Virus Hemagglutinin Research Articles

In silico prediction of influenza vaccine effectiveness by sequence analysis

The effectiveness of seasonal influenza vaccines varies with the matching of vaccine strains to circulating strains. Based on the genetic distance of hemagglutinin and neuraminidase gene of the influenza viruses to vaccine strains, we statistically quantified the relationship between the genetic mismatch and vaccine effectiveness (VE) for influenza A/H1N1pdm09, A/H3N2 and B. We also proposed a systematic approach to integrate multiple genes and influenza types for overall VE estimation. Evident linear relationships were identified and validated in independent data. The modelling framework may enable in silico prediction for VE on a real-time basis and inform the influenza vaccine selection strategy.

Repositioning Drugs on Human Influenza A Viruses Based on a Novel Nuclear Norm Minimization Method.

Influenza A viruses, especially H3N2 and H1N1 subtypes, are viruses that often spread among humans and cause influenza pandemic. There have been several big influenza pandemics that have caused millions of human deaths in history, and the threat of influenza viruses to public health is still serious nowadays due to the frequent antigenic drift and antigenic shift events. However, only few effective anti-flu drugs have been developed to date. The high development cost, long research and development time, and drug side effects are the major bottlenecks, which could be relieved by drug repositioning. In this study, we proposed a novel antiviral Drug Repositioning method based on minimizing Matrix Nuclear Norm (DRMNN). Specifically, a virus-drug correlation database consisting of 34 viruses and 205 antiviral drugs was first curated from public databases and published literature. Together with drug similarity on chemical structure and virus sequence similarity, we formulated the drug repositioning problem as a low-rank matrix completion problem, which was solved by minimizing the nuclear norm of a matrix with a few regularization terms. DRMNN was compared with three recent association prediction algorithms. The AUC of DRMNN in the global fivefold cross-validation (fivefold CV) is 0.8661, and the AUC in the local leave-one-out cross-validation (LOOCV) is 0.6929. Experiments have shown that DRMNN is better than other algorithms in predicting which drugs are effective against influenza A virus. With H3N2 as an example, 10 drugs most likely to be effective against H3N2 viruses were listed, among which six drugs were reported, in other literature, to have some effect on the viruses. The protein docking experiments between the chemical structure of the prioritized drugs and viral hemagglutinin protein also provided evidence for the potential of the predicted drugs for the treatment of influenza.

Establishment of a genetically engineered chicken DF-1 cell line for efficient amplification of influenza viruses in the absence of trypsin

BackgroundThe initial step of influenza infection is binding of the virus to specific sialic acid receptors expressed by host cells. This is followed by cell entry via endocytosis. Cleavage of the influenza virus hemagglutinin (HA) protein is critical for infection; this is performed by host cell proteases during viral replication. In cell culture systems, HA is cleaved by trypsin added to the culture medium. The vast majority of established cell lines are mammalian.ResultsIn the present study, we generated genetically engineered chicken DF-1 cell lines overexpressing transmembrane protease, serine 2 (TMPRSS2, which cleaves HA), ST3 beta-galactoside alpha-2,3-sialyltransferase 1 (ST3GAL1, which plays a role in synthesis of α-2,3 linked sialic acids to which avian-adapted viruses bind preferentially), or both. We found that overexpression of TMPRSS2 supports the virus life cycle by cleaving HA. Furthermore, we found that overexpression of ST3GAL1 increased the viral titer. Finally, we showed that overexpression of both TMPRSS2 and ST3GAL1 increased the final viral titer due to enhanced support of viral replication and prolonged viability of the cells. In addition, overexpression of these genes of interest had no effect on cell proliferation and viability.ConclusionsTaken together, the results indicate that these engineered cells could be used as a cell-based system to propagate influenza virus efficiently in the absence of trypsin. Further studies on influenza virus interactions with chicken cell host factors could be studied without the effect of trypsin on cells.

