To measure the concentration of serum amyloid A (SAA) protein in horses a sensitive and highly reproducible sandwich (ELISA) was established, using affinity purified SAA antibody. Results of the ELISA were found to have a high correlation (r = 0.95) with those of the single radial immunodiffusion test. Equine SAA concentration was measured by use of this ELISA. In clinically normal horses, the concentration of SAA was high immediately after birth to 2 weeks of age. After that, SAA concentration had periodic fluctuations in the range of approximately 10 to 30 microgram/ml. Mean (+/- SD) concentrations of SAA in foals (less than or equal to 12 months old) and adult horses (greater than or equal to 18 months old) were 21.23 +/- 12.20 and 14.93 +/- 9.07 microgram/ml, respectively. In mares during the perinatal period, SAA concentration remained stable within the reference range before parturition. It increased quickly after delivery, and reached a peak value of 101.29 +/- 98.82 microgram/ml on postpartum day 3, then began to decrease, at postpartum week 2, to the reference range by the end of postpartum month 1. In horses with experimentally induced inflammation, SAA concentration increased quickly and reached approximately four- to 40-fold increase over the pretreatment value on day 1 and remained high on days 2 to h after treatment. It then returned to the baseline value by 2 to 4 weeks in association with disappearance of local signs of inflammation. The SAA concentration was high in most horses with clinical signs of inflammation. It was concluded from these data that this ELISA is sensitive and reliable for measuring SAA in horses.

The vaccine efficacy of a genetically engineered deletion mutant strain of pseudorabies virus, strain 783, was compared with that of the conventionally attenuated Bartha strain. Strain 783 has deletions in the genes coding for glycoprotein I and thymidine kinase. In experiment 1, which had a 3-month interval between vaccination and challenge exposure, strain 783 protected pigs significantly (P < 0.05) better against virulent virus challenge exposure than did the Bartha strain. The growth of pigs vaccinated with strain 783 was not arrested, whereas that of pigs vaccinated with the Bartha strain was arrested for 7 days. Of 8 pigs given strain 783, 4 were fully protected against challenge exposure; none of the pigs given strain Bartha was fully protected. In experiment 2, which had a 3-week interval between vaccination and challenge exposure, the growth of pigs vaccinated with strain 783 was arrested for 3.5 days, whereas that of pigs vaccinated with the Bartha strain was arrested for 6 days. In experiment 3, pigs with moderate titer of maternal antibodies were vaccinated twice IM or once intranasally with either strain 783 or Bartha and were challenge-exposed 3 months after vaccination. Pigs given strain 783 twice IM were significantly (P < 0.05) better protected than were the other pigs. They had growth arrest of only 6 days, compared with 9 days for pigs of other groups, and shed less virus after challenge exposure. Results of this study indicate that the vaccine based on the deletion mutant strain 783 is more efficacious than is the Bartha strain of pseudorabies virus.

Neutrophils were purified from blood of dexamethasone-treated (0.04 mg/kg of body weight) and untreated calves. Cells were untreated (controls) or cultured in media containing 5 or 10 ng of bovine recombinant granulocyte-macrophage colony-stimulating factor (rbGM-CSF)/ ml for 10 to 12 hours before being tested for various functions. Dexamethasone treatment of calves decreased luminol-dependent chemiluminescence, decreased phagocytosis of Pasteurella multocida and several Staphylococcus spp by various degrees, and decreased antibody-dependent cell-mediated cytotoxocity against bovine herpesvirus-infected cells by 26 to 32%. The percentage phagocytosis of coagulase-positive S aureus and S intermedius was higher than that of coagulase-negative S epidermidis for neutrophils from all calves. Culture of neutrophils with rbGM-CSF significantly increased (P < 0.05) all of the aforementioned functions, compared with control neutrophils; however, rbGM-CSF-induced increases in function tended to be higher in neutrophils from dexamethasone-treated calves than in neutrophils from untreated calves.

Oestrus resynchronisation (RES, Resynch) programmes for non-pregnant cows allow shortening the period between an unsuccessful insemination and the next attempt on the same cow. The protocol of oestrus RES may be started after ruling out pregnancy by means of ultrasonography carried out 28 days after insemination or after performing a test for pregnancy-specific glycoproteins (PAG) in blood or milk. The Resynch protocol can be based on a double application of prostaglandins, the OvSynch protocol, or hormonal therapy with exogenous sources of progesterone (CIDR intravaginal devices). The efficiency of the method depends on the functional state of the ovaries, the diameter of the corpus luteum, external factors, and the health and maturity of the cows. The present paper constitutes a comparison of research findings concerning the effectiveness of RES programmes.

Proper conformational arrangement of the E2 molecules of bovine viral diarrhoea-mucosal disease virus (BVD-MDV) is crucial to obtain an effective recombinant vaccine candidate against the disease. In this study, we characterised a new molecule composed of two distinct sequences of the E2 glycoprotein of BVD-MDV and the Fc fragment of human immunoglobulin (BVDE2Fc). The chimaeric protein was expressed in mammalian cell lines of different species by adenoviral transduction and purified by immobilised metal-affinity chromatography. The N-glycans were profiled by HPLC, and the BVDE2Fc immunogenicity was assessed in male mice. The antigen-antibody reactions were evaluated by ELISA. The MDBK cell line was selected from among five for the final production of BVDE2Fc. After purification to over 90%, the N-glycan profile showed neutral and complex oligosaccharides. The mouse immunisation induced a strong humoral response, which produced antibodies able to attach to conformational epitopes on E2 molecules, while the Fc fragment barely contributed to the immune response. Additionally, BVDE2Fc attached to antibodies from bovine sera positive to distinct BVD-MDV subtypes, whereas the loss of BVDE2Fc structure during the deglycosylation process considerably diminished those interactions. These results demonstrate that the structure of E2 molecules arranged in tandem and attached to an Fc fragment could represent a viable design for future vaccine candidates against BVD-MD.

