Secondary heterologous dengue infection risk: Disequilibrium between immune regulation and inflammation?

Increased serum levels of cytokines released by cells of the immune response have been detected in patients suffering from dengue disease. Likewise, secondary infections by a different dengue virus serotype result in a highest risk of development of the severe dengue disease. Both findings suggest that the memory immune response is one of the key players in the pathogenesis of this disease. Here we take advantage of the particular Cuban epidemiological situation in dengue to analyze a broad spectrum of cell-mediated immune response mediators at mRNA and protein level. Evidences for a regulatory immune pattern in homologous (TGF-beta, IL-10) vs. pro-inflammatory pattern (IFN-gamma, TNF-alpha) in heterologous dengue virus re-challenge were found, suggesting a possible association with the higher incidence of severe dengue cases in the latter case.

Dengue: where are we today?

Dengue is considered the main arthropod-borne viral disease of humans. In the last few years, an increasing number of reports of mild and severe cases have been reported. The growing dengue incidence observed in recent years has been accompanied by reports of new observations, findings and global initiatives with an improvement in our understanding of this phenomenon. The epidemiology and new clinical classification of dengue, advances in the diagnostic and pathogenesis knowledge, and vaccine development as well as control methods including new global initiatives are summarised here.

Dengue haemorrhagic fever integral hypothesis: confirming observations, 1987-2007.

In 1987, Kouri et al. published in Transactions their integral hypothesis to explain the development of dengue haemorrhagic fever (DHF) epidemics (Kouri, G.P., Guzmán, M.G., Bravo, J.R., 1987. Why dengue haemorrhagic fever in Cuba? 2. An integral analysis. Trans. R. Soc. Trop. Med. Hyg. 81, 821-823). Based on observations carried out during the 1981 Cuban DHF epidemic, the authors integrated in one model the most advanced knowledge at that time. Observations in the last 20 years confirm the importance of this multifactorial and unifying view of the problem, where the interaction between the host, the virus and the vector in an epidemiological and ecosystem setting might determine DHF as a final outcome. Investigations on the interaction among host, virus and mosquito with an ecosystemic view are needed.

An integrated ecosystem approach for sustainable prevention and control of dengue in Central Havana.

The authors developed and evaluated a comprehensive participatory ecosystem health approach for preventing the transmission of dengue, the most prevalent vector-borne disease in Cuba and the Latin America-Caribbean region. The integrated surveillance system central to this initiative encompassed three main subsystems (environmental; entomological; clinical-epidemiologic), relying on extensive community involvement. The study was conducted in Central Havana, Cuba. Indicators from each subsystem were selected and mapped using a GIS procedure providing instant visualization by city block in the municipality. To elucidate the factors affecting control and prevention efforts, perceived needs and risks, as well as knowledge, attitudes, and behaviors related to dengue, were assessed. Specific factors associated with the presence of mosquito breeding sites and risks of dengue were examined in a case-control study.

Ethnicity and difference in dengue virus-specific memory T cell responses in Cuban individuals.

The different risk factors associated with dengue hemorrhagic fever pathogenesis needs yet additional clarification. The exceptional epidemiological circumstances in Cuba allow their evaluation in a well-defined situation. In the present study the memory T cell response of 80 Cuban donors previously infected with dengue-1 and dengue-2 during the 1977 and 1981 epidemics, and belonging to different ethnic groups, was examined. White people showed, in contrast to black people, stronger and remarkably cross-reactive dengue virus-specific memory CD4(+) T lymphocyte proliferation and interferon-gamma release. The observed variation in T cell response according to ethnicity could be related to the immunopathogenesis of dengue hemorrhagic fever, and may partially explain the epidemiological evidence that black individuals are at lower risk for the most severe dengue clinical course compared with white individuals.

Dengue hemorrhagic Fever caused by sequential dengue 1-3 virus infections over a long time interval: Havana epidemic, 2001-2002.

A dengue epidemic caused by dengue virus 3 (DENV-3) occurred in Cuba in 2001-2002. It included cases of dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). We report neutralizing antibody studies on sera from 54 of 78 DHF/DSS patients that provide evidence of infections occurring in the sequence DENV-1 followed by DENV-3. No sera showed infection in the sequence DENV-2 followed by DENV-3. Some sera showed a pattern of infection in the sequence DENV-1 followed by DENV-2 and then DENV-3. However definitive categorization of a tertiary infection was not possible because of broadly reactive antibodies, which could have been raised by infections in the sequence DENV-1 then DENV-3.

