The four serotypes of dengue virus (DENV) cause the most important and rapidly emerging arboviral diseases in humans. The recent phase 2b and 3 studies of a tetravalent dengue vaccine reported a moderate efficacy despite the presence of neutralizing antibodies, highlighting the need for a better understanding of neutralizing antibodies in polyclonal human sera. Certain type-specific (TS) antibodies were recently discovered to account for the monotypic neutralizing activity and protection after primary DENV infection. The nature of neutralizing antibodies after secondary DENV infection remains largely unknown. In this study, we examined sera from 10 vaccinees with well-documented exposure to first and second DENV serotypes through heterotypic immunization with live-attenuated vaccines. Higher serum IgG avidities to both exposed and nonexposed serotypes were found after secondary immunization than after primary immunization. Using a two-step depletion protocol to remove different anti-envelope antibodies, including group-reactive (GR) and complex-reactive (CR) antibodies separately, we found GR and CR antibodies together contributed to more than 50% of neutralizing activities against multiple serotypes after secondary immunization. Similar findings were demonstrated in patients after secondary infection. Anti-envelope antibodies recognizing previously exposed serotypes consisted of a large proportion of GR antibodies, CR antibodies, and a small proportion of TS antibodies, whereas those recognizing nonexposed serotypes consisted of GR and CR antibodies. These findings have implications for sequential heterotypic immunization or primary immunization of DENV-primed individuals as alternative strategies for DENV vaccination. The complexity of neutralizing antibodies after secondary infection provides new insights into the difficulty of their application as surrogates of protection. IMPORTANCEThe four serotypes of dengue virus (DENV) are the leading cause of arboviral diseases in humans. Despite the presence of neutralizing antibodies, a moderate efficacy was recently reported in phase 2b and 3 trials of a dengue vaccine; a better understanding of neutralizing antibodies in polyclonal human sera is urgently needed. We studied vaccinees who received heterotypic immunization of live-attenuated vaccines, as they were known to have received the first and second DENV serotype exposures. We found anti-envelope antibodies consist of group-reactive (GR), complex-reactive (CR), and type-specific (TS) antibodies, and that both GR and CR antibodies contribute significantly to multitypic neutralizing activities after secondary DENV immunization. These findings have implications for alternative strategies for DENV vaccination. Certain TS antibodies were recently discovered to contribute to the monotypic neutralizing activity and protection after primary DENV infection; our findings of the complexity of neutralizing activities after secondary immunization/infection provide new insights for neutralizing antibodies as surroga...

The levels of neutralizing antibody to a pathogen are an effective indicator to predict efficacy of a vaccine in trial. And yet not all the trial vaccines are in line with the theory. Using dengue virus (DENV) to investigate the viral morphology affecting the predictive value, we evaluated the viral morphology in acute dengue plasma compared to that of Vero cells derived DENV. The virions in plasma were infectious and heterogeneous in shape with a “sunny-side up egg” appearance, viral RNA was enclosed with CD61+ cell-derived membrane interspersed by the viral envelope protein, defined as dengue vesicles. The unique viral features were also observed from ex vivo infected human bone marrow. Dengue vesicles were less efficiently neutralized by convalescent patient serum, compared to virions produced from Vero cells. Our results exhibit a reason why potencies of protective immunity fail in vivo and significantly impact dengue vaccine and drug development.

Dengue is becoming recognized as one of the most important vector-borne human diseases. It is predominant in tropical and subtropical zones but its geographical distribution is progressively expanding, making it an escalating global health problem of today. Dengue presents with spectrum of clinical manifestations, ranging from asymptomatic, undifferentiated mild fever, dengue fever (DF), to dengue hemorrhagic fever (DHF) with or without shock (DSS), a life-threatening illness characterized by plasma leakage due to increased vascular permeability. Currently, there are no antiviral modalities or vaccines available to treat and prevent dengue. Supportive care with close monitoring is the standard clinical practice. The mechanisms leading to DHF/DSS remains poorly understood. Multiple factors have been attributed to the pathological mechanism, but only a couple of these hypotheses are popular in scientific circles. The current discussion focuses on underappreciated factors, temperature, natural IgM, and endotoxin, which may be critical components playing roles in dengue pathogenesis.