Temperature-sensitive mutants of influenza virus XVI. Transfer of the two ts lesions present in the Udorn/72-ts- 1 A2 donor virus to the Victoria/3/75 wild-type virus

Previously, the influenza A/Hong Kong/68-ts-1[E] virus and its recombinants (38° shutoff temperature of plaque formation) were shown to be insufficiently attenuated for persons who lacked immunity to both the hemagglutinin and neuraminidase surface glycoproteins, i.e., doubly seronegative individuals. To meet the need for immunization of such individuals, a virus more defective than the ts-1[E] recombinants was produced. The resulting virus, designated Udorn/72-ts-1A2, possessed is mutations in genes represented by complementation-recombination groups 1 and 5 and was more restricted in replication in vitro at 37° and in vivo in the hamster's lungs and nasal turbinates than the Udorn/72ts-1[E] virus. These properties suggested that the Udorn/72 ts-1A2 virus might serve as a donor of its two ts lesions to new variants of influenza A virus to produce attenuated strains for use in doubly seronegative individuals. The Udorn/72-ts-1A2 virus was mated with the influenza A/Victoria/3/75 wild-type virus, and six is recombinant viruses bearing the Vic/75 hemagglutinin were isolated. Five of these Vic/75-ts-1A2 recombinants had a 37° shutoff temperature and two ts lesions like their Udorn/72-s-1A2 parent. Each of these clones replicated to low titer or not at all in the hamster's lungs and were 100-fold restricted in the nasal turbinates like their ts parental virus. Each of 158 isolates recovered from hamsters infected with the parental Udorn/72-ts-1A2 virus or one of its Vic/75 ts double lesion recombinants retained the ts phenotype. One Vic/75-ts-1A2 recombinant clone that had a 38°hutoff temperature, and only the group 1 ts lesion replicated in the hamster's lungs to a level intermediate between that of the parental ts-1A2 and wild-type virus. All isolates recovered from hamsters infected with this recombinant were also ts. Infection of hamsters with a Vic/75-ts-1A2 recombinant, bearing the group 1 and group 5 ts mutations, induced resistance to homologous wild-type virus challenge. Since the parental and double lesion recombinant ts-1A2 viruses had similar restriction of replication in vitro at 37° and in vivo, it is likely that the two ts-1A2 is lesions will effect a similar level of attenuation following transfer into other antigenic variants of influenza A virus.