Pseudorabies virus expressing bovine herpesvirus 1 glycoprotein B exhibits altered neurotropism and increased neurovirulence.
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Herpesvirus glycoproteins play dominant roles in the initiation of infection of target cells in culture and thus may also influence viral tropism in vivo. Whereas the relative contribution of several nonessential glycoproteins to neurovirulence and neurotropism of Pseudorabies virus (PrV), an alphaherpesvirus which causes Aujeszky's disease in pigs, has recently been uncovered in studies using viral deletion mutants, the importance of essential glycoproteins is more difficult to assess. We isolated an infectious PrV mutant, PrV-9112C2, which lacks the gene encoding the essential PrV glycoprotein B (gB) but stably carries in its genome and expresses the homologous gene of bovine herpesvirus 1 (BHV-1) (A. Kopp and T. C. Mettenleiter, J. Virol. 66:2754-2762, 1992). Apart from exhibiting a slight delay in penetration kinetics, PrV-9112C2 was similar in its growth characteristics in cell culture to wild-type PrV. To analyze the effect of the exchange of these homologous glycoproteins in PrV's natural host, swine, 4-week-old piglets were intranasally infected with 10(6) PFU of either wild-type PrV strain Kaplan (PrV-Ka), PrV-9112C2, or PrV-9112C2R, in which the PrV gB gene was reinserted instead of the BHV-1 gB gene. Animals infected with PrV-Ka and PrV-9112C2R showed a similar course of disease, i.e., high fever, marked respiratory symptoms but minimal neurological disorders, and excretion of high amounts of virus. All animals survived the infection. In contrast, animals infected with PrV-9112C2 showed no respiratory symptoms and developed only mild fever. However, on day 5 after infection, all piglets developed severe central nervous system (CNS) symptoms leading to death within 48 to 72 h. Detailed histological analyses showed that PrV-9112C2R infected all regions of the nasal mucosa and subsequently spread to the CNS preferentially by the trigeminal route. In contrast, PrV-9112C2 primarily infected the olfactory epithelium and spread via the olfactory route. In the CNS, more viral antigen and significantly more pronounced histological changes resulting in more severe encephalitis were found after PrV-9112C2 infection. Thus, our results demonstrate that replacement of PrV gB by the homologous BHV-1 glycoprotein resulted in a dramatic increase in neurovirulence combined with an alteration in the route of neuroinvasion, indicating that the essential gB is involved in determining neurotropism and neurovirulence of PrV. (+info)
Role of pseudorabies virus Us9, a type II membrane protein, in infection of tissue culture cells and the rat nervous system.
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The protein product of the pseudorabies virus (PRV) Us9 gene is a phosphorylated, type II membrane protein that is inserted into virion envelopes and accumulates in the trans-Golgi network. It is among a linked group of three envelope protein genes in the unique short region of the PRV genome which are absent from the attenuated Bartha strain. We found that two different Us9 null mutants exhibited no obvious phenotype after infection of PK15 cells in culture. Unlike those of gE and gI null mutants, the plaque size of Us9 null mutants on Madin-Darby bovine kidney cells was indistinguishable from that of wild-type virus. However, both of the Us9 null mutants exhibited a defect in anterograde spread in the visual and cortical circuitry of the rat. The visual system defect was characterized by restricted infection of a functionally distinct subset of visual projections involved in the temporal organization of behavior, whereas decreased anterograde spread of virus to the cortical projection targets was characteristic of animals receiving direct injections of virus into the cortex. Spread of virus through retrograde pathways in the brain was not compromised by a Us9 deletion. The virulence of the Us9 null mutants, as measured by time to death and appearance of symptoms of infection, also was reduced after their injection into the eye, but not after cortical injection. Through sequence analysis, construction of revertants, measurement of gE and gI protein synthesis in the Us9 null mutants, and mixed-infection studies of rats, we conclude that the restricted-spread phenotype after infection of the rat nervous system reflects the loss of Us9 and is not an indirect effect of the Us9 mutations on expression of glycoproteins gE and gI. Therefore, at least three viral envelope proteins, Us9, gE, and gI, function together to promote efficient anterograde transneuronal infection by PRV in the rat central nervous system. (+info)
Antiretroviral resistance mutations in human immunodeficiency virus type 1 reverse transcriptase and protease from paired cerebrospinal fluid and plasma samples.
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Twenty-four adults infected with human immunodeficiency virus type 1 (HIV-1) with central nervous system symptoms were studied for antiretroviral resistance mutations in HIV-1 RNA obtained from paired cerebrospinal fluid (CSF) and plasma samples. Paired sequences were obtained from 21 and 13 patients for reverse transcriptase (RT) and for protease, respectively. Mutations conferring resistance to the RT inhibitors zidovudine, lamivudine, or nevirapine were detected in 14 patients, including 11 pretreated and 3 drug-naive subjects. The mutation patterns in the 2 compartments were different in most patients. Genotypic resistance to protease inhibitors was detected in both plasma and CSF from 1 patient treated with multiple protease inhibitors. However, accessory protease inhibitor resistance mutations at polymorphic sites were different in plasma and CSF in several patients. Partially independent evolution of viral quasispecies occurs in plasma and CSF, raising the possibility that compartmentalization of drug resistance may affect response to antiretroviral treatment. (+info)
Synaptic pathology in Borna disease virus persistent infection.
