Dr. Kawaoka gave a talk at the Institut Pasteur as part of the 9th Louis Pasteur conference on Emerging Infectious Diseases. No title or abstract were provided. Lots of data were flashed up; not easy to keep up with the tempo.
• Started with pandemic 2009 influenza A virus (pH1N1) and selected escape mutants with polyclonal sera. Classic. There was little diversity and was disappointed.
• Took gene segment corresponding to amino acid residues 71-270 of the hemagglutinin (HA), performed error prone PCR, re-expressed as mutant library in H1N1 backbone by reverse genetics.
• Took 14 convalescent sera and isolated 50 escape mutants. Found 9 amino acid sites of variation.
• Factored in a few other sites (didn’t catch how) and ended up with 15 key sites in the HA sequence
• Performed saturation codon mutagenesis on these sites in the gene and ended up with 2 amino acid site, 3 site, 4 site and 5 site virus variants that were further refined by mutagenesis. 4 and 5 site mutants completely escaped vaccine-induced antibodies.
They worked hard to get a vaccine resistant virus. Apparently the work was performed at biosafety level 2 (BSL2).