On the Path Toward a Universal Influenza Vaccine

Inactivated influenza vaccines target the head domain of the viral hemagglutinin (HA), a highly mutable region that allows the virus to escape

Comparative analysis of sialic acid α2–3/6 galactose glycan-binding proteins in human saliva and serum

It has been discovered that SAα2–6 Gal glycans prevailingly distribute in human upper respiratory tract while a few SAα2–3 Gal glycans distribute in human lower respiratory tract. In this study, we aimed to separate and identify SAα2–3/6 Gal glycan-binding proteins (GBPs) in human saliva and serum, since that these GBPs may possess anti-influenza A virus (anti-IAVs) protective effects by binding to receptor glycoconjugates earlier than virus hemagglutinin (HA). By using SAα2–3/6 Gal glycan-magnetic particles complexes, a total of 116 GBPs (33 SAα2–3 Gal GBPs, S3GBPs; 22 SAα2–6 Gal GBPs, S6GBPs; and 61 SAα2–3/6 Gal GBPs, SGBPs) in saliva and 144 GBPs (29 S3GBPs, 27 S6GBPs and 88 SGBPs) in serum were identified. Through emPAI analysis, for 61 saliva SGBPs, 16.39% showed higher extracting amount by SAα2–3 Gal, and 57.38% showed higher extracting amount by SAα2–6 Gal; for 88 serum SGBPs, only 2.27% showed higher extracting amount by SAα2–3 Gal, and up to 89.77% showed higher extracting amount by SAα2–6 Gal. A computational docking approach verified the docking between cystatin/apolipoprotein A-I and SAα2–3/6 Gal. Our study may supply useful information to help understand the nature protective role of saliva, and the anti-IAVs double effects of proteins that simultaneously play sialoglycoprotein and SAα2–3/6 Gal GBP roles.

Development of Neutralizing Nanobodies to the Hemagglutinin Stem Domain of Influenza A Viruses.

The influenza virus infection claims ~650,000 lives annually. Taking into account the evolving resistance of the pathogen to antiviral drugs and the waning effectiveness of vaccination among certain populations, new approaches to the treatment of influenza are needed. The current study is aimed at obtaining single-domain antibodies (Nanobodies®) to the highly conserved stem domain of influenza A virus hemagglutinin by phage display. Two high-affinity neutralizing clones of Nanobodies® with a particular specificity were selected; they ensured 100% neutralization of the H1N1 and H5N2 influenza viruses in vivo. The obtained data demonstrate that it is possible to develop highly effective VHH-based drugs for the treatment of influenza.

Peculiarities of the Presentation of the Encephalitogenic MBP Peptide by HLA-DR Complexes Providing Protection and Predisposition to Multiple Sclerosis.

Predisposition to multiple sclerosis (MS), a chronic autoimmune disease of the central nervous system, is due to various factors. The genetic component is considered one of the most important factors. HLA class II genes contribute the most to the development of MS. The HLA-DRB1*15 allele group is considered one of the main genetic risk factors predisposing to MS. The group of HLA-DRB1*01 alleles was shown to have a protective effect against this disease in the Russian population. In this work, we compared the binding of the encephalitogenic fragment of the myelin basic protein (MBP) to two HLA-DR complexes that provide protection against and predisposition to MS: HLA-DR1 (HLA-DRB1*0101) and HLA-DR15 (HLA-DRB1*1501), respectively. We found that the myelin peptide MBP88-100 binds to HLA-DR1 at a rate almost an order of magnitude lower than the viral peptide of hemagglutinin (HA). The same was true for the binding of MBP85-97 to HLA-DR15 in comparison with viral pp65. The structure of the C-terminal part of the peptide plays a key role in the binding to HLA-DR1 for equally high-affinity N-terminal regions of the peptides. The IC50 of the myelin peptide MBP88-100 competing with viral HA for binding to HLA-DR1 is almost an order of magnitude higher than that of HA. As for HA, the same was also true for the binding of MBP85-97 to HLA-DR15 in comparison with viral pp65. Thus, autoantigenic MBP cannot compete with the viral peptide for binding to protective HLA-DR1. However, it is more competitive than viral peptide for HLA-DR15.

