentatum, and Trichuris suis; worm counts of T. suis and mite counts of Sarcoptes scabiei were reduced 90.1% on Day 21 and 99.5% on Day 28 following treatment, respectively.
The most probable number (MPN) method combined with a nested polymerase chain reaction (nested PCR) for the detection and enumeration of enterotoxigenic Clostridium perfringens in the intestinal contents of cattle, pig and chicken was examined. Ten-fold serial dilutions of samples were added to three tubes of enrichment medium, which were incubated at 37°C for 20-24 hr, and the C. perfringens enterotoxin gene was detected by nested PCR from the enrichment culture without isolating the organism. The results obtained by this method with artificially contaminated intestinal contents were significantly correlated with those obtained by a plate count method. When the method was applied to the detection and enumeration of indigenous enterotoxigenic C. perfringens, the organism was found in two, two and three samples of 10 intestinal contents of cattle, pig and chicken, respectively. Most of the positive samples contained fewer than 10 MPN/g of enterotoxigenic C. perfringens, except one sample of chicken, which contained 1.5 × 102 MPN/g. The MPN method combined with nested PCR is easy to perform and may be a useful tool for the detection and enumeration of enterotoxigenic C. perfringens in intestinal contents.
Ovariectomized goats were implanted with the electrode arrays for monitoring the electrophysiological manifestation of the activity of the hypothalamic gonadotropin-releasing hormone (GnRH) pulse generator, namely multiple-unit activity (MUA) volleys associated with pulsatile luteinizing hormone secretion. They were then subjected to i.v. challenges of lipopolysaccharide (LPS) at the dose of 200 or 400 ng/kg. The interval between the MUA volleys was significantly prolonged by higher dose of LPS whereas neither amplitude nor duration of the MUA volleys was altered. These results suggest that immunological disturbance as evoked by LPS administration directly affects the hypothalamic GnRH pulse generator by slowing down the pulse frequency, and thereby lowers gonadotropin secretion from the anterior pituitary gland, which would culminate in gonadal suppression.
The effects of different concentrations of etoposide and cycloheximide (ETO-CHXM), used for chemical enucleation of mouse oocytes, on polar body extrusion and chromatin expulsion were tested. The developmental ability of blastomeres of late 2-cell stage embryos fused to chemically enucleated oocytes of different ages or cytoplasts from different sources was also examined in vitro. Metaphase I oocytes cultured in different concentrations of ETO-CHXM (10-50 μg/ml each) extruded polar bodies at rates similar to those cultured without ETO-CHXM (58.5-65.9% and 64.6%, respectively). However, low percent of the oocytes (1.7-6.2%) expressed signs of meiotic perturbation, which was manifested by blebbing of the cytoplasmic membrane and extrusion of two or more polar body-like fragments. Twenty-three percent of the chemically enucleated oocytes cultured in ETO-CHXM-free medium spontaneously fused to their polar bodies. The rates of total chromatin expulsion were similar when ETO-CHXM concentrations were 36 and 50 μg/ml (93.5 and 98%, respectively). The results also showed that the cleavage rates of reconstituted embryos were significantly (P<0.001) affected by the age of the chemically enucleated oocytes. Cytoplasts of bisected oocytes that matured in vivo supported the development of 31.7% of the reconstituted embryos to the blastocyst stage. However, both cytoplasts of chemically enucleated oocytes and in vitro matured oocytes did not support development to the blastocyst stage. A high percentage (85.5%) of the reconstituted embryos with chemically enucleated recipients displayed abnormality of the metaphase plate. These results suggest that concentrations of etoposide between 36 and 50 μg/ml are optimum for enucleation of mouse oocytes. Furthermore, increasing the age or reducing the cytoplasmic volume of the chemically enucleated oocytes did not improve the development of the reconstituted embryos to the blastocyst stage.
A canine calicivirus (CaCV) isolated in Japan, designated as CaCV No. 48 strain, was propagated in MDCK cells and purified by CsCl equilibrium gradient centrifugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of the purified samples revealed the presence of only one major species of viral protein of about 60 kilodaltons after Coomassie staining. The same band, presumably that of the capsid protein, was detected by western blotting using a mouse hyperimmune serum. This capsid protein was synthesized in MDCK cells as early as 2 hr post-inoculation. Experimental infection of dogs resulted in the production of anti-CaCV antibodies which were detected by microneutralization test and western blotting. Likewise, serosurvey revealed not only the presence of neutralizing antibodies but also reactivity of the field sera against the capsid protein of the purified virus. These results indicate that the capsid protein of CaCV No. 48 strain is immunogenic and could be detected by antibodies in western blotting.
To obtain monoclonal antibodies (MAbs) specific to feline panleukopenia virus (FPLV) and mink enteritis virus (MEV), 15 hybridomas secreting MAbs against MEV-Abashiri were established and the properties of the MAbs were analyzed. The cross-reactivity of MAbs revealed that one MAb, P2-215 was specific for FPLV and MEV, whereas the remaining fourteen MAbs reacted with canine parvovirus (CPV), FPLV, and MEV. Epitope analyses using various CPV/MEV chimeric viruses revealed that the MAb P2-215 recognized the epitope comprised of amino acid 93-Lys in VP2, which is known to be FPLV and MEV-specific.