Mapping the Human Memory B Cell and Serum Neutralizing Antibody Responses to Dengue Virus Serotype 4 Infection and Vaccination

ABSTRACT The four dengue virus (DENV) serotypes are mosquito-borne flaviviruses responsible for dengue fever and dengue hemorrhagic fever. People exposed to DENV develop antibodies (Abs) that strongly neutralize the serotype responsible for infection. Historically, infection with DENV serotype 4 (DENV4) has been less common and less studied than infections with the other three serotypes. However, DENV4 has been responsible for recent large and sustained epidemics in Asia and Latin America. The neutralizing antibody responses and the epitopes targeted against DENV4 have not been characterized in human infection. In this study, we mapped and characterized epitopes on DENV4 recognized by neutralizing antibodies in people previously exposed to DENV4 infections or to a live attenuated DENV4 vaccine. To study the fine specificity of DENV4 neutralizing human antibodies, B cells from two people exposed to DENV4 were immortalized and screened to identify DENV-specific clones. Two human monoclonal antibodies (MAbs) that neutralized DENV4 were isolated, and their epitopes were finely mapped using recombinant viruses and alanine scan mutation array techniques. Both antibodies bound to quaternary structure epitopes near the hinge region between envelope protein domain I (EDI) and EDII. In parallel, to characterize the serum neutralizing antibody responses, convalescence-phase serum samples from people previously exposed to primary DENV4 natural infections or a monovalent DENV4 vaccine were analyzed. Natural infection and vaccination also induced serum-neutralizing antibodies that targeted similar epitope domains at the EDI/II hinge region. These studies defined a target of neutralizing antigenic site on DENV4 targeted by human antibodies following natural infection or vaccination. IMPORTANCE The four serotypes of dengue virus are the causative agents of dengue fever and dengue hemorrhagic fever. People exposed to primary DENV infections develop long-term neutralizing antibody responses, but these principally recognize only the infecting serotype. An effective vaccine against dengue should elicit long-lasting protective antibody responses to all four serotypes simultaneously. We and others have defined antigenic sites on the envelope (E) protein of viruses of dengue virus serotypes 1, 2, and 3 targeted by human neutralizing antibodies. The epitopes on DENV4 E protein targeted by the human neutralizing antibodies and the mechanisms of serotype 4 neutralization are poorly understood. Here, we report the properties of human antibodies that neutralize dengue virus serotype 4. People exposed to serotype 4 infections or a live attenuated serotype 4 vaccine developed neutralizing antibodies that bound to similar sites on the viral E protein. These studies have provided a foundation for developing and evaluating DENV4 vaccines.

Download Full-text

Tracking the polyclonal neutralizing antibody response to a dengue virus serotype 1 type-specific epitope across two populations in Asia and the Americas

Scientific Reports ◽

10.1038/s41598-019-52511-z ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 2

Author(s):

Daniela V. Andrade ◽

Colin Warnes ◽

Ellen Young ◽

Leah C. Katzelnick ◽

Angel Balmaseda ◽

...

Keyword(s):

Dengue Virus ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Sri Lankan ◽

Human Monoclonal Antibodies ◽

Virus Serotype ◽

Serotype 1 ◽

Dengue Virus Serotypes ◽

Kinetics Of ◽

Two Populations

Abstract The four dengue virus serotypes (DENV1-4) cause major public health problems worldwide. Highly neutralizing type-specific human monoclonal antibodies (hmAbs) target conformation-dependent epitopes on the DENV envelope protein, including 1F4, a DENV1 type-specific hmAb. Using a recombinant DENV2 virus displaying the DENV1 1F4 epitope (rDENV2/1), we measured the proportion and kinetics of DENV1 neutralizing antibodies targeting the 1F4 epitope in individuals living in Asia and the Americas where different DENV1 genotypes were circulating. Samples from 20 individuals were analyzed 3 and 18 months post-primary DENV1 infection, alongside samples from 4 individuals collected annually for four years post-primary DENV1 infection, from two studies in Nicaragua. We also analyzed convalescent post-primary DENV1 plasma samples from Sri Lankan individuals. We found that neutralizing antibodies recognizing the 1F4 epitope vary in prevalence across both populations and were detected from 20 days to four years post-infection. Additionally, both populations displayed substantial variability, with a range of high to low proportions of DENV1 type-specific neutralizing antibodies recognizing the 1F4 epitope seen across individuals. Thus, the 1F4 epitope is a major but not exclusive target of type-specific neutralizing antibodies post-primary infection with different DENV1 genotypes in Asia and Latin America, and additional epitopes likely contribute to type-specific neutralization of DENV1.

