Isolation and characterization of influenza virus recombinants with high and low neuraminidase activity: Use of 2-(3′-methoxyphenyl)- N-acetylneuraminic acid to identify cloned populations

Abstract

Two antigenically identical HON2 influenza virus recombinants derived from A/England/42/72 (H3N2) and A/PR8/34 (HON1) were separated from a mixed population by the use of a staining procedure specific for neuraminidase. This procedure employing an artificial neuraminidase substrate (2-(3′-methoxyphenyl)- N-acetylneuraminic acid, MPN) permitted the identification of deeply stained (MPN +) and poorly stained (MPN −) clones of virus in clone 1-5C-4 cells. Upon isolation and purification of the 2 variants, they were found to have eightfold differences in the ratios of neuraminidase/hemagglutination activity. Analysis by polyacrylamide gel electrophoresis demonstrated that the differences in neuraminidase activity reflected differences in the amount of neuraminidase per virion. The MPN − variant which had low neuraminidase content replicated to equal titer in eggs and formed larger plaques than MPN + variant on clone 1-5C-4 cells. Attempts to isolate clones of MPN − virus from wild type A/England/42/72 virus have not been successful so far.