Classical dengue hemorrhagic fever resulting from two dengue infections spaced 20 years or more apart: Havana, Dengue 3 epidemic, 2001-2002.

OBJECTIVE: To report the clinical, laboratory and sonographic findings in 76 adult cases of dengue hemorrhagic fever (DHF). PATIENTS AND METHODS: A dengue 3 epidemic occurred in Havana City from June 2001 to March 2002. 12,889 cases were reported, with 81 DHF cases. From this, 76 serologically confirmed cases were studied descriptively. RESULTS: Bronchial asthma and white race were important risk factors for the severe form of the disease. Fever (100%), headache (92.1%), myalgia (76.3%), arthralgia (73.7%) and retro-orbital pain (57.7%) were the most frequent general symptoms. Vomiting and abdominal pain were observed in 59.2% and 48.6% of cases, respectively. The most common bleeding site was the vagina (64%), followed by the skin (55.2%). Eighteen patients (23.6%) had shock syndrome. Laboratory findings included thrombocytopenia (100%), hemoconcentration (93.4%), an increase in liver enzymes (82.8%), and leukopenia (71%). Ultrasound detected thickening of the gallbladder wall in 35.1%, pleural effusion in 20.3%, and splenomegaly in 12.9% of cases. CONCLUSION: These findings contribute to a better understanding of the clinical aspects of DHF in adult patients due to the dengue 3 virus.

Dengue: an update.

This review is an update of dengue and dengue haemorrhagic fever (DHF) based on international and Cuban experience. We describe the virus characteristics and risk factors for dengue and DHF, and compare incidence and the case fatality rates in endemic regions (southeast Asia, western Pacific, and the Americas). The clinical picture and the pathogenesis of the severe disease are explained. We also discuss the viral, individual, and environmental factors that determine severe disease. Much more research is necessary to clarify these mechanisms. Also reviewed are methods for viral isolation and the serological, immunohistochemical, and molecular methods applied in the diagnosis of the disease. We describe the status of vaccine development and emphasise that the only alternative that we have today to control the disease is through control of its vector Aedes aegypti.

Dengue and dengue hemorrhagic fever in the Americas: lessons and challenges.

The incidence of dengue and dengue hemorrhagic fever (DF/DHF) has increased significantly over the last decades. Yearly, an estimated 50-100 million cases of DF and about 250000-500000 cases of DHF occur worldwide. The epidemiological situation in Latin America now resembles that in Southeast Asia. Here, the main clinical, epidemiological and virological observations in the American region are presented and compared with those previously reported from Southeast Asia. During 2002, more than 30 Latin American countries reported over 1000000 DF cases. DHF occurred in 20 countries with more than 17000 DHF cases, including 225 fatalities. The co-circulation of multiple serotypes has been reported from many countries. In the Americas, DHF is observed both in children and adults; secondary infection by a different dengue virus serotype has been confirmed as an important risk factor for this severe form of the disease. However, some new risk factors such as the interval of dengue virus infections and the ethnicity and underlying chronic conditions of the patient have also been identified. The sequence of dengue virus infections and association with certain genotypes are further factors of importance. We also discuss the control and prevention strategies. In conclusion, without urgent action for the prevention and control of dengue/DHF and its vector, the current situation will worsen and, more dramatical, there is a risk of the urbanization of yellow fever.

Oxidative stress in adult dengue patients.

An association between viral diseases and increased oxidative stress has been suggested. The time course of serum levels of total antioxidant status (TAS), peroxidation potential (PP), glutathione (GSH), lipid peroxidation measured as hydroperoxides, and malondyaldehyde and 4-hydroxyalkenals (MDA + 4-HDA), as well as antioxidant enzymatic activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx), were measured in 22 serologically confirmed dengue patients. Most of the patients had dengue fever and three of them had dengue hemorrhagic fever. The redox parameters were compared with those of age- and sex- matched controls. No significant difference was observed for levels of GSH and TAS between patients and controls. Levels of PP, MDA + 4-HDA, and SOD were significantly higher. Levels of GPx and total hydroperoxides were significantly lower in patients in comparison with controls. These findings suggest that the alteration in redox status could result of increased oxidative stress and it may play a role in the pathogenesis of the disease.

Dengue vaccine: priorities and progress.

