Sutee Yoksan

Dengue: a continuing global threat

Dengue fever and dengue haemorrhagic fever are important arthropod-borne viral diseases. Each year, there are ∼50 million dengue infections and ∼500,000 individuals are hospitalized with dengue haemorrhagic fever, mainly in Southeast Asia, the Pacific and the Americas. Illness is produced by any of the four dengue virus serotypes. A global strategy aimed at increasing the capacity for surveillance and outbreak response, changing behaviours and reducing the disease burden using integrated vector management in conjunction with early and accurate diagnosis has been advocated. Antiviral drugs and vaccines that are currently under development could also make an important contribution to dengue control in the future.

Evaluation of diagnostic tests: dengue

Dengue is an arthropod-borne flavivirus that comprises four distinct serotypes (DEN-1, DEN-2, DEN-3 and DEN-4) that constitute an antigenic complex of the genus flavivirus, family Flaviviridae. Infection by one serotype induces life-long immunity against reinfection by the same serotype, but only transient and partial protection against infection with the other serotypes1,2. Dengue virus infections can result in a range of clinical manifestations from asymp tomatic infection to dengue fever (DF) and the severe disease dengue haemorrhagic fever/dengue shock syndrome (DHF/ DSS). Most dengue infections are asymptomatic or cause mild symptoms, which are characterized by undifferentiated fever with or without rash. Typical DF is characterized by high fever, severe headache, myalgia, arthralgia, retro-orbital pain and maculopapular rash. Some patients show petechiae, bruising or thrombocytopenia. The clinical presentation of acute dengue infection is non-specific but 5–10% of patients progress to severe DHF/DSS, which can result in death if it is not managed appropriately. Plasma extravasation is the main pathophysiological finding of DHF/ DSS, which differentiates it from DF. DHF/ DSS is characterized by high fever, bleeding, thrombocytopenia and haemoconcentration (an increase in the concentration of blood cells as a result of fluid loss). Approximately 3–4 days after the onset of fever, patients can present with petechiae, rash, epistaxis, and gingival and gastrointestinal bleeding. Pleural effusion and ascites are common. Some patients develop circulatory failure (DSS), presenting with a weak and fast pulse, narrowing of pulse pressure or hypotension, cold and moist skin and altered mental state. Although there are no specific antiviral treatments for dengue infection, patients usually recover when the need for fluid management is identified early and electrolytes are administered3. It has been proposed that the classification of dengue disease should be simplified as severe and non-severe dengue. This simplified classification would make patient management and surveillance easier4. There is a need for specific, inexpensive dengue diagnostic tests that can be used for clinical management, surveillance and outbreak investigations and would permit early intervention to treat patients and prevent or control epidemics. Progress is being made in primary prevention, with several candidate dengue vaccines in late phases of development as well as improved vector control measures. Additionally, new techniques for the early detection of severe disease such as the use of biomarkers have the potential to decrease morbidity and

A variant in the CD209 promoter is associated with severity of dengue disease

Dengue fever and dengue hemorrhagic fever are mosquito-borne viral diseases. Dendritic cell–specific ICAM-3 grabbing nonintegrin (DC-SIGN1, encoded by CD209), an attachment receptor of dengue virus, is essential for productive infection of dendritic cells. Here, we report strong association between a promoter variant of CD209, DCSIGN1-336, and risk of dengue fever compared with dengue hemorrhagic fever or population controls. The G allele of the variant DCSIGN1-336 was associated with strong protection against dengue fever in three independent cohorts from Thailand, with a carrier frequency of 4.7% in individuals with dengue fever compared with 22.4% in individuals with dengue hemorrhagic fever (odds ratio for risk of dengue hemorrhagic fever versus dengue fever: 5.84, P = 1.4 × 10−7) and 19.5% in controls (odds ratio for protection: 4.90, P = 2 × 10−6). This variant affects an Sp1-like binding site and transcriptional activity in vitro. These results indicate that CD209 has a crucial role in dengue pathogenesis, which discriminates between severe dengue fever and dengue hemorrhagic fever. This may have consequences for therapeutic and preventive strategies.

Chimeric Live, Attenuated Vaccine against Japanese Encephalitis (ChimeriVax-JE): Phase 2 Clinical Trials for Safety and Immunogenicity, Effect of Vaccine Dose and Schedule, and Memory Response to Challenge with Inactivated Japanese Encephalitis Antigen

ChimeriVax-JE is a live, attenuated vaccine against Japanese encephalitis, using yellow fever (YF) 17D vaccine as a vector. In a double-blind phase 2 trial, 99 adults received vaccine, placebo, or YF 17D vaccine (YF-VAX). ChimeriVax-JE was well tolerated, with no differences in adverse events between treatment groups. Viremias resulting from administration of ChimeriVax-JE and YF-VAX were of short duration and low titer; 82 (94%) of 87 subjects administered graded doses (1.8-5.8 log(10)) of ChimeriVax-JE developed neutralizing antibodies. A second dose, administered 30 days later, had no booster effect. Previous inoculation with YF did not interfere with ChimeriVax-JE, but there was a suggestion (not statistically significant) that ChimeriVax-JE interfered with YF-VAX administered 30 days later. A separate study explored immunological memory both in subjects who had received ChimeriVax-JE 9 months before and in ChimeriVax-JE-naive subjects challenged with inactivated mouse-brain vaccine (JE-VAX). Anamnestic responses were observed in preimmune individuals. ChimeriVax-JE appears to be a safe vaccine that provides protective levels of neutralizing antibody after a single dose.

