Anthony C. Minson

The order Herpesvirales

The taxonomy of herpesviruses has been updated by the International Committee on Taxonomy of Viruses (ICTV). The former family Herpesviridae has been split into three families, which have been incorporated into the new order Herpesvirales. The revised family Herpesviridae retains the mammal, bird and reptile viruses, the new family Alloherpesviridae incorporates the fish and frog viruses, and the new family Malacoherpesviridae contains a bivalve virus. Three new genera have been created in the family Herpesviridae, namely Proboscivirus in the subfamily Betaherpesvirinae and Macavirus and Percavirus in the subfamily Gammaherpesvirinae. These genera have been formed by the transfer of species from established genera and the erection of new species, and other new species have been added to some of the established genera. In addition, the names of some nonhuman primate virus species have been changed. The family Alloherpesviridae has been populated by transfer of the genus Ictalurivirus and addition of the new species Cyprinid herpesvirus 3. The family Malacoherpesviridae incorporates the new genus Ostreavirus containing the new species Ostreid herpesvirus 1.

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.

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.

Excretion of Non-infectious Virus Particles Lacking Glycoprotein H by a Temperature-sensitive Mutant of Herpes Simplex Virus Type 1: Evidence that gH Is Essential for Virion Infectivity

A temperature-sensitive mutant of herpes simplex virus type 1, tsQ26, was shown to contain an amino acid substitution in glycoprotein H (gH). The mutant entered cells efficiently at the non-permissive temperature and replicated to give nearly normal yields of intracellular infectivity. The intracellular virions contained, predominantly, an immature form of gH and no gH was found on the surface of infected cells. Excreted virions were devoid of gH and were not infectious. Virions excreted at the permissive temperature were infectious and contained gH and no loss of gH resulted from incubation of these virions at the non-permissive temperature. The temperature-sensitive phenotype apparently results from the loss of gH from virions during their transport to the cell surface, and since loss of gH is accompanied by loss of infectivity we conclude that gH is an essential component of the infectious virion.

Characterisation and physical mapping of an HSV-1 glycoprotein of approximately 115 × 103 molecular weight

A type-specific monoclonal antibody that efficiently neutralises HSV-1 immunoprecipitated a glycoprotein of slightly greater electrophoretic mobility than gB from HSV-1 infected cells. Pulse and pulse chase experiments indicate that this glycoprotein is distinct from HSV-1 glycoproteins gB, gC, gD, and gE. This was confirmed by the reactions of LP11 with a series of intertypic recombinants the results of which indicate that the LP11 target gene is located close to the HSV-1 thymidine kinase gene between map positions 0.28 and 0.31. In accordance with the presently agreed convention this glycoprotein should be designated gH-1, and it may correspond to the 110K glycoprotein described by S. D. Showalter, M. Zweig, and B. Hampar (1981), Infect. Immun. 34, 684-692. Antibody LP11 inhibits plaque formation when added to cell monolayers after infection suggesting that gH-1 may play a role in cell-to-cell spread of infectious virus.

Analysis of the L1 Gene Product of Human Papillomavirus Type 16 by Expression in a Vaccinia Virus Recombinant

The L1 open reading frame of human papillomavirus type 16 (HPV16) has been expressed in vaccinia virus under the control of both the 7.5K early and late promoter, and the 4b major late promoter. Antibodies to a beta-galactosidase fusion protein containing a C-terminal portion of the HPV16 L1 gene product were used to compare the levels of L1 expression in the two recombinants, and showed that greater levels of expression were obtained when the gene was placed under the control of the 4b late promoter. Immunofluorescence studies revealed a nuclear location of the L1 gene product when expressed in vaccinia virus. Antibodies to the beta-galactosidase fusion protein detected a major polypeptide species of 57K and a minor species of 64K in Western blots of recombinant-infected cell lysates. The 64K species was not detected when cells were infected in the presence of tunicamycin, indicating that the primary translation product of the HPV16 L1 open reading frame is modified by N-linked glycosylation when expressed in vaccinia virus. Whereas antibodies to HPV16 L1 fusion proteins and to a peptide containing amino acids from the C terminus of HPV16 L1 reacted well in Western blots with the HPV16 L1 target expressed in vaccinia virus, no reactivity was observed with antibodies to bovine papillomavirus type 1 particles or to a HPV6b fusion protein.

