Progress 10/01/99 to 09/30/04
Outputs
Bovine herpesvirus type 5 (BHV-5) is an alphaherpesvirus that causes fatal encephalitis in calves. Envelope glycoproteins E (gE) and gI of alphaherpesviruses are important for the pathogenesis in vivo. Previously we determined that BHV-5 gE is important for BHV-5 neurovirulence. To determine the role of gI in BHV-5 neurovirulence, we have constructed gI-deleted and gI-revertant BHV-5 and analyzed their neuropathogenic properties in a rabbit seizure model. Following intranasal infection, 40% of the rabbits infected with the gI-deleted virus showed severe neurological signs. BHV-5 gI-deleted virus invaded all the CNS structures invaded by the gI-revertant BHV-5, however the number of neurons infected by the gI-deleted virus was similar or slightly reduced (2-4 fold). Thus, the gI-deleted virus retained significant neurovirulence and/or neuroinvasive properties when compared with the gE-deleted BHV-5. Pulse-Chase analysis revealed that the gE of gI-deleted virus
was processed to a larger and a diffused 94-100 kD protein (instead of 94 kD). The 94-100 kD protein was processed in the Golgi with a delayed kinetics but it was EndoH resistant. In cells infected with gI-deleted virus, there was a reduction in cell-surface gE expression compared to wild-type which correlated to reduced amount of gE processed in the Golgi. Based on these results, we believe that in the absence of gI, BHV-5 gE is sufficient for BHV-5 neurovirulence.
Impacts
BHV-5 is naturally attenuated for respiratory disease and BHV-1 is naturally attenuated for neurological disease. These findings suggest that an excellent vaccine candidate for both BHV-1 and BHV-5 infections could be developed from BHV-5 that is attenuated for neurovirulence and neuroinvasion. We have learned that in the case of BHV-5, gE alone determines the virulence. BHV-5 gI-deleted viruses retains significant neurovirulence. This is different from what has been reported for other herpesvirus.
Publications
- Al-Mubarak, A. and Chowdhury, S.I. 2004. Glycoprotein I (gI)-deleted BHV-5 retains significant neurovirulence. J. NeuroVirol. 10: 233-243.
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Progress 01/01/03 to 12/31/03
Outputs
Bovine herpesvirus type 5 (BHV-5) is a neurovirulent alphaherpesvirus that causes fatal encephalitis in calves. Glycoprotein E (gE) is important for neurovirulence of BHV-5. We initiated this study to investigate the role of BHV-5 gE cytoplasmic tail in the BHV-5 gE processing, envelope incorporation and neuropathogenesis. We constructed one amber mutation (Am 480) in the cytoplasmic tail of gE. Am 480 encoded only 480 amino acids (out of total 598 amino acids upstream of the first YXXL motif. Preliminary data indicate that gE Am480 is incorporated within the virion envelope, however incorporation of gE Am 480 was at a significantly lower level when compared with the wild-type gE. Interestingly, gE Am 480 was processed to a lower protein when compared with the wild-type. Construction of an amber mutation immediately upstream of the gE acidic domain is in progress. We have also generated entire gE ectodomain-specific goat antibody. Using this antibody gE endocytosis,
gE surface expression studies are currently being conducted. Preliminary neuropathogenesis studies in our rabbit seizure model indicate that neurovirulence of BHV-5 gE Am 480 is reduced. Further in vivo studies are in progress.
Impacts
BHV-5 is naturally attenuated for respiratory disease and BHV-1 is naturally attenuated for neurological disease. These findings suggest that an excellent vaccine candidate for both BHV-1 and BHV-5 infections could be developed from BHV-5 that is attenuated for neurovirulence and neuroinvasion 1. BHV-1 gE-deleted BHV-1 mutants is significantly attenuated. However, compared to gC-deleted BHV-1 it is less efficacious as a vaccine. An ideal vaccine virus strain should be attenuated, yet would maintain the protective gE antigenic region. The gE cytoplasmic tail truncated mutants may serve that objective.
Publications
- Al-Mubarak, A. and Chowdhury, S.I. 2003. Glycoprotein I (gI)-deleted BHV-5 retains significant neurovirulence (Accepted pending minor revision J. Neurovirology).
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Progress 01/01/02 to 12/31/02
Outputs
We have constructed gI-deleted and gI-revertant BHV-5 recombinants and analyzed their neuropathogenic properties in a rabbit seizure model. Following intranasal infection, 48% of the rabbits infected with the gI-deleted virus showed severe neurological signs. BHV-5 gI-deleted virus invades all the CNS structures invaded by the gI-revertant BHV-5; however, the number of neurons infected by the gI-deleted virus is reduced 2-4 fold. Thus, the gI-deleted virus retained significant neurovirulence and/or neuroinvasive properties when compared with the gE-deleted BHV-5 Pulse-Chase and biochemical analyses revealed that the gE of gI-deleted virus is processed to a considerably larger molecular mass (100 kD instead of 94 kD) and the processed form of gE is endoglycosidase H resistant. Confocal microscopy showed that the epitope-deleted gE is expressed on the cell surface but in a reduced amount when compared with the wild-type gE surface expression. Since the gI-deleted virus
retained significant neurovirulence in rabbits, we believe that in the absence of gI, BHV-5 gE alone is sufficient for BHV-5 neurovirulence.
