Infection of pigs by Aujeszky's disease virus via the breath of intranasally inoculated pigs

Aujeszky's disease is a worldwide problem in the pig industry. In this experiment, four pigs chosen to act as shedder pigs were intranasally infected with Aujeszky's disease virus. Next, on three consecutive days, eight recipient pigs were exposed to the breath of a pair of shedder pigs via a mask-to-mask module. Except for the virtual absence of CNS signs, shedder pigs expressed clinical signs that were similar to pigs infected naturally or experimentally. Only mild respiratory signs occurred in recipient pigs, but all were infected by aerosols of Aujeszky's disease virus as evidenced by seroconversion. The pig is a much more sensitive indicator of airborne virions than our aerosol collection methods. We conclude that the mild respiratory disease acquired by the aerogenous route in recipient pigs is an easily managed model for studying the transmission of airborne respiratory infections and the immune responses to this type of infection.     

Serological response of pigs infected with Aujeszky's disease virus

The temporal development of antibody in four groups of pigs infected with different Aujeszky's disease virus isolates was examined. The enzyme-linked immunosorbent assay detected antibody by five to six days after infection and the antibody-dependent cell-mediated cytotoxicity assay detected antibody seven to nine days after infection. Neutralising antibody was first detected nine to 10 days after infection, whereas assays measuring complement mediated antibody lysis did not detect antibody until 10 days after infection. These results are discussed in terms of their importance to the diagnosis of and recovery from Aujeszky's disease.     

Horizontal transmission of Aujeszky's disease virus from sheep to pigs

Eight 2-month-old merino lambs were inoculated intranasally with different (10(2.0)-10(5.0)TCID50) amounts of Aujeszky's disease virus (ADV). Electron microscopic studies indicated that ADV replicated in extra-neural sites, in the epithelial cells of the mucosa of the upper and lower respiratory tract. Although the virus was excreted continuously in nasal discharges, horizontal transmission to contact lambs failed. The surviving exposed and contact lambs had no demonstrable antibodies against ADV and they were susceptible when challenged by ADV. However, the virus was transmitted to susceptible pigs in contact with the exposed lambs. One of the five contact pigs showed characteristic clinical signs of Aujeszky's disease, developed a nonsuppurative meningoencephalomyelitis and ADV was recovered from the brain, nasal discharge and other organs. Restriction enzyme analysis of DNA from this virus confirmed the sheep origin of the isolate. The other 4 pigs seroconverted. ADV infection in sheep is therefore a possible source of infection for pigs, but the lack of horizontal transmission in sheep was confirmed.     

Detection of Aujeszky's disease virus in nasal cells of fattening pigs by immunoassay

Both conventional and specific pathogen free pigs were inoculated intranasally with a strain of Aujeszky's disease virus (ADV). Nasal cells were collected daily by swab, aspiration or wash. The nasal cells were examined for ADV by isolation on cell culture, direct or indirect immunofluorescence and immunoperoxidase staining by monoclonal antibodies. The infected pigs were studied for nasal shedding of infected cells until 30 days after infection. The study was also extended to naturally infected farm pigs. Swabbing, washing and aspiration proved effective methods of collecting between 10(5) and 10(8) pavement or columnar epithelial cells and non-epithelial cells. Macrophages and polymorphonuclear leucocytes were also identified. Infected nasal cells were detected by immunofluorescence and immunoperoxidase from one to 21 days after infection. The viral antigen was detected in both epithelial and non-epithelial cells, the fluorescence was nuclear and, or, 'cytoplasmic', in the latter case only the cell membrane was stained. ADV antigens were detected in nasal cavity cells in pigs infected with a virulent and a hypovirulent strain. Nasal swabs proved effective in confirming infection both by virus isolation and immunological assay, and the latter was shown to be a useful experimental tool for the rapid diagnosis of Aujeszky's disease virus infection in fattening pigs suffering from acute respiratory distress.     

