PubMed Health⌕ Search

Biomedical subjects

S Efstathiou

Publications and source records attributed to S Efstathiou.

52 records · Page 3Linked to original sources
PubMed1994

Vaccine potential of a herpes simplex virus type 1 mutant with an essential glycoprotein deleted.

Several approaches to the production of vaccines to human herpesviruses have been proposed. Subunit vaccines, subunits delivered by live vectors, and rationally attenuated vaccines have all been shown to be efficacious in animal models but suffer from uncertainties as to the roles of individual genes involved in pathogenesis and the most relevant components of the immune response required for protection in humans and the target antigens involved. With these problems in mind, we examined the vaccine potential of a fully disabled herpes simplex virus type 1 mutant that is capable of only a single round of replication, since a virus of this type should induce the full spectrum of immune responses but has no pathogenic potential. A virus has been described which lacks essential glycoprotein H (gH) and can be propagated in a cell line which supplies gH in trans (A. Forrester, H. Farrell, G. Wilkinson, J. Kaye, N. Davis-Poynter, and T. Minson, J. Virol. 66:341-348, 1992). Infection of normal cells with this mutant is indistinguishable from a wild-type infection, except that the resulting progeny are gH negative and noninfectious: the virus is self-limiting. Infection of mice by the ear pinna route was similarly self-limiting in that input infectivity decreased rapidly at the inoculation site and no infectivity was detected in sensory ganglia. Animals given a wide range of doses of the gH-negative mutant produced both humoral and T-cell responses to herpes simplex virus type 1 and proved solidly resistant to challenge with a high dose of wild-type virus. The gH-negative mutant is presumably capable of establishing a latent infection, but since no infectious virus was detected in numerous attempts to reactivate the mutant, the risk of a pathogenic outcome is minimal.

Animals↗
PubMed1993

Interactions of murine gammaherpesvirus 68 with B and T cell lines.

Murine gammaherpesvirus is a natural pathogen of wild rodents. We have established that in vivo the virus persists in B lymphocytes in a latent form and therefore has similar biological properties to Epstein-Barr virus and related gamma-I-herpesviruses. In this report we have established a persistent infection in mouse myeloma (B) cells (NSO cell line), but not in mouse thymoma (T) cells (BW 5147 cell line). The virus persists indefinitely in myeloma cells, without any apparent cytopathic effect, but with the production of infectious virus. We demonstrate that ACV abolishes the productive infection, but large numbers of cells harbor the virus in a latent form, as determined by an infectious center assay. Analysis of the viral DNA has shown that during a persistent infection linear virus genomes predominated, with low levels of circular DNA also present. Treatment of cells with ACV results in a significant reduction of linear genomes, but has no effect on the level of circular DNA molecules. These data provide further evidence to support our earlier observations on B cells as the site of latency and provides an in vitro model with which to study the molecular basis of MHV-68 latency/persistence.

Animals↗
PubMed1993

Intranuclear foci containing low abundance herpes simplex virus latency-associated transcripts visualized by non-isotopic in situ hybridization.

During latent infection of neurons with herpes simplex virus type 1 (HSV-1), several RNA transcripts of varying abundance arise from a single locus within the virus repeats. The functions of latency-associated transcripts (LATs) are unknown and the relationship between the various RNA species requires further clarification. Reported here is a novel approach to the study of HSV transcripts during latency, based on the increasing realization that cellular and viral RNAs are synthesized and processed by macromolecular complexes that occupy discrete compartments within the nucleoplasm of a cell. High resolution non-isotopic in situ hybridization was used to study the intranuclear topology of HSV-1 LATs in primary sensory neurons of latently infected mice and humans. Low abundance (minor) LATs were localized to sharply defined intranuclear foci of 1 to 3 microns in diameter. On average, there were 2.6 to 2.8 foci/LAT+ neuronal profile (5 microns), representing 13 to 14 foci/cell. In contrast to the focal deployment of minor LATs, the more abundant latency-associated RNAs were distributed diffusely throughout the nucleoplasms of latency infected neurons, with prominent sparing of nucleolar regions. These data establish a foundation for studying the synthesis, processing and transport of LATs in vivo. It should now be possible to investigate the nature of those cellular products which associate with HSV-1 encoded LATs in vivo and thereby determine whether minor LATs are associated with previously characterized macromolecular complexes, such as those responsible for processing of pre-messenger RNA.

