ATTENUATED TEMPERATURE-SENSITIVE RESPIRATORY SYNCYTIAL VIRUS MUTANTS GENERATED BY COLD ADAPTATION

Two strains of respiratory syncytial virus (RSV), RSV 2B and RSV 3A (r epresenting subgroup B and A virus respectively) were cold-adapted by passaging in Vero cells for up to 42 weeks at successively lower tempe ratures down to 20 degrees C. Successful cold adaptation of the virus population was dependent on the amount of time the cultures were maint ained at the various low temperatures, as well as on the strain of vir us used. Temperature-sensitive (TS) mutants appeared in the cold passa ged virus populations; however, the majority of the virus variants rem ained predominantly non-TS. Four RSV 2B and three RSV 3A TS mutants we re selected for further characterization. These seven TS mutants retai ned their fusion phenotype and two major neutralizing antibody epitope s, and displayed varying levels of temperature sensitivity. Six of the seven mutants had a cold-adapted (CA) phenotype. All of the RSV 2B mu tants were highly attenuated in cotton rats and two of the mutants eli cited relatively high levels of neutralizing antibody and were able to protect rats against virus challenge. The RSV 3A TS mutants grew well in the nose but poorly in the cotton rat lungs, as did the parental 3 A virus. All 3A mutants elicited high titers of neutralizing antibody and provided complete protection against virus challenge. These mutant s showed varying levels of temperature sensitivity in vitro and attenu ation in vivo and represent potential vaccine candidates.