Objective-To test horses for serologic evidence of an association between vesiviral antibodies and abortion. Sample Population-Sera from 141 horses. Procedures-2 experiments were conducted. Experiment 1 comprised sera obtained in 2001 and 2002 from 3 groups of horses (58 mares from farms with a history of abortion problems, 25 mares between 3 and 13 years of age with unknown reproductive histories that were sold at auction breeding-age control mares, and 29 mixed-age males and yearling females sold at auction negative control population). Experiment 2 comprised sera from 3 groups of pregnant mares (10 pregnant mares fed Eastern tent caterpillars ETCs, 9 pregnant mares fed ETC frass only, and 10 pregnant control mares). Sera were analyzed for antibodies against vesivirus by use of a validated recombinant vesivirus-specific peptide antigen in an indirect ELISA. Results-For experiment 1, 37 of 58 (63.8%) mares from farms with abortion problems were seropositive for vesivirus antibodies, whereas 10 of 25 (40%) breeding-age control mares were seropositive. All 29 mixed-age males and yearling females were seronegative for vesivirus antibodies. For experiment 2, 17 of 29 mares aborted (some from each group). Seropositive status for vesivirus antibodies increased from 47.1% (8/17) to 88.2% (15/17) for the pregnant mares that aborted during the experiment. Conclusion and Clinical Relevance-Significant association was detected between seropositive status for vesivirus and abortion in mares; consequently, vesivirus appears to be a pathogenic virus associated with abortion in mares. These data support adding vesivirus antibody testing into diagnostic screening to determine the cause for abortion in mares.

Recombinant equine interferon-beta 1 (reqIFN-beta 1) induces an antiviral state in blood mononuclear cells (BMC) of horses. Maximal protection against replication of vesicular stomatitis virus is achieved 6 hours after treatment with IFN in vitro and in vivo. Duration of the protective effect depends on the dose of IFN in vitro and in vivo. Availability of reqIFN-beta 1 in cultures of BMC for up to 48 hours does not prolong the antiviral state. The protective effect on BMC after treatment with IFN has similar duration in vivo and in vitro. Monitoring of the effect of IFN in vivo is, thus, simplified because the antiviral state may be recorded by testing cells twice (ie, before and 6 hours after application of interferon). All further tests may be performed in vitro. Multiple administrations of reqIFN-beta 1 do not prolong duration of the protective phases after each administration. Duration of the antiviral state depends only on the dose of reqIFN-beta 1.

Vaccina virus (VV) infection induces specific antibodies and cytotoxic T cells in various animal species. Therefore, helper T cells also should be induced that stimulate the humoral and cellular immune responses. We determined such helper T-cell activity in 2 species after VV infection. Rabbits and rhesus macaques were infected with the Copenhagen strain of VV or with recombinant VV expressing retroviral proteins. Animals of both species developed antibodies and specific proliferative T-cell response. This reactivity could be enhanced by booster infection with VV. The proliferating macaque cells were CD4+ and major histocompatability complex class II-restricted. These data confirm the broad immunogenicity of VV. Expression of additional polypeptides expressed from a recombinant VV does not lead to altered immune response to VV antigens. However, strength of the helper T-cell response, as well as clinical reactions, differed between macaques and rabbits. Infection with recombinant VV as delivery vectors offers the opportunity for combined vaccination against recombinant proteins and does not diminish cellular and humoral immune responses to VV itself.

Acyloxyacyl hydrolase (AOAH) is a lysosomal enzyme found in neutrophils and macrophages that acts to partially deacylate the lipid-A component of the endotoxin of gram-negative bacteria rendering it less toxic, yet maintaining much of its immunostimulatory potential. We have found that the activity of neutrophil AOAH per cell increased during localized inflammation. The purpose of this study was to determine the mechanism(s) responsible for these increases in neutrophil AOAH activity. Because changes in neutrophil maturity commonly are associated with inflammation, intravascular infusion of purified gram-negative bacterial lipopolysaccharide and SC injection of bovine recombinant granulocyte colony-stimulating factor was used to induce large numbers of circulating immature neutrophils. Immature neutrophils were found to have AOAH activity equal to that of mature cells; however, when neutrophils were stimulated in vitro with known activators, AOAH activity of activated cells was more than that of unstimulated cells. The increase in AOAH activity was inversely related to prestimulation activity. Increases in AOAH activity after neutrophil activation were not a result of de novo synthesis of the enzyme, because cycloheximide did not prevent activation-induced increases in activity.