Animals↗
PubMed1992

Murine gammaherpesvirus 68 establishes a latent infection in mouse B lymphocytes in vivo.

Murine gammaherpesvirus 68 (MHV-68) is able to persist in spleen cells of infected mice. To determine the cell type harbouring persistent virus, spleen cells from infected animals were separated into immunoglobulin (Ig)-positive (B cell-enriched), Ig-negative (T cell-enriched) and plastic-adherent (macrophage-enriched) fractions. These cells were co-cultivated with permissive BHK-21 cells in an infectious centre assay. The consistent recovery and enrichment of infectious centres in the Ig-positive fraction clearly demonstrates that B cells are a major site of virus persistence/latency. This observation indicates that MHV-68 is biologically similar to Epstein-Barr virus and other members of the B cell lymphotropic gammaherpesvirus 1 subgroup.

Acute Disease↗
PubMed1992

Two patterns of persistence of herpes simplex virus DNA sequences in the nervous systems of latently infected mice.

The number of herpes simplex virus (HSV) genome equivalents recovered from latently infected mouse spinal ganglia was compared with the proportion of neurons containing latency-associated transcripts (LATs). Two distinct patterns of HSV persistence were observed, depending on the anatomical location of ganglia with respect to the site of cutaneous inoculation. The location of the bulk of latent viral DNA did not correspond with the highest prevalence of LAT+ neurons. Viral DNA was most abundant in spinal ganglia directly innervating the inoculation site and the amount recovered, which was similar to that found previously in human trigeminal ganglia, suggested that LAT+ neurons each contain hundreds of copies of HSV DNA. In stark contrast, although LAT+ neurons were most abundant in neighbouring ganglia, viral DNA was scarce (approx. 20 copies/LAT+ cell). These data indicate that amplification of HSV DNA sequences is greatest in ganglia previously shown to be associated with viral antigen expression during the productive phase of primary infection.

Animals↗
PubMed1992

Identification of homologues to the human cytomegalovirus US22 gene family in human herpesvirus 6.

The sequence of 10079 bp corresponding to the overlapping SalI H and SmaI G restriction fragments of the genome of human herpesvirus 6 (HHV-6) strain U1102 was determined. The sequence contains six complete open reading frames (ORFs) and two incomplete ORFs located at the 5' and 3' ends of the SalI H and SmaI G fragments respectively. Seven of these ORFs have recognizable homologues only in the beta-herpesvirus human cytomegalovirus (HCMV), no obvious counterparts being detectable in the genomes of the human alpha-herpesviruses, varicella-zoster virus and herpes simplex virus type 1 or the gamma-herpesvirus Epstein-Barr virus. The DNA sequenced is located proximal to the left repeat of the HHV-6 genome outside the well recognized region encompassing conserved herpesvirus gene blocks. A close colinear relationship is evident between the HHV-6 ORFs identified in this study and their counterparts in HCMV, ORFs UL23, UL24 and UL27 to UL31. Four of the HHV-6 ORFs, SHL1, SHL2, SFL1 and SSL2, are related to members of the HCMV US22 family of proteins, which are themselves tandemly arranged and located predominantly within the unique short and the left end of the unique long region of the prototype HCMV strain AD169 genome. Two adjacent HHV-6 ORFs, SSL1 and SHL3, are related to HCMV UL27. The identification of this gene set in addition to the HHV-6 ORFs with amino acid sequence similarity to the HCMV US22 family indicates a particularly close relationship between these two human herpesviruses, and suggests that the clustering of these related tandemly arranged genes may be a general feature of beta-herpesvirus-type genomes.

Amino Acid Sequence↗
PubMed1992

Virological and pathological features of mice infected with murine gamma-herpesvirus 68.

