DELETING VALINE-125 AND CYSTEINE-126 IN GLYCOPROTEIN-GI OF PSEUDORABIES VIRUS-STRAIN NIA-3 DECREASES PLAQUE SIZE AND REDUCES VIRULENCE IN MICE

We investigated the function of antigenic domains on gI in virulence a nd immunogenicity. Three PRV gI mutants were constructed by deleting n ucleotides coding for the following amino acids: valine-125 and cystei ne-126, located in a discontinuous antigenic domain (M 303); glycine-5 9 and aspartic acid-60 located in a continuous antigenic domain (M 304 ); and arginine-67 and alanine-68, located in a discontinuous antigeni c domain (M 305). Mismatch primers in the polymerase chain reaction we re used to introduce the deletions. Anti-gI monoclonal antibodies were used in an immunoperoxidase monolayer assay to distinguish PRV gI mut ants from wild-type PRV. The gI mutant viruses were tested for their g rowth in vitro and for their virulence in mice. The growth properties of PRV gI mutant virus M 303 were comparable to the growth properties of a PRV gI-negative mutant (M 301): both mutants produced small plaqu es in various cells, and when grown on swine kidney cells and chicken embryo fibroblasts, their growth was disadvantaged compared to wild-ty pe PRV. However, in embryonal Balb/c mouse cells expressing gI, gI mut ant viruses and wild-type PRV produced plaques of the same size, confi rming that the mutations in gI are responsible for the small plaque ph enotype. The growth properties of PRV gI mutant viruses M 304 and M 30 5 were comparable to the growth properties of wild-type PRV. When the mean time to death was used as the criterion, the gI mutant viruses M 301 and M 303 were significantly less virulent in mice than wild-type PRV. Four other, independently obtained, PRV mutants all carrying the valine-125 and cysteine-126 deletion (M 308, M 309, M 310 and M 311 re spectively) exhibit the same phenotype. Our results show that deleting valine-125 and cysteine-126 in gI decreases plaque size and reduces v irulence in mice to the same degree as deleting the gI protein.