Genetic recombination of pseudorabies virus: evidence that homologous recombination between insert sequences is less frequent than between autologous sequences.
We studied in vivo recombination between a thymidine kinase (TK) negative, glycoprotein E (gE) negative, attenuated strain and a virulent strain of pseudorabies virus (PRV) in pigs. To simplify the detection of recombination we inserted different but overlapping (375 bp) parts of the E1 gene of classical swine fever virus into the gG locus of both virus strains. Recombination between the E1 sequences of these viruses results in reconstitution of the complete E1 coding sequence and expression of the E1 protein. Since E1 is highly immunogenic, we expected to detect in vivo recombination in co-inoculated pigs by the presence of serum antibodies against E1. However, after co-inoculation of pigs with high doses of both virus strains, we were unable to detect antibodies against E1, suggesting that in vivo recombination did not occur or remained below the detection limit. Analysis of individual progeny viruses showed that 13 out of 995 (1.3%) possessed a recombinant TK-negative gE-positive phenotype. In contrast, no E1-positive viruses were detected among 5000 analyzed. This result showed that in vivo recombination between the two virus strains did occur, but was much more frequent between the TK and gE loci than between the E1 sequences. Similar results were obtained in in vitro recombination experiments in which possible growth differences between the various virus strains were excluded. The different recombination frequencies could not be attributed to the difference in distance of the genetic loci since recombination between mutations at a distance of 266 bp in the TK gene occurred as frequent as recombination between the TK and gE genes which are separated by approximately 60 kilobasepairs. These results indicate that some property of the E1 sequence and/or the location of the E1 sequence within the PRV genome affects the frequency of recombination.
Glazenburg
K L
KL
Institute for Animal Science and Health (ID-DLO), Department of Virology, Lelystad, The Netherlands.
Moormann
R J
RJ
Kimman
T G
TG
Gielkens
A L
AL
Peeters
B P
BP
D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis; D002460:Cell Line; D011558:Herpesvirus 1, Suid / Q000235:genetics* / Q000276:immunology; D051379:Mice; D016296:Mutagenesis; D011557:Pseudorabies / Q000276:immunology / Q000821:virology; D011995:Recombination, Genetic; D013552:Swine; D013937:Thymidine Kinase / Q000235:genetics; D014759:Viral Envelope Proteins / Q000235:genetics
D000914:Antibodies, Viral; D014759:Viral Envelope Proteins; C089580:glycoprotein E, Suid herpesvirus 1; D013937:Thymidine Kinase