Characterization of,and immune responses of mice to,the purified OmpA-equivalent outer membrane protein of Pasteurella multocida serotype A:3 (Omp28)

Pasteurella multocida A:3 is a major cause of bovine pneumonia. A major antigenic heat-modifiable 28kDa outer membrane protein (Omp28) was previously identified. The purpose of this study was to purify and characterize Omp28 immunologically and structurally. Omp28 was extracted from N-lauroylsarcosine-insoluble protein preparations by a combination of detergent fractionation with Zwittergent 3-14 and chromatography. Partial N-terminal amino acid sequence confirmed Omp28 as a member of the OmpA-porin family. However, porin activity could not be demonstrated in a lipid-bilayer assay. Heat modifiability of purified Omp28 was demonstrated, and Omp28 was found in outer membrane fraction of P. multocida. Surface exposure of Omp28 was demonstrated by partial protease digestion of intact bacteria, by binding of anti-Omp28 polyclonal ascites fluid to the bacterial surface, and by partial inhibition of anti-outer membrane antiserum binding by previous incubation of the bacteria with anti-Omp28 serum. CD-1 mice vaccinated with purified Omp28 developed a significant antibody titer (P<0.05) compared to the control treatment group but were not protected from a homologous intraperitoneal bacterial challenge. By contrast, treatment groups vaccinated with P. multocida outer membrane, formalin-killed P. multocida or a commercial vaccine were significantly protected from challenge. In vitro complement-mediated killing of P. multocida was observed in post-vaccination sera of outer membrane, formalin-killed P. multocida, and commercial vaccine-treatment groups, but not with sera from the Omp28-treatment group. In conclusion, although Omp28 is surface exposed and antigenic, it may not be a desirable immunogen for stimulating immunity to P. multocida.     

Genetic and antigenic diversity in avian infectious bronchitis virus isolates of the 1940s

In order to verify a commonly held assumption that only Massachusetts (Mass) serotype of infectious bronchitis virus (IBV) was prevalent in the United States between the 1930s (when IBV was first isolated) and the 1950s (when the use of commercial IBV vaccines began), we examined 40 IBV field isolates from the 1940s. Thirty-eight of those isolates were recognized as Mass serotype viruses based on their reactivity to Mass-specific monoclonal antibody (Mab) and neutralization by Mass-specific chicken serum. The remaining two isolates, N-M24 and N-M39, that did not react with Mass-specific Mab, resisted neutralization by Mass-specific chicken serum, and were neutralized only by homologous chicken antibody were identified as non-Mass IBV. When the first 900 nucleotides (nt) from the 5'-end of the spike (S1) glycoprotein gene and their deduced amino acid (aa) sequences were compared, the two non-Mass isolates differed from each other by 24% and 28%, respectively. In a similar comparison, the non-Mass viruses N-M24 and N-M39 differed from M28, a Mass-type isolate from the 1940s, by 21% and 22% (nt) and 28% and 27% (aa), respectively. These data indicate that antigenic and genetic diversity among IBV isolates existed even in the 1940s. Interestingly, when the N-terminal region of the S1 of M28 was compared to that of M41, a prototype Mass virus that has undergone countless number of in vivo and in vitro host passages, the two viruses differed by only 2% (nt) and 4% (aa). This finding suggests that frequent genetic changes are not inherent in all IBV genomes.     

Effects of dietary ingredients on manure characteristics and odorous emissions from swine

Two feeding studies were conducted to examine the impact of dietary inclusion of specific feed ingredients on manure characteristics and manure odor. In one study, 72 finishing pigs were used to evaluate the effects of distillers dried grains with solubles (DDGS) on pig performance, manure characteristics, and odorous emissions. Three diets containing 0, 5, and 10% DDGS were fed during six 4-wk feeding periods. Week 1 served as a dietary adjustment period. Animals were housed in two feeding rooms (six pigs/room) with one treatment/room. A new group of animals (average initial BW = 85.8 kg) was used for each feeding period. Diets were replicated four times. Rooms were equipped with individual shallow manure storage pits that were cleaned once weekly (d 7). On d 4 and 7 of each week, manure pit samples, for chemical analyses, and air samples, for olfactometry analysis, were collected from each room. Odor dilution threshold was greater on d 7 than on d 4 of manure storage across all treatments (P < 0.01). No treatment differences in manure composition were noted. In the second study, weaned pigs (approximately 5 wk old) were fed isonitrogenous diets containing 0, 1.5, or 3% bloodmeal. Pigs were housed by diet (three pigs/diet) in one of four individual feeding rooms. A new group of pigs was used for each of the two, 4-wk feeding periods. During period 1, the 3% bloodmeal diet was fed in two of the four rooms; the 0% bloodmeal diet was fed in two rooms during period 2. Manure samples, for chemical analyses, and air samples, for olfactometry analysis, were collected 2 d per week (d 4 and d 6) from each room during wk 2 through 4. No significant treatment differences were observed for odor dilution threshold (P = 0.30). Longer manure storage time, 6 d vs 4 d, resulted in a larger odor dilution ratio (P < 0.01). Manure composition was unaltered by storage time. Results suggest that odor intensifies during storage.     

