Neutralizing and nonneutralizing antibodies to bovine viral diarrhea (BVD) virus were detected in 3 cows persistently infected with noncytopathic BVD virus after vaccination with modified-live cytopathic BVD virus. Neutralizing antibodies detected in serum samples from each persistently infected cow at 3 weeks after vaccination were highly specific for certain isolates of cytopathic BVD virus and reacted only with a viral protein with a molecular weight of 53,000. Neutralizing antibodies to 1 of 3 isolates of noncytopathic BVD virus were detected in a serum sample obtained at 12 weeks after vaccination from 1 of 3 persistently infected cows. Nonneutralizing antibodies were detected in all cows at 7 to 12 weeks after vaccination.The nonneutralizing antibodies were less specific for isolates of BVD virus and reacted with viral proteins with molecular weights of 115,000, 80,000, 53,000, and 47,000.

The first outbreak of aujeszky's disease (AD) was identified from piggery located at the southern part of Korea in July, 1987. This piggery suffered from a significant economic loss caused by unexpected piglet mortality and reproductive failure. Etiologic viral agents were isolated from tonsil and spleen of the infected piglets, and the isolates produced a typical cytopathic effect of herpesvirus with giant cell formation when inoculated in many different cells. Subsequently the field isolates were characterized as suid herpesvirus I by cross-neutralization test and indirect fluorescence assay utilizing specific monoclonal antibody, and proved to be a pathogenic strain of AD virus(ADV).

OBJECTIVE To establish a method for evaluation of the efficacy of a classical swine fever virus (CSFV) subunit vaccine in rabbits as determined via humoral immune responses to the virus. ANIMALS 40 specific pathogen–free rabbits. PROCEDURES Rabbits were randomly assigned to 4 groups (10 rabbits/group) for SC injection of 0.05, 0.1, and 0.2 mL of a CSFV subunit E2 vaccine (representing 1.15, 2.3, or 4.6 μg of E2 protein/dose, respectively) or saline (0.9% NaCl) solution. Blood samples were collected 21 days after vaccination for measurement of the antibody response against CSFV via ELISA and virus neutralization methods. On the same day, the CSFV Chinese (C) strain was injected into an ear vein. Vaccine efficacy was determined by monitoring of rabbits for pyrexia for 4 days and measurement of viral copies in spleen lysates at the end of the study. Reproducibility of the antibody response was tested with 2 other batches of the vaccine at the minimum immunization dose identified for the initially tested batch. RESULTS The E2 protein dose of the initially tested vaccine was positively correlated with the antibody response and protection rate in rabbits. The identified minimum immunization dose per rabbit was 0.1 mL, representing an E2 protein content of approximately 2.3 μg, and reproducibility of the antibody response to vaccination with the 2 other batches at this dose was good. CONCLUSIONS AND CLINICAL RELEVANCE A method was established in rabbits for evaluation of the efficacy of a CSFV subunit vaccine that could help in the optimization of later large-scale vaccine production and quality control processes as well as in the clinical application of the vaccine.

OBJECTIVE To compare antibody responses of horses naturally infected with West Nile virus (WNV) and those vaccinated against WNV, to identify whether vaccination interferes with the ability to diagnose WNV infection, and to determine the duration of antibody responses after vaccination. SAMPLE Sera from horses naturally infected with WNV (n = 10) and adult WNV-naïve horses before and after vaccination with a live canarypox virus–vectored vaccine (7) or a killed virus vaccine (8). PROCEDURES An established WNV IgM capture ELISA was used to measure IgM responses. Newly developed capture ELISAs were used to measure responses of 8 other WNV-specific immunoglobulin isotypes. A serum neutralization assay was used to determine anti-WNV antibody titers. RESULTS WNV-specific IgM responses were typically detected in the sera of WNV-infected horses but not in sera of horses vaccinated against WNV. Natural infection with and vaccination against WNV induced an immunoglobulin response that was primarily composed of IgG1. West Nile virus–specific IgG1 was detected in the sera of most horses 14 days after vaccination. Serum anti-WNV IgG1 and neutralizing antibody responses induced by the killed-virus vaccines were higher and lasted longer than did those induced by the live canarypox virus–vectored vaccine. CONCLUSIONS AND CLINICAL RELEVANCE On the basis of these findings, we recommend that horses be vaccinated against WNV annually near the beginning of mosquito season, that both IgM and IgG1 responses against WNV be measured to distinguish between natural infection and vaccination, and that a WNV IgG1 ELISA be used to monitor anti-WNV antibodies titers in vaccinated horses.

