J P Vergnes

Bio: J P Vergnes is an academic researcher from University of Pittsburgh. The author has contributed to research in topics: Promoter & Gene. The author has an hindex of 1, co-authored 1 publications receiving 44 citations.

Transcriptional mapping of the varicella-zoster virus regulatory genes encoding open reading frames 4 and 63

Abstract: Four of the 68 varicella-zoster virus (VZV) unique open reading frames (ORFs), i.e., ORFs 4, 61, 62, and 63, encode proteins that influence viral transcription and are considered to be positional homologs of herpes simplex virus type 1 (HSV-1) immediate-early (IE) proteins. In order to identify the elements that regulate transcription of VZV ORFs 4 and 63, the encoded mRNAs were mapped in detail. For ORF 4, a major 1.8-kb and a minor 3.0-kb polyadenylated [poly(A)+] RNA were identified, whereas ORF 63-specific probes recognized 1.3- and 1.9-kb poly(A)+ RNAs. Probes specific for sequences adjacent to the ORFs and mapping of the RNA 3' ends indicated that the ORF 4 RNAs were 3' coterminal, whereas the RNAs for ORF 63 represented two different termination sites. S1 nuclease mapping and primer extension analyses indicated a single transcription initiation site for ORF 4 at 38 bp upstream of the ORF start codon. For ORF 63, multiple transcriptional start sites at 87 to 95, 151 to 153, and (tentatively) 238 to 243 bp upstream of the ORF start codon were identified. TATA box motifs at good positional locations were found upstream of all mapped transcription initiation sites. However, no sequences resembling the TAATGARAT motif, which confers IE regulation upon HSV-1 IE genes, were found. The finding of the absence of this motif was supported through analyses of the regulatory sequences of ORFs 4 and 63 in transient transfection assays alongside those of ORFs 61 and 62. Sequences representing the promoters for ORFs 4, 61, and 63 were all stimulated by VZV infection but failed to be stimulated by coexpression with the HSV-1 transactivator Vmw65. In contrast, the promoter for ORF 62, which contains TAATGARAT motifs, was activated by VZV infection and coexpression with Vmw65. These results extend the transcriptional knowledge for VZV and suggest that ORFs 4 and 63 contain regulatory signals different from those of the ORF 62 and HSV-1 IE genes.

Varicella-zoster virus.

Abstract: Varicella-zoster virus (VZV) is a ubiquitous human alphaherpesvirus that causes varicella (chicken pox) and herpes zoster (shingles). Varicella is a common childhood illness, characterized by fever, viremia, and scattered vesicular lesions of the skin. As is characteristic of the alphaherpesviruses, VZV establishes latency in cells of the dorsal root ganglia. Herpes zoster, caused by VZV reactivation, is a localized, painful, vesicular rash involving one or adjacent dermatomes. The incidence of herpes zoster increases with age or immunosuppression. The VZV virion consists of a nucleocapsid surrounding a core that contains the linear, double-stranded DNA genome; a protein tegument separates the capsid from the lipid envelope, which incorporates the major viral glycoproteins. VZV is found in a worldwide geographic distribution but is more prevalent in temperate climates. Primary VZV infection elicits immunoglobulin G (IgG), IgM, and IgA antibodies, which bind to many classes of viral proteins. Virus-specific cellular immunity is critical for controlling viral replication in healthy and immunocompromised patients with primary or recurrent VZV infections. Rapid laboratory confirmation of the diagnosis of varicella or herpes zoster, which can be accomplished by detecting viral proteins or DNA, is important to determine the need for antiviral therapy. Acyclovir is licensed for treatment of varicella and herpes zoster, and acyclovir, valacyclovir, and famciclovir are approved for herpes zoster. Passive antibody prophylaxis with varicella-zoster immune globulin is indicated for susceptible high-risk patients exposed to varicella. A live attenuated varicella vaccine (Oka/Merck strain) is now recommended for routine childhood immunization.

Attenuation of the Vaccine Oka Strain of Varicella-Zoster Virus and Role of Glycoprotein C in Alphaherpesvirus Virulence Demonstrated in the SCID-hu Mouse