Download Full-text

Broad and potent neutralizing human antibodies to tick-borne flaviviruses protect mice from disease

Journal of Experimental Medicine ◽

10.1084/jem.20210236 ◽

2021 ◽

Vol 218 (5) ◽

Author(s):

Marianna Agudelo ◽

Martin Palus ◽

Jennifer R. Keeffe ◽

Filippo Bianchini ◽

Pavel Svoboda ◽

...

Keyword(s):

Antibody Response ◽

Neutralizing Antibodies ◽

Natural Infection ◽

Specific Treatment ◽

Neutralizing Antibody ◽

Hemorrhagic Fever ◽

Human Antibody ◽

Lateral Ridge ◽

Tick Borne Encephalitis Virus ◽

Domain Iii

Tick-borne encephalitis virus (TBEV) is an emerging human pathogen that causes potentially fatal disease with no specific treatment. Mouse monoclonal antibodies are protective against TBEV, but little is known about the human antibody response to infection. Here, we report on the human neutralizing antibody response to TBEV in a cohort of infected and vaccinated individuals. Expanded clones of memory B cells expressed closely related anti-envelope domain III (EDIII) antibodies in both groups of volunteers. However, the most potent neutralizing antibodies, with IC50s below 1 ng/ml, were found only in individuals who recovered from natural infection. These antibodies also neutralized other tick-borne flaviviruses, including Langat, louping ill, Omsk hemorrhagic fever, Kyasanur forest disease, and Powassan viruses. Structural analysis revealed a conserved epitope near the lateral ridge of EDIII adjoining the EDI–EDIII hinge region. Prophylactic or early therapeutic antibody administration was effective at low doses in mice that were lethally infected with TBEV.

Download Full-text

Elicitation of potent serum neutralizing antibody responses in rabbits by immunization with an HIV-1 clade C trimeric Env derived from an Indian elite neutralizer

10.1101/2020.09.15.297697 ◽

2020 ◽

Author(s):

Rajesh Kumar ◽

Suprit Deshpande ◽

Leigh M. Sewall ◽

Gabriel Ozorowski ◽

Christopher A. Cottrell ◽

...

Keyword(s):

Electron Microscopy ◽

Neutralizing Antibodies ◽

Natural Infection ◽

Neutralizing Antibody ◽

Antibody Responses ◽

Clade C ◽

Immunogen Design ◽

Env Protein ◽

Immunogenic Properties ◽

Hiv 1

AbstractEvaluating the structure-function relationship of viral envelope (Env) evolution and the development of broadly cross-neutralizing antibodies (bnAbs) in natural infection can inform rational immunogen design. In the present study, we examined the magnitude and specificity of autologous neutralizing antibodies induced in rabbits by a novel HIV-1 clade C Env protein (1PGE-THIVC) vis-à-vis those developed in an elite neutralizer from whom the env sequence was obtained that was used to prepare the soluble Env protein. The thermostable 1PGE-THIVC Env displayed a native like pre-fusion closed conformation in solution as determined by small angle X-ray scattering (SAXS) and negative stain electron microscopy (EM). This closed spike conformation of 1PGE-THIVC Env trimers was correlated with weak or undetectable binding of non-neutralizing monoclonal antibodies (mAbs) compared to neutralizing mAbs. Furthermore, 1PGE-THIVC SOSIP induced potent neutralizing antibodies in rabbits to autologous virus variants. The autologous neutralizing antibody specificity induced in rabbits by 1PGE-THIVC was mapped to the C3/V4 region (T362/P401) of viral Env. This observation agreed with electron microscopy polyclonal epitope mapping (EMPEM) of the Env trimer complexed with IgG Fab prepared from the immunized rabbit sera. While the specificity of antibodies elicited in rabbits associated with neutralizing autologous viruses were distinct to those developed in the elite neutralizer, EMPEM analysis demonstrated significant changes to Env conformations when incubated with polyclonal antibody sera from the elite neutralizer, suggesting these antibodies lead to the destabilization of Env trimers. Our study not only shows distinct mechanisms associated with potent neutralization of sequence matched and unmatched autologous viruses by antibodies induced in rabbits and in the elite neutralizer, but also highlights how neutralizing antibodies developed during the course of natural infection can impact viral Env conformations.Author SummaryThe interplay between circulating virus variants and broadly cross neutralizing polyclonal antibodies developed in a subset of elite neutralizers is widely believed to provide strategies for rational immunogen design. In the present study, we studied the structural, antigenic and immunogenic properties of a thermostable soluble trimeric protein with near native pre-fusion conformation prepared using the primary sequence of an HIV-1 clade C env isolated from the broadly cross neutralizing plasma of an elite neutralizer. This novel SOSIP Env trimer demonstrated comparable antigenic, structural and immunogenic properties that favoured several ongoing subunit vaccine design efforts. The novel clade C SOSIP induced polyclonal neutralizing antibody response developed in rabbits not only differed in its epitope specificity compared to that elicited in natural infection in presence of pool of viral quasispecies but also showed how they differ in their ability to influence Env structure and conformation. A better understanding of how vaccine-induced polyclonal neutralizing antibody responses compares to responses that developed in natural infection will improve our knowledge in designing better vaccine design strategies.

