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Biomedical subjects

T G Kimman

Publications and source records attributed to T G Kimman.

At least 73 records · Page 4Linked to original sources
PubMed1994

Virulence and pathogenesis of non-virulent and virulent strains of pseudorabies virus expressing envelope glycoprotein E1 of hog cholera virus.

Pseudorabies virus (PRV) expressing the envelope glycoprotein E1 (E1) of hog cholera virus (HCV) was used as a model to study the potential risks connected with the use of a live herpesvirus vaccine expressing a foreign gene. The gene encoding E1 was inserted into the glycoprotein X (gX) locus of both a virulent PRV strain and a non-virulent PRV strain in which the virulence genes encoding glycoprotein I (gI) and thymidine kinase (TK) had been inactivated. We investigated whether strain M205 (gI-, TK-, gX-, E1+) had a changed cell or host tropism or virulence compared with strain M206 (gI-, TK-, gX-) in pigs, rabbits, hamsters, rats, mice and rhesus monkeys. The insertion of E1 into this non-virulent PRV strain caused no change in cell or host tropism. However, pigs inoculated with M205 shed less virus over a shorter period than pigs inoculated with M206. Theoretically, virulent PRV strains expressing E1 (gX-, E1+) could arise through transfer of the E1 gene of M205 to a virulent PRV strain. Therefore, we inoculated pigs with strain M12 (gX-, E1+) or the control strain M104 (gX-) and compared the virulence and pathogenesis. M12 and M104 were of approximately equal virulence and the pathogenesis of both strains was similar. We concluded that incorporating E1 of HCV into the gX locus of PRV did not change cell or host tropism, nor did it change the virulence of either non-virulent or virulent PRV.

Animals↗
PubMed1994

Glycoprotein gE-negative pseudorabies virus has a reduced capability to infect second- and third-order neurons of the olfactory and trigeminal routes in the porcine central nervous system.

We investigated the spread of glycoprotein gE (gE)-negative pseudorabies virus (PRV) and its rescued 'wild-type' strain into and within the central nervous system (CNS) of 3- and 10-week-old pigs. This is the first study that demonstrates PRV invasion of the porcine CNS via the synaptically linked neurons of the olfactory and trigeminal routes and that demonstrates the role of gE in this invasion. After intranasal inoculation with high doses of virus, gE-negative PRV replicated less efficiently in peripheral tissues. The titres of the gE-negative virus in the oropharyngeal mucosa, olfactory epithelium, draining lymph nodes and trigeminal ganglion were approximately 100-fold lower in 3-week-old pigs and 10-fold lower in 10-week-old pigs than titres of the 'wild-type' virus. In contrast to the 'wild-type' virus, titres of the gE-negative virus were very low or undetectable in the olfactory bulb, brain stem and other tissues of the CNS. Viral antigen of rescued 'wild-type' PRV and of gE-negative PRV was detected immunohistochemically in the olfactory epithelium and in neurons of the trigeminal ganglion, and also in the olfactory and trigeminal axons leading towards the CNS. But, in contrast to 'wild-type' virus, no viral antigen of the gE-negative virus was detected in second- or third-order neurons in the olfactory bulb or in the brain stem. We conclude that gE-negative PRV can infect first-order neurons of the olfactory and trigeminal routes and is able to spread via their axons towards the CNS. Yet, gE-negative PRV has a greatly reduced capacity to infect second- or third-order neurons. Finally, we report lateral spread of 'wild-type' PRV in the trigeminal ganglion, i.e. nonsynaptic transport from neuron to neuron. Possible mechanisms that could explain the reduced levels of the gE-negative virus in the CNS are discussed.

Aging↗
PubMed1994

Epitope-specific antibody response against glycoprotein E of pseudorabies virus.