Mutations in the genome of avian influenza viruses of the H1 and H5 subtypes responsible for adaptation to mammals

Avian influenza viruses of the H1 and H5 subtypes were involved in the formation of highly pathogenic viruses that caused pandemics and panzootics in the 20th–21st centuries. In order to assess the zoonotic potential of viruses of these subtypes, two viruses of the H1N1 and H5N3 subtypes have been isolated from wild ducks in Moscow and adapted for growth in mouse lungs. Their phenotypic properties were studied, and the genetic changes that occurred during adaptation were identified. The original A/duck/Moscow/4970/2013 (H1N1) and A/duck/Moscow/4182-C/2010 (H5N3) viruses were apathogenic for mice but became pathogenic after 7–10 passages in mouse lungs. Complete genome sequencing revealed 2 amino acid substitutions in the proteins of the H1N1 mouse-adapted variant (Glu627Lys in PB2 and Asp35Asn in hemagglutinin (HA) – numbering according to H3) and 6 mutations in the proteins of H5N3 virus (Glu627lys in PB2, Val113Ala in PB1, Ser82Pro in PB1-F2, Lys52Arg in HA2, Arg65Lys in NP, and Ser-59Ile in NA). The increase in virulence is most likely due to a Glu627Lys substitution in the protein PB2 found in both viruses. The replacement Asp35Asn in HA of the mouse-adapted H1N1 virus is associated with an increase in the pH value of the HA transition to 5.5 versus 5.0 for that of the wild virus. The mutations found in the HA, NA, and PB1-F2 proteins of the adapted H5N3 variant are unique. The mutations Glu627Lys in PB2, Arg65Lys in NP, and Val113Ala in PB1 are most likely host adaptive.

Длительное переживание вирусов гриппа А в зоокультурах водных беспозвоночных

In experiments to infect aquatic invertebrates in the zooculture, we used influenza A viruses, namely, to infect crustaceans Daphnia magna Straus, 1826 – human influenza virus, Hong Kong strain 1569/79 (H3N2), and to infect molluscs Anodonta cygnea Linné, 1758 – influenza virus A birds, Strain Rostok 1/34 (Hav1Neq1) – the so-called true bird plague virus. As a result of a series of experiments, found that influenza A viruses persist in the water for no more than 3 days, while in the gills and mantle of molluscs the virus is isolated on chicken embryos for at least another 35 days after contact with virus-containing water (a total of 70 individuals were studied). From the body Daphnia magna, to isolate the human influenza A virus on chicken embryos was possible within 14 days after infection through water (examined 6,800 individuals), by the method of immunofluorescence the influenza virus was determined in the intestines of crustaceans during the entire period of observation – 70 days from the time of infection. Influenza A viruses do not have a harmful effect on crustaceans or molluscs, infected animals also develop and reproduce, as well as individuals of control groups. Interesting is the fact that we have established the possibility of the loss of agglutination of red blood cells of chickens as a result of the reproduction of the human influenza A virus in the body of invertebrate Daphnia magna, which indicates a change in the viral protein hemagglutinin. The use of aquatic invertebrate zooculture can help in the study of the circulation of influenza A viruses in nature, as well as in the study of the variability of influenza A viruses.

An automated detection of influenza virus based on 3-D magnetophoretic separation and magnetic label.

Automated detection of the influenza virus is important for the prevention of infectious viruses. Herein, assisted by three-dimensional (3-D) magnetophoretic separation and magnetic label, an automated detection device was constructed for H7N9 influenza virus hemagglutinin. Multi-layer glass slides were used to generate a 3-D microchannel network with two-level channels, realizing 3-D magnetophoretic separation with a magnetic field in the vertical direction to microchannels for the sample treatment. After the immunomagnetic separation, a magnetic-tagged complex was captured in an antibody-modified glass capillary, where magnetic beads further as a label could cause the voltage change of the miniature tube liquid sensor to obtain the detection signal. Moreover, the whole detection process and detection results were controlled and read through a liquid crystal display (LCD) screen to improve the automation. Finally, the detection limit was calculated to be 8.4 ng mL-1 for H7N9 hemagglutinin and had good specificity and reproducibility. These results indicate that this detection device proposes promising automated avenues for the early detection of infectious diseases.

