Whereas the clinical efficacy of vaccination against pseudorabies has been studied extensively, methods to evaluate the influence of vaccination on pseudorabies virus (PRV) transmission have only recently become available. In this study, PRV transmission and growth performance in finishing pigs vaccinated either once or twice were compared. The incidence of PRV infections was significantly (P = 0.039) higher in the group vaccinated once (38%) than in the group vaccinated twice (10%). The reproduction ratio R, which is defined as the average number of new infections caused by 1 infectious individual, was estimated in both groups. This ratio was also significantly (P = 0.025) higher among single vaccinated pigs (R = 3.4) than among pigs that had received double vaccination (R = 1.5). In compartments where serologic evidence of PRV introduction was observed, the mean daily weight gain was significantly (P = 0.029) lower in pigs vaccinated once (698 g/d) than in pigs vaccinated twice (721 g/d). Results of this study document the possibility to objectively evaluate the effect of vaccination on PRV transmission under field conditions. From the results, we concluded that double vaccination is advantageous in populations of finishing pigs at risk for PRV introduction. However, even among pigs vaccinated twice, extensive spread of PRV can occur.

The Rice strain of pseudorabies virus (PRV) was intranasally instilled in pigs that were seronegative to PRV. Cells were scraped or brushed from tonsillar surfaces biweekly until pigs were euthanatized at either 10 or 16 weeks after infection. The DNA extracted from tonsillar cells or parenchyma were subjected to polymerase chain reaction analysis, using either a single set of oligonucleotide primers or nested primers from the PRV gII glycoprotein gene. Pigs became seropositive to PRV by 3 weeks after infection. The virus was isolated from the trigeminal ganglia and tonsils of pigs that were euthanatized or died 1 to 2 weeks after infection, but not from pigs that were euthanatized 10 or 16 weeks after infection. The PRV gene products were consistently detected in trigeminal ganglia and tonsils of all pigs at 1, 10, and 16 weeks after infection, and sporadically in the nasal mucosa, lymph nodes, and lungs of pigs that were euthanatized or died during the first 2 weeks after infection. Cells collected biweekly from tonsillar surfaces were mostly nucleated, squamous epithelial cells with fewer lymphocytes and neutrophils. Polymerase chain reaction analysis of DNA extracted from these cells revealed PRV DNA in a large proportion of the samples when sufficient cells were collected to provide 1 microgram of extracted DNA for use in the reaction mixtures. A second group of pigs had PRV strain 4892 intranasally instilled. The virus was isolated from tonsillar swab specimens until 3 weeks after infection. Tonsillar brushing specimens were collected biweekly until 14 weeks after infection. Some brushing specimens contained all nucleated, squamous epithelial cells, whereas other specimens contained a mixture of epithelial cells and up to 15% neutrophils, lymphocytes, and small mononuclear cells. Results of polymerase chain reaction analysis of DNA extracted from tonsillar cells collected 5, 11, and 14 weeks after infection were consistently positive for PRV gene products. Intact cells collected from tonsillar surfaces were placed in polymerase chain reaction mixtures with nested oligonucleotide primers from the PRV gII glycoprotein gene and were subjected to multiple amplification cycles. Afterward, the specificity of the amplified PRV gene products was determined by hybridization procedures, using a virus-specific oligonucleotide probe. Most nucleated, squamous epithelial cells stained positive for PRV DNA, suggesting that these cells were the primary source of PRV gene products in tonsillar brushing specimens.

