Interestingly, most previous studies42C45 that have reported selecting single mutations with large effects (???10-fold) on neutralization by anti-stalk antibodies have used group 2 (e

Interestingly, most previous studies42C45 that have reported selecting single mutations with large effects (???10-fold) on neutralization by anti-stalk antibodies have used group 2 (e.g., H3 or H7) HAs rather than group 1 HAs like the one used in our work,?although at least one study has selected a large-effect escape mutation to a broad anti-stalk antibody in an H5 group 1 HA61. addition, all of this code as well as the manuscript itself and other data are available on GitHub at https://github.com/jbloomlab/HA_antibody_ease_of_escape. Finally, the dms_tools2 software76 that performs most of the analysis is available at https://jbloomlab.github.io/dms_tools2/. The authors declare that all other data supporting the findings of this study are available within the article and its Supplementary Information files, or are available from the authors upon request Abstract Influenza virus can escape most antibodies with single mutations. However, rare antibodies broadly neutralize many viral strains. It is unclear how easily influenza virus might escape such antibodies if there was strong pressure to do so. Here, we map all single amino-acid mutations that increase resistance to broad antibodies to H1 hemagglutinin. Our approach not only identifies antigenic mutations but also quantifies their effect sizes. All antibodies select mutations, but the effect sizes vary widely. The virus can escape a broad antibody to hemagglutinins receptor-binding site the same way it escapes narrow strain-specific antibodies: via single mutations with huge effects. In contrast, broad antibodies to hemagglutinins stalk only select mutations with small effects. Therefore, among the antibodies we examine, breadth is an imperfect indicator of the potential for viral escape via single mutations. Antibodies targeting the H1 hemagglutinin stalk are quantifiably Doripenem harder to escape than the other antibodies tested here. Influenza A virus can escape antibodies, but it is unclear how the ease of CCR2 escape depends on the epitope targeted by an Doripenem antibody. Here, the authors show that neutralization breadth is an imperfect indicator of the ease of viral escape by single mutations, and that antibodies targeting the stalk of hemagglutinin are harder to escape. Introduction Nearly all viruses show some antigenic variation. However, the extent of this variation ranges widely. For instance, although both measles virus1,2 and polio virus3C5 exhibit antigenic variation, the magnitude of this variation is small. Therefore, immunity of these viruses is lifelong6,7. In contrast, human influenza virus exhibits much more antigenic variation. So although infection with an influenza virus strain provides long-term immunity to that exact strain8C10, the viruss rapid antigenic evolution erodes the effectiveness of this immunity to that strains descendants within 5 years11,12. One possible reason that viruses exhibit different amounts of antigenic variation is that they have disparate evolutionary capacities to escape the immunodominant antibodies generated by natural immune responses13C15. According to this explanation, human influenza virus undergoes rapid antigenic drift because most neutralizing antibodies target epitopes on the viral hemagglutinin (HA) protein that are highly tolerant of mutational change. This explanation is supported by classic experiments showing that it is easy to select viral mutants that escape most antibodies16,17, as well as by the observation that mutations that alter antigenicity arise frequently during influenzas evolution globally18C22 and within individual humans with long-term infections23. A corollary of this explanation is that influenza viruss capacity for antigenic drift would be reduced if most antibodies instead targeted epitopes that were less mutationally tolerant. Verifying this corollary has become of practical importance with the discovery of broadly neutralizing antibodies against influenza virus. These antibodies typically target conserved epitopes in HAs stalk24C26 or receptor-binding site27C29, and neutralize a wide range of viral strains. Broad antibodies are usually less abundant in human serum than Doripenem antibodies to antigenically variable epitopes on the head of HA30,31. However, major efforts are underway to elicit broad antibodies by vaccination or administer them directly as therapeutics32,33. If these efforts succeed, the epitopes of broad antibodies could come under stronger antigenic selection in human influenza virus. Might such selection then drive antigenic variation in these epitopes? Doripenem There is precedent for the idea that the immune status of the host population can shape influenza virus evolution: the virus undergoes faster antigenic drift in long-lived humans that accumulate immune memory than in short-lived swine that are mostly naive34,35, and poultry vaccination may accelerate antigenic drift of avian influenza36,37. But alternatively, perhaps broad antibodies are broad because the Doripenem virus has difficulty escaping them regardless of selection from host immunity. So far, there is limited data to distinguish between these possibilities. Several studies have shown that the head domain of HA is more mutationally tolerant than the stalk domain where many broad antibodies bind38C40. However, these studies did not select for antibody escape, so.