In combination with computational analyses and other methods28, selection of potential antigenic drift mutants may improve the selection of vaccine seed viruses. Methods Ethics Human blood was collected from a volunteer by following a protocol approved by the Research Ethics Review Committee of the Institute of Medical Science, the University or college of Tokyo (approval number 25-58-1205), and all experiments in this manuscript were performed in accordance with the University or college of Tokyos guidelines and regulations. that occurred in epidemic strains, other HA mutations can confer resistance to antibodies that identify the K166 area, leading to emergence of epidemic strains with such mutations. Introduction The first influenza pandemic of the 21st century began in 2009 2009 with the emergence of the A(H1N1)pdm09 computer virus, which replaced the previous seasonal H1N1 (sH1N1) computer virus1,2. Surveillance of circulating A(H1N1)pdm09 viruses has revealed some genetic variations in the viral surface glycoproteins, hemagglutinin (HA) and neuraminidase (NA)3,4. However, until recently, the antigenicity of the circulating A(H1N1)pdm09 viruses was similar to the vaccine strain (A/California/7/2009) in assays with panels of antiserum obtained from infected ferrets3. Therefore, the World Health Organization (WHO) recommended using A/California/7/2009-like computer virus as a vaccine seed computer virus until the 2016C2017 Northern hemisphere influenza season3. However, human sera distinguished the antigenicity of recent A(H1N1)pdm09 viruses from that of the A/California/7/2009-like vaccine computer virus, whereas ferret antisera failed to detect this antigenic difference5,6. Accordingly, since the 2017 Southern Hemisphere influenza season, the WHO has recommended A/Michigan/45/2015-like computer virus be used as the vaccine RGX-104 free Acid seed computer virus5,6. The HA protein is the major influenza viral antigen and the primary target of neutralizing antibodies7. A(H1N1)pdm09-HA has five major antigenic sites, which were identified by studies using A/Puerto Rico/8/34 (H1N1)8C11. Two immunodominant sites (Sa and Sb) are located proximal to the receptor-binding pocket and elicit high potency neutralizing antibodies8,10. The Ca sites (Ca1 and Ca2) are at the subunit interface, and the Cb site is usually close to the stalk region of HA.8,10. A(H1N1)pdm09 viruses isolated after the 2012C2013 influenza season, which are classified into the genetic RGX-104 free Acid group 6B, possess a lysine-to-glutamine substitution at position 166 (K166Q, H3 numbering) within the Sa antigenic site12. This mutation affects the antigenicity of recent A(H1N1)pdm09 viruses13. In the 2016C2017 influenza season, A(H1N1)pdm09 viruses classified into genetic group 6B.1 circulated among humans5. The 6B.1 viruses obtained a serine-to-asparagine substitution at position 165 (S165N) in the Sa antigenic site that resulted in the generation of an N-glycosylation site4,14. Previous reports have explained several human monoclonal antibodies that identify an epitope around position 166 of A(H1N1)pdm09-HA and neutralize A(H1N1)pdm09 and sH1N1 viruses. However, these antibodies failed to neutralize A(H1N1)pdm09 viruses isolated after the 2012C2013 season, which possessed the K166Q mutation, or sH1N1 viruses isolated between 1986 and 2008, which experienced a potential glycosylation site (129-NHT-131) masking the epitope around position 16615C17. Middle-aged adults RGX-104 free Acid (i.e., given birth to between 1965 and 1979) were reported to have a high antibody titer against the epitope around position 166 of the HA of A(H1N1)pdm09 viruses that were circulating before 2012, since they had been exposed to sH1N1 viruses that were circulating before 1985 and whose HA lacked the 129-NHT-131 glycosylation site. These middle-aged adults suffered from A(H1N)pdm09 computer virus infection with substantial morbidity and mortality during the 2013C2014 influenza season because of low neutralization antibody titers against the viruses possessing the K166Q substitution12. These reports demonstrate that this epitope around position 166 plays a role in the antigenic drift recently detected in assays with human, but not ferret, sera. Another group confirmed this phenomenon by using a panel of human monoclonal antibodies from a middle-aged adult17. selection of Agt escape mutants from these monoclonal antibodies revealed that a mutation at position 166 of HA was responsible for the resistance to neutralization, suggesting that this antigenic drift was caused by the selective pressure of the human antibodies realizing the epitope around position 16617. Here, we established two human anti-A(H1N1)pdm09 HA monoclonal antibodies that acknowledged the epitope around position 166. We then attempted to obtain escape mutants possessing an amino acid substitution other than at position 166 and compared their growth ability.
In combination with computational analyses and other methods28, selection of potential antigenic drift mutants may improve the selection of vaccine seed viruses