The rapid detection of Aujeszky's disease virus in pigs by direct immunoperoxidase labelling

Direct immunoperoxidase labelling on impression smears of brain and pharynx was compared with virus isolation and direct immunofluorescence for the detection of Aujeszky's disease virus in experimentally-infected pigs. Immunoperoxidase labelling was as sensitive as immunofluorescence and more sensitive than virus isolation for tissue that had been stored at room temperature (approximately 20 degrees C) for up to 144 h.     

Antibodies to Aujeszky's disease virus in pigs immunized with purified virus glycoproteins

Antibodies to Aujeszky's disease virus (ADV) glycoproteins gII, gIII, and gp50 were compared using four in vitro tests. Antibodies generated by vaccination with a modified-live vaccine (MLV) were also compared. The serological assays employed were: serum neutralization test (SNT), complement facilitated serum neutralization test (C'SNT), complement-mediated cytolysis and antibody dependent cellular cytotoxicity (ADCC). Pigs were immunized with single glycoproteins twice 14 days apart, or once with the modified-live vaccine. Fourteen days after the second immunization, sera were collected. Virus neutralizing activity (SNT) was demonstrated in the sera from all pigs immunized with gp50 and in one out of three immunized with gIII. Sera from the MLV group all had neutralization titers higher than animals immunized with single glycoproteins. Addition of guinea pig complement to the serum neutralization test (i.e., C'SNT) produced an enhancement of antibody titers in all groups except the pigs immunized with gIII. The complement-mediated cytolysis test rendered antibody titers similar in magnitude for all pigs immunized with single glycoproteins, but slightly lower than values for MLV vaccinated pigs. ADCC activity was clearly displayed in sera from pigs immunized with gIII or vaccinated with MLV, whereas sera from pigs immunized with gII or gp50 had a minimal response. The results indicate that the relative efficiency of antibodies against ADV glycoproteins in protection should be considered for selecting or producing gene-deleted strains for use in vaccine production.     

Local humoral and cellular responses in Aujeszky's disease virus infection in pigs

Mucosal and tracheal washings from pigs vaccinated parenterally and intranasally with Aujeszky's disease virus were tested for specific anti-Aujeszky's disease virus responses. Antibody tests included complement dependent antibody lysis, antibody dependent cellular cytotoxicity, virus neutralisation, and anti-Aujeszky's disease virus IgA and IgG levels. There was no correlation between the levels of these antibodies and protection from subsequent challenge. Direct lymphocyte cytotoxicity against cells infected with Aujeszky's disease virus was found in lymph nodes draining the tonsillar area.     

Tonsillar changes in pigs given pseudorabies (Aujeszky's disease) virus

All of the eight 5-day-old pigs orally given pseudorabies (Aujeszky's disease) virus developed tonsillitis. The initial changes occurred in the subepithelial area between the lymphoid nodule and the crypt epithelium, showing a characteristic pattern of necrosis. The necrosis became more severe and gained access into the lymphoid nodule and crypt epithelium. Coincident with the histopathologic changes, numerous specific immunofluorescences were detected, first in the nucleus and in some parts of the cytoplasm of cells distributed in the subepithelial area. The fluorescence subsequently spread into adjacent lymphoid nodules and crypt epithelial cells. Ultrastructurally, many enveloped virus particles were detected in the center of the necrosis. Thereafter, the crypt epithelial cells also underwent degeneration, and a small number of virus particles were detected in the nucleus of the degenerating epithelial cells. In the more advanced stage, the enveloped virus particles were discharged into the crypt lumen.     