Impacts
We have determined that BHV-5 glycoprotein gE is important for neuroinvasiveness and neurovirulence in the olfactory pathway of the rabbit. The high degree of homology between BHV-1 and BHV-5 offers us a powerful system to analyze functional differences in the gE between a neuropathogenic and non neuropathogenic herpesvirus. BHV-5 is naturally attenuated for respiratory disease and BHV-1 is naturally attenuated for neurological disease. These findings suggest that an excellent vaccine candidate for both BHV-1 and BHV-5 infections could be developed from BHV-5 that is attenuated for neurovirulence and neuroinvasion.
Publications
- Al-Mubarak, A. and Chowdhury, S.I.. 2002. Glycoprotein I (gI)-deleted BHV-5 retains significant neurovirulence (In submission, J. Neurovirology).
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Progress 01/01/01 to 12/31/01
Outputs
We have determined that the gI-deleted virus retained significant neurovirulence and/or neuroinvasive properties when compared with the wild-type BHV-5 and gE-deleted BHV-5. To understand the role of gE/gI in BHV-5 neuropathogenesis, we are characterizing the processing of gE and gI in the absence of gI and gE, respectively. Pulse-chase analysis revealed that gE in the gI-deleted BHV-5 is processed to its mature form with reduced efficiency. In addition, the processed gE in the gI-deleted virus has a higher molecular weight when compared with the wild-type gE. Since, the gI-deleted BHV-5 retained significant neurovirulence, we believe that in the absence of gI the reduced amount of mature gE is sufficient for BHV-5 neurovirulence.
Impacts
We have determined that BHV-5 glycoprotein gE is important for neuroinvasiveness and neurovirulence in the olfactory pathway of the rabbit. The high degree of homology between BHV-1 and BHV-5 offers us a powerful system to analyze functional differences in the gE between a neuropathogenic and non neuropathogenic herpesvirus. BHV-5 is naturally attenuated for respiratory disease and BHV-1 is naturally attenuated for neurological disease. These findings suggest that an excellent vaccine candidate for both BHV-1 and BHV-5 infections could be developed from BHV-5 that is attenuated for neurovirulence and neuroinvasion.
Publications
- No publications reported this period
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Progress 01/01/00 to 12/31/00
Outputs
Herpesvirus glycoprotein C (gC) is one of the major virus attachment proteins. To determine the contribution of the BHV-5 glycoprotein gC (gC5) to the neuropathogenesis of BHV-5, we have constructed two BHV-5 recombinants: gC-deleted BHV-5 (BHV-5gC-deleted) and BHV-5 expressing BHV1 gC (BHV-5gC1). Neurovirulence properties of these viruses were analyzed using a rabbit seizure model that distinguishes BHV-1 and -5 based on their differential neuropathogeneses. Immuno-histochemistry results showed that the number of infected neurons was two- to fourfold less with the gC-deleted BHV-5 than with the wild type BHV-5. The gC-deleted BHV-5 did not invade the hippocampus but invaded additional sites not invaded by wild- type BHV-5. Similarly, the BHV-5gC1 virus failed to invade the hippocampus, but it did not invade the additional sites. Virus isolation results suggest that these recombinants replicate less efficiently in the brain than the wild-type and gC-revertant viruses.
However, compared to the gC-deleted BHV-5, the gC-exchanged BHV-5gC1 replicated better within the CNS.
Impacts
These results indicate that gC regulates BHV-5 neurotropism in some areas of the olfactory pathway. Additionally, gC is important for BHV-5 neurovirulence in the olfactory pathway but it is not essential.
Publications
- Chowdhury, SI., Lee,, BJ., Onderci, M., Weiss, ML. and Mosier, D. (2000). Neurovirulence of glycoprotein C-deleted Bovine Herpesvirus type-5 (BHV-5) in a rabbit seizure model. J. NeuroVirol. 6: 284-295.
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Progress 01/01/99 to 12/31/99
Outputs
To determine the neurovirulence of BHV-1 and BHV-5 in the olfactory pathway after intracerebral injection, rabbits were injected by steriotaxis injection into the bulb with each viruses separately (5x105 PFUs). Our results indicate that both the viruses can produce neurological disease when injected directly into the bulb. However, BHV-1 seems to be transported less efficiently than the BHV-5. Future studies will involve injection of recombinant gE-deleted and gE-replaced BHV-5 viruses into the bulb.
Impacts
We determined that viral glycoprotein genes play important roles in neuroinvasion of BHV-5. The viral envelope glycoprotein genes gE/gI together play a significant role in BHV-5 neuropathogenesis.
Publications
- No publications reported this period
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