Respiratory syncytial virus (RSV) SH and G proteins are not essential for viral replication in vitro: clinical evaluation and molecular characterization of a cold-passaged, attenuated RSV subgroup B mutant.

A live, cold-passaged (cp) candidate vaccine virus, designated respiratory syncytial virus (RSV) B1 cp-52/2B5 (cp-52), replicated efficiently in Vero cells, but was found to be overattenuated for RSV-seronegative infants and children. Sequence analysis of reverse-transcription-PCR-amplified fragments of this mutant revealed a large deletion spanning most of the coding sequences for the small hydrophobic (SH) and attachment (G) proteins. Northern blot analysis of cp-52 detected multiple unique read-through mRNAs containing SH and G sequences, consistent with a deletion mutation spanning the SH:G gene junction. Immunological studies confirmed that an intact G glycoprotein was not produced by the cp-52 virus. Nonetheless, cp-52 was infectious and replicated to high titer in tissue culture despite the absence of the viral surface SH and G glycoproteins. Thus, our characterization of this negative-strand RNA virus identified a novel replication-competent deletion mutant lacking two of its three surface glycoproteins. The requirement of SH and G for efficient replication in vivo suggests that selective deletion of one or both of these RSV genes may provide an alternative or additive strategy for developing an optimally attenuated vaccine candidate.

Identification and characterization of a new base substitution in the vaccine strain of Sabin 3 poliovirus.

The complete RNA sequence of Sabin 3 (LED3) used in vaccine in the United States has been determined. The LED3 Sabin 3 sequence contains the attenuating mutations at bases 472 and 2034 but differs from that published by Stanway et al. (Nucleic Acids Res., 11, 5629-5643, 1983) at two other base positions, 2493 and 6061. The change at base 6061 is silent and does not affect amino acid composition. The other base, a C at position 2493, is contained in the viral capsid protein VP1 and predicts a new Sabin 3 specific amino acid change of a threonine instead of an isoleucine at amino acid 6 of the protein [corrected]. Reversion of this base to that present in the pathogenic progenitor strain, Leon, is observed to occur after replication of vaccine virus in the gut of primary vaccines and in nervous tissue of neurovirulence test monkeys. Passage conditions have been identified that lead to the reversion of base 2493 as well as the reversion of the attenuated base to the parental base (Leon) at position 472 in the 5' noncoding region. The observation that these two bases delta position are found to revert during passage suggests that there is a selective advantage for virus containing the parental bases at these positions.