Rules of co-occurring mutations characterize the antigenic evolution of human influenza A/H3N2, A/H1N1 and B viruses.
ABSTRACT: The human influenza viruses undergo rapid evolution (especially in hemagglutinin (HA), a glycoprotein on the surface of the virus), which enables the virus population to constantly evade the human immune system. Therefore, the vaccine has to be updated every year to stay effective. There is a need to characterize the evolution of influenza viruses for better selection of vaccine candidates and the prediction of pandemic strains. Studies have shown that the influenza hemagglutinin evolution is driven by the simultaneous mutations at antigenic sites. Here, we analyze simultaneous or co-occurring mutations in the HA protein of human influenza A/H3N2, A/H1N1 and B viruses to predict potential mutations, characterizing the antigenic evolution.We obtain the rules of mutation co-occurrence using association rule mining after extracting HA1 sequences and detect co-mutation sites under strong selective pressure. Then we predict the potential drifts with specific mutations of the viruses based on the rules and compare the results with the "observed" mutations in different years.The sites under frequent mutations are in antigenic regions (epitopes) or receptor binding sites.Our study demonstrates the co-occurring site mutations obtained by rule mining can capture the evolution of influenza viruses, and confirms that cooperative interactions among sites of HA1 protein drive the influenza antigenic evolution.
Project description:To examine whether positive selection operates on the hemagglutinin 1 (HA1) gene of human influenza A viruses (H1 subtype), 21 nucleotide sequences of the HA1 gene were statistically analyzed. The nucleotide sequences were divided into antigenic and nonantigenic sites. The nucleotide diversities for antigenic and nonantigenic sites of the HA1 gene were computed at synonymous and nonsynonymous sites separately. For nonantigenic sites, the nucleotide diversities were larger at synonymous sites than at nonsynonymous sites. This is consistent with the neutral theory of molecular evolution. For antigenic sites, however, the nucleotide diversities at nonsynonymous sites were larger than those at synonymous sites. These results suggest that positive selection operates on antigenic sites of the HA1 gene of human influenza A viruses (H1 subtype).
Project description:The antigenic variability of influenza viruses has always made influenza vaccine development challenging. The punctuated nature of antigenic drift of influenza virus suggests that a relatively small number of genetic changes or combinations of genetic changes may drive changes in antigenic phenotype. The present study aimed to identify antigenicity-associated sites in the hemagglutinin protein of A/H1N1 seasonal influenza virus using computational approaches. Random Forest Regression (RFR) and Support Vector Regression based on Recursive Feature Elimination (SVR-RFE) were applied to H1N1 seasonal influenza viruses and used to analyze the associations between amino acid changes in the HA1 polypeptide and antigenic variation based on hemagglutination-inhibition (HI) assay data. Twenty-three and twenty antigenicity-associated sites were identified by RFR and SVR-RFE, respectively, by considering the joint effects of amino acid residues on antigenic drift. Our proposed approaches were further validated with the H3N2 dataset. The prediction models developed in this study can quantitatively predict antigenic differences with high prediction accuracy based only on HA1 sequences. Application of the study results can increase understanding of H1N1 seasonal influenza virus antigenic evolution and accelerate the selection of vaccine strains.
Project description:BACKGROUND: In pandemic and epidemic forms, avian and human influenza viruses often cause significant damage to human society and economics. Gradually accumulated mutations on hemagglutinin (HA) cause immunologically distinct circulating strains, which lead to the antigenic drift (named as antigenic variants). The "antigenic variants" often requires a new vaccine to be formulated before each annual epidemic. Mapping the genetic evolution to the antigenic drift of influenza viruses is an emergent issue to public health and vaccine development RESULTS: We developed a method for identifying antigenic critical amino acid positions, rules, and co-mutated positions for antigenic variants. The information gain (IG) and the entropy are used to measure the score of an amino acid position on hemagglutinin (HA) for discriminating between antigenic variants and similar viruses. A position with high IG and entropy implied that this position is highly correlated to an antigenic drift. Nineteen positions with high IG and high genetic diversity are identified as antigenic critical positions on the HA proteins. Most of these antigenic critical positions are located on five epitopes or on the surface based on the HA structure. Based on IG values and entropies of these 19 positions on the HA, the decision tree was applied to create a rule-based model and to identify rules for predicting antigenic variants of a given two HA sequences which are often a vaccine strain and a circulating strain. The predicting accuracies of this model on two sets, which consist of a training set (181 hemagglutination inhibition (HI) assays) and an independent test set (31,878 HI assays), are 91.2% and 96.2% respectively. CONCLUSION: Our method is able to identify critical positions, rules, and co-mutated positions on HA for predicting the antigenic variants. The information gains and the entropies of HA positions provide insight to the antigenic drift and co-evolution positions for influenza seasons. We believe that our method is robust and is potential useful for studying influenza virus evolution and vaccine development.