Altered NKp46 Recognition and Elimination of Influenza B Viruses

Every year, millions of people worldwide are infected with influenza, causing enormous health and economic problems. The most common type of influenza is influenza A. It is known that Natural Killer (NK) cells play an important role in controlling influenza A infection, mostly through the recognition of the viral protein hemagglutinin (HA) by the activating receptor, NKp46. In contrast, little is known regarding NK cell recognition of influenza B viruses, even though they are responsible for a third of all pediatric influenza deaths and are therefore included in the seasonal vaccine each year. Here we show that NKp46 also recognizes influenza B viruses. We show that NKp46 binds the HA protein of influenza B in a sialic acid-dependent manner, and identified the glycosylated residue in NKp46, which is critical for this interaction. We discovered that this interaction has a binding affinity approximately seven times lower than NKp46 binding of influenza A’s HA. Finally, we demonstrated, using mice deficient for the mouse orthologue of NKp46, named NCR1, that NKp46 is not important for influenza B elimination. These findings enable us to better understand the interactions between the different influenza viruses and NK cells that are known to be crucial for viral elimination.

Glycosylation generates an efficacious and immunogenic vaccine against H7N9 influenza virus.

Zoonotic avian influenza viruses pose severe health threats to humans. Of several viral subtypes reported, the low pathogenic avian influenza H7N9 virus has since February 2013 caused more than 1,500 cases of human infection with an almost 40% case-fatality rate. Vaccination of poultry appears to reduce human infections. However, the emergence of highly pathogenic strains has increased concerns about H7N9 pandemics. To develop an efficacious H7N9 human vaccine, we designed vaccine viruses by changing the patterns of N-linked glycosylation (NLG) on the viral hemagglutinin (HA) protein based on evolutionary patterns of H7 HA NLG changes. Notably, a virus in which 2 NLG modifications were added to HA showed higher growth rates in cell culture and elicited more cross-reactive antibodies than did other vaccine viruses with no change in the viral antigenicity. Developed into an inactivated vaccine formulation, the vaccine virus with 2 HA NLG additions exhibited much better protective efficacy against lethal viral challenge in mice than did a vaccine candidate with wild-type (WT) HA by reducing viral replication in the lungs. In a ferret model, the 2 NLG-added vaccine viruses also induced hemagglutination-inhibiting antibodies and significantly suppressed viral replication in the upper and lower respiratory tracts compared with the WT HA vaccines. In a mode of action study, the HA NLG modification appeared to increase HA protein contents incorporated into viral particles, which would be successfully translated to improve vaccine efficacy. These results suggest the strong potential of HA NLG modifications in designing avian influenza vaccines.

The cytoplasmic tail of influenza A/H1N1 virus hemagglutinin is β-structural

Influenza A/H1N1 virus hemagglutinin (HA) is an integral type I glycoprotein that contains a large glycosylated ectodomain, a transmembrane domain, and a cytoplasmic tail (CT) of 10-14 amino acid residues. There are absolutely no data on the secondary or tertiary structure of the HA CT, which is important for virus pathogenesis. Three highly conserved cysteines are post-translationally modified by the attachment of fatty acid residues that pin the CT to the lipid membrane inside the virion. We applied circular dichroism (CD) and fluorescence spectroscopy analysis to examine four synthetic peptides corresponding to 14-15 C-terminal residues of H1 subtype HA (NH2-WMCSNGSLQCRICI-COOH; NH2-FWMCSNGSLQCRICI-COOH), with free or acetaminomethylated cysteines, in the reduced or non-reduced state, at various pH values and temperatures. The CD analysis detected the formation of a β-structure (30–65% according to the new BeStSel algorithm), in addition to an unstructured random coil, in every peptide in various conditions. It was completely or partially recognized as an antiparallel β-structure that was also confirmed by the multi-bounce Horizontal Attenuated Total Reflectance Fourier Transformed Infrared (HATR-FTIR) spectroscopy analysis. According to the experimental data, as well as 3 D modeling, we assume that the amino acid sequence corresponding to the HA CT may form a short antiparallel β-structure under the lipid membrane within a virion. Communicated by Ramaswamy H. Sarma