The primary infection of BALB/c mice with murine herpesvirus 68 (MHV-68) was investigated. When the virus was introduced intranasally, the lung was the main tissue infected, the virus being associated with alveolar epithelium and mononuclear cells. A productive infection lasted for 10 days, after which viral DNA could be detected by in situ hybridization up to 30 days after infection. At that time lymphoproliferative accumulations were also observed in the lung, with formation of germinal centres. Virus could also be recovered from the heart, kidney, adrenal gland and spleen during the primary infection. In addition, the spleen appeared to be the major site of virus persistence, with latently infected cells detected up to 90 days post-infection. During the primary infection, there was atrophy of the thymus and spleen of clinically sick animals. In contrast, lymphoproliferative responses, typified by splenomegaly, were frequently seen in asymptomatic animals. The pattern of infection observed in MHV-68-infected mice is similar to that seen in infectious mononucleosis of man following Epstein-Barr virus infection. The model described in this paper may prove to be useful in studying natural gamma-herpesvirus infections of man and domestic animals.

Acute Disease↗
PubMed1991

Acquisition of the human adeno-associated virus type-2 rep gene by human herpesvirus type-6.

Human herpesvirus type-6 (HHV-6) is a recently isolated herpesvirus which is highly prevalent in adult populations around the world. HHV-6 was first isolated from the peripheral blood of six individuals with lymphoproliferative disorders, two of whom were also infected with human immunodeficiency virus. HHV-6, in common with other herpesviruses, transactivates the HIV long terminal repeat linked to reporter genes and has in addition been shown to accelerate HIV gene expression and CD4 cell death in cultures co-infected with both viruses. The virus is tropic for CD4+ lymphocytes and persists in the peripheral blood of most seropositive individuals. We have now identified a gene in HHV-6 encoding a 490-amino-acid polypeptide homologous to the human adeno-associated virus type-2 (AAV-2) rep gene. This gene has an essential role in AAV-2 DNA replication, can trans-regulate homologous and heterologous gene expression, and inhibits cellular transformation. The acquisition of rep by HHV-6 could be due to natural transfer of genetic information between DNA viruses of eukaryotes and is likely to have important consequences for the life-cycle of HHV-6 and for the host CD4 cell.

Amino Acid Sequence↗
PubMed1991

Investigation of herpes simplex virus type 1 genes encoding multiply inserted membrane proteins.

The herpes simplex virus type 1 genome contains four open reading frames (ORFs) which are predicted to encode hydrophobic proteins with the potential to cross a membrane several times. The products of these genes (genes UL10, UL20, UL43 and UL53) have not previously been identified. To investigate the role of these proteins in the virus life cycle, we attempted to inactivate the genes individually by inserting the lacZ gene from Escherichia coli within the ORFs. Using this approach we have isolated insertion mutants for UL10 and UL43, as well as a deletion mutant lacking the majority of the UL43 ORF. The growth of the UL10-lacZ virus was slightly impaired in tissue culture compared to that of the wild-type virus parent, whereas the growth of the UL43 mutants was indistinguishable from that of wild-type virus. Furthermore, deletion of the majority of the UL43 ORF did not impair the ability of the virus to replicate in vivo at the periphery, or to spread to and replicate within the nervous system, in a mouse ear model. Repeated attempts to isolate lacZ insertion mutants for UL20 and UL53 were unsuccessful, suggesting that these genes may be essential for virus growth, at least in tissue culture. Using antipeptide sera, the products of genes UL10 and UL20 have been detected.

Animals↗
PubMed1990

Cloning and molecular characterization of the murine herpesvirus 68 genome.

Murine herpesvirus 68 (MHV-68) is a naturally occurring herpesvirus of small free-living rodents. In order to facilitate the molecular characterization of the virus genome, a library of cloned restriction fragments has been produced and restriction enzyme cleavage maps deduced for the enzymes BamHI, EcoRI and HindIII. The MHV-68 genome comprises a region of unique DNA of approximately 118 kbp which is flanked by variable numbers of a 1.23 kb repeat unit. The organization of the MHV-68 genome is, therefore, most similar to that of the lymphotropic gamma 2 group of herpesviruses which include herpesvirus saimiri and herpesvirus ateles.

Animals↗
PubMed1990

Murine herpesvirus 68 is genetically related to the gammaherpesviruses Epstein-Barr virus and herpesvirus saimiri.