Immunogenicity of herpes simplex virus type 1 glycoproteins expressed in vaccinia virus recombinants

Vaccinia virus recombinants expressing glycoproteins B (vgB11), D (VgD52), E (gE/7.5 and gE/4B), G (gG-vac), H (gH-vac), and I (gI-vac) of HSV-1 were used to compare the protective response to these individual glycoproteins in the mouse. Glycoprotein D induced the best neutralizing antibody titers and the most increased rates of HSV clearance from the ear as well as good protection from the establishment of latent HSV infections in the sensory ganglia. Glycoprotein B also induced good neutralizing antibody titers and as great a protection from the establishment of latency as gD although the rate of virus clearance from the ear was not as great as after immunization with gD. Glycoprotein E induced weak neutralizing antibody but gG, gH, and gI did not show a neutralizing antibody response. At higher challenge doses of virus (10(6) PFU HSV-1 in the ear), gE induced a protective response by increasing the rate of virus clearance and reducing the acute infection of ganglia as compared to negative control immunized mice. However there was no protection from the establishment of latent infections after immunization with gE. No protective response was seen to gG, gH, or gl.

Induction of protective immunity with antibody to herpes simplex virus type 1 glycoprotein H (gH) and analysis of the immune response to gH expressed in recombinant vaccinia virus

Passive administration of neutralizing monoclonal antibody (MAb) to glycoprotein H (gH) of herpes simplex virus type 1 (HSV-1) was found to protect mice from an HSV-1 strain SC16 challenge infection. To investigate further the protective potential of gH, recombinant vaccinia viruses were constructed which expressed the HSV-1 gH open reading frame under the control of the vaccinia virus 7.5K early/late promoter or the 4b late promoter. Immunization with recombinant viruses, however, did not induce the production of neutralizing antisera and the mice were not protected from zosteriform spread or the establishment of latent infection following viral challenge. The gH produced by the recombinant vaccinia viruses differed in electrophoretic mobility and antigenicity from authentic HSV-1 gH. Only one of three neutralizing MAbs specific for conformational epitopes on gH was able to immunoprecipitate gH synthesized in recombinant vaccinia virus-infected cells. In addition cell surface expression of gH was not detected in cells infected with the recombinant vaccinia viruses.

The herpes simplex virus type 1 Fc receptor discriminates between IgG1 allotypes.

Herpes simplex virus type 1 (HSV‐1) expresses a complex of two virally encoded glycoproteins, gE and gI, which is capable of binding nonimmune human IgG. The gE‐gI complex has thus become known as an Fc receptor (FcR), which reportedly binds human IgG subclasses in the order IgG4 > IgG1 ≥ IgG2 and does not bind IgG3 from many individuals. There is, however, allelic variation in the genes encoding the human IgG1 heavy chain constant region and this gives rise to allotypes of IgG1. Using recombinant monoclonal IgG molecules of known isotype and mutants thereof we have unexpectedly discovered that the HSV‐1 FcR discriminates between IgG1 allotypes. This is evidence of functional differences between IgG1 allotypes that may account for their distribution in populations. Furthermore, these findings suggest HSV‐1 FcR binding sites on the IgG molecule some distance from the proposed binding site in the CH2‐CH3 domain interface.

Characterization of herpes simplex virus type 1 recombinants with mutations in the cytoplasmic tail of glycoprotein H

Herpes simplex virus (HSV) type 1 glycoprotein H is essential for fusion of virus envelopes with cellular membranes and for the fusion of an infected cell membrane with an uninfected neighbour. Previous studies have pointed to a requirement for certain amino acid residues of the cytoplasmic tail of gH in these processes. Results from transient transfection experiments suggested that the serine-valine-proline (SVP) motif in the cytoplasmic tail may be important for gH-mediated fusion. HSV recombinants expressing gH molecules with mutations in the cytoplasmic tail were constructed and analysed in terms of their abilities to fuse cellular membranes and to function in virus entry. Viruses containing a deletion of the SVP motif, or in which the valine residue of this triplet was replaced by alanine, entered cells less efficiently than wild-type virus and were unable to induce syncytium formation on Vero cells.