Antibodies That Engage the Hemagglutinin Receptor-Binding Site of Influenza B Viruses.

We describe cross-reactive human antibodies recognizing influenza B viruses spanning nearly 80 years of antigenic drift. Structures show that they engage the receptor-binding site (RBS) of the viral hemagglutinin with strong similarities to their influenza A counterparts, despite structural differences between the RBS of influenza A and B. Our data show that these antibodies readily cross-react with both influenza B Victoria and Yamagata lineages. We also note that all antibodies are encoded by IGHV3-9/IGK1-33. Future research will provide insight into the prevalence of these antibodies in the human population.

Perspectives of treatment of influenza and influenza-like illnesses with Enisamium yodide – Antiviral with anti-inflammatory effect

Background: Enisamium iodide (EI) is a Russian drug whose molecule was developed in the Institute of Pharmacology and Toxicology of Ukraine as an anti-inflammatory and antipyretic agent. Later, other properties of EI were found: inhibition of the influenza virus hemagglutinin, increasing the production of interferon-γ, level of Th1, blood antioxidant activity, inhibition TNF-α and other pro-inflammatory cytokines, activation of macrophages and lysozyme activity. The aim of study was to evaluate the efficacy of antiviral therapy with EI in out-patient patients with influenza and influenza-like illnesses (ILI) without risk factors for severe course of disease. Methods and materials: 124 patients aged 18–55 years with influenza and ILI within 6–48 h of symptoms onset were randomized into 2 groups. 1st group (n = 66) was treated with EI 500 mg tid 5 days, 2nd group (n = 63) received placebo according to the same scheme. In the 1nd group the proportion between influenza and other respiratory viruses was 32.2%: 67.8%, in the 2nd group 31.7%: 68.3%, accordingly. Duration of disease and main clinical symptoms, frequency of complications and speed of the virus elimination from the nasopharynx were estimated. Results: The number of cases with complete recovery within 96 h was 56 (84.8% ± 4.2%) in 1st group and 44 (69.8% ± 5.8%) in 2nd group (p = 0.047). The duration of fever was 68.0 ± 2.8 h in 1th group and 77.1 ± 2.9 h in 2nd group (p = 0.044), muscle pain – 52.3 ± 2.3 and 60.6 ± 2.6 h (p = 0.041), headache-52.1 ± 2.6 and 65.1 ± 2.5 h (p = 0.032), weakness – 75.5 ± 2.3 and 90.4 ± 2.5 h (p = 0.001), accordingly. On the 4th day of treatment the viruses were isolated in 37.8 ± 5.9% of cases in the 1st group and 57.1 ± 6.2% in 2nd group; p = 0.048. Bacterial complications were observed in 3.0 ± 2.1% and 12.6 ± 4.2% accordingly, p = 0.048. Conclusion: The study is demonstrated the effectiveness of EI in the treatment of influenza and ILI in adult outpatient patients. The antiviral and anti-inflammatory effects of the drug, which was administered within 48 h of the symptoms onset were demonstrated by a more rapid reduction of clinical symptoms, frequency of bacterial complications and reduction in the time of virus elimination. Continuation of EI clinical trials are necessary for more full evaluation of its effectiveness.

Structural intermediates in the low pH-induced transition of influenza hemagglutinin.