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Borna disease virus (BDV) infection of newborn rats leads to a persistent infection of the brain, which is associated with behavioral and neuroanatonomical abnormalities. These disorders occur in the absence of lymphoid cell infiltrates, and BDV-induced cell damage is restricted to defined brain areas. To investigate if damage to synaptic structures anteceded neuronal loss in BDV neonatally infected rats, we analyzed at different times postinfection the expression levels of growth-associated protein 43 and synaptophysin, two molecules involved in neuroplasticity processes. We found that BDV induced a progressive and marked decrease in the expression of these synaptic markers, which was followed by a significant loss of cortical neurons. Our findings suggest that BDV persistent infection interferes with neuroplasticity processes in specific cell populations. This, in turn, could affect the proper supply of growth factors and other molecules required for survival of selective neuronal populations within the cortex and limbic system structures. (+info)
Control of Sindbis virus infection by antibody in interferon-deficient mice.
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Antibodies clear Sindbis virus from infected animals through an unknown mechanism. To determine whether interferon-induced pathways are required for this clearance, we examined mice which are unable to respond to alpha/beta interferon or gamma interferon. Although extremely susceptible to infection, such mice survived and completely cleared virus if antibodies against Sindbis virus were given. (+info)
A molecular clone of simian-human immunodeficiency virus (DeltavpuSHIV(KU-1bMC33)) with a truncated, non-membrane-bound vpu results in rapid CD4(+) T cell loss and neuro-AIDS in pig-tailed macaques.
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We report on the role of vpu in the pathogenesis of a molecularly cloned simian-human immunodeficiency virus (SHIV(KU-1bMC33)), in which the tat, rev, vpu, env, and nef genes derived from the uncloned SHIV(KU-1b) virus were inserted into the genetic background of parental nonpathogenic SHIV-4. A mutant was constructed (DeltavpuSHIV(KU-1bMC33)) in which 42 of 82 amino acids of Vpu were deleted. Phase partitioning studies revealed that the truncated Vpu was not an integral membrane protein, and pulse-chase culture studies revealed that cells inoculated with DeltavpuSHIV(KU-1bMC33) released viral p27 into the culture medium with slightly reduced kinetics compared with cultures inoculated with SHIV(KU-1bMC33). Inoculation of DeltavpuSHIV(KU-1bMC33) into two pig-tailed macaques resulted in a severe decline of CD4(+) T cells and neurological disease in one macaque and a more moderate decline of CD4(+) T cells in the other macaque. These results indicate that a membrane-bound Vpu is not required for the CD4(+) T cell loss and neurological disease in SHIV-inoculated pig-tailed macaques. Furthermore, because the amino acid substitutions in the Tat and Rev were identical to those previously reported for the nonpathogenic SHIV(PPc), our results indicate that amino acid substitutions in the Env and/or Nef were responsible for the observed CD4(+) T cell loss and neurological disease after inoculation with this molecular clone. (+info)
Detection of Herpes simplex virus DNA by real-time PCR.
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Molecular detection of herpes simplex virus (HSV) DNA is recognized as the reference standard assay method for the sensitive and specific diagnosis of central nervous system infections caused by HSV. In this study, a molecular assay based on real-time PCR on the LightCycler (LC) instrument was evaluated and compared with a home-brew molecular assay. The detection limit of the LC assay was determined with 10-fold dilutions of plasmid pS4 with the SalI restriction fragment of the DNA polymerase gene and with the First European Union Concerted Action HSV Proficiency Panel. A total of 59 cerebrospinal fluid (CSF) specimens were investigated for the comparative study. With plasmid pS4, the detection limit of the LC assay was found to be 10(4) copies per ml, i.e., 12.5 copies per run. When samples of the First European Union Concerted Action HSV Proficiency Panel were tested, 2x10(3) to 5x10(3) HSV type 1 genome equivalents (GE) per ml, i.e., 2.5 to 6.3 GE per run, could consistently be detected. There was a correlation between the LC assay and the home-brew assay in 55 of 59 specimens. In conclusion, the LC assay allows very rapid detection of HSV DNA in CSF. It was found to be laborsaving and showed sufficient sensitivity. (+info)
Neurological symptoms during primary human immunodeficiency virus (HIV) infection correlate with high levels of HIV RNA in cerebrospinal fluid.
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This analysis involves 22 patients with diagnosed symptomatic human immunodeficiency virus (HIV) infection. Neurologic symptoms were present in 11 patients, ranging from severe and persistent headache to clinical signs suggestive of meningitis. A strong correlation between neurological symptoms and cerebrospinal fluid (CSF) viral load was found. The mean CSF HIV ribonucleic acid (RNA) level was 4. 12 log for patients with neurological symptoms and 2.58 log for patients without neurological symptoms (P<.00001). Plasma viral load alone does not correlate or predict central nervous system (CNS) involvement. In our sample of patients, HIV RNA levels could be detected in most patients regardless of the presence of neurological symptoms. Moreover, early treatment including drugs with high levels of penetration in the CNS must be considered for patients with primary HIV infection. (+info)