Experimental Aujeszky's disease in pigs: excretion, survival and transmission of the virus

Airborne Aujeszky's disease virus was recovered from looseboxes containing groups of pigs infected with virus strains from England, Northern Ireland and Denmark from days 1 to 7 after infection. Pigs sampled individually excreted most airborne virus on days 2 and 3 after infection. On a 24 hour basis the maximum amount of airborne virus excreted per pig was log10 5.3 TCID50. Subclinical infection was transmitted from a clinically affected group of pigs to a seronegative group held in separate looseboxes when air was drawn through ducting connecting one box with the other. Tissues taken from pigs killed at varying times after infection showed that the main sites of virus replication were in the head and neck region. Aujeszky's disease virus was detected for up to 40 days in a range of tissues taken from pigs at the acute stage of disease and stored at -20 degrees C.     

Vaccination of pigs against Aujeszky's disease by the intradermal route using live attenuated and inactivated virus vaccines

A study was undertaken of the protection induced by inactivated and live Aujeszky's disease virus vaccines. The vaccines were administered using a special device which, without the use of a needle, delivered the preparation intradermally. The trials were performed on 88 pigs which were vaccinated at the beginning of the fattening period both in experimental conditions and in pig herds. All the pigs were challenged at the end of the fattening period in isolation units. The results obtained were compared with those obtained using the same vaccines injected intramuscularly. It was shown that vaccination via the intradermal route induced good protection in the vaccinated animals and was similar to that conferred by live virus vaccine injected intramuscularly. The results, with the inactivated virus vaccine, were not so good when it was injected via the intradermal route. Studies with intradermal vaccination showed no local lesion or very small nodules strictly localized to the dermis. The results also confirmed that the effects of challenge exposure depended on the health status of animals prior to infection and show the necessity to use a synthetic value (delta G) to interpret the data and mainly to compare the results objectively. In fattening pigs this vaccination procedure is attractive because (i) less animal constraint is needed than would be for intramuscular injections, (ii) injection can be checked by the presence of a visible papula at the site of inoculation and, (iii) pigs can be vaccinated in the ham while they are feeding. Injection without a needle also contributes to avoiding bacterial contamination under practical farm conditions of vaccination.     

Rapid diagnosis of Aujeszky's disease in pigs by immunofluorescence

Direct immunofluorescence on impression smears of brain and pharynx was compared with virus isolation in cell culture for the diagnosis of Aujeszky's disease in experimentally and naturally infected pigs. Pharyngeal impression smears were more sensitive than virus isolation in two pigs killed 10 and 12 days after experimental infection. Both methods were of similar sensitivity in the detection of virus from field cases of disease. Smears of brain and pharynx were more sensitive than virus isolation for tissue which had been stored at room temperature (approximately 20 degrees C) for up to 48 hours. Some reduction in the amounts of virus recovered from tissues and the intensity of fluorescent staining occurred in these samples.     

Studies on immunisation of pigs with the Bartha strain of Aujeszky's disease virus.

The K strain of Aujeszky's disease virus (ADV) grown in Vero cells was used to vaccinate pigs. Following intramuscular inoculation, the pigs remained healthy, no vaccine virus was excreted and virus could be detected only at the inoculation site. One inoculation gave good protection against challenge with a virulent strain of ADV, and the amount of virulent ADV excreted was geatly curtailed. Following vaccination only low leads of serum neutralizing antibody were detected (geometric mean titre 1/2), but three weeks after challenge very high levels were found (GMT 1/1773). Intranasal vaccination gave similar results. There was minimal excretion of vaccine virus. The clinical reaction on challenge was less severe than in the intramuscularly challenged group, although lower antibody levels were detected three wekks following challenge (GMT 1/483). A field trial, using this strain given subcutaneously, indicated that one inoculation of this vaccine is effective.     

Functional antibody responses in pigs vaccinated with live and inactivated Aujeszky's disease virus

Functional antibody tests, including virus neutralising activity of serum, antibody dependent cellular cytotoxicity and complement mediated lysis, were used to measure the response of pigs given either live or inactivated Aujeszky's disease virus vaccines. Pigs were then challenged with virulent Aujeszky's disease virus and antibody responses were analysed and found not to correlate with protection. Reasons for this lack of correlation are discussed and it is suggested that these results indicate that more emphasis should be placed on measuring the local immune response.