The hemagglutinin (HA) glycoproteins of influenza viruses play a key role in binding host cell receptors and in mediating virus-host cell membrane fusion during virus infection. Upon virus entry, HA is triggered by low pH and undergoes large structural rearrangements from a prefusion state to a postfusion state. While structures of prefusion state and postfusion state of HA have been reported, the intermediate structures remain elusive. Here, we report two distinct low pH intermediate conformations of the influenza virus HA using cryo-electron microscopy (cryo-EM). Our results show that a decrease in pH from 7.8 to 5.2 triggers the release of fusion peptides from the binding pockets and then causes a dramatic conformational change in the central helices, in which the membrane-proximal ends of the central helices unwind to an extended form. Accompanying the conformational changes of the central helices, the stem region of the HA undergoes an anticlockwise rotation of 9.5 degrees and a shift of 15 Å. The HA head, after being stabilized by an antibody, remains unchanged compared to the neutral pH state. Thus, the conformational change of the HA stem region observed in our research is likely to be independent of the HA head. These results provide new insights into the structural transition of HA during virus entry.

Hemagglutinating properties of bovine parainfluenza-3, coronavirus and rotavirus infections’ pathogens in cattle

The purpose of the work was to determine the hemagglutinating activity of different strains of bovine parainfluenza-3, coronavirus and rotavirus infections’ pathogens in cattle. Accumulation of hemagglutinins of viruses was carried out by infecting cell cultures with viruses, determining their infectious activity by cytopathic action, followed by the establishment of hemagglutinating activity with erythrocytes of different species of animals. It has been established that within one type of virus there may be strains that have different hemagglutinating activity. It has been proved that within one type of virus there may be strains that have different hemagglutinating activity. It has been established that for the detection of coronavirus hemagglutinins mouse erythrocytes are the most suitable, bovine parainfluenza-3 virus hemagglutinins — guinea pig erythrocytes, rotavirus hemagglutinins — rooster erythrocytes. The relationship between the infectious activity of parainfluenza-3 virus, corona-, rotaviruses and their hemagglutinating properties has been established — the highest hemagglutinin titer was observed under the conditions of infectious activity of bovine parainfluenza-3 virus, corona-, rotaviruses in 7.0 lg TCD50/cm3 and higher. Long-term storage of bovine parainfluenza-3, coronavirus and rotavirus infections’ pathogens at a temperature of minus 18 °С and lower did not have a negative effects on their hemagglutinating properties, in contrast to the storage of these viruses at a temperature of minus 4 °С or repeated defrosting

Development and biochemical characterization of the monoclonal antibodies for specific detection of the emerging H5N8 and H5Nx avian influenza virus hemagglutinins.

The highly pathogenic avian influenza (HPAI) H5N8 virus has been detected in wild birds and poultry worldwide. The threat caused by HPAI H5N8 virus still exists with concerns for human infection. The preparedness for epidemic prevention and decreasing the agricultural and economic lost is extremely important. Hemagglutinin (HA), a surface glycoprotein of influenza viruses, is considered as the major target for detection of the influenza virus subtype in the infected samples. In this study, the recombinant H5N8 HA1 and HA2 proteins were expressed in Escherichia coli, and were utilized to generate two monoclonal antibodies, named 7H6C and YC8. 7H6C can bind the HA proteins of H5N1 and H5N8, but cannot bind the HA proteins of H1N1, H3N2, and H7N9, indicating that it has H5-subtype specificity. In contrast, YC8 can bind the HA proteins of H1N1, H5N1, and H5N8, but cannot bind the HA proteins of H3N2 and H7N9, indicating that it has H1-subtype and H5-subtype specificity. The epitope sequences recognized by 7H6C are located in the head domain of H5N8 HA, and are highly conserved in H5 subtypes. The epitope sequences recognized by YC8 are located in the stalk domain of H5N8 HA, and are highly conserved among the H1 and H5 subtypes. 7H6C and YC8 can be applied for specific detection of the HA proteins of H5N8 and H5Nx avian influenza viruses. KEY POINTS: • The mAb 7H6C or YC8 was generated by using the HA1 or HA2 of the HPAI H5N8 virus as the immunogen. • 7H6C recognized the head domain of H5N8 HA, and YC8 recognized the stalk domain of H5N8 HA. • 7H6C and YC8 can detect the HA proteins of H5Nx subtypes specifically.

