Dengue Virus (DENV) Neutralizing Antibody Kinetics in Children After Symptomatic Primary and Postprimary DENV Infection.

The immune response to dengue virus (DENV) infection is complex and not fully understood. Using longitudinal data from 181 children with dengue in Thailand who were followed for up to 3 years, we describe neutralizing antibody kinetics following symptomatic DENV infection. We observed that antibody titers varied by serotype, homotypic vs heterotypic responses, and primary versus postprimary infections. The rates of change in antibody titers over time varied between primary and postprimary responses. For primary infections, titers increased from convalescence to 6 months. By comparing homotypic and heterotypic antibody titers, we saw an increase in type specificity from convalescence to 6 months for primary DENV3 infections but not primary DENV1 infections. In postprimary cases, there was a decrease in titers from convalescence up until 6 months after infection. Beginning 1 year after both primary and postprimary infections, there was evidence of increasing antibody titers, with greater increases in children with lower titers, suggesting that antibody titers were boosted due to infection and that higher levels of neutralizing antibody may be more likely to confer a sterilizing immune response. These findings may help to model virus transmission dynamics and provide baseline data to support the development of vaccines and therapeutics.

Revealing the microscale spatial signature of dengue transmission and immunity in an urban population.

It is well-known that the distribution of immunity in a population dictates the future incidence of infectious disease, but this process is generally understood at individual or macroscales. For example, herd immunity to multiple pathogens has been observed at national and city levels. However, the effects of population immunity have not previously been shown at scales smaller than the city (e.g., neighborhoods). In particular, no study has shown long-term effects of population immunity at scales consistent with the spatial scale of person-to-person transmission. Here, we use the location of dengue patients' homes in Bangkok with the serotype of the infecting pathogen to investigate the spatiotemporal distribution of disease risk at small spatial scales over a 5-y period. We find evidence for localized transmission at distances of under 1 km. We also observe patterns of spatiotemporal dependence consistent with the expected impacts of homotypic immunity, heterotypic immunity, and immune enhancement of disease at these distances. Our observations indicate that immunological memory of dengue serotypes occurs at the neighborhood level in this large urban setting. These methods have broad applications to studying the spatiotemporal structure of disease risk where pathogen serotype or genetic information is known.

Dengue--how best to classify it.

Dengue has emerged as a major public health problem worldwide. Dengue virus infection causes a wide range of clinical manifestations. Since the 1970s, clinical dengue has been classified according to the World Health Organization guideline as dengue fever and dengue hemorrhagic fever. The classification has been criticized with regard to its usefulness and its applicability. In 2009, the World Health Organization issued a new guideline that classifies clinical dengue as dengue and severe dengue. The 2009 classification differs significantly from the previous classification in both conceptual and practical levels. The impacts of the new classification on clinical practice, dengue research, and public health policy are discussed.

Is dengue severity related to nutritional status?

A retrospective review of dengue patients admitted to Queen Sirikit National Institute of Child Health (previously known as Children's Hospital) from 1995 to 1999 revealed 4,532 confirmed cases of dengue infection; 80.9% were dengue hemorrhagic fever (DHF) and 19.1% were dengue fever cases (DF). Among the DHF patients; 30.6% had shock. The majority of them, 66.6%, had a normal nutritional status, while 9.3% were malnourished and 24.2% had obesity as classified by weight for age. Compared with control patients with other diagnoses (excluding HIV/AIDS patients), malnourished children had a lower risk of contracting dengue infection (odds ratio = 0.48, 95% Cl = 0.39-0.60, p = 0.000) while obese children had a greater risk of infection with dengue viruses (odds ratio = 1.96, 95% Cl = 1.55-2.5, p = 0.000). The clinical signs, symptoms and laboratory findings of dengue were almost the same among the 3 groups of malnourished, normal, and obese patients. The minor differences observed were that in obese children liver enlargement was found less often; maculopapular/convalescence rash and elevations of alanine aminotransferase were found more often. Malnourished patients had a higher risk of developing shock (37.8%) than normal (29.9%) and obese patients (30.2%) (p = 0.000). Obese patients had more unusual presentations: encephalopathy (1.3%) and associated infections (4.8%), than normal (0.5% and 2.7%) and malnourished patients (1.2% and 3.1%). Complications of fluid overload were found more in obese patients (6.5%) compared to normal (3.2%) and malnourished patients (2.1%) (p = 0.000). The case-fatality rates (CFR) in malnourished patients and obese patients were 0.5% and 0.4%, respectively, while in normal patients the CFR was 0.07%. Under and over nutrition DHF patients had either a greater risk of shock or unusual presentations and complications, which can lead to severe disease or complications and probably a higher CFR.

Serotype-specific dengue virus circulation and dengue disease in Bangkok, Thailand from 1973 to 1999.

Dengue virus circulation and association with epidemics and severe dengue disease were studied in hospitalized children with suspected dengue at the Queen Sirikit National Institute of Child Health in Bangkok, Thailand, from 1973 to 1999. Dengue serology was performed on all patients and viral isolation attempted on laboratory-confirmed patients. Acute dengue was diagnosed in 15,569 children and virus isolated from 4,846. DEN-3 was the most frequent serotype in primary dengue (49% of all isolates), DEN-2 in secondary and in dengue hemorrhagic fever (37% and 35%, respectively). The predominant dengue serotype varied by year: DEN-1 from 1990-92, DEN-2 from 1973-86 and 1988-89; DEN-3 in 1987 and 1995-99; and DEN-4 from 1993-94. Only DEN-3 was associated with severe outbreak years. Our findings illustrate the uniqueness of each serotype in producing epidemics and severe disease and underscore the importance of long-term surveillance of dengue serotypes in understanding the epidemiology of these viruses.

Clinical features of diphtheria in thai children: a historic perspective.

To study the clinical and laboratory features of childhood diphtheria, the cases of 381 children with clinical and bacteriological diagnoses of diphtheria who were treated at the Children's Hospital between 1976 and 1985 were reviewed. Of these, 191 were males and 190 were females. The mean age was 4.6 years. Approximately 75% of the patients had no history of immunization. Common manifestations of diphtheria included patch (100%), fever (92.4%), upper respiratory tract infection (91.6%), upper airway obstruction (42.3%), hoarseness (36.7%), and bull neck (11.3%). The mean duration of fever prior to admission was 3.3 days with a range of 0-11 days. