A Genome-Wide Comparative Evolutionary Analysis of Herpes Simplex Virus Type 1 and Varicella Zoster Virus

Herpes simplex virus type 1 (HSV-1) and varicella zoster virus (VZV) are closely related viruses causing lifelong infections. They are typically associated with mucocutaneous or skin lesions, but may also cause severe neurological or ophthalmic diseases, possibly due to viral- and/or host-genetic factors. Although these viruses are well characterized, genome-wide evolutionary studies have hitherto only been presented for VZV. Here, we present a genome-wide study on HSV-1. We also compared the evolutionary characteristics of HSV-1 with those for VZV. We demonstrate that, in contrast to VZV for which only a few ancient recombination events have been suggested, all HSV-1 genomes contain mosaic patterns of segments with different evolutionary origins. Thus, recombination seems to occur extremely frequent for HSV-1. We conclude by proposing a timescale for HSV-1 evolution, and by discussing putative underlying mechanisms for why these otherwise biologically similar viruses have such striking evolutionary differences

Glycoprotein G of Herpes simplex virus type 2. Antigenicity and genetic variability

Herpes simplex virus type 2 (HSV-2) causes genital lesions, meningitis and occasionally, severe neonatal infections. Globally HSV-2 infection is one of the most common sexually transmitted diseases. As HSV-2 is frequently acquired and sustained asymptomatically, diagnosis during this phase of infection is essential both for the patient, and for preventive and epidemiological work. Type-specific serology is the diagnostic method of choice for this situation. The viral envelope glycoprotein G-2 (gG-2) exhibits two features unique in HSV-2 proteins. First, precursor gG-2 is cleaved to a secreted portion (sgG-2) and to a cell- and virion-associated, highly O-glycosylated mature portion (mgG-2). Second, mgG-2 is the only known HSV-2 glycoprotein which can be used as an antigen for type-discriminating serodiagnosis. In the present work we produced monoclonal antibodies (MAbs) against sgG-2 and mgG-2, and localized linear epitopes to the carboxy-terminal part of both proteins by a pepscan technique. Human anti-mgG-2 antibodies also recognized linear epitopes within mgG-2, localized to three regions of the protein of which one was found to be immunodominant. A synthetic peptide representing the immunodominant region was used successfully as antigen for detection of human anti-mgG-2 antibodies. The anti-mgG-2 MAbs as well as the human anti-mgG-2 antibodies identified type-specific epitopes and showed no cross-reactivity to HSV-1 antigen.One of the anti-mgG-2 MAbs was used for typing of 2,400 clinical HSV-2 isolates. The antibody was reactive with all except 13 of the HSV-2 isolates, and displayed no reactivity with clinical HSV-1 isolates indicating that the MAb was suitable for typing of HSV-2. Five of these 13 isolates presented a single frameshift mutation within the gG-2 gene with subsequent inactivation of the protein expression in four of the five isolates. The other eight isolates harbored a single missense mutation localized within the epitope which explained the loss of binding. These mutations did not diminish the antibody response to mgG-2 in the respective patients. Nucleotide sequencing of the gG-2 gene segment coding for mgG-2 among clinical HSV-2 isolates revealed that the epitope regions in mgG-2 were highly conserved.In conclusion, we have shown that mgG-2 is highly immunogenic for the human host and is a suitable antigen for type-discriminating serology, as well as for typing of HSV-2 isolates. The sgG-2 protein exhibited exclusively type-specific epitopes and may prove to be a novel antigen with serodiagnostic potential. The anti-gG-2 MAbs and HSV-2 mutants described here provide promising tools for further study on the function of the gG-2 proteins

Genotyping of Clinical Herpes Simplex Virus Type 1 Isolates by Use of Restriction Enzymes

Recently, three distinct genotypes of clinical herpes simplex virus type 1 (HSV-1) isolates were identified based on DNA sequence information and phylogenetic analysis of clinical isolates and laboratory strains. We utilized single-nucleotide polymorphism within the genes coding for glycoproteins G and I for rapid genotype classification by PCR and restriction enzyme cleavage. The method is suitable for high-scale genotyping of clinical HSV-1 isolates and for the detection of recombinants

Secreted Portion of Glycoprotein G of Herpes Simplex Virus Type 2 Is a Novel Antigen for Type-Discriminating Serology

The secreted portion of glycoprotein G (sgG-2) of herpes simplex virus type 2 (HSV-2) was evaluated as a novel antigen in an enzyme-linked immunosorbent assay (ELISA) format for detection of type-specific immunoglobulin G (IgG) antibodies in HSV-2-infected patients. The results were compared with those obtained by a commercially available assay, the HerpeSelect 2 ELISA (the FOCUS2 assay). Five different panels of sera were analyzed: panel A consisted of 109 serum samples from patients with a culture-proven HSV-1 infection that were Western blotting (WB) negative for HSV-2; panel B consisted of 106 serum samples from patients with a culture-proven recurrent HSV-2 infection that were WB positive for HSV-2; panel C consisted of 100 serum samples with no detectable IgG antibodies against HSV-1 and HSV-2; panel D consisted of 70 HSV-2 negative “tricky” serum samples containing antinuclear IgG antibodies or IgM antibodies against other viruses or bacteria; and panel E consisted of consecutive serum samples from 21 patients presenting with a first episode of HSV-2-induced lesions. When sera in panels A to C were analyzed, the sgG-2 ELISA and the FOCUS2 assay both showed sensitivities and specificities of ≥98%. In total, among the samples in panel D, 13 serum samples (19%) were false positive by the FOCUS2 assay and 1 serum sample (1.4%) was false positive by the sgG-2 ELISA. When the sera in panel E were analyzed, the sgG-2 ELISA detected seroconversion somewhat later than WB or the FOCUS2 assay did. We conclude that sgG-2 induces an HSV-2 type-specific antibody response and can be used for type-discriminating serology

Dichotomy of Glycoprotein G Gene in Herpes Simplex Virus Type 1 Isolates

Herpes simplex virus type 1 (HSV-1) encodes 11 envelope glycoproteins, of which glycoprotein G-1 (gG-1) induces a type-specific antibody response. Variability of the gG-1 gene among wild-type strains may be a factor of importance for a reliable serodiagnosis and typing of HSV-1 isolates. Here, we used a gG-1 type-specific monoclonal antibody (MAb) to screen for mutations in the immunodominant region of this protein in 108 clinical HSV-1 isolates. Of these, 42 isolates showed no reactivity to the anti-gG-1 MAb. One hundred five strains were further examined by DNA sequencing of the middle part of the gG-1 gene, encompassing 106 amino acids including the immunodominant region and epitope of the anti-gG-1 MAb. By phylogenetic comparisons based on the sequence data, we observed two (main) genetic variants of the gG-1 gene among the clinical isolates corresponding to reactivity or nonreactivity to the anti-gG-1 MAb. Furthermore, four strains appeared to be recombinants of the two gG-1 variants. In addition, one strain displayed a gG-1-negative phenotype due to a frameshift mutation, in the form of insertion of a cytosine nucleotide. When immunoglobulin G reactivity to HSV-1 in sera from patients infected with either of the two variants was investigated, no significant differences were found between the two groups, either in a type-common enzyme-linked immunosorbent assay (ELISA) or in a type-specific gG-1 antigen-based ELISA. Despite the here-documented existence of two variants of the gG-1 gene affecting the immunodominant region of the protein, other circumstances, such as early phase of infection, might be sought for explaining the seronegativity to gG-1 commonly found in a proportion of the HSV-1-infected patients