LIMITED GENETIC CHANGES IN THE SABIN-1 STRAIN OF POLIOVIRUS OCCURRINGIN THE CENTRAL-NERVOUS-SYSTEM OF MONKEYS

Replication of attenuated poliovirus strains results in their partial deattenuation. Recently we identified mutations accumulating in the Sa bin 1 poliovirus in cell cultures. Here we report genetic changes occu rring in this virus during replication in the central nervous system ( CNS) of monkeys. Viruses isolated from different parts of the CNS of r hesus monkeys (inoculated into the spinal cord) were screened for sequ ence heterogeneities and newly identified mutations were independently confirmed and quantified using mutant analysis by PCR and restriction enzyme cleavage (MAPREC). All consistently accumulating mutations ide ntified in this study were located in untranslated regions: GU --> AU or GU --> GC substitution at a complementary pair formed by nucleotide s 480 and 525, U --> C substitution at nucleotide 612, and GU --> AU o r GU --> GC substitution of a base pair formed by the nucleotides 7427 /7441 immediately preceding the poly(A) tract. All these mutations exc ept one (7427) were previously identified in cell culture passages or stool isolates from vaccinees. Sequencing of 11 CNS isolates also iden tified a few random silent mutations that accumulated as neutral 'pass engers', passively coselected with genuinely selectable mutations pres ent on the same RNA molecule. One isolate also contained the wild-type base at nucleotide 2741 (Ala(88) --> Thr in VPI). Our results demonst rate a remarkable genetic stability of the Sabin 1 poliovirus in the C NS of monkeys, suggesting that deattenuation is determined by a very l imited number of mutations. These mutations can be assayed by MAPREC t o monitor the consistency of oral poliovirus vaccine (OPV) production.