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M J Willemse

Publications and source records attributed to M J Willemse.

7 recordsLinked to original sources

Transcriptional analysis of the short segment of the feline herpesvirus type 1 genome and insertional mutagenesis of a unique reading frame.

Transcription mapping was performed in the short region of the feline herpesvirus type 1 (FHV-1) genome as a first approach to the functional analysis of open reading frames encoding the homologs of the herpes simplex virus type 1 (HSV-1) gD, gl, gE, US9, and probably also US8.5. All reading frames appeared to be transcribed. Transcripts were grouped into two nested RNA sets; namely, the coterminal transcripts of gD and gl and the coterminal transcripts of gE, US8.5, and US9. This situation was similar to that found in the equivalent region of HSV-1. The FHV-1 ORFs US8.5 and US9 have not been described previously. Sequence analysis and comparison of the putative polypeptide encoded by US8.5 revealed that this ORF was unique to FHV-1. However, US8.5 of FHV-1 might be functionally related to its positional homologs in HSV-1 and equine herpesvirus type 1. In all three viruses, US8.5 does not seem to be critical for virus propagation in cell culture. This was shown for FHV-1 by isolating a mutant containing an insertion in US8.5 and comparing its growth properties in cell culture to those of the parent virus G2620. With regard to US9, conscientious amino acid sequence alignment of FHV-1 US9 and homologs in related herpesviruses suggests that this particular protein could contribute to the virus infectivity in vivo. This speculation was based on the highly conserved C-terminus of US9, starting with a characteristic YYSES motif and followed by a nuclear target sequence and a transmembrane region.

Alphaherpesvirinae

The gene downstream of the gC homologue in feline herpes virus type 1 is involved in the expression of virulence.

Feline herpesvirus type 1 (FHV-1) mutants were constructed, carrying a beta-galactosidase marker gene integrated into the region downstream of the gene encoding the homologue of glycoprotein C (gC) of herpes simplex virus type 1. In cell culture, no differences in replication were observed between mutants and the parent FHV-1 strain. However, in experimentally infected cats, mutants caused fewer clinical signs after oronasal administration although they replicated to the same extent as the parental strain. Sequence analysis in the region of the UL segment surrounding the insertion site revealed an open reading frame (ORF 2) encoding a putative polypeptide of 21K. RNA analysis indicated a corresponding transcript of 0.8 kb that was detected late after infection of cells in culture. This particular UL locus downstream of the gC gene has not been thoroughly investigated in any of the herpesviruses. The putative gene product showed only limited evolutionary conservation since similarity could be found only with the assumed homologue of equine herpesvirus type 1. Further characterization of this newly identified FHV-1 gene involved in virulence may provide insight into the development of disease owing to herpesvirus infection.

Algorithms

Avian herpesvirus as a live viral vector for the expression of heterologous antigens.

Control of Marek's disease in the poultry industry has been successfully achieved for several decades by large-scale vaccination of day-old chickens with live herpesvirus of turkeys (HVT) strains. Several features of this virus including lack of pathogenicity and long-term immune protection due to a persistent viraemic infection made us decide to use HVT as a live viral vector for the expression of foreign antigens. Potential sites for the integration of foreign DNA in the unique short region of the HVT genome were identified by the insertion of a beta-galactosidase expression cassette. Vaccination trials with recombinant virus strains indicated that the marker gene was expressed and stably maintained during animal passage. Based on an insertion site mapping in one of the open reading frames of the unique short region, a general recombination vector was designed for the integration of foreign genes into HVT. Recombinant virus-directed expression of individual antigens from Newcastle disease virus was driven by a strong promoter element derived from the lung terminal repeat sequence of Rous sarcoma virus.

Amino Acid Sequence