Project description:A new antigenic variant of H1N1 swine influenza virus A (Sw/QC/5393/91 [QC/91]) has been found to be associated with porcine proliferative and necrotizing pneumonia. Analysis of its genomic RNA by T1 oligonucleotide mapping revealed that considerable genomic divergence exists between QC/91 and the swine influenza viruses currently circulating in North American swine herds. Analysis of the nucleotide sequence of the HA1 region of the hemagglutinin RNA of QC/91, in comparison with those of most common H1N1 human and swine influenza A viruses, showed the presence of multiple point mutations. Two amino acid substitutions appeared to be located in antigenic sites Sb and Ca. This correlates with antigenic variations demonstrated between A/NJ/8/76, A/Sw/WI/49/76, and Québec isolate A/Sw/QC/5393/91 of swine influenza virus A. Another mutation was responsible for the loss of a glycosylation site, which may have also affected the antigenicity. The other mutations seem to have been accumulated progressively over time. This significant constancy in the fixation of mutations with time suggests that genetic diversity of these viruses may best be interpreted as the result of drifts in the population of circulating swine influenza viruses in Québec.
Project description:BACKGROUND:The genetic and antigenic characteristics of influenza A viruses (IAV) within and between species change over time due to antigenic shift and drift. Although pigs are known to play a key role in the epidemiology of IAV between species, little is known about the molecular evolution of IAV hemagglutinin (HA) in pigs. OBJECTIVES:The aim of this study was to evaluate the HA drift of an H1N1 IAV after infecting weaned pigs with or without maternally derived passive immunity. METHODS:Three- to four-week-old piglets born either to vaccinated or unvaccinated sows were contact-infected upon exposure with an IAV-infected pig. Nasal swabs were collected daily from each pig and tested for IAV by RRT-PCR. Full-length HA sequences were obtained directly from positive nasal swabs and compared between groups. RESULTS:Synonymous and non-synonymous mutations were detected in pigs with and without passive immunity. Most of the non-synonymous mutations occurred within the HA1 region of the HA. Changes within HA1 region were only identified in antigenic site B in pigs without passive immunity and in antigenic sites A, B, and D in pigs with passive immunity. However, there was no association between the immune status of the pig and the amino acid substitutions observed. CONCLUSIONS:Overall, we demonstrated that amino acid substitutions within antigenic sites can happen in weaned pigs with or without passive immunity shortly after infection.
Project description:Influenza is notable for its evolutionary capacity to escape immunity targeting the viral hemagglutinin. We used deep mutational scanning to examine the extent to which a high inherent mutational tolerance contributes to this antigenic evolvability. We created mutant viruses that incorporate most of the ?10(4) amino-acid mutations to hemagglutinin from A/WSN/1933 (H1N1) influenza. After passaging these viruses in tissue culture to select for functional variants, we used deep sequencing to quantify mutation frequencies before and after selection. These data enable us to infer the preference for each amino acid at each site in hemagglutinin. These inferences are consistent with existing knowledge about the protein's structure and function, and can be used to create a model that describes hemagglutinin's evolution far better than existing phylogenetic models. We show that hemagglutinin has a high inherent tolerance for mutations at antigenic sites, suggesting that this is one factor contributing to influenza's antigenic evolution.
Project description:Each year, influenza viruses cause epidemics by evading pre-existing humoral immunity through mutations in the major glycoproteins: the haemagglutinin (HA) and the neuraminidase (NA). In 2004, the antigenic evolution of HA of human influenza A (H3N2) viruses was mapped (Smith et al., Science 305, 371-376, 2004) from its introduction in humans in 1968 until 2003. The current study focused on the genetic evolution of NA and compared it with HA using the dataset of Smith and colleagues, updated to the epidemic of the 2009/2010 season. Phylogenetic trees and genetic maps were constructed to visualize the genetic evolution of NA and HA. The results revealed multiple reassortment events over the years. Overall rates of evolutionary change were lower for NA than for HA1 at the nucleotide level. Selection pressures were estimated, revealing an abundance of negatively selected sites and sparse positively selected sites. The differences found between the evolution of NA and HA1 warrant further analysis of the evolution of NA at the phenotypic level, as has been done previously for HA.