Pregnant sows, immune against pseudorabies after vaccination, were inoculated at 70 days of gestation either with autologous blood mononuclear cells that had been infected in vitro with pseudorabies virus (PRV) or with cell-free PRV. The infected cells or cell-free PRV were inoculated surgically into the arteria uterina. Eight sows (A to H) had been vaccinated with an inactivated vaccine. The titer of seroneutralizing antibodies in their serum varied between 12 and 48. Five sows (A to E) were inoculated with autologous mononuclear cells, infected either with a Belgian PRV field strain or with the Northern Ireland PRV strain NIA. These 5 sows aborted their fetuses: 2 of them (B and C) 3 days after inoculation, and the other 3 (A, D, and E) 10, 11, and 12 days after inoculation, respectively. Sows F, G, and H were inoculated with a cell-free PRV field strain. They farrowed healthy Utters after normal gestation. Neutralizing antibodies were absent against PRV in the sera of the newborn pigs, which were obtained prior to the uptake of colostrum. The 23 fetuses that were aborted in sows B and C 3 days after the inoculation were homogeneous in appearance and size. Foci of necrosis were not detected in the liver. Viral antigens were located by immunofluorescence in individual cells in lungs, liver, and spleen of 15 fetuses. Virus was isolated from the liver, lungs, or body fluids of 12 fetuses. The 39 fetuses that were aborted in sows A, D, and E between 10 and 12 days after inoculation were of 2 types: 17 were mummified and 22 were normal-appearing. Foci of necrosis were found in the liver of all mummified fetuses and 13 of the normal-appearing fetuses. In fetuses with foci of necrosis in the liver, viral antigens were located in groups of cells in the liver, lungs, and spleen. Virus was isolated from 16 normal-appearing fetuses and from 11 mummified fetuses. Pseudorabies virus was isolated from vaginal excretions of sows A and D until 1 and 2 days after abortion, respectively, and of sows B and C until 4 and 5 days after abortion, respectively. Virus was not isolated from sow E. It was concluded that PRV can reach the uterine and fetal tissues, via infected mononuclear cells, in the presence of circulating antibodies induced on vaccination. This cell-associated spread led to abortion. Cell-free virus did not induce abortion under similar circumstances.

Various procedures of vaccination for pseudorabies were compared for their effects on shedding, latency, and reactivation of attenuated and virulent pseudorabies virus. The study included 6 groups: group 1 (10 swine neither vaccinated nor challenge-exposed), group 2 (20 swine not vaccinated, but challenge-exposed), and groups 3 through 6 (10 swine/group, all vaccinated and challenge-exposed). Swine were vaccinated with killed virus IM (group 3) or intranasally (group 4), or with live virus IM (group 5) or intranasally (group 6). The chronologic order of treatments was as follows: vaccination (week 0), challenge of immunity by oronasal exposure to virulent virus (week 4), biopsy of tonsillar tissue (week 12), treatment with dexamethasone in an attempt to reactivate latent virus (week 15), and necropsy (week 21). Vaccination IM with killed or live virus and vaccination intranasally with live virus mitigated clinical signs and markedly reduced the magnitude and duration of virus shedding after challenge exposure. Abatement of signs and shedding was most pronounced for swine that had been vaccinated intranasally with live virus. All swine, except 4 from group 2 and 1 from group 4, survived challenge exposure. Only vaccination intranasally with live virus was effective in reducing the magnitude and duration of virus shedding after virus reactivation. Vaccination intranasally with killed virus was without measurable effect on immunity. Of the 55 swine that survived challenge exposure, 54 were shown subsequently to have latent infections by use of dexamethasone-induced virus reactivation, and 53 were shown to have latent infections by use of polymerase chain reaction (PCR) with trigeminal ganglia specimens collected at necropsy. Fewer swine were identified by PCR as having latent infections when other tissues were examined; 20 were identified by testing specimens of olfactory bulbs, 4 by testing tonsil specimens collected at necropsy, and 4 by testing tonsillar biopsy specimens. Eighteen of the 20 specimens of olfactory bulbs and 3 of the 4 tonsil specimens collected at necropsy in which virus was detected by PCR were from swine without detectable virus-neutralizing antibody at the time of challenge exposure. One pig that had been vaccinated intranasally with live virus shed vaccine virus from the nose and virulent virus from the pharynx concurrently after dexamethasone treatment. Evaluation of both viral populations for unique strain characteristics failed to provide evidence of virus recombination.