Identification of monoclonal antibodies that cross-react with cytokines from different animal species

Eleven monoclonal antibodies specific for ovine, bovine and human cytokines were investigated by flow cytometry for cross-reactivities with cytokines produced by peripheral blood mononuclear cells (PBMCs) from sheep, cattle, goat, swine, horse, dog, mink, rabbit and human. Four antibodies specific for IL-4, IL-8, IFN-gamma and TNF-alpha cross-reacted with cytokines from a majority of the species investigated. These antibodies can be applied to flow cytometric studies of cytokine production by PBMCs from several veterinary species. Another five antibodies specific for IL-2, IL-6, GM-CSF and IFN-gamma (two antibodies) cross-reacted weakly and with a variable number of animal species. These antibodies could in certain situations be useful in flow cytometry. In a number of cases the immunological cross-reactivities were confirmed by Western blot analyses. Overall, the results of this study will remedy some of the lack of species-specific anti-cytokine antibodies in veterinary research.     

Comparison between meloxicam and carprofen for postoperative analgesia after feline ovariohysterectomy

Eighty female cats presented for ovariohysterectomy were randomly allocated to one of two treatment groups in this assessor-blinded trial. After pre-anaesthetic assessment, the cats were premedicated with acepromazine (0.1 mg/kg). Anaesthesia was induced with thiopentone and maintained with halothane in oxygen. Forty cats received carprofen (4 mg/kg subcutaneously) and 40 received meloxicam (0.3 mg/kg subcutaneously) after anaesthetic induction. Following routine flank ovariohysterectomy the cats were assessed using visual analogue scale scores for pain and sedation over a 20-hour study period. Blood samples were taken before sedation and at 20 hours for serum biochemistry (urea, creatinine, alanine aminotransferase and aspartate aminotransferase). There were no significant differences between the groups for pain and sedation scores. Serum biochemistry values were similar between the groups, with some differences within groups between the pre-sedation and 20-hour values. One cat in the carprofen group and two cats in the meloxicam group required rescue analgesia with intramuscular morphine (0.2 mg/kg).     

Identification of immunoreactive polypeptides of Babesia equi parasite during immunization

This study was carried out to identify immunoreactive polypeptides in Babesia equi merozoite antigen. Three fractions of killed B. equi merozoite antigen viz.; whole merozoite (WM), cell membrane (CM) and high speed supernatant (HSS) antigens were prepared from the parasite infected erythrocytes. These antigenic preparations along with ghost antigen from non-infected erythrocytes were fractionated on 12% sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotted with sera showing high antibody titres. On SDS-PAGE, 16 polypeptides with molecular weight (Mr) in the range of 112-17kDa were obtained from the WM and CM antigens. But only six polypeptides were detected (96.5-28kDa) in the HSS antigen. On immunoblotting with high titred serum collected from donkeys following two immunizations with a killed B. equi merozoite immunogen, 11 polypeptides were observed in the WM and CM antigens (Mr 112-18kDa). Of these, four polypeptides (Mr 112, 45, 33 and 18kDa) were identified as most immunoreactive. Besides these, a 28kDa was observed as strong immunoreactive protein in WM and CM antigens. The HSS antigen showed only six polypeptides and one peptide (28kDa) was identified as immunoreactive. When high titred serum collected from immunized donkeys following challenge with B. equi infected blood and was used for immunoblotting, the protein profile of WM and CM antigens remained the same. However, three additional polypeptides (Mr 81, 54.5 and 39kDa) were detected in HSS antigen.     