In this study we investigated the epitope-specific antibody response against glycoprotein E (gE) of pseudorabies virus. Epitope-specific antibody responses were investigated by enzyme-linked immunoperoxidase monolayer assays. In a vaccinated crossbred pig population, most pigs responded to antigenic domain E and to a lesser degree to antigenic domains C and D. Only few pigs responded to antigenic domains F, A, and B. Using vaccinated pigs, we investigated the influence of two different pseudorabies virus strains and the genetic background of the host on the epitope-specific antibody response. More pigs infected with the virulent NIA-3 strain had a detectable antibody response against antigenic domains C, F and A than did pigs infected with the mildly virulent Sterksel strain (P < or = 0.05; Fisher's exact test). No differences in the epitope-specific antibody responses of two genetically different pig breeding lines were observed (P > or = 0.1; Fisher's exact test). In both breeding lines the incidence of the epitope-specific antibody response was comparable to that in the crossbred pig population. In addition, we studied the epitope-specific antibody responses were strikingly different and indicated that genetic background influenced the epitope-specific antibody response. Of the serum samples of mice with C57BL and a BALB background, 40 and 17% respectively, were positive in the one of the epitope-specific immunoassays. In contrast to pigs, mice responded predominantly to antigenic domain D and to a lesser degree to antigenic domains E and B. Only few mice had a detectable antibody response against antigenic domains C and A, and none had a detectable antibody response against antigenic domain F.

Animals↗
PubMed1994

Glycoprotein I of pseudorabies virus (Aujeszky's disease virus) determines virulence and facilitates penetration of the virus into the central nervous system of pigs.

In the present study we investigated the virulence and neural spread of pseudorabies virus (PRV) strains with mutations in the gene encoding glycoprotein I (gI) in 3-week-old pigs which were intranasally infected. Mutant M303 (lambda 125, 126) lacks amino acids valine-125 and cysteine-126 in an immunodominant antigenic region of gI which contains 2 discontinuous antigenic domains, whereas mutant virus M304 (lambda 59, 60) lacks amino acids glycine-59 and aspartic acid-60 in a continuous antigenic domain. Mutant M301 contains a frame shift mutation. Both mutants M301 (gI-) and M303 (lambda 125, 126) were not virulent for pigs, whereas mutant M304 (lambda 59, 60) was as virulent as wild-type PRV. All gI mutant viruses replicated in the oropharyngeal mucosa, although M304 (lambda 59, 60) and wild-type PRV replicated to higher titres than M303 (lambda 125, 126) and M301 (gI-). In contrast to M304 (lambda 59, 60) and wild-type PRV, both mutant viruses M301 (gI-) and M303 (lambda 125, 126) were not recovered from any part of the central nervous system at day 6 after infection. To study the spread of M301 (gI-) in the central nervous system in more detail, a second experiment was done in which 100-fold more virus was intranasally administered and virus was recovered from various tissues at day 4 after infection. Again, no gI-negative virus was isolated from the central nervous system. We concluded that deleting the amino acids valine-125 and cysteine-126 decreases virulence and reduces neurotropism to the same degree as deleting the gI protein. In addition, gI-negative virus does not spread in the central nervous system of pigs, probably because the transport of the virus across the synapse is hampered.

Animals↗
PubMed1994

Reduction of the prevalence of pseudorabies virus-infected breeding pigs by use of intensive regional vaccination.

The influence of an intensive vaccination program on prevalence of pseudorabies virus (PRV)-infected breeding pigs was examined in a region where the disease was enzootic. In a 2-year period, significantly (P < 0.005) greater decrease in the prevalence of PRV-infected breeding pigs was observed in herds participating in the regional vaccination program and in herds outside the trial region that intensified PRV vaccination during the study than in herds that applied routine PRV control measures. In the regional vaccination program, introduction of breeding stock from outside the area was significantly (P < 0.05) associated with higher prevalence of PRV-infected pigs at the end of the study. These results indicate that transmission of PRV can be markedly reduced by use of an accurately applied intensive vaccination program.

Animals↗
PubMed1993

Use of colostrum to detect antibodies against glycoprotein I of Aujeszky's disease virus.

