In a search of viral agents in pulmonary macrophages of horses with chronic pulmonary disease, equine herpesvirus 2 was found to be unique. In 8 of 9 horses with chronic pulmonary disease, antigens of equine herpesvirus 2 were detected by indirect immunofluorescence staining of scattered foamy macrophages immediately after harvesting by bronchoalveolar lavage and fractionation on metrizamide gradients. In a healthy horse, antigens were not found. After 1 week of cultivation of bronchoalveolar lavage cells from a second group of 9 horses with chronic pulmonary disease, viral antigens were detected in 90% of the adherent pulmonary macrophages. In 2 of 3 healthy horses, viral antigens also were found in 90% of the adherent pulmonary macrophages. Antigens of equine herpesvirus 1, equine herpesvirus 4, parainfluenza virus 3, or adenovirus were not detected. Antigens of the 5 investigated viruses also were not detected in lung tissue slices from a third group of 14 horses, 4 healthy; 7 with varying degrees of bronchiolitis, 2 of which also had chronic intestitial pneumonia; 2 with eosinophilic bronchitis; and 1 with pulmonary hemorrhage. The exclusive presence of equine herpesvirus 2 in pulmonary macrophages was confirmed qualitatively by isolation of infective virus by cocultivation. In a fourth group of 12 horses with chronic pulmonary disease, infective virus could be isolated from pulmonary macrophages of 3 horses and from mixed-blood leukocytes of 5 horses. Virus isolations from 2 healthy horses were not successful from pulmonary macrophages, whereas 1 isolation was obtained from mixed-blood leukocytes. Other viruses were not detected by cocultivation.

Biological responses to recombinant DNA-derived bovine interferon alpha (rBoIFN-alpha I1) by bovine alveolar macrophages were examined by measuring viral yield reduction and 2',5'-oligoadenylate synthetase (2',5'-OAS) production by IFN-treated cells. In vitro IFN pretreatment of alveolar macrophages reduced viral yield in cultures challenged exposed with parainfluenza-3 virus, compared with control cultures. In vitro treatment of alveolar macrophages with IFN also resulted in increased 2',5'-OAS activity. The 2',5'-OAS activity was measured in alveolar macrophages and blood mononuclear leukocytes of calves injected im with 3.6 X 10(6) U of rBoIFN-alpha I1/kg of body weight. The IFN action was monitored by measuring 2',5'-OAS activity of blood mononuclear leukocytes beginning 6 days before and ending 24 hours after IFN treatment. The 2',5'-OAS activity in the blood mononuclear leukocytes sharply increased 24 hours after IFN treatment, indicating response to IFN. The alveolar macrophages collected from the same calves 24 hours after IFN administration also had increased 2',5'-OAS activity, compared with alveolar macrophages from the same calves collected 6 days before treatment. Increased 2',5'-OAS activity indicates: a possible mechanism of IFN action in cattle that may be responsible for viral yield reduction; potential use of high enzyme activity as a marker for IFN induction; and potential use of 2',5'-OAS activity as a marker for determining effects of IFN on bovine macrophages and other cells of the bovine immune system.

Pulmonary lavage samples were collected from 90- to 130-day-old calves before and 6 days after aerosol inoculation with bovine herpesvirus-1 (BHV-1) or parainfluenza-3 (PI3) virus. Alveolar lining material was separated from lavage fluids by high-speed centrifugation and phospholipids were extracted from alveolar lining material and analyzed by high-performance liquid chromatography. Phosphatidylcholine and phosphatidylethanolamine were 74.2 +/- 6.5% and 13.3 +/- 2.8%, respectively, of the total phospholipid content in the surfactant obtained from calves before virus inoculation. Other phospholipids were represented by substantially lower percentages. Infection with either of the 2 viruses caused a significant (P < 0.05) decrease in the percentage of phosphatidylcholine to 66.0 +/- 8.0% and 65.1 +/- 10.8% in the calves inoculated with BHV-1 and PI3 virus, respectively. A significant (P < 0.05) increase in the percentage of phosphatidylethanolamine to 18.1 +/- 2.2% and 17.8 +/- 4.5% developed in calves inoculated with BHV-1 and PI3 virus, respectively. Infection with BHV-1 also induced an increase (not significant) in the percentage of phosphatidylinositol from 5.5 +/- 2.8% to 7.8 +/- 2.2%. A similar, but not significant, increase in the percentage of phosphatidylinositol was also seen in the calves inoculated with PI3 virus. Less substantial changes in the percentage of other phospholipids were detected after virus infection.