Recombinant human interleukin-2 (rhIL-2) was evaluated for its influence on total and differential WBC counts lymphocyte blastogenic responsiveness to mitogens, and several measurements of neutrophil function in clinically normal and in dexamethasone-treated cattle. A single dose of rhIL-2 (2.5 X 10(7) U) given SC had no influence on the total or differential WBC count; however, it did cause an inhibition of neutrophil random migration. The other measurements of neutrophil function (Staphylococcus aureus ingestion, cytochrome C reduction, iodination, and antibody-dependent and antibody-independent cell-mediated cytotoxicity) evaluated were not significantly altered. The rhIL-2 treatment was associated with a significant (P < 0.01) decrease in uptake of [3H]thymidine in unstimulated lymphocytes and a tendency toward enhanced blastogenesis of lymphocytes stimulated with phytohemagglutinin. This enhancement was significant (P < 0.05) only when the results were expressed as a stimulation index. Lymphocyte responsiveness to concanavalin A and pokeweed mitogen was not significantly influenced by rhIL-2 administration. Dexamethasone (0.04 mg/kg) administered every 24 hours for 3 consecutive days altered the WBC count and several measurements of lymphocyte and neutrophil function. The administration of a single dose of rhIL-2 (2.5 x 10(7) U) 8 hours after the first dose of dexamethasone did not alter the influence of dexamethasone on any of the measurements. These results indicated that rhIL-2 has some biologic activity in cattle, but when used as administered here, did not overcome the influence of dexamethasone on the in vitro measurements of lymphocyte and neutrophil function that were evaluated.

Programmed cell death protein 1 (PD-1), a costimulatory molecule of the CD28 family, has 2 ligands, PD-L1 and PD-L2. Our previous studies showed that the expression of PD-1 and PD-L1 is up-regulated during viral infection in pigs. Extensive studies have shown that blockade of the PD-1/PD-L1 pathways by anti-PD-L1 antibody or soluble PD-1 restores exhausted T-cells in humans and mice. In the present study the extracellular domains of PD-1 and PD-L1 were used to evaluate the binding of PD-1 and PD-L1 with peripheral blood mononuclear cells (PBMCs). We amplified the cDNA encoding the extracellular domains of PD-1 and PD-L1 to construct recombinant expression plasmids and obtain soluble recombinant proteins, which were then labeled with fluorescein isothiocyanate (FITC). The His-ExPD-1 and His-ExPD-L1 recombinant proteins were expressed in the form of inclusion bodies with a relative molecular weight of 33.0 and 45.0 kDa, respectively. We then prepared polyclonal antibodies against the proteins with a multi-antiserum titer of 1:102 400. Binding of the proteins with PBMCs was evaluated by flow cytometry. The fluorescence signals of His-ExPD-1-FITC and His-ExPD-L1-FITC were greater than those for the FITC control. These results suggest that the soluble recombinant proteins may be used to prepare monoclonal antibodies to block the PD-1/PD-L1 pathway.

Iridovirus is an icosahedral cytoplasmic double-stranded DNA virus with a genome size of 170-200kb. Outbreaks of fish iridovirus infection are characterized by their wide geographic distribution and broad host spectrum, especially in water temperatures of 22-27℃ Recently, the causative agent of high mortalities in flounder (Paralichthys olivaceus) was identified as fish iridovirus in Korea. Iridoviral infection repeatedly occurs in the same area for long periods, suggesting the possibility of viral infection in nursery.

Actinobacillus pleuropneumoniae is the causative agent of a porcine contagious pleuropneumonia. Among several virulence factors including exotoxin (Apx toxins), LPS, transferrin-binding proteins, OMPs, and some proteases, Apx toxins have been major targets for the protection study. In this study, cloning and expression of A. pleuropneumoniae Apx I and Apx Ⅱ toxin, which are produced by all highly virulent strains, were performed by Escherichia coli expression system. Genes coding Apx I and Ⅱ toxin were amplified from the A. pleuropneumoniae serotype 5 genomic DNA using polymerase chain reaction and cloned to a prokaryotic expression vector, pRSET.

The role of interleukin 1 (IL-1) as an inflammatory mediator during mastitis and the therapeutic effect of recombinant human IL-1 receptor antagonist (IL-1ra) for bovine mastitis was studied. Cows were intramammarily infused with lipopolysaccharide (25 g) in 1 mammary gland. Half the cows also received infusions of 5 mg of IL-1ra into the same mammary gland just prior to endotoxin infusion and 4, 8, and 12 hours later. After endotoxin infusion, tumor necrosis factor and high IL-1 bioactivity were detected in whey from infused glands. Vascular permeability changes and neutrophil accumulation in milk paralleled the appearance of cytokines. A systemic reaction, characterized by pyrexia and an increase in blood cortisol concentration, also were observed. Milk yield was inhibited and milk composition was altered in infused and noninfused glands. The increase in IL-1 bioactivity in milk after endotoxin infusion was almost completely prevented in glands receiving IL-1ra. However, IL-1ra had no effect on local inflammation, systemic reaction, or impairment in productive performance. These results indicate that IL-1 does not mediate its effect within the milk compartment, and suggest either that IL-1 is not critical to the mastitic response or that intramammary infusion of IL-1ra does not place the antagonist where IL-1 interacts with its receptor.