Short nucleotide sequence analysis of seven restriction fragments of murine herpesvirus 68 (MHV-68) DNA has been undertaken and used to determine the overall genome organization and relatedness of this virus to other well characterized representatives of the alpha-, beta- and gammaherpesvirus subgroups. Nine genes have been identified which encode amino acid sequences with greater similarity to proteins of the gammaherpesvirus Epstein-Barr virus (EBV) than to the homologous products of the alphaherpesviruses varicella-zoster virus and herpes simples virus type 1 or the betaherpesvirus human cytomegalovirus. In addition, the genome organization of MHV-68 is shown to have an overall collinearity with that of the gammaherpesviruses EBV and herpesvirus saimiri. In common with these viruses, dinucleotide frequency analysis of MHV-68 coding sequences reveals a marked reduction in CpG dinucleotide frequency thus implicating a dividing cell population as the site of latency in vivo.

Amino Acid Sequence↗
PubMed1989

Brief report: primary human herpesvirus 6 infection in a patient following liver transplantation from a seropositive donor.

A virus, isolated from peripheral blood lymphocytes of a patient 23 days after liver transplantation, was identified by negative contrast and thin-section electron microscopy, DNA hybridisation, and immunofluorescence as human herpesvirus 6 (HHV-6). The patient was seronegative for antibodies to HHV-6 before receiving the liver from a seropositive donor. Seroconversion for IgM and IgG antibodies occurred at the same time as the viraemia, indicating a primary response to HHV-6 infection that was probably acquired from the donor liver. The relationship of the infection to the patient's subsequent disease episodes is discussed.

Adult↗
PubMed1989

The role of herpes simplex virus type 1 thymidine kinase in pathogenesis.

A genetically engineered herpes simplex virus type 1 (HSV-1)thymidine kinase (TK) deletion mutant has been constructed and used to investigate the role of this gene in pathogenesis. Inoculation of mice with the HSV TK deletion mutant resulted in the establishment of latent ganglionic infection as demonstrated by superinfection of explanted ganglia with wild-type (wt) virus but not by routine explant culture suggesting that the virus-encoded TK is not essential for the establishment of latent infection but may be necessary for either reactivation or virus replication following reactivation. In addition, Southern blot hybridization has been used to demonstrate in vivo complementation of this mutant by wt virus in both peripheral and central nervous system tissues of mice during acute infection and to show that such complementation can result in the establishment and reactivation of latent TK- infection.

Acute Disease↗
PubMed1986

Detection of herpes simplex virus-specific DNA sequences in latently infected mice and in humans.

Herpes simplex virus-specific DNA sequences have been detected by Southern hybridization analysis in both central and peripheral nervous system tissues of latently infected mice. We have detected virus-specific sequences corresponding to the junction fragment but not the genomic termini, an observation first made by Rock and Fraser (Nature [London] 302:523-525, 1983). This "endless" herpes simplex virus DNA is both qualitatively and quantitatively stable in mouse neural tissue analyzed over a 4-month period. In addition, examination of DNA extracted from human trigeminal ganglia has shown herpes simplex virus DNA to be present in an "endless" form similar to that found in the mouse model system. Further restriction enzyme analysis of latently infected mouse brainstem and human trigeminal DNA has shown that this "endless" herpes simplex virus DNA is present in all four isomeric configurations.

Adult↗
PubMed1984

A quantitative study of the effects of several nucleoside analogues on established herpes encephalitis in mice.

Mice with established herpes encephalitis were used to compare the effects of chemotherapy using three different nucleoside analogues. Encephalitis was produced by intranasal inoculation of a type 1 strain of herpes simplex virus. Without chemotherapy all mice died within 5 to 7 days of inoculation. Oral acyclovir (ACV) was a successful preventative measure if commenced within 2 days of inoculation but much less effective if the onset of treatment was further delayed. From the third day, when central nervous system infection had definitely become established, ACV only reduced mortality if given intraperitoneally (i.p.) at regular 6-hourly intervals. Comparison with bromovinyldeoxyuridine (BVdU) and the new nucleoside analogue dihydroxypropoxymethylguanine (DHPG) using the same 6-hourly i.p. regimen revealed that BVdU was poorly effective, despite better activity in vitro, whereas DHPG was the most successful. Virus was rapidly eradicated from all parts of the brain by DHPG therapy, and by day 10, no infectious virus remained in the brains of treated mice, no virus antigens were observed and no trace of virus DNA could be detected in neural tissues by Southern blotting.

Acyclovir↗