Characterization of recombinant influenza B viruses with key neuraminidase inhibitor resistance mutations.
OBJECTIVE
An influenza B virus plasmid-based rescue system was used to introduce site-specific mutations, previously observed in neuraminidase (NA) inhibitor-resistant viruses, into the NA protein of six recombinant viruses. Three mutations observed only among in vitro selected zanamivir-resistant influenza A mutants were introduced into the B/Beijing/1/87 virus NA protein, to change residue E116 to glycine, alanine or aspartic acid. Residue E116 was also mutated to valine, a mutation found in the clinic among oseltamivir-resistant viruses. An arginine to lysine change at position 291 (292 N2 numbering) mimicked that seen frequently in influenza A N2 clinical isolates resistant to oseltamivir. Similarly, an arginine to lysine change at position 149 (152 in N2 numbering) was made to reproduce the change found in the only reported zanamivir-resistant clinical isolate of influenza B virus. In vitro selection and prolonged treatment in the clinic leads to resistance pathways that require compensatory mutations in the haemagglutinin gene, but these appear not to be important for mutants isolated from immunocompetent patients. The reverse genetics system was therefore used to generate mutants containing only the NA mutation.


CONCLUSIONS
With the exception of a virus containing the E116G mutation, mutant viruses were attenuated to different levels in comparison with wild-type virus. This attenuation was a result of altered NA activity or stability depending on the introduced mutation. Mutant viruses displayed increased resistance to zanamivir, oseltamivir and peramivir, with certain viruses displaying cross-resistance to all three drugs.
Jackson
David
D
School of Animal and Microbial Sciences, University of Reading, Whiteknights, PO Box 228, Reading RG6 6AJ.
Barclay
Wendy
W
Zürcher
Thomas
T
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D004285:Dogs; D024882:Drug Resistance, Viral / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D009981:Influenza B virus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009439:Neuraminidase / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D011995:Recombination, Genetic; D014709:Vero Cells
D004791:Enzyme Inhibitors; D009439:Neuraminidase