OBJECTIVE To compare pregnancy-associated glycoprotein 1 (PAG1) concentrations in maternal (jugular vein) and fetal (uterine vein) circulations and amniotic fluid samples between pregnant ewes that were and were not experimentally infected with bovine viral diarrhea virus (BVDV). ANIMALS 11 healthy pregnant yearling ewes. PROCEDURES Before study initiation, all ewes were naïve to BVDV and confirmed pregnant by transabdominal ultrasonography at approximately 60 days of gestation. At 65 days of gestation, ewes were intranasally inoculated with a noncytopathic BVDV type 1b strain (concentration, 107 TCID50/mL; 2 mL/nostril; n = 6) or an equal volume of BVDV-free viral culture medium (control; 5). A blood sample was collected for measurement of PAG1 concentration before inoculation. At 80 days of gestation, each ewe was anesthetized and underwent an ovariohysterectomy. While sheep were anesthetized, blood samples from the jugular and uterine veins and an amniotic fluid sample were collected for measurement of PAG1 concentration. Fetal tissues underwent real-time PCR analysis for BVDV RNA, and placental specimens underwent histologic evaluation and immunohistochemical staining for BVDV antigen. RESULTS At 80 days of gestation, BVDV RNA in fetal tissues and mild placentitis were detected in 5 of 6 BVDV-inoculated ewes. Mean PAG1 concentrations in the maternal and fetal circulations of BVDV-inoculated ewes were significantly less than those in control ewes. Mean amniotic fluid PAG1 concentration did not differ significantly between the 2 groups. CONCLUSIONS AND CLINICAL RELEVANCE Concentration of PAG1 in the maternal circulation may be a useful biomarker for determining placental health in sheep after viral infection of the reproductive tract.

Objective: To evaluate urine concentrations of glycosaminoglycans, Tamm-Horsfall glycoprotein, and nephrocalcin in cats fed a diet formulated to prevent calcium oxalate uroliths. Animals: 10 cats with calcium oxalate urolithiasis. Procedures: In a previous study conducted in accordance with a balanced crossover design, cats were sequentially fed 2 diets (the diet each cat was consuming prior to urolith detection and a diet formulated to prevent calcium oxalate uroliths). Each diet was fed for 8 weeks. At the end of each 8-week period, a 72-hour urine sample was collected. Concentrations of glycosaminoglycans, Tamm-Horsfall glycoprotein, and the 4 isoforms of nephrocalcin in urine samples collected during that previous study were measured in the study reported here. Results: Diet had no effect on the quantity of Tamm-Horsfall glycoprotein and nephrocalcin in urine. However, the urine concentration of glycosaminoglycans was significantly higher during consumption of the urolith prevention diet. Conclusions and Clinical Relevance: Feeding a urolith prevention diet increased the urine concentration of glycosaminoglycans, which are glycoprotein inhibitors of growth and aggregation of calcium oxalate crystals.

Porcine reproductive and respiratory syndrome (PRRS) is characterized by a delayed and defective adaptive immune response. The viral nonstructural protein 1 (NSP1) of the PRRS virus (PRRSV) is able to suppress the type I interferon (IFN) response in vitro. In this study, recombinant adenoviruses (rAds) expressing NSP1 (rAd-NSP1), glycoprotein 5 (GP5) (rAd-GP5), and the NSP1-GP5 fusion protein (rAd-NSP1-GP5) were constructed, and the effect of NSP1 on immune responses was investigated in pigs. Pigs inoculated with rAd-NSP1 or rAd-NSP1-GP5 had significantly lower levels of IFN-γ and higher levels of the immunosuppressive cytokine IL-10 than pigs inoculated with rAd-GP5, wild-type adenovirus, or cell culture medium alone. The antibody response to vaccination against classic swine fever virus (CSFV) was significantly decreased by inoculation of NSP1 7 d after CSFV vaccination in pigs. Thus, NSP1-mediated immune suppression may play an important role in PRRSV pathogenesis.

Glycoprotein 5 (GP5) of porcine reproductive and respiratory syndrome virus (PRRSV) has been studied extensively as a target for vaccine development. This study evaluated the serodiagnostic application of PRRSV GP5 by enzyme-linked immunosorbent assay (ELISA). Two immunodominant peptides (VR #1 and VR #2) and two neutralizing ectodomain-containing peptides (Ecto #1 and Ecto #2), as well as recombinant GP5 (rGP5) as a control, were prepared. Serum from unvaccinated pigs was screened for the antibodies that bind to these peptide and protein antigens. The results were compared with those from a commercially available diagnostic ELISA kit (HerdChek), which uses the nucleocapsid (N) protein as an antigen. Only VR #1+#2 showed a result statistically similar to that of N protein. Ecto #1 and Ecto #2 had a lower sensitivity than VR #1+#2 and rGP5. The peptides and rGP5 showed significant associations with the N protein (P < 0.05 or 0.01), which suggests that GP5 may also be a candidate serodiagnostic antigen. Since antibodies against GP5 persist much longer than those against the N protein, GP5 itself and some of its fragments are thought to be good targets for serodiagnosis. In addition, the presence of antibodies against the PRRSV structural antigens showed significant antigen-dependent differences.