Pigs [9+/-1] weeks old) were inoculated with Streptococcus suis type 2, pseudorabies virus (PRV) or both. For each pig of groups A, B, and C the inoculum of S suis was 10(9) colony-forming units. For each pig of groups A, B, and D the inoculum of PRV was 5 X 10(3) TCID50 of either PRV strain 4892 (group A, n = 9) or PRV isolate B (group B, n = 9). The PRV strain 4892 is a highly virulent strain; isolate B causes mild clinical signs of infection in inoculated pigs. Group-C pigs (n = 9) were given S suis alone, and group-D pigs (n = 3) were inoculated only with PRV isolate B. Clinical signs of infection and development of lesions were readily seen in pigs of groups A, B, and C. Duration and severity of clinical signs of disease and lesions were reduced in pigs of group C, compared with those of the other 2 groups. Lesions, such as polyarthritis and fibrinous pericarditis, were more abundant and acute in the groups of pigs given mixed challenge exposure, compared with pigs inoculated exclusively with S suis type 2 (group C). The group of pigs inoculated with PRV isolate B alone did not manifest clinical signs of disease or lesions. Average daily gain for group-C pigs was higher, compared with that of other groups; the difference was statistically significant at P < 0.02 and P < 0.05 for groups B and D, respectively. Spread of S suis within the tissues of infected pigs was higher in pigs of groups A and B, compared with pigs of group C. Total number of isolations was 8, 15, and 7 for groups A, B, and C, respectively; S suis was isolated from more than 1 tissue specimen from some pigs. The rate of pigs carrying S suis was 4 of 4 in group-A, 7 of 9 in group-B, and 5 of 9 in group-C pigs. It was concluded that clinical disease associated with S suis type 2 was enhanced by concomitant infection with PRV and such effect was common to both PRV strains tested, the highly virulent strain and the strain with low virulence.

To study the antibody response to glycoprotein I (gI) of pseudorabies virus (PRV) in maternally immune pigs, 3 groups of 6 pigs were given low doses of the mildly virulent Sterksel strain of PRV at 3 and 11 weeks of age. Group A consisted of seronegative pigs; groups B and C consisted of pigs with maternal antibodies deficient of antibodies to gI. At 3 weeks of age, 3 pigs of each group were inoculated intranasally with 10(2.5) plaque-forming units (groups A and B), or with 10(3.5) plaque-forming units (group C) of PRV. The 3 other pigs in each group were contact-exposed to the inoculated pigs. In group A, 4 of 6 pigs shed virus and all developed antibodies to gI of PRV and produced PRV-specific IgM and virus-neutralizing antibodies. In groups B and C, 10 pigs shed virus and all developed low and inconsistent titers of gI antibodies, whereas only 3 pigs produced PRV-IgM antibodies with low titers. Thus, after PRV infection of pigs with high concentrations of maternal antibodies deficient of gI antibodies, the antibody responses to PRV were severely inhibited. The pigs were reinoculated with 10(3) plaque-forming units of the same virus 8 weeks after the first inoculation. The pigs in group A did not respond at all, as they were immune. The pigs in groups B and C shed considerable amounts of virus. Three pigs had a clear secondary antibody response to gI, whereas the others developed an early to normal antibody response to gI. None of the pigs mounted a secondary neutralizing antibody response to PRV. We concluded that inoculation of pigs having high titers of gI-deficient maternal antibodies with low doses of a mildly virulent PRV, induced low to undetectable concentrations of antibodies to gI.

A fully age-structured deterministic model of the population biology of a pseudorabies epizootic in a farrow-to-finish operation was used to examine the disease-related change in productivity following the initial disease episode. A strategy involving continual sow vaccination was compared with various strategies involving the vaccination of growing pigs, as well as sows. The model suggests that vaccinating growing pigs, in addition to the breeding herd, results in only a relatively small improvement in long-term productivity following a pseudorabies epizootic.

