MUTATIONS AFFECTING THE UL21 GENE CONTRIBUTE TO AVIRULENCE OF PSEUDORABIES VIRUS-VACCINE STRAIN BARTHA

Analysis of the live attenuated pseudorabies virus (PrV) vaccine strai n Bar?ha indicated location of a major determinant for PrV neurovirule nce within the genomic BamHI fragment 4 (B. Lomniczi ei al., 1984, J. Virol. 52, 198-205). To more precisely localize the defect, marker res cue experiments were performed using cloned subfragments of BamHI-4. R escuants were analyzed after intracerebral infection for their virulen ce in chicken, as well as after intranasal infection for virulence in pigs. We show that the defect associated with attenuation in strain Ba rtha is located in a 3.8-kb subfragment of BamHI-4 which encompasses t he PrV UL20 and UL21 genes and a putative origin of replication (B. Kl upp, H. Kern, and T. C. Mettenleiter, 1992, Virology 191, 900-908). Se quence analysis of this region of the strain Bartha genome and compari son with the corresponding region in wild-type PrV strain Ka revealed the presence of eight point mutations. Four nucleotide exchanges resid e within the UL21 gene with three of them leading to amino acid substi tutions; one is located in the intergenic region between the UL20 and UL21 genes and three are localized downstream from the UL21 gene. Neit her the UL20 gene nor the putative origin sequence was affected. Inser tional inactivation of the UL21 gene in wild-type PrV strain Ka led to a marked attenuation of the virus for pigs infected by the intranasal route. In summary, our data show that the PrV UL21 gene is a major de terminant of PrV virulence and that point mutations affecting the UL21 gene of live vaccine strain Bartha contribute to its attenuated pheno type. (C) 1995 Academic Press, Inc.