Clinical proof of principle for ChimeriVax™: recombinant live, attenuated vaccines against flavivirus infections

ChimeriVax is a live, attenuated recombinant virus constructed from yellow fever (YF) 17D in which the envelope protein genes of YF 17D are replaced with the corresponding genes of another flavivirus. A ChimeriVax vaccine was developed against Japanese encephalitis (JE). A randomized, double-blind, outpatient study was conducted to compare the safety and immunogenicity of ChimeriVax-JE and YF 17D. Six YF immune and six non-immune adults were randomized to receive a single SC inoculation of ChimeriVax-JE (5log(10)PFU), ChimeriVax-JE (4log(10)PFU) or YF-VAX((R)) (5log(10)PFU). Mild, transient injection site reactions and flu-like symptoms were noted in all treatment groups, with no significant difference between the groups. Nearly all subjects inoculated with ChimeriVax-JE at both dose levels developed a transient, low-level viremia which was similar in magnitude and duration to that following YF-VAX). Neutralizing antibody seroconversion rates to ChimeriVax-JE was 100% in the high and low dose groups in both naïve and YF immune subjects; seroconversion to wild-type JE strains was similar or lower than to the homologous (vaccine) virus. Mean neutralizing antibody responses were higher in the ChimeriVax-JE high dose groups (naïve subjects LNI 1.55, PRNT(50) 254; YF immune subjects LNI 2.23, PRNT(50) 327) than in the low dose groups (naïve subjects 1.38, PRNT(50) 128; YF immune subjects LNI 1.62, PRNT(50) 270). JE antibody levels were higher in YF immune than in naïve subjects, dispelling concerns about anti-vector immunity. The safety and immunogenicity profile of ChimeriVax-JE vaccine appears to be similar to that of YF 17D. The new vaccine holds promise for prevention of JE in travelers and residents of endemic countries. The ChimeriVax technology platform is being exploited for development of new vaccines against dengue and West Nile.

Live Attenuated Chimeric Yellow Fever Dengue Type 2 (ChimeriVax™-DEN2) Vaccine: Phase I Clinical Trial for Safety and Immunogenicity: Effect of Yellow Fever Pre-immunity in Induction of Cross Neutralizing Antibody Responses to All

A randomized double-blind Phase I Trial was conducted to evaluate safety, tolerability, and immunogenicity of a yellow fever (YF)-dengue 2 (DEN2) chimera (ChimeriVax™-DEN2) in comparison to that of YF vaccine (YF-VAX®). Forty-two healthy YF naïve adults randomly received a single dose of either ChimeriVax™-DEN2 (high dose, 5 log plaque forming units [PFU] or low dose, 3 log PFU) or YF-VAXâ by the subcutaneous route (SC). To determine the effect of YF pre-immunity on the ChimeriVaxTM-DEN2 vaccine, 14 subjects previously vaccinated against YF received a high dose of ChimeriVax™-DEN2 as an open-label vaccine. Most adverse events were similar to YF-VAX® and of mild to moderate intensity, with no serious side-effects. One hundred percent and 92.3% of YF naïve subjects inoculated with 5.0 and 3.0 log10 PFU of ChimeriVaxTM-DEN2, respectively, seroconverted to wt DEN2 (strain 16681); 92% of subjects inoculated with YF-VAX® seroconverted to YF 17D virus but none of YF naïve subjects inoculated with ChimeriVax-DEN2 seroconverted to YF 17D virus. Low seroconversion rates to heterologous DEN serotypes 1, 3, and 4 were observed in YF naïve subjects inoculated with either ChimeriVax™-DEN2 or YF-VAX®. In contrast, 100% of YF immune subjects inoculated with ChimeriVax™-DEN2 seroconverted to all 4 DEN serotypes. Surprisingly, levels of neutralizing antibodies to DEN 1, 2, and 3 viruses in YF immune subjects persisted after 1 year. These data demonstrated that 1) the safety and immunogenicity profile of the ChimeriVax™-DEN2 vaccine is consistent with that of YF-VAX®, and 2) pre-immunity to YF virus does not interfere with ChimeriVaxTM-DEN2 immunization, but induces a long lasting and cross neutralizing antibody response to all 4 DEN serotypes. The latter observation can have practical implications toward development of a dengue vaccine.