Serum and colostrum samples taken from 499 sows from five herds of pigs endemically infected and vaccinated against Aujeszky's disease virus, were used to investigate whether colostrum could be used to detect antibodies against glycoprotein I (gI) of the virus. Using serum as the reference, the test applied to colostrum had a sensitivity of 97 per cent and a specificity of 88 per cent. When samples were taken from 50 sows from a gI seronegative vaccinated herd, one colostrum sample was gI-positive, giving a specificity for the test of 98 per cent. The mean gI antibody titres in colostrum were about six times higher than in serum. Samples of colostrum were also taken from 132 sows from eight unvaccinated herds free of Aujeszky's disease virus. All these samples were gI-negative, giving a specificity of 100 per cent. Colostrum samples can be stored for at least six weeks at -20 degrees C without compromising the test results, and the repeatability and reproducibility of the test applied to colostrum were good.

Animals↗
PubMed1993

Deleting valine-125 and cysteine-126 in glycoprotein gI of pseudorabies virus strain NIA-3 decreases plaque size and reduces virulence in mice.

We investigated the function of antigenic domains on gI in virulence and immunogenicity. Three PRV gI mutants were constructed by deleting nucleotides coding for the following amino acids: valine-125 and cysteine-126, located in a discontinuous antigenic domain (M 303); glycine-59 and aspartic acid-60 located in a continuous antigenic domain (M304); and arginine-67 and alanine-68, located in a discontinuous antigenic domain (M305). Mismatch primers in the polymerase chain reaction were used to introduce the deletions. Anti-gI monoclonal antibodies were used in an immunoperoxidase monolayer assay to distinguish PRV gI mutants from wild-type PRV. The gI mutant viruses were tested for their growth in vitro and for their virulence in mice. The growth properties of PRV gI mutant virus M303 were comparable to the growth properties of a PRV gI-negative mutant (M301): both mutants produced small plaques in various cells, and when grown on swine kidney cells and chicken embryo fibroblasts, their growth was disadvantaged compared to wild-type PRV. However, in embryonal Balb/c mouse cells expressing gI, gI mutant viruses and wild-type PRV produced plaques of the same size, confirming that the mutations in gI are responsible for the small plaque phenotype. The growth properties of PRV gI mutant viruses M 304 and M 305 were comparable to the growth properties of wild-type PRV. When the mean time to death was used as the criterion, the gI mutant viruses M 301 and M 303 were significantly less virulent in mice than wild-type PRV. Four other, independently obtained, PRV mutants all carrying the valine-125 and cysteine-126 deletion (M 308, M 309, M 310 and M 311 respectively) exhibit the same phenotype. Our results show that deleting valine-125 and cysteine-126 in gI decreases plaque size and reduces virulence in mice to the same degree as deleting the gI protein.

Amino Acid Sequence↗
PubMed1993

Deleting two amino acids in glycoprotein gI of pseudorabies virus decreases virulence and neurotropism for pigs, but does not affect immunogenicity.

The virulence, pathogenicity and immunogenicity of two pseudorabies virus (PRV) variants were investigated in 3-week-old pigs that had been intranasally infected. Variant M303 (delta 125,126) lacked amino acids valine (125) and cysteine(126) in an immunodominant antigenic region of glycoprotein I (gI) containing two discontinuous antigenic domains, whereas M304 (delta 59,60) lacked amino acids glycine(59) and aspartic acid(60) in a continuous antigenic domain. M303 (delta 125,126) was not virulent for pigs, but M304 (delta 59,60) was as virulent as wild-type PRV: all pigs died within 8 days of infection. Both gI mutant viruses replicated in the oropharyngeal mucosa, although M304 (delta 59,60) replicated to higher virus titres than M303 (delta 125,126), and virus was recovered from various tissues. However, in contrast to M304 (delta 59,60), M303 (delta 125,126) was not recovered from any central nervous system (CNS) tissues examined. Thus, the tendency of PRV to locate in the CNS was markedly reduced by deleting amino acids valine(125) and cysteine(126) of gI. Pigs immunized with M303 were completely protected against challenge infection; no clinical signs of disease were detected, no virus was shed, and no secondary antibody response was detected. Thus, deleting amino acids valine(125) and cysteine(126) in gI decreases virulence and neurotropism and does not affect immunogenicity.