Pseudorabies is a porcine herpesvirus of major importance in the swine industry. Isoprinosine is an immunomodulating drug that has been shown to be beneficial in treating herpesvirus infections. Twenty-four 7-week-old pigs were allotted within litters to 1 of 4 groups: control, isoprinosine (ISO), pseudorabies virus (PRV), or isoprinosine and pseudorabies virus (ISO-PRV). Isoprinosine was administered daily for 16 days to the ISO and ISO-PRV groups (75 mg/kg of body weight/day, PO). Immunity in pigs in the PRV and ISO-PRV groups was challenged with pseudorabies virus (10(5) TCID50 units) on day 4. Rectal temperatures and viral excretion were monitored daily; total and differential leukocyte counts, lymphocyte response to mitogens, and interleukin-2 production were monitored every 4 days. Pigs challenge-inoculated with pseudorabies virus became ill, with the ISO-PRV group most severely affected. Rectal temperatures were high (P less than 0.05) in virally challenged pigs on days 5 to 12 and 14 to 16; isoprinosine did not alter this effect. Pseudorabies virus-infected pigs had leukocytosis (P less than 0.05) on days 12 and 16, primarily caused by neutrophilia. Concanavalin A-stimulated lymphocyte proliferation was decreased (P less than 0.06) in both PRV and ISO-PRV groups on day 12, compared with control pigs, but only in the PRV group on day 16. Pokeweed mitogen-stimulated lymphocyte proliferation was decreased (P less than 0.02) in ISO-PRV pigs on day 8 of the experiment. Interleukin-2 concentrations, pooled over all sampling days, were decreased (P less than 0.03) in pseudorabies virus-infected pigs. Viral excretion was not altered by isoprinosine treatment. These data suggest that pseudorabies virus infection decreased lymphocyte proliferative responses and interleukin-2 prodcution in pigs, and that isoprinosine did not mitigate these effects.

A field trial was conducted on a commercial swine farm quarantined because of infection with pseudorabies virus. The purpose was to investigate, in growing pigs born to hyperimmunized sows, the immunogenicity of a vaccine with a glycoprotein I (gE) deletion. One hundred twenty pigs were assigned at random to 1 of 3 vaccination schedules at ages: 8 and 12 weeks; 8, 12, and 14 weeks; and 8, 12, and 16 weeks. Immune response was measured at 8, 12, 14, 16, and 18 weeks, using the serum neutralization test, a screening ELISA, and assays of IgG and IgA in serum and nasal secretions. Results of the serum neutralization test and the screening ELISA indicated that, for pigs vaccinated only at 8 and 12 weeks, the percentage of pigs with pseudorabies virus serum antibodies decreased substantially by 18 weeks; for pigs given a booster at 14 or 16 weeks, the prevalence of serum antibodies at 18 weeks was higher, with 16-week booster vaccination eliciting the best response. At each age, nasal IgA and IgG values were highly correlated (r greater than or equal to 0.70), as were serum IgA and IgG values; correlations of serum with nasal IgA and IgG values were somewhat lower (approx range, r = 0.40 to 0.70). Nevertheless, an increase in serum IgA or IgG values on vaccination was no guarantee of an increase in nasal IgA or IgG values. For serum and nasal mucosal antibodies, a poor immune response was associated with high quantities of maternally derived antibodies. Vaccination at 16 weeks was necessary to ensure eliciting of an immune response in almost all pigs.

Use of a combination of an effective gE gene-deleted pseudorabies virus (PRV) vaccine with a companion diagnostic kit for PRV glycoprotein gE has proven successful in several pseudorabies-eradication programs. To produce a large quantity of functional gE protein for development of a PRV-gE diagnostic kit, an Escherichia coli expression system containing the distal region of the PRV-gE gene of a PRV strain CF was constructed. The expressed protein contained 134 amino acids of gE protein (amino acids 77-210) fused to a 19-amino acids tag containing 6 histidine residues. After induction, a truncated PRV-gE polypeptide of 18-kd was expressed to about 20% of the total E coli proteins. Results of immunoblot analysis indicated that this E coli-produced PRV-gE protein reacted specifically with serum from PRv-hyperimmunized pigs and from field PRv-infected pigs, but not with serum samples from specific-pathogen-free pigs or pigs inoculated with gE-deleted PRV vaccine. These data indicate that, although the recombinant gE protein is produced in E coli, it still retains the antigenicity of the viral gE glycoprotein. Comparison between the recombinant gE protein, using immunoblot analysis with a commercial gE ELISA containing natural PRV-gE protein, revealed comparable test performance. This finding indicated that recombinant gE protein produced by E coli can be used for development of a companion serologic assay for a PRV-gE gene-deleted vaccine.