Diagnostic accuracy of NS1 ELISA and lateral flow rapid tests for dengue sensitivity, specificity and relationship to viraemia and antibody responses.

Background Dengue is a public health problem in many countries. Rapid diagnosis of dengue can assist patient triage and management. Detection of the dengue viral protein, NS1, represents a new approach to dengue diagnosis. Methodology/Principal Findings The sensitivity and specificity of the Platelia NS1 ELISA assay and an NS1 lateral flow rapid test (LFRT) were compared against a gold standard reference diagnostic algorithm in 138 Vietnamese children and adults. Overall, the Platelia NS1 ELISA was modestly more sensitive (82%) than the NS1 LFRT (72%) in confirmed dengue cases. Both ELISA and LFRT assays were more sensitive for primary than secondary dengue, and for specimens collected within 3 days of illness onset relative to later time points. The presence of measurable DENV-reactive IgG and to a lesser extent IgM in the test sample was associated with a significantly lower rate of NS1 detection in both assays. NS1 positivity was associated with the underlying viraemia, as NS1-positive samples had a significantly higher viraemia than NS1-negative samples matched for duration of illness. The Platelia and NS1 LFRT were 100% specific, being negative in all febrile patients without evidence of recent dengue, as well as in patients with enteric fever, malaria, Japanese encephalitis and leptospirosis. Conclusions/Significance Collectively, these data suggest NS1 assays deserve inclusion in the diagnostic evaluation of dengue patients, but with due consideration for the limitations in patients who present late in their illness or have a concomitant humoral immune response.

A Novel Tetravalent Dengue Vaccine Is Well Tolerated and Immunogenic against All 4 Serotypes in Flavivirus-Naive Adults

BACKGROUND. Sanofi Pasteur has developed a tetravalent dengue vaccine (TDV) against the worlds most common arbovirus infection. METHODS. We assessed the safety and immunogenicity of the TDV in healthy adults randomized into 2 groups. Group 1 received 3 TDV injections at months 0, 4, and 12-15; group 2 received saline placebo at month 0 and then 2 TDV injections at months 4 and 12-15. Adverse events were recorded, and biological parameters and viremia levels were measured. Neutralizing antibodies against 4 World Health Organization (WHO) reference strains were measured before and after vaccinations. RESULTS. A total of 33 participants were enrolled in each group. Demographic characteristics were comparable. No vaccine-related serious adverse event was reported. The most common systemic reactions were headache, malaise, and myalgia. Low viremia levels were detected, mainly of serotype 4. Immune response increased with successive vaccine doses. All participants seroconverted against all 4 serotypes after receiving 3 doses at 0, 4, and 12-15-months, and almost all seroconverted after 2 doses given 8-11 months apart. CONCLUSIONS. Sanofi Pasteurs TDV was well tolerated and induced full seroconversion against all WHO reference strain serotypes after 3 doses.

Evaluation of Commercially Available Anti–Dengue Virus Immunoglobulin M Tests

Anti–dengue virus immunoglobulin M kits were evaluated. Test sensitivities were 21%–99% and specificities were 77%–98% compared with reference ELISAs. False-positive results were found for patients with malaria or past dengue infections. Three ELISAs showing strong agreement with reference ELISAs will be included in the World Health Organization Bulk Procurement Scheme.

Safety and immunogenicity of a three dose regimen of two tetravalent live-attenuated dengue vaccines in five- to twelve-year-old Thai children.

Objective. The safety and immunogenicity of tetravalent live-attenuated dengue vaccines after a three dose vaccination series were evaluated in Thai children. Method. One hundred three healthy flavivirus-seronegative schoolchildren ages 5 to 12 years were randomized to receive either dengue vaccine containing 3, 2, 1 and 2 log10 of the 50% cell culture infective dose, respectively, of the live-attenuated dengue vaccine serotypes 1, 2, 3 and 4 per dose (F3212; n = 40) or 3, 3, 1 and 3 log10 of the 50% cell culture infective dose (F3313; n = 42) or purified Vero cell rabies vaccine (control group; n = 21) given in a two dose schedule (3 to 5 months apart). A third dose was administered 8 to 12 months after the second dose to 90 subjects. Safety and immunogenicity were evaluated within 28 days after each injection. Results. No serious adverse event related to the vaccines occurred. Most children experienced mild to moderate fever, rash, headache and myalgia occurring within 12 days after Dose 1 and generally lasting 3 days or less. One subject in Group F3212 had a 1-week dengue-like fever. Reactogenicity was minimal after Doses 2 and 3. Transient mild variations in liver enzymes and hematologic indices were noted mainly after Dose 1. After the third dose 89% of the subjects in Group F3212 seroconverted (neutralizing antibody response, ≥10) to all four serotypes, and all children in Group F3313 seroconverted. Conclusion. This study demonstrates a moderate although improvable reactogenicity and high seroconversion rates against the four serotypes of dengue after a three dose schedule of tetravalent live-attenuated dengue vaccine in children.