Computed tomographic osteoabsorptiometry of the elbow joint in clinically normal dogs

OBJECTIVE: To evaluate subchondral bone density patterns in elbow joints of clinically normal dogs by use of computed tomographic (CT) osteoabsorptiometry. SAMPLE POPULATION: 20 cadaver forelimbs from 10 clinically normal dogs. PROCEDURE: Each elbow joint was imaged in parasagittal and transverse planes of 1.5-mm thickness. Slice data were converted to dipotassium phosphate equivalent density (PPED) values. Sagittal, parasagittal, and transverse medial coronoid process topographic maps were constructed. Defined zones were created for each of the 3 CT planes, and confluence and peak PPED values were determined. RESULTS: The lowest PPED value was 340 mg/ml (articular and subchondral confluence), and the highest was 1780 mg/ml (peak subchondral density). Detectable effects of joint laterality were not found in the confluence or peak PPED measurements or in the peak-to-confluence PPED ratio for all 3 CT planes. Significant differences were found among zones in all 3 planes for confluence and peak PPED measurements and between sagittal and transverse planes for peak-to-confluence PPED ratios. Subjectively, the pattern of density distribution among dogs was fairly consistent for the sagittal and parasagittal slices. Three specific patterns of density distribution were apparent on the transverse topographic maps of the medial coronoid process that corresponded to conformational differences. CONCLUSIONS AND CLINICAL RELEVANCE: The use of CT osteoabsorptiometry provides a repeatable technique that can be used to noninvasively examine bone density and the effects of stress acting on joints in vivo. Variability in density values for any of the CT planes was not identified among clinically normal dogs.     

Immunohistochemical characterisation of the lesions of canine idiopathic pericarditis

Pericardial tissue was obtained from 14 dogs with idiopathic pericarditis, and from three dogs with pericardial effusion associated with neoplastic disease, for histopathological assessment and characterisation of infiltrating leucocytes by immunohistochemistry. The major pathological change was extensive pericardial fibrosis which was generally accompanied by a mixed inflammatory response that was of greatest intensity at the cardiac surface of the tissue. Perivascular lymphoplasmacytic aggregates were present at the pleural surface and within the fibrosed pericardium. There were no features that clearly distinguished the samples from dogs with neoplastic disease from dogs with idiopathic pericarditis. The pericardial infiltrates were dominated by MAC 387+ monocyte-macrophages and plasma cells expressing immunoglobulin (Ig)A or IgG. CD3+ T lymphocytes and major histocompatibility complex (MHC) class II+ macrophages were less common, although the perivascular aggregates were mixtures of T and B lymphocytes and a proportion of fibroblasts expressed MHC class II. There was no vascular pathology or deposition of immunoglobulin or complement within vessel walls. These findings are consistent with an immune response dominated by humoral effector mechanisms (Th2 immunity) but do not clearly support a primary immune-mediated pathogenesis for idiopathic pericarditis.     

Associations between age or sex and prevalence of gastric ulceration in Standardbred racehorses in training

OBJECTIVE: To determine associations between age, sex, or medical treatment and prevalence and severity of gastric ulceration in Standardbred racehorses in training. DESIGN: Cross-sectional study. ANIMALS: 224 Standardbred racehorses in training. PROCEDURE: Gastroscopy was performed on each horse, and mucosal ulceration was graded from 0 (normal mucosa, no lesions) to 3 (extensive, often coalescing, lesions with areas of deep ulceration). Associations between age, sex, or treatment and prevalence and severity of ulcers were evaluated. RESULTS: Prevalence of gastric ulceration was 87%. Although there was little association between age and prevalence of ulcers, there was an association between age and severity of ulcers. Most 2-year-old horses (57.7%) had an ulcer score of 0 or 1. In all other age groups, most (58% to 82.61%) of horses had an ulcer score of 2 or 3. Although overall prevalence of ulceration was comparable among sex groups, the relative risk for gastric ulceration increased with age in castrated males, whereas it decreased in females and sexually intact males. CONCLUSIONS AND CLINICAL RELEVANCE: Gastric ulceration is common in Standardbred horses in race training. Severity is higher in horses > or = 3 years of age than in 2-year-old horses. Relative risk for ulceration increases with age in castrated males.