Objective: To determine the frequency of spontaneous canine herpesvirus-1 (CHV-1) reactivation and ocular viral shedding in latently infected dogs and the effect of topical ocular administration of cyclosporine. Animals: 8 mature Beagles with experimentally induced latent CHV-1 infection. Procedures: Following induction of primary ocular CHV-1 infection, the presence of reactivatable CHV-1 latency was confirmed by systemically administering prednisolone to the dogs. Dogs were then monitored for 36 weeks via clinical examination and conjunctival sample CHV-1 PCR assay performed at 4-day intervals and CHV-1 virus neutralization antibody assay performed at 2-week intervals. During weeks 16 to 32, dogs were administered 0.2% cyclosporine ointment in both eyes twice daily and blood cyclosporine concentrations were monitored. During weeks 33 to 36, the presence of reactivatable CHV-1 latency was reconfirmed via systemic administration of prednisolone. Results: Reactivation of latent CHV-1 was not detected via clinical examination or viral shedding during the initial 32 weeks, including before and during topical ocular administration of cyclosporine, and there were no significant differences in CHV-1 virus neutralization titer increases between the study periods. Blood cyclosporine concentrations were less than assay detection limits in all dogs on the sampling days. Systemic administration of corticosteroids repeatedly resulted in ocular disease and viral shedding. Conclusions and Clinical Relevance: Spontaneous CHV-1 reactivation did not occur frequently in latently infected mature dogs, and this was not altered by topical ocular administration of cyclosporine. This characteristic may be a factor contributing to the lower frequency of recurrent herpetic ocular disease in dogs relative to other host species and their associated alphaherpesviruses.

Objective--To evaluate antimicrobial activity of bovine bactericidal permeability-increasing protein (bBPI)-derived synthetic peptides against mastitis-causing gram-negative bacteria. Sample Population--Bacterial isolates from the milk of cows with clinical mastitis. Procedures--3 peptides were synthesized with sequences corresponding to amino acids 65 to 99 (bBPI6599) or 142 to 169 (bBPI142169) or the combination of amino acids 90 to 99 and 148 to 161 (bBPI9099,148161) of bBPI. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of these peptides against bacterial isolates from cows with mastitis were determined by use of a standardized broth microdilution assay. The ability of these peptides to retain their antimicrobial activity in serum and milk was also evaluated. Finally, bacterial lipopolysaccharide (LPS)-neutralizing activity of these peptides was assayed with the Limulus amebocyte lysate test. Results--Of the 3 peptides tested, bBPI9099,148161 had the widest spectrum of antimicrobial activity, with MIC and MBC values ranging from 16 to 64 Mg/mL against Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp and from 64 to 128 Mg/mL against Pseudomonas aeruginosa. None of the peptides had any growth-inhibitory effect on Serratia marcescens. The antimicrobial activity of bBPI9099,148161 was inhibited in milk, but preserved in serum. Finally, bBPI142169 and bBPI9099,148161 completely neutralized LPS. Conclusions and Clinical Relevance--bBPI9099,148161 is a potent neutralizer of the highly proinflammatory molecule bacterial LPS and has antimicrobial activity against a variety of gram-negative bacteria. The ability of bBPI9099,148161 to retain antimicrobial activity in serum suggests a potential therapeutic application for this peptide in the management of gram-negative septicemia.

Six tests for the detection of West Nile virus (WNV) antibodies in the serum of experimentally infected chickens were compared. The tests included the hemagglutination-inhibition test (HIT), immunoglobulin M (IgM)-capture enzyme-linked immunosorbent assay (ELISA) with WNV-infected mouse brain antigen, immunoglobulin G (IgG) indirect ELISA with tickborne encephalitis viral antigen, the microtitre virus neutralization test, the standard plaque reduction neutralization test (PRNT), and the microtitre PRNT (micro-PRNT). Thirty adult chickens, intravenously and intramuscularly inoculated with 107 plaque-forming units (PFU) of WNV strain Egypt 101, were bled and given a booster of 107 PFU at 7, 15, and 21 d postinoculation; the final blood collection was on day 28. Although the micro-PRNT is capable of detecting the highest antibody titres during both early and late infection, because of the technical complexity and time requirements of this test a combination of IgM and IgG ELISAs is recommended for serologic screening. Serum samples that give positive results in the ELISAs can then be tested by the micro-PRNT to determine the specificity of antibodies to WNV.

The ability of viral glycoproteins (VP) VP4/VP7 reassortant swine rotaviruses (RV) to induce cross-neutralizing antibody against parental serotypes was investigated in guinea pigs. Using selective culture conditions, we produced 10 reassortant viruses that contained gene segment 4 of the OSU RV strain and gene segment 9 of the Gottfried RV strain. These reassortant RV grew to high titer in cell culture and were neutralized by monospecific antisera against both parental RV strains. The reassortant RV were chemically inactivated with binary ethylenimine, adjuvanted with aluminum hydroxide, and used to produce antisera in guinea pigs. The hyperimmune antisera had high neutralization titer against both parent RV strains. These results indicate that several of the reassortant RV may be capable of inducing neutralizing antibodies to VP4 and VP7 and may have future use as bivalent vaccine strains.

Tissue specimens and serum samples obtained from adult budgerigars in various stages of reproduction housed in an aviary with enzootic avian polyomavirus (APV) disease were examined by means of polymerase chain reaction techniques for APV DNA. Although the birds were apparently healthy, APV DNA could be detected in all 40 birds examined (inapparent infection rate, 100%). Viral DNA was found in most organ systems examined. Analysis of data suggested that organ virus concentrations were lower in breeding than in nonbreeding birds. Serum samples from 144 birds were examined for virus-neutralizing (VN) antibody. All serum samples had detectable VN antibody titers. Determining VN titer had a sensitivity of 100% for detection of APV infection in birds and was more sensitive than analysis of droppings by use of polymerase chain reaction techniques to detect APV infection in 6-month-old birds. Analysis of the data suggested that lower VN antibody titers were associated with longer duration of continuous breeding.