Mechanisms of Antiviral Activity of Cistus Salviifolius Extract Against Human Respiratory Viruses

Influenza and ARVIs are the most common forms of infectious respiratory diseases in humans. In this regard, the search and development of means for the prevention and treatment of viral infections is a high priority task. The aim of this study was to assess the mechanisms of the antiviral activity of sage-leaved rock-rose extract (Cistus salviifolius) against the causative agents of influenza and ARVIs in humans. In the course of the study, it was shown that C.salviifolius extract inhibits reproduction of influenza viruses A(H1N1), A (H1N1)pdm09, A(H3N2), A(H5N2), A(H7N9) and influenza B virus. The extract showed maximum virus-inhibiting activity at the early stages of the viral cycle (0–2 hours after infection). C.salviifolius extract significantly reduced the hemagglutinating activity of the virus, and at the same time did not affect the fusogenic properties of viral hemagglutinin. Transmission electron microscopy was used to demonstrate that the cistus extract prevents the absorption of influenza virions on the surface of cells in culture. The inhibitory activity of the extract against other human respiratory viruses, parainfluenza virus and adenovirus, was also shown. The protective activity of C.salviifolius extract was demonstrated when applied intranasally during the experiments on a model of influenza pneumonia in mice. The degree of this activity was in inverse proportion to the time window between the application of the extract and the infection of the animals. The virus, pre-incubated with C.salviifolius extract, did not cause death in the animals. The data obtained indicate that C.salviifolius extract serves as an effective and broad-range means of preventing respiratory viral infections in humans.

Characterization of Novel Cross-Reactive Influenza B Virus Hemagglutinin Head Specific Antibodies That Lack Hemagglutination Inhibition Activity.

Humoral immune responses to influenza virus vaccines in elderly individuals are poorly adapted toward new antigenically drifted influenza virus strains. Instead, older individuals respond in an original antigenic sin fashion and produce much more cross-reactive but less potent antibodies. Here, we investigated four influenza B virus hemagglutinin (HA) head specific, hemagglutination inhibition-inactive monoclonal antibodies (MAbs) from elderly individuals. We found that they were broadly reactive within the B/Victoria/2/1987-like lineage, and two were highly cross-reactive with B/Yamagata/16/1988-like lineage viruses. The MAbs were found to be neutralizing, to utilize Fc effector functions, and to be protective against lethal viral challenge in a mouse model. In order to identify residues on the influenza B virus hemagglutinin interacting with the MAbs, we generated escape mutant viruses. Interestingly, escape from these MAbs led to numerous HA mutations within the head domain, including in the defined antigenic sites. We observed that each individual escape mutant virus was able to avoid neutralization by its respective MAb along with other MAbs in the panel, although in many cases binding activity was maintained. Point mutant viruses indicated that K90 is critical for the neutralization of two MAbs, while escape from the other two MAbs required a combination of mutations in the hemagglutinin. Three of four escape mutant viruses had increased lethality in the DBA2/J mouse model. Our work indicates that these cross-reactive antibodies have the potential to cause antigenic drift in the viral population by driving mutations that increase virus fitness. However, binding activity and cross-neutralization were maintained by a majority of antibodies in the panel, suggesting that this drift may not lead to escape from antibody-mediated protection.IMPORTANCE Understanding the immune response that older individuals mount to influenza virus vaccination and infection is critical in order to design better vaccines for this age group. Here, we show that older individuals make broadly neutralizing antibodies that have no hemagglutination-inhibiting activity and are less potent than strain-specific antibodies. These antibodies could drive viral escape from neutralization but did not result in escape from binding. Given their different mechanisms of action, they might retain protective activity even against escape variants.