Project description:The hemagglutinin protein of H3N2 influenza viruses is the major target of neutralizing antibodies induced by infection and vaccination. However, the virus frequently escapes antibody-mediated neutralization due to mutations in the globular head domain. Five topologically distinct antigenic sites in the head domain of H3 hemagglutinin, A to E, have been previously described by mapping the binding sites of monoclonal antibodies, yet little is known about the contribution of each site to the immunogenicity of modern H3 hemagglutinins, as measured by hemagglutination inhibition activity, which is known to correlate with protection. To investigate the hierarchy of antibody immunodominance, five ?1 recombinant influenza viruses expressing hemagglutinin of the A/Hong Kong/4801/2014 (H3N2) strain with mutations in single antigenic sites were generated. Next, the ?1 viruses were used to determine the hierarchy of immunodominance by measuring the hemagglutination inhibition reactivity of mouse antisera and plasma from 18 human subjects before and after seasonal influenza vaccination in 2017-2018. In both mice and humans, mutations in antigenic site B caused the most significant decrease in hemagglutination inhibition titers compared to wild-type hemagglutinin. This study revealed that antigenic site B is immunodominant in the H3N2 influenza virus strain included in the current vaccine preparations.IMPORTANCE Influenza viruses rapidly evade humoral immunity through antigenic drift, making current vaccines poorly effective and antibody-mediated protection short-lived. The majority of neutralizing antibodies target five antigenic sites in the head domain of the hemagglutinin protein that are also the most sequence-variable regions. A better understanding of the contribution of each antigenic site to the overall antibody response to hemagglutinin may help in the design of improved influenza virus vaccines.
Project description:BACKGROUND: The influenza A(H1N1)2009 virus has been the dominant type of influenza A virus in Finland during the 2009-2010 and 2010-2011 epidemic seasons. We analyzed the antigenic characteristics of several influenza A(H1N1)2009 viruses isolated during the two influenza seasons by analyzing the amino acid sequences of the hemagglutinin (HA), modeling the amino acid changes in the HA structure and measuring antibody responses induced by natural infection or influenza vaccination. METHODS/RESULTS: Based on the HA sequences of influenza A(H1N1)2009 viruses we selected 13 different strains for antigenic characterization. The analysis included the vaccine virus, A/California/07/2009 and multiple California-like isolates from 2009-2010 and 2010-2011 epidemic seasons. These viruses had two to five amino acid changes in their HA1 molecule. The mutation(s) were located in antigenic sites Sa, Ca1, Ca2 and Cb region. Analysis of the antibody levels by hemagglutination inhibition test (HI) indicated that vaccinated individuals and people who had experienced a natural influenza A(H1N1)2009 virus infection showed good immune responses against the vaccine virus and most of the wild-type viruses. However, one to two amino acid changes in the antigenic site Sa dramatically affected the ability of antibodies to recognize these viruses. In contrast, the tested viruses were indistinguishable in regard to antibody recognition by the sera from elderly individuals who had been exposed to the Spanish influenza or its descendant viruses during the early 20(th) century. CONCLUSIONS: According to our results, one to two amino acid changes (N125D and/or N156K) in the major antigenic sites of the hemagglutinin of influenza A(H1N1)2009 virus may lead to significant reduction in the ability of patient and vaccine sera to recognize A(H1N1)2009 viruses.
Project description:The HA1 domain of HA, the major antigenic protein of influenza A viruses, contains all of the antigenic sites of HA and is under continual immune-driven selection. To resolve controversies on whether only a few or many residue sites of HA1 have undergone positive selection, whether positive selection at HA1 is continual or punctuated, and whether antigenic change is punctuated, we introduce an approach to analyze 2,248 HA1 sequences collected from 1968 to 2005. We identify 95 substitutions at 63 sites from 1968 to 2005 and show that each substitution occurred very rapidly. The rapid substitution and the fact that 57 of the 63 sites are antigenic sites indicate that hitchhiking plays a minor role and that most of these sites, many more than previously found, have undergone positive selection. Strikingly, 88 of the 95 substitutions occurred in groups, and multiple mutations at antigenic sites sped up the fixation process. Our results suggest that positive selection has been ongoing most of the time, not sporadic, and that multiple mutations at antigenic sites cumulatively enhance antigenic drift, indicating that antigenic change is less punctuated than recently proposed.