Amino Acids↗
PubMed1993

Characterization of the pseudorabies virus-specific immunoglobulin M response and evaluation of its diagnostic use in pigs with preexisting immunity to the virus.

Despite preexisting immunity to pseudorabies virus (PRV), pigs may become infected and may or may not show clinical signs of disease. To investigate whether detection of immunoglobulin M (IgM) antibodies to PRV is suitable for diagnosis of recent infection in pigs with (or without) preexisting immunity, the IgM responses of pigs were examined after both experimental and natural infections. Upon inoculation of seronegative pigs with a low dose of a mildly virulent strain of PRV, IgM was first detectable at day 7 postinoculation (p.i.), reached a maximum at day 14 p.i., and became undetectable again at about days 32 to 36 p.i. In inoculated pigs with maternal antibodies against PRV, the IgM response began later and ended sooner, and peak titers were also lower. In immune pigs with maternally derived antibodies, there was apparently no correlation between the virulence of the inoculated strain and the IgM response. The suitability of the IgM enzyme-linked immunosorbent assay (ELISA) for detection of recent infection in the field was compared with that of the virus neutralization (VN) assay and with an ELISA which specifically detects antibodies directed to glycoprotein I (gI) of PRV. Paired sera were obtained from pigs suspected of PRV infection in an area endemic for PRV infection in which vaccination against PRV is often applied. Practically all pigs had antibodies to PRV in the acute phase of the disease. Compared with the VN assay, the specificity of the IgM ELISA was high but its sensitivity was low. However, all three serotests apparently failed to detect some PRV infections. The IgM ELISA appeared to be especially useful as a diagnostic aid for detection of recent infections in pigs with high levels of neutralizing and gI antibodies, probably maternally derived, in the acute phase of the disease. Such pigs may fail to develop a significant rise in VN antibody titer. The IgM ELISA may be the only serotest for monitoring infections in such pigs.

Animals↗
PubMed1993

Cellular immune response to hog cholera virus (HCV): T cells of immune pigs proliferate in vitro upon stimulation with live HCV, but the E1 envelope glycoprotein is not a major T-cell antigen.

T-cell responses of pigs to hog cholera virus (HCV) have reportedly been absent or difficult to detect. Therefore, little is known about cellular immunity to HCV. In this study, we used an attenuated strain of pseudorabies virus expressing the envelope glycoprotein E1 of HCV and purified recombinant E1 to examine whether the E1 protein is a target antigen recognized by the T cells of HCV-immune pigs. We were unable to identify the E1 protein as a major target antigen recognized by the T cells of HCV-immune animals. However, such cells proliferated in vitro upon stimulation with viable HCV antigen. The lymphoproliferative response to HCV was strictly time and dose dependent and could be induced upon stimulation by live but not by UV light-inactivated HCV. Depletion studies demonstrated that lymphoproliferation depended on the presence of CD2+CD8bright+ lymphocytes, but CD2+CD4+ cells also contributed to the lymphoproliferative response. The primary lymphoproliferative response in animals inoculated with 10(7) 50% tissue culture infective doses of strain Brescia 2.1.1 was stronger than that observed in animals inoculated with 10(3) 50% tissue culture infective doses of the Cedipest strain. A remarkable finding was the increase in non-antigen-specific lymphoproliferation upon inoculation of the animals with HCV strains. This immunological phenomenon may mask a specific T-cell response to the virus.

Animals↗
PubMed1992

Measurement of isotype-specific antibody responses to Aujeszky's disease virus in sera and mucosal secretions of pigs.

