CHARACTERIZATION OF A TEMPERATURE-SENSITIVE INFLUENZA-B VIRUS MUTANT DEFECTIVE IN NEURAMINIDASE

ts5, a temperature-sensitive mutant of influenza B virus, belongs to o ne of seven recombination groups. When the mutant infected MDCK cells at the nonpermissive temperature (37.5-degrees-C), infectious virus wa s produced at very low levels compared with the yield at the permissiv e temperature (32-degrees-C) and hemagglutinating and enzymatic activi ties were undetectable. However, viral protein synthesis and transport of hemagglutinin (HA) and neuraminidase (NA) to the cell surface were not affected. The NA was found as a monomer within cells even at 32-d egrees-C, in contrast to wild-type virus NA, existing mostly as an oli gomer, but the mutant had oligomeric NA, like the wild-type virus. Its enzymatic activity was more thermolabile than that of wild-type virus . Despite the low yield, large aggregates of progeny virus particles w ere found to accumulate on the cell surface at the nonpermissive tempe rature, and these aggregates were broken by treatment with bacterial n euraminidase, with the concomitant appearance of hemagglutinating acti vity, suggesting that NA prevents the aggregation of progeny virus by removal of neuraminic acid from RA and cell receptor, allowing its rel ease from the cells. Further treatment with trypsin resulted in the re covery of infectivity. When bacterial NA was added to the culture earl y in infection, many hemagglutinable infectious virus was produced. We also suggest that the removal of neuraminic acid from RA by NA is ess ential for the subsequent cleavage of HA by cellular protease. Nucleot ide sequence analysis of RNA segment 6 revealed that ts5 encoded five amino acid changes in the NA molecule but not in NB.