Dengue transmission has increased considerably in the past 20 years. Currently, it can only be reduced by mosquito control; however, the application of vector-control methods are labor intensive, require discipline and diligence, and are hard to sustain. In this context, a safe dengue vaccine that confers long-lasting protection against infection with the four dengue viruses is urgently required. This review will discuss the requirements of a dengue vaccine, problems, and advances that have been made. Finally, new targets for research will be presented.

Dengue diagnosis, advances and challenges.

Dengue diagnosis was one of the topics discussed at the symposium 'The Global Threat of Dengue - Desperately Seeking Solutions' organized during the 10th International Congress of Infectious Diseases held in Singapore in 2002. In this paper, a review is presented focusing on the main advances, problems and challenges of dengue diagnosis.IgM capture ELISA, virus isolation in mosquito cell lines and live mosquitoes, dengue specific monoclonal antibodies and PCR have all represented major advances in dengue diagnosis. However, an appropriate rapid, early and accessible diagnostic method useful both for epidemiological surveillance and clinical diagnosis is still needed. Also, tools that suggest a prognosis allowing for better management are also needed. Finally, laboratory infrastructure, technical expertise and research capacity must be improved in endemic countries in order to positively influence dengue surveillance, clinical case management and the development of new approaches to dengue control.

Effect of age on outcome of secondary dengue 2 infections.

OBJECTIVE: Dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) is a growing global health problem. It is not known how age affects the outcome of secondary dengue infections. In an island setting, a large DHF/DSS outbreak in Cuba occurred in 1981. Involved were individuals, 3-40 year old, whose only lifetime dengue exposure was to DEN-1 in 1977 and DEN-2 in 1981. In this report we calculate age-specific DHF/DSS hospitalization and death rates based on secondary DEN 2 infections. METHODS: Published and unpublished hospital and seroepidemiologic data from the 1981 DHF/DSS outbreak were used for the analysis. RESULTS: Children, aged 3 and 4 years, with secondary DEN-2 infections were found to have a high death rate (25.4/10 000 secondary DEN-2 infections). The death rate fell with increasing age, being 15.9-fold lower in the 10-14-year age group. The death rate for children aged 3-14 years was 14.5-fold higher than in young adults aged 15-39 years. The death rate rose somewhat in adults aged 50 years and older. DHF/DSS hospitalization rates showed the same trend as death rates. CONCLUSIONS: Age is an important variable in the outcome of secondary DEN-2 infections. DHF/DSS case fatality and hospitalization rates are highest in young infants and the elderly. The risk that a child will die during a secondary DEN-2 infection is nearly 15-fold higher than the risk in adults.

Epidemiological studies on dengue virus type 3 in Playa municipality, Havana, Cuba, 2001-2002.

OBJECTIVES: Recognizing the uniqueness of secondary dengue virus (DENV)-1/3 dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) cases at an interval of 24 years, we sought to estimate DENV infections as well as the ratios between mild disease and DHF/DSS by DENV infection sequence in Playa District (Havana, Cuba) during the 2001-2002 outbreak of dengue virus type 3 (DENV-3). METHODS: A retrospective seroepidemiological study was conducted in 2003 in Playa District. Blood samples were collected from a 1% random sample of residents and were studied for the prevalence of dengue neutralizing antibodies. RESULTS: DENV-3 was found to have infected 7.2% (95% confidence interval (95% CI) 6.0-8.4%) of susceptible individuals (the entire cohort), the majority of whom experienced silent infections. Virtually every individual who had a secondary infection in the sequence DENV-1 then DENV-3 became ill, with a ratio of severe to mild cases of 1:35 (95% CI 1:67-1:23). Secondary infections in the sequence DENV-2/3 were less pathogenic than DENV-1/3. Mild disease accompanying secondary DENV2/3 occurred at a ratio of 1:4.49 infections (95% CI 1:5.77-1:3.42) secondary infections. CONCLUSIONS: The results obtained highlight the role of the infecting serotype and also the sequence of the viral infection in the clinical outcome of a dengue infection.

Prevalence of febrile syndromes in dengue surveillance, havana city, 2007.