Download Full-text

Antibody responses induced by SHIV infection are more focused than those induced by soluble native HIV-1 envelope trimers in non-human primates

PLoS Pathogens ◽

10.1371/journal.ppat.1009736 ◽

2021 ◽

Vol 17 (8) ◽

pp. e1009736

Author(s):

Jelle van Schooten ◽

Marlies M. van Haaren ◽

Hui Li ◽

Laura E. McCoy ◽

Colin Havenar-Daughton ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Natural Infection ◽

Neutralizing Antibody ◽

Antibody Responses ◽

Broadly Neutralizing Antibodies ◽

Antibody Diversity ◽

Infected Infant ◽

Immunodeficiency Virus ◽

Shiv Infection ◽

Hiv 1

The development of an effective human immunodeficiency virus (HIV-1) vaccine is a high global health priority. Soluble native-like HIV-1 envelope glycoprotein trimers (Env), including those based on the SOSIP design, have shown promise as vaccine candidates by inducing neutralizing antibody responses against the autologous virus in animal models. However, to overcome HIV-1’s extreme diversity a vaccine needs to induce broadly neutralizing antibodies (bNAbs). Such bNAbs can protect non-human primates (NHPs) and humans from infection. The prototypic BG505 SOSIP.664 immunogen is based on the BG505 env sequence isolated from an HIV-1-infected infant from Kenya who developed a bNAb response. Studying bNAb development during natural HIV-1 infection can inform vaccine design, however, it is unclear to what extent vaccine-induced antibody responses to Env are comparable to those induced by natural infection. Here, we compared Env antibody responses in BG505 SOSIP-immunized NHPs with those in BG505 SHIV-infected NHPs, by analyzing monoclonal antibodies (mAbs). We observed three major differences between BG505 SOSIP immunization and BG505 SHIV infection. First, SHIV infection resulted in more clonal expansion and less antibody diversity compared to SOSIP immunization, likely because of higher and/or prolonged antigenic stimulation and increased antigen diversity during infection. Second, while we retrieved comparatively fewer neutralizing mAbs (NAbs) from SOSIP-immunized animals, these NAbs targeted more diverse epitopes compared to NAbs from SHIV-infected animals. However, none of the NAbs, either elicited by vaccination or infection, showed any breadth. Finally, SOSIP immunization elicited antibodies against the base of the trimer, while infection did not, consistent with the base being placed onto the virus membrane in the latter setting. Together these data provide new insights into the antibody response against BG505 Env during infection and immunization and limitations that need to be overcome to induce better responses after vaccination.

Download Full-text

Addition of Partial Envelope Domain II into Envelope Domain III of Dengue Virus Antigen Potentiates the Induction of Virus-Neutralizing Antibodies and Induces Protective Immunity

Vaccines ◽

10.3390/vaccines8010088 ◽

2020 ◽

Vol 8 (1) ◽

pp. 88 ◽

Cited By ~ 3

Author(s):

Jisang Park ◽

Hyun-Young Lee ◽

Ly Tuan Khai ◽

Nguyen Thi Thu Thuy ◽

Le Quynh Mai ◽

...

