To study the relationships between herpesviruses recently isolated from different pinniped species, antigenic and genetic analyses were performed. First, herpesviruses isolated from North American harbour seals (Phoca vitulina), a Californian sea lion (Zalophus californianus) and a European grey seal (Halichoerus grypus) were examined in an enzyme immunoassay (EIA) with a panel of monoclonal antibodies which had previously been shown to allow typing of herpesviruses from European harbour seals into two distinct virus types: phocid herpesvirus type-1 and type-2 (PhHV-1 and PhHV-2). The EIA data showed that all but one of the isolates from seals ranging in United States coastal waters were PhHV-2-1ike while the European grey seal herpesvirus was PhHV-l-like. Genetic characterization was facilitated by PCR analysis using primers based on conserved regions of the glycoprotein B and D (gB and gD) genes of the antigenically closely related canid (CHV) and felid (FHV) herpesviruses. Specific amplified products were obtained with five isolates antigenically characterized as PhHV-l-like but not with five PhHV-2-like isolates. Sequence analysis of the PCR products confirmed greatest similarity to members of the genus Varicellovirus of the Alphaherpesvirinae and in particular to CHV. Sequence analysis of two EcoRI fragments of the PhHV-2 genome (European isolate 7848) revealed greatest similarity to gammaherpesviruses and in particular equine herpesvirus-2. Although an unambiguous subgrouping was not feasible, this is the first evidence that PhHV-2 may be a putative gammaherpesvirus of pinnipeds.
SUMMARYThe immunogenic proteins of cells infected with the alpha-or the gamma-herpesvirus of seals, phocid herpesvirus-1 and -2 (PhHV-1, -2), were examined in radioimmunoprecipitation assays as a further step towards the development of a PhHV-1 vaccine. With sera obtained from convalescent seals of different species or murine monoclonal antibodies (Mabs), at least seven virus-induced glycoproteins were detected in lysates of PhHV-1-infected CrFK cells. A presumably disulphide-linked complex composed of glycoproteins of 59, 67 and 113/120 kDa, expressed on the surface of infected cells, was characterized as a major immunogenic infected cell protein of PhHV-1. This glycoprotein complex has previously been identified as the proteolytically cleavable glycoprotein B homologue of PhHV-1 (14). At least three distinct neutralization-relevant epitopes were operationally mapped, by using Mabs, on the glycoprotein B of PhHV-1. Among the infected cell proteins of the antigenically closely related feline and canine herpesvirus, the glycoprotein B equivalent proved to be the most highly conserved glycoprotein. Sera obtained from different seal species from Arctic, Antarctic, and European habitats did not precipitate uniform patterns of infected cell proteins from PhHV-1-infected cell lysates although similar titres of neutralizing antibodies were displayed. Thus, antigenic differences among the alphaherpesvirus species prevalent in the different pinniped populations cannot be excluded. PhHV-2 displayed a different pattern of infected cell proteins and only limited cross-reactivity to PhHV-1 at the protein level was detected, which is in line with its previous classification as a distinct species, based on nucleotide sequence analysis, of the gammaherpesvirus lineage. A Mab raised against PhHV-2 and specific for a major glycoprotein of 117 kDa, cross reacted with the glycoprotein B of PhHV-1. The 117-kDa glycoprotein could represent the uncleaved PhHV-2 glycoprotein B homologue.
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