Characterization of in vitro and in vivo phenotypes of poliovirus type 1 mutants with reduced viral protein synthesis activity

Abstract

Sabin vaccine strains of poliovirus (PV) contain major attenuation determinants in the internal ribosomal entry site (IRES), an area that directs viral protein synthesis. To examine the effect of reduced viral protein synthesis on PV neurovirulence, spacer sequences, consisting of short open reading frames of different lengths, were introduced between the IRES and the initiation codon of viral polyprotein, resulting in PV mutants with reduced viral protein synthesis. These PV mutants had a viral protein synthesis activity 8.8-55 % of that of the parental Mahoney strain as measured in HeLa S3 cells. Only viruses with more than 28 % of the wild-type activity had intact spacer sequences following plaque purification. Mutants with 17 % or 21 % of the wild-type activity were unstable and a mutant with 8.8 % was lethal. The neurovirulence of PV mutants was evaluated in transgenic mice carrying the human PV receptor gene. In this test, mutants with more than 28 % of the wild-type activity remained neurovirulent, while a mutant with 17 % of wild-type activity exhibited a partially attenuated phenotype. This mutant stably replicated in the spinal cord; however, the stability was severely affected during the course of virus infection from the cerebrum to the spinal cord. These results suggest that reduced viral protein synthesis activity as measured in cultured cells (17-55 % of the wild-type activity) is not the main determinant of PV attenuation.