Abstract: The SCID-hu mouse implanted with human fetal tissue is a novel model for investigating human viral pathogenesis. Infection of human skin implants was used to investigate the basis for the clinical attenuation of the varicella-zoster virus (VZV) strain, V-Oka, from which the newly licensed vaccine is made. The pathogenicity of V-Oka was compared with that of its parent, P-Oka, another low-passage clinical isolate, strain Schenke (VZV-S), and VZV-Ellen, a standard laboratory strain. The role of glycoprotein C (gC) in infectivity for human skin was assessed by using gC-negative mutants of V-Oka and VZV-Ellen. Whereas all of these VZV strains replicated well in tissue culture, only low-passage clinical isolates were fully virulent in skin, as shown by infectious virus yields and analysis of implant tissues for VZV DNA and viral protein synthesis. The infectivity of V-Oka in skin was impaired compared to that of P-Oka, providing the first evidence of a virologic basis for the clinical attenuation of V-Oka. The infectivity of V-Oka was further diminished in the absence of gC expression. All strains except gC-Ellen retained some capacity to replicate in human skin, but cell-free virus was recovered only from implants infected with P-Oka or VZV-S. Although VZV is closely related to herpes simplex virus type 1 (HSV-1) genetically, experiments in the SCID-hu model revealed differences in tropism for human cells that correlated with differences in VZV and HSV-1 disease. VZV caused extensive infection of epidermal and dermal skin cells, while HSV-1 produced small, superficial lesions restricted to the epidermis. As in VZV, gC expression was a determinant for viral replication in skin. VZV infects human CD4+ and CD8+ T cells in thymus/liver implants, but HSV-1 was detected only in epithelial cells, with no evidence of lymphotropism. These SCID-hu mouse experiments show that the clinical attenuation of the varicella vaccine can be attributed to decreased replication of V-Oka in skin and that tissue culture passage alone reduces the ability of VZV to infect human skin in vivo. Furthermore, gC, which is dispensable for replication in tissue culture, plays a critical role in the virulence of the human alphaherpesviruses VZV and HSV-1 for human skin.

Live attenuated varicella vaccine

Abstract: Varicella-zoster virus (VZV) is a ubiquitous human pathogen that causes varicella, commonly called chicken pox; establishes latency; and reactivates as herpes zoster, referred to as shingles. A live attenuated varicella vaccine, derived from the Oka strain of VZV has clinical efficacy for the prevention of varicella. The vaccine induces persistent immunity to VZV in healthy children and adults. Immunization against VZV also has the potential to lower the risk of reactivation of latent virus. The varicella vaccine may eventually reduce or eliminate herpes zoster, which is a serious problem for elderly and immunocompromised individuals.

The transcriptional regulatory proteins encoded by varicella-zoster virus open reading frames (ORFs) 4 and 63, but not ORF 61, are associated with purified virus particles.

Abstract: Of the five varicella-zoster virus (VZV) open reading frames (ORFs) known to encode proteins which influence viral transcriptional events, two (ORFs 10 and 62) encode proteins associated with the tegument of virus particles, where they may function during the immediate-early events of infection. In this study, antibodies which recognize the products of the three additional VZV ORFs, ORFs 4, 61, and 63, were made and used to characterize their association with virus particles. ORF 4 encoded a 52-kDa polypeptide, and antibodies to ORF 63 reacted with polypeptides of 47 and 28 kDa. Antibodies to ORF 61 recognized heterogeneous polypeptides of 62 to 66 kDa in cells infected with a vaccinia virus recombinant expressing ORF 61 and in VZV-infected melanoma cells but reacted very weakly with polypeptides of VZV-infected human foreskin fibroblasts, suggesting that cell-specific factors were involved in ORF 61 protein accumulation. Analysis of virus particles purified from melanoma cells indicated that a 52-kDa polypeptide from ORF 4 and the 47-kDa polypeptide from ORF 63, but not any from ORF 61, were associated with virus particles. The virion proteins were likely components of the tegument, as they were not solubilized by treatment of virus with mild detergents and were completely resistant to trypsin digestion unless prior envelope solubilization was performed. The products of ORFs 4 and 63 were not found in purified VZV nucleocapsids. These results suggest that forms of the ORF 4- and ORF 63-encoded transcriptional regulatory proteins are also structural and may also have roles in the immediate-early events of infection.

Use of polymerase chain amplification reaction for the detection of adenoviruses in ocular swab specimens

Abstract: Purpose To evaluate the application of polymerase chain reaction (PCR) methodology as a potential diagnostic tool for the detection of adenovirus DNA in ocular swab samples. Methods Oligonucleotides derived from the adenovirus hexon gene were used to amplify a 306-base pair (bp) product by PCR. Radiolabeled oligonucleotides derived from sequences within the amplified product were used as specific probes. Specificity was determined against DNA of 13 adenovirus serotypes (types 1 to 11, inclusive, and types 19 and 37) and from nonadenoviral DNAs. Limits of detection were determined by PCR amplification of known amounts of purified adenovirus serotype 2 DNA. The assay was tested on 107 ocular swab samples and correlated to results obtained from tissue culture and a commercial immunoassay (Adenoclone). Results The 306-bp PCR product was amplified from all adenovirus serotypes tested, but not from negative control DNAs. As little as 15 fg of adenovirus type 2 DNA could be detected by PCR and ethidium bromide stain. Using a simplified sample preparation procedure, 46 of 58 adenovirus culture-positive but Adenoclone-negative swabs were positive by PCR (79% sensitivity). All (11 of 11) Adenoclone-positive clinical eye swabs tested were positive by PCR (100% sensitivity). Only 1 of 38 nonadenoviral ocular swab samples was positive by PCR (97% specificity). Conclusions PCR appeared to be highly suitable for the diagnosis of adenovirus in ocular swabs, offering important improvements in speed over tissue culture isolation and in sensitivity over immunoassay.