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Evidence for the existence of porcine gammaherpesviruses was obtained by PCR and sequence analysis. Initially, samples of peripheral blood mononuclear cells (PBMC), spleens, lungs, kidneys and livers of pigs from Germany and Spain were tested with a PCR assay which targets conserved regions of the herpesvirus DNA polymerase gene with degenerate and deoxyinosine-substituted primers. Amplicons of identical sequence were obtained from one spleen and two PBMC samples. This sequence showed a high percentage of identity with the DNA polymerase genes of herpesviruses of the oncogenic subfamily Gammaherpesvirinae. Alignment of amino acid sequences showed the highest identity values with bovine gammaherpesviruses, namely alcelaphine herpesvirus type 1 (68 %), ovine herpesvirus type 2 (68 %) and bovine lymphotropic herpesvirus (67 %). Comparison with pseudorabies virus and porcine cytomegalovirus, which are the only porcine herpesvirus species presently known, showed values of only 41 %. PCR analysis of PBMC (n l 39) and spleen (n l 19) samples from German pigs, using primers specific for the novel sequence, revealed a prevalence of 87 and 95 %, respectively. In this analysis, three out of eight spleen samples from Spanish pigs were also positive. Subsequent sequencing of the amplicons revealed the presence of two closely related gammaherpesvirus sequences, differing from each other by 8 % at the amino acid level. The putative novel porcine herpesviruses, from which these sequences originated, were tentatively designated porcine lymphotropic herpesvirus type 1 and type 2 (PLHV-1 and PLHV-2). When using pig organs for xenotransplantation, the presence of these viruses has to be considered.

A novel porcine gammaherpesvirus was detected in the blood of domestic pigs by PCR. With degenerate-primer PCR and subsequent long-distance PCR approaches a 60-kbp genome stretch was amplified. Sequence analysis revealed the presence of the gammaherpesvirus ORFs 03 to 46 as well as a putative chemokine receptor and a v-bcl-2 gene. The 60-kbp sequence was compared with the corresponding sequence of the porcine lymphotropic herpesvirus 1 (PLHV-1) published recently and the sequence of PLHV-2, which was amplified from porcine tonsil. Considerable sequence differences (amino acid identities: 49-89%) were found between the novel virus and PLHV-1 as well as PLHV-2, which were very closely related to each other (amino acid identities: 85-98%). The novel virus had essentially the same genome organization as PLHV-1 and -2 and was therefore designated PLHV-3. Like PLHV-1 and -2, PLHV-3 was frequently found in the blood and in lymphoid organs of domestic and feral pigs from different geographic locations. In the blood, the PLHVs were detected predominantly in B-cells. Indication for latent as well as productive PLHV-3 infection was found in the porcine B-cell line L23. It can be concluded that the PLHVs are widespread and are likely to cause a persistent B-lymphotropic infection. Since PLHV-1 has been implicated in the development of porcine posttransplantation lymphoproliferative disease, all porcine lymphotropic gammaherpesviruses are of concern when pigs are used as donors in xenotransplantation.

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Equine herpesvirus type 2 (EHV-2) is a slow-growing, cytopathogenic gammaherpesvirus, which is suggested to be ubiquitous in the equine population. However, its precise role as a pathogen and its tissue tropism remains uncertain. To estimate the prevalence of EHV-2 in Germany and to investigate the possible pathogenicity of the virus, peripheral blood leucocytes (PBL) from 172 horses were examined for EHV-2 DNA by a sensitive and specific nested PCR based on the EcoRI-N genomic fragment and by classical cocultivation. PBL samples from 51% of the horses were positive by PCR and virus was isolated from 31% of the horses by cocultivation. However, almost all animals were seropositive for EHV-2. This may indicate that PBL do not harbour EHV-2 indefinitely after infection. Furthermore, a correlation between clinical signs and EHV-2 as a causative agent could not be determined. Nevertheless, the prevalence of virus was high among horses with upper respiratory tract disease, abortion and severe ataxia. The products of the second round of the PCR reactions showed size polymorphism. Sequencing of the products revealed that these size differences were due to repetition of the motif (AGACAGGGGCCATGCTGGC) between 9-16 times depending on the isolate, suggesting that the nested PCR might be a useful tool for the differentiation of EHV-2 isolates.