Enzyme-linked immunosorbent assays (ELISAs) for the detection of porcine IgM, IgA, IgG1 and IgG2 antibodies directed against Aujeszky's disease virus (ADV) are described. ADV-specific IgA and IgM were detected in an antibody capture assay, and ADV-specific IgG1 and IgG2 were detected in an indirect double antibody sandwich assay. A selected set of samples was tested in the four ELISAs and in a 24 h virus neutralization assay. Comparison of the results showed that the ELISAs were isotype-specific, sensitive, and reproducible. Samples with ADV antibody of one isotype showed that ADV-specific IgG1, IgG2 and IgM were able to neutralize the virus in vitro. In vitro neutralization of virus can be enhanced by complement. ADV-specific IgA neutralized virus only weakly. ADV-infected cells activated complement in the absence of antibody. Specific IgG2 and IgM enhanced complement activation. Analysis of the time course of antibody responses after infection or vaccination revealed that the isotype-specific ELISAs are suitable to study the humoral antibody response of pigs to the virus in mucosal secretions. Wild-type virus (strain NIA-3) and an attenuated vaccine strain (Bartha) administered intranasally induced mucosal IgM and IgA responses to the virus. In contrast, a killed vaccine (Nobivac) administered intramuscularly induced only weak mucosal IgM responses. The attenuated vaccine strain primed for a mucosal IgA memory response evoked upon challenge infection with wild-type virus.

Animals↗
PubMed1992

Comparative efficacy of three doses of the genetically engineered Aujeszky's disease virus vaccine strain 783 in pigs with maternal antibodies.

Pigs with low levels of maternally derived antibodies were vaccinated twice intramuscularly with 10(5), 10(5.5), or 10(6) plaque forming units (p.f.u.) of the genetically engineered Aujeszky's disease virus (ADV) vaccine strain 783. Strain 783 has deletions in genes encoding glycoprotein gI and thymidine kinase. All vaccinated pigs showed a high level of protection against clinical disease after challenge infection with virulent ADV. Vaccination also reduced virus excretion. The daily mean virus excretion and the mean number of days with virus excretion, fever, mild clinical signs, and growth retardation were higher in pigs vaccinated with 10(5) than in pigs vaccinated with 10(5.5) or 10(6) p.f.u. of strain 783.

Animals↗
PubMed1992

Role of different genes in the virulence and pathogenesis of Aujeszky's disease virus.

In this study the role of different genes located in the unique short region of the genome of Aujeszky's disease virus was examined. Inactivation of the genes encoding the protein kinase (PK), gp63, and gI reduced virulence of the virus for pigs, in contrast to inactivation of the genes encoding the 28 kDa protein, and gX. There was no correlation between virulence and virus multiplication in vitro or in the oropharynx in vivo. The morphogenesis of the PK mutant was altered. The gI mutant replicated to normal titres in the oropharynx and could be recovered from the trigeminal ganglia but not from other parts of the central nervous system, suggesting that gI facilitates the spread of the virus from neuron to neuron. All mutants induced neutralizing antibody and complete or partial protection against a challenge infection. PK and gp63 were required for the induction of complete protection, although these proteins are reportedly not targets for neutralizing antibody or cytotoxic T cells.

Animals↗
PubMed1992

Round table on control of Aujeszky's disease and vaccine development based on molecular biology.

A summary is given on the 4 topics which were discussed during the round table and which represent current knowledge on the molecular biology of Aujeszky's disease (pseudorabies) virus. They include a review on 1. the genome and gene products of the virus; 2. the viral genes associated with virulence; 3. the immunological role of the viral gene products and 4. studies intended to compare the efficacy of several commercially available vaccines and to establish a possible correlation between antibodies against individual structural viral proteins and degree of protection. It was concluded that gI deleted vaccines appear to be the best choice for use in intensive vaccination programmes towards eradication of Aujeszky's disease virus. However, there remains a need for development of more potent vaccines which induce strong humoral and cell mediated immune responses and afford complete protection, virological protection included. It is often observed that live vaccine strains which are completely avirulent lose much capacity to replicate and spread within the vaccinated animal. It is, however, not excluded that a certain degree of dissemination may be needed to be fully efficacious. Loss of virulence may thus be accompanied by too much loss of immunogenicity. An improved genetic stability of live vaccine strains when they are obtained by genetic manipulation, possibly justifies a more widespread dissemination of the vaccine strain in the body compared to that with conventionally developed strains or compared to what is presently allowed.

Animals↗
PubMed1992

Risks connected with the use of conventional and genetically engineered vaccines.