Bovine parainfluenza virus type 5 (bPIV5) was isolated from cattle with downer cow syndrome in 2012, and included both respiratory and neurotropic pathogens from a variety of animals. In the current study, we conducted serosurveillance using sera obtained from seven Korean farms and optimized a reverse transcription-polymerase chain reaction (RT-PCR) assay to detect bPIV5. The overall seropositive rate for Korean cattle was 21.4% (163/760). A farm located near the city of Milyang in Gyeoungnam province had a markedly elevated seropositive rate for bPIV5 compared to that of the other six farms. The regional seropositive rates were 4.2% (8/192) for Haman, 19.5% (18/ 55) for Hwasung, 73.9% (65/88) for Milyang, 26.0% (50/192) for Namwon, 1.0% (1/96) for Uljin, 13.5% (13/96) for Yeongju, and 32.7% (8/41) for Yongin. The sensitivity and specificity of three RT-PCR primer sets used to amplify the conserved fusion gene of bPIV5 were also evaluated. An RT-PCR assay using the bPIVFR3 primer set was 10- fold more sensitive than the assays using the two other primer sets and did not result in non-specific amplification. These results demonstrated that the bPIFR3 primer set can be used to detect bPIV5.

OBJECTIVE To determine titers of serum antibodies against 3 genotypes of bovine parainfluenza 3 virus (BPI3V) in unvaccinated ungulates in Alabama. ANIMALS 62 cattle, goats, and New World camelids from 5 distinct herds and 21 captured white-tailed deer. PROCEDURES Serum samples were obtained from all animals for determination of anti-BPI3V antibody titers, which were measured by virus neutralization assays that used indicator (reference) viruses from each of the 3 BPI3V genotypes (BPI3V-A, BPI3V-B, and BPI3V-C). The reference strains were recent clinical isolates from US cattle. Each sample was assayed in triplicate for each genotype. Animals with a mean antibody titer = 2 for a particular genotype were considered seronegative for that genotype. RESULTS Animals seropositive for antibodies against BPI3V were identified in 2 of 3 groups of cattle and the group of New World camelids. The geometric mean antibody titer against BPI3V-B was significantly greater than that for BPI3V-A and BPI3V-C in all 3 groups. All goats, captive white-tailed deer, and cattle in the third cattle group were seronegative for all 3 genotypes of the virus. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that BPI3V-A may no longer be the predominant genotype circulating among ungulates in Alabama. This may be clinically relevant because BPI3V is frequently involved in the pathogenesis of bovine respiratory disease complex, current vaccines contain antigens against BPI3V-A only, and the extent of cross-protection among antibodies against the various BPI3V genotypes is unknown.

Calves, 90 to 130 days old, were inoculated with bovine herpesvirus-1 (BHV-1) or parainfluenza-3 (PI-3) virus. Pulmonary lavage specimens obtained from calves before virus inoculation contained 98% alveolar macrophages (AM) and 1% neutrophils. Six days after inoculation, the mean percentage of neutrophils in lavage specimens had significantly increased to 7.9 +/- 6.0% in BHV-1-inoculated calves and to 18.3 +/- 9.9% in PI-3 virus-inoculated calves, reflecting viral-induced pulmonary inflammation that was confirmed histologically. Approximately 75% of AM obtained before virus inoculation had Fc surface receptors, and 60% had C3b receptors. Six days after inoculation, the percentage of AM with Fc and C3b receptors was significantlyreduced to 69.7 +/- 8.6% and 27.1 +/- 19.8%, respectively, in BHV-1-inoculated calves and to 67.8 +/- 15.4% and 38.8 +/- 23.2%, respectively, in PI-3 virus-inoculated calves. Alveolar macrophages obtained after virus inoculation were significantly impaired in their ability to phagocytize opsonized Staphylococcus epidermidis, but were able to kill ingested bacteria. Alveolar macrophage dysfunctions caused by BHV-1 or PI-3 respiratory infection did not differ appreciably.