Keyword(s):

Dengue Virus ◽

Dengue Fever ◽

Neutralizing Antibodies ◽

Subunit Vaccine ◽

Secondary Infection ◽

Febrile Illness ◽

Hemorrhagic Fever ◽

Denv Infection ◽

Severe Dengue ◽

Domain Iii

Dengue virus (DENV) comprises four serotypes in the family Flaviviridae and is a causative agent of dengue-related diseases, including dengue fever. Dengue fever is generally a self-limited febrile illness. However, secondary infection of patients with a suboptimal antibody (Ab) response provokes life-threatening severe dengue hemorrhagic fever or dengue shock syndrome. To develop a potent candidate subunit vaccine against DENV infection, we developed the EDII-cEDIII antigen, which contains partial envelope domain II (EDII) including the fusion loop and BC loop epitopes together with consensus envelope domain III (cEDIII) of all four serotypes of DENV. We purified Ab from mice after immunization with EDII-cEDIII or cEDIII and compared their virus neutralization and Ab-dependent enhancement of DENV infection. Anti-EDII-cEDIII Ab showed stronger neutralizing activity and lower Ab-dependent peak enhancement of DENV infection compared with anti-cEDIII Ab. Following injection of Ab-treated DENV into AG129 mice, anti-EDII-cEDIII Ab ameliorated DENV infection in tissues with primary and secondary infection more effectively than anti-cEDIII Ab. In addition, anti-EDII-cEDIII Ab protected against DENV1, 2, and 4 challenge. We conclude that EDII-cEDIII induces neutralizing and protective Abs, and thus, shows promise as a candidate subunit vaccine for DENV infection.

Download Full-text

SARS-CoV-2 antibodies remain detectable 12 months after infection and antibody magnitude is associated with age and COVID-19 severity

10.1101/2021.04.27.21256207 ◽

2021 ◽

Author(s):

Eric D. Laing ◽

Nusrat J. Epsi ◽

Stephanie A. Richard ◽

Emily C. Samuels ◽

Wei Wang ◽

...

Keyword(s):

Symptom Onset ◽

Neutralizing Antibodies ◽

Natural Infection ◽

Geometric Mean ◽

Neutralizing Antibody ◽

Antibody Responses ◽

Serum Samples ◽

Military Hospitals ◽

Igg Binding ◽

One Year

ABSTRACTImportanceThe persistence of SARS-CoV-2 antibodies may be a predictive correlate of protection for both natural infections and vaccinations. Identifying predictors of robust antibody responses is important to evaluate the risk of re-infection / vaccine failure and may be translatable to vaccine effectiveness.ObjectiveTo 1) determine the durability of anti-SARS-CoV-2 IgG and neutralizing antibodies in subjects who experienced mild and moderate to severe COVID-19, and 2) to evaluate the correlation of age and IgG responses to both endemic human seasonal coronaviruses (HCoVs) and SARS-CoV-2 according to infection outcome.DesignLongitudinal serum samples were collected from PCR-confirmed SARS-CoV-2 positive participants (U.S. active duty service members, dependents and military retirees, including a range of ages and demographics) who sought medical treatment at seven U.S. military hospitals from March 2020 to March 2021 and enrolled in a prospective observational cohort study.ResultsWe observed SARS-CoV-2 seropositivity in 100% of inpatients followed for six months (58/58) to one year (8/8), while we observed seroreversion in 5% (9/192) of outpatients six to ten months after symptom onset, and 18% (2/11) of outpatients followed for one year. Both outpatient and inpatient anti-SARS-CoV-2 binding-IgG responses had a half-life (T1/2) of >1000 days post-symptom onset. The magnitude of neutralizing antibodies (geometric mean titer, inpatients: 378 [246-580, 95% CI] versus outpatients: 83 [59-116, 95% CI]) and durability (inpatients: 65 [43-98, 95% CI] versus outpatients: 33 [26-40, 95% CI]) were associated with COVID-19 severity. Older age was a positive correlate with both higher IgG binding and neutralizing antibody levels when controlling for COVID-19 hospitalization status. We found no significant relationships between HCoV antibody responses and COVID-19 clinical outcomes, or the development of SARS-CoV-2 neutralizing antibodies.Conclusions and RelevanceThis study demonstrates that humoral responses to SARS-CoV-2 infection are robust on longer time-scales, including those arising from milder infections.However, the magnitude and durability of the antibody response after natural infection was lower and more variable in younger participants who did not require hospitalization for COVID-19. These findings support vaccination against SARS-CoV-2 in all suitable populations including those individuals that have recovered from natural infection.