A review is given of real and potential risks connected with the use of conventional and genetically engineered live and dead vaccines. Special attention is given to live carrier vaccines expressing one or more heterologous genes of other microorganisms. Because most carrier vaccines are still in an experimental phase, there is only limited experience with the risks of carrier vaccines. There are three potential risks of live carrier vaccines which will be discussed: 1. Changes in cell, tissue, of host tropism, and virulence of the carrier through the incorporation of foreign genes. 2. Exchange of genetic information with other vaccine or wild-type strains of the carrier organism. 3. Spread in the environment. Only limited experimental data are available on changes in biological behaviour of microorganisms through the incorporation of foreign genes. For example, there are indications that vaccinia virus carrying the attachment protein G of respiratory syncytial virus (RSV) replicates better in lungs of mice than vaccinia virus carrying other genes of RSV. Poxviruses carry genes that probably determine their replication in different hosts. Exchange of such host tropism genes might alter their host spectrum. Recombination between herpesvirus vaccine or wild-type strains may lead to the appearance of virulent strains with of without heterologous genes. Before carrier vaccines are applied, these risks must be thoroughly evaluated case-by-case. Potential methods for the design of safe carrier vaccines are discussed.

Animals↗
PubMed1992

Contribution of single genes within the unique short region of Aujeszky's disease virus (suid herpesvirus type 1) to virulence, pathogenesis and immunogenicity.

Pigs (3 and 10 weeks old) were infected intranasally with Aujeszky's disease virus (ADV) mutants that functionally lacked one of the non-essential genes in the unique short region of the genome (except the gene encoding the 11K protein). Virus excretion in oropharyngeal fluid and disease symptoms were monitored. Some pigs were killed to study pathogenesis, whereas others were challenged with virulent ADV 8 weeks after the primary infection. Mutants lacking protein kinase, or glycoproteins gp63 or gI showed reduced virulence, but mutants lacking gX or the 28K protein showed normal virulence. Glycoprotein gI appears to affect the tissue tropism of ADV in pigs, presumably by facilitating the spread of the virus through the central nervous system. In this study, there was no correlation between virulence and virus multiplication in either cultured cells or in the oropharynx in vivo. All mutants induced neutralizing antibody and complete or partial protection against challenge infection. Complete protection was obtained by inoculation with the gI and gX mutants, whereas incomplete protection was obtained using gp63 and protein kinase mutants. Complete clinical and virological protection was associated with the absence of secondary antibody responses in the serum.

Animals↗
PubMed1992

Role of memory B-cell responses in serum and mucosal fluids of swine for protective immunity against pseudorabies virus.

We examined primary and memory isotype-specific antibody responses directed against pseudorabies virus in serum and mucosal fluids of pigs with and without passively acquired maternal antibody, and we studied the relationship between these responses and protection against virus challenge. Pigs were inoculated intranasally with the virulent NIA-3 strain or the avirulent Bartha strain, or they were inoculated IM with an inactivated vaccine containing the Phylaxia strain. Ten weeks later, all pigs were challenge-exposed intranasally with strain NIA-3. Only pigs that were without passively acquired antibody at the time they were inoculated with virulent virus appeared to have complete protective immunity against challenge exposure, as evidenced by lack of clinical signs of pseudorabies and lack of virus excretion. In contrast, pigs inoculated with strain Bartha or with the inactivated vaccine developed fever, had a period of growth arrest, and excreted virus after challenge exposure. In pigs without passively acquired antibody, intranasal inoculation with strains NIA-3 or Bartha was followed by primary IgM and IgA responses in serum and in oropharyngeal fluid as well as primary IgG1 and IgG2 responses in serum. Intramuscular inoculation with the inactivated vaccine induced primary serum IgM, IgG1, and IgG2 responses, but no mucosal responses. Challenge exposure of pigs that had been inoculated with the Bartha strain or the inactivated vaccine was followed by clear memory responses in serum and in oropharyngeal fluid. In contrast, challenge exposure of pigs that had been inoculated by the virulent NIA-3 strain was not followed by memory responses.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