OBJECTIVE: Determine point prevalence of febrile syndromes and compare with prevalence reported by habitual clinical and seroepidemiologic dengue surveillance system in Havana City. METHODS: In October 2007, a descriptive, cross-sectional study was carried out in a representative sample, calculating prevalences of febrile syndromes and undifferentiated febrile syndromes. Chi-square analysis was used for rate comparisons. RESULTS: Point prevalences of febrile syndromes and undifferentiated febrile syndromes were 352.6 and 144.2 times greater, respectively, than those reported by the habitual clinical and seroepidemiologic dengue surveillance system; these differences were statistically significant (p < 0.001). CONCLUSIONS: Point prevalence of febrile syndromes was far greater than prevalence reported by the habitual clinical and seroepidemiologic dengue surveillance system, an indication of underreporting.

Virus role during intraepidemic increase in dengue disease severity.

Dengue epidemics in Cuba have repeatedly demonstrated a month-to-month increase in clinical severity during secondary infections. The dengue 2 outbreak that occurred in Santiago de Cuba in 1997 was accompanied by the most severe intraepidemic increase in disease severity reported to date. It was initially proposed that the appearance of neutralization escape mutants during the course of the epidemic might explain this phenomenon. Recent studies have revealed that during the course of this epidemic, nucleotide substitutions appeared only in nonstructural (NS) genes, most of which were silent, except for one change in the NS1 gene. To study whether or not variation in the NS1 gene might be associated with increased disease severity during the epidemic, this gene was partially sequenced from 15 isolates obtained at different times during the 1997 epidemic. Early epidemic isolates differed from those obtained later by replacement only of threonine with serine at position 164 in the NS1 protein, an amino acid rarely found in any genotype of dengue 2 virus. All viruses isolated from patients located in Health Districts, where dengue 2 transmissions occurred late in the epidemic, contained Serine at position 164, indicating that this change was fixed within a few months. Here we argue that this single mutation contributes to viral survival or replication efficiency, resulting in enhanced infection in the presence of enhancing antibodies, a phenomenon that we term increased virus "fitness" in contrast to "virulence," an intrinsic property of the virus.

The role of disease-specific research: a developing-country perspective.

Over a decade ago, the Commission on Health Research for Development published a pioneering study on the gross imbalance between the world's research funding priorities and global disease burden distribution. This study reported that less than 10% of world resources for health research were being applied to the health problems of developing countries, where 90% of the avoidable burden of ill-health was to be found. What became known as the "10/90 gap" has captured the attention and imagination of those committed to global health equity everywhere, searching for ways to incline the research agenda towards "diseases of the poor".

Dual infection with dengue virus 3 and human immunodeficiency virus 1 in Havana, Cuba.

Although dengue virus (DEN) endemic regions overlap with human immunodeficiency virus 1 (HIV) high incidence areas, little has been documented on HIV and DEN mixed infection. Here we report DEN/HIV concurrent infections recorded during the DEN-3 epidemic in 2001-2002 in Havana. Serologic-confirmed DEN is described in two HIV-infected subjects with dengue fever symptoms. Although patients had dengue disease, the CD4+ cells remained within normal levels and no accelerated progression of HIV disease was observed. To our knowledge, DEN cases caused by DEN-3 in HIV-infected individuals have not been reported previously. Further research is needed to diagnose this likely underreported mixed viral infection in DEN endemic areas.

Randomized, double-blind, placebo-controlled trial to evaluate the safety and immunogenicity of live oral cholera vaccine 638 in Cuban adults.

A randomized, double-blind, placebo-controlled clinical trial was conducted to evaluate the safety, reactogenicity and the immunogenicity of a 2 x 10(9)CFU dose of the 638 lyophilized live attenuated cholera vaccine for oral administration, formulated and produced at Finlay Institute, City of Havana, Cuba. Thirty-six healthy female and male adult volunteers from 18 to 40 years old were involved, clinically examined and laboratory tested after the informed consent signature. Adverse events were monitored and seroconversion rates and geometrical mean titer (GMT) of vibriocidal antibodies were tested in volunteer's sera samples. Neither serious adverse events nor other damages to the volunteers due to vaccine or placebo feeding were reported during the clinical follow-up period of this study; none of the adverse events registered within the first 72 h after inoculation were life-threatening for volunteers. Neither severe nor moderate adverse events were reported. Sixty-one percent of subjects showed mild expected adverse events in an interval lower than 24h up to the first 72 h, 75% of these in the vaccinated group and 18% in the placebo group. Fourteen days after inoculation the GMT of vibriocidal antibodies in the vaccine group significantly increased in comparison to the placebo group. All subjects in the vaccine group (24) seroconverted (100%). Results show that this vaccine is safe, well tolerated and immunogenic in healthy female and male volunteers.