Download Full-text

A high-throughput cell- and virus-free assay shows reduced neutralization of SARS-CoV-2 variants by COVID-19 convalescent plasma

Science Translational Medicine ◽

10.1126/scitranslmed.abi8452 ◽

2021 ◽

pp. eabi8452

Author(s):

Craig Fenwick ◽

Priscilla Turelli ◽

Céline Pellaton ◽

Alex Farina ◽

Jérémy Campos ◽

...

Keyword(s):

High Throughput ◽

Neutralizing Antibodies ◽

Competitive Inhibition ◽

Natural Infection ◽

Neutralizing Antibody ◽

Antibody Responses ◽

Spike Protein ◽

Serum Samples ◽

Live Virus ◽

Protein Variants

The detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibodies in the serum of an individual indicates prior infection or vaccination. However, it provides limited insight into the protective nature of this immune response. Neutralizing antibodies recognizing the viral spike protein are more revealing, yet their measurement traditionally requires virus- and cell-based systems that are costly, time-consuming, inflexible, and potentially biohazardous. Here, we present a cell-free quantitative neutralization assay based on the competitive inhibition of trimeric SARS-CoV-2 spike protein binding to the angiotensin converting enzyme 2 (ACE2) receptor. This high-throughput method matches the performance of the gold standard live virus infection assay, as verified with a panel of 206 seropositive donors with varying degrees of infection severity and virus-specific IgG titers, achieving 96.7% sensitivity and 100% specificity. Furthermore, it allows for the parallel assessment of neutralizing activities against multiple SARS-CoV-2 spike protein variants of concern. We used our assay to profile serum samples from 59 patients hospitalized with coronavirus disease 2019 (COVID-19). We found that, although most sera had high activity against the 2019-nCoV parental spike protein and, to a lesser extent, the α (B.1.1.7) variant, only 58% of serum samples could efficiently neutralize a spike protein derivative containing mutations present in the β (B.1.351) variant. Thus, we have developed an assay that can evaluate effective neutralizing antibody responses to SARS-CoV-2 spike protein variants of concern after natural infection and that can be applied to characterize vaccine-induced antibody responses or to assess the potency of monoclonal antibodies.

Download Full-text

Immunogenicity of Multiple Doses of pDNA Vaccines against SARS-CoV-2

Pharmaceuticals ◽

10.3390/ph14010039 ◽

2021 ◽

Vol 14 (1) ◽

pp. 39

Author(s):

Iman Almansour ◽

Nabela Calamata Macadato ◽

Thamer Alshammari

Keyword(s):

Immune Responses ◽

Neutralizing Antibodies ◽

Natural Infection ◽

Neutralizing Antibody ◽

Serum Levels ◽

Antibody Responses ◽

Limited Data ◽

Effective Measure ◽

Vaccine Candidates ◽

Cellular Immune

Since its identification in Wuhan, China, in December 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has resulted in 46 million cases and more than one million deaths worldwide, as of 30 October 2020. Limited data exist on the magnitude and durability of antibodies generated by natural infection with SARS-CoV-2 and whether they can provide long-lasting immunity from reinfection. Vaccination has proven the most effective measure for controlling and preventing pandemics and, thus, development of a vaccine against COVID-19 is a top priority. However, the doses required to induce effective, long-lasting antibody responses against SARS-CoV-2 remain undetermined. Here, we present the development of SARS-CoV-2 vaccine candidates encoding the viral spike (S) gene, generated using plasmid (p)DNA technology, and we demonstrate the eliciting of S-specific antibodies in mice after three and four doses. The magnitude of binding and neutralizing antibody responses with three doses of synthetic, codon-optimized, full-length S (S.opt.FL) vaccine is comparable to that generated after four doses, suggesting that three doses are sufficient to elicit robust immune responses. Conversely, four doses of S1.opt pDNA vaccine, containing the S globular head, are required to elicit high levels of neutralizing antibodies. Furthermore, the S.opt.FL pDNA vaccine induces the highest serum levels of interferon (IFN)-γ, a marker for activation of cellular immune responses. Overall, our data show that three doses of S.FL pDNA vaccine elicit potent neutralizing antibody responses, with preclinical data that support the immunogenicity of these COVID-19 vaccine candidates and provide justification for further translational studies.

Download Full-text

The Chimeric Protein Domain III-Capsid of Dengue Virus Serotype 2 (DEN-2) Successfully Boosts Neutralizing Antibodies Generated in Monkeys upon Infection with DEN-2

Clinical and Vaccine Immunology ◽

10.1128/cvi.00382-10 ◽

2011 ◽

Vol 18 (3) ◽

pp. 455-459 ◽

Cited By ~ 17

Author(s):

Iris Valdés ◽

Lázaro Gil ◽

Yaremis Romero ◽

Jorge Castro ◽

Pedro Puente ◽

...

Keyword(s):

Dengue Virus ◽

Neutralizing Antibodies ◽

Chimeric Protein ◽

Neutralizing Antibody ◽

Protein Domain ◽

Homologous Virus ◽

Virus Serotype ◽

Attenuated Virus ◽

Domain Iii ◽

Dengue Virus Serotype 2

ABSTRACTUse of a heterologous prime-boost strategy based on a combination of nonreplicative immunogens and candidate attenuated virus vaccines against dengue virus in the same schedule is an attractive approach. These combinations may result in a condensed immunization regime for humans, thus reducing the number of doses with attenuated virus and the time spacing. The present work deals with the evaluation of the heterologous prime-boost strategy combining a novel chimeric protein (domain III-capsid) of dengue virus serotype 2 (DEN-2) and the infective homologous virus in the same immunization schedule in monkeys. Primed monkeys received one dose of infective DEN-2 and were then vaccinated with the recombinant protein. We found that animals developed a neutralizing antibody response after the infective dose and were notably boosted with a second dose of the chimeric protein 3 months later. The neutralizing antibodies induced were long lasting, and animals also showed the ability to induce a specific cellular response 6 months after the booster dose. As a conclusion, we can state that the domain III region, when it is properly presented as a fusion protein to the immune system, is able to recall the neutralizing antibody response elicited following homologous virus infection in monkeys. Further prime-boost approaches can be performed in a condensed regime combining the chimeric domain III-capsid protein and candidate live attenuated vaccines against DEN-2.

Download Full-text

A New Quaternary Structure Epitope on Dengue Virus Serotype 2 Is the Target of Durable Type-Specific Neutralizing Antibodies

mBio ◽

10.1128/mbio.01461-15 ◽

2015 ◽

Vol 6 (5) ◽

Cited By ~ 55

Author(s):

E. N. Gallichotte ◽

D. G. Widman ◽

B. L. Yount ◽

W. M. Wahala ◽

A. Durbin ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Dengue Virus ◽

Neutralizing Antibodies ◽

Quaternary Structure ◽

Natural Infection ◽

Viral Envelope ◽

Protective Response ◽

Denv Serotypes ◽

Virus Serotype ◽

Dengue Virus Serotype 2

ABSTRACT Dengue virus serotype 2 (DENV2) is widespread and responsible for severe epidemics. While primary DENV2 infections stimulate serotype-specific protective responses, a leading vaccine failed to induce a similar protective response. Using human monoclonal antibodies (hMAbs) isolated from dengue cases and structure-guided design of a chimeric DENV, here we describe the major site on the DENV2 envelope (E) protein targeted by neutralizing antibodies. DENV2-specific neutralizing hMAb 2D22 binds to a quaternary structure epitope. We engineered and recovered a recombinant DENV4 that displayed the 2D22 epitope. DENV2 neutralizing antibodies in people exposed to infection or a live vaccine tracked with the 2D22 epitope on the DENV4/2 chimera. The chimera remained sensitive to DENV4 antibodies, indicating that the major neutralizing epitopes on DENV2 and -4 are at different sites. The ability to transplant a complex epitope between DENV serotypes demonstrates a hitherto underappreciated structural flexibility in flaviviruses, which could be harnessed to develop new vaccines and diagnostics. IMPORTANCE Dengue virus causes fever and dengue hemorrhagic fever. Dengue serotype 2 (DENV2) is widespread and frequently responsible for severe epidemics. Natural DENV2 infections stimulate serotype-specific neutralizing antibodies, but a leading DENV vaccine did not induce a similar protective response. While groups have identified epitopes of single monoclonal antibodies (MAbs), the molecular basis of DENV2 neutralization by polyclonal human immune sera is unknown. Using a recombinant DENV displaying serotype 2 epitopes, here we map the main target of DENV2 polyclonal neutralizing antibodies induced by natural infection and a live DENV2 vaccine candidate. Proper display of the epitope required the assembly of viral envelope proteins into higher-order structures present on intact virions. Despite the complexity of the epitope, it was possible to transplant the epitope between DENV serotypes. Our findings have immediate implications for evaluating dengue vaccines in the pipeline as well as designing next-generation vaccines.

Download Full-text