Significance of a newly identified attenuating mutation in Sabin 3 oral poliovirus vaccine.

Sequence analysis of poliovirus vaccine RNA has resulted in the identification of a new point mutation at position 2493 in Sabin 3. cDNA-derived Sabin 3 virus whose genome includes the new mutation has phenotypic properties expected for a vaccine strain. Virus engineered to contain a single base substitution at 2493 has lost the small plaque phenotype, and low neurovirulence characteristics normally associated with Sabin poliovirus strains. The significance of these observations and a potential relationship to the frequency of vaccine-associated poliomyelitis are described.

Oral poliovirus vaccine in the United States: molecular characterization of Sabin type 3 after replication in the gut of vaccinees.

Derivatives of Sabin 3 shed from recipients of oral poliovirus vaccine in the United States (U.S.) were examined for genetic changes identified in strains excreted by vaccinees in the United Kingdom [U.K.; Evans et al., 1985; Cammack et al., 1988, Macadam et al., 1989]. Among the eight primary vaccinees studied, the duration of excretion and molecular evolution of type 3 strains varied greatly. The period of virus excretion after vaccination ranged from as few as 2 days to as many as 36 days. Nucleotide sequence analysis of viral RNAs extracted from shed virus indicated that only fifty percent of the vaccinees exclusively excreted strains in which the attenuating mutation at nucleotide 472 in the 5' noncoding region of the genome had reverted from uracil (U) to cytosine (C), the nucleotide found in neurovirulent strains. Compared to the wild-type Leon strain, the low activity of stool isolate KW4 in a complete monkey neurovirulence test demonstrated that presence of C at 472 does not render a type 3 strain pathogenic. Conversely, an isolate was identified which efficiently replicated in monkey nervous tissue and maintained the attenuated U at 472. Oligonucleotide fingerprinting and sequence analysis of viral RNAs from stool isolates indicated that one vaccinee (KW) eventually excreted intertypic recombinant strains consistent with those reported in the U.K. studies. Unique to this study, one vaccinee (KS) excreted nonrecombinant virus possessing U at 472 for up to 21 days. The significance of the KS strain profile in relation to differences in the U.S. vaccine compared to the vaccine distributed in the U.K. and other countries is discussed.

A convenient procedure for rapid release of protoplasts from Ustilago maydis.

Filtrates from cultures of Trichoderma harzianum, grown on a liquid mineral salts medium containing vitamin and trace element supplements, 0.5 g/l glucose and 2 g/l of dried residue of ethanol-extracted commercial mushrooms (Agaricus brunnescens), are a good source of enzymes for the release of protoplasts from the sporidia of Ustilago maydis. When concentrated 50-fold by (NH4)2SO4 precipitation, the enzymes liberated 95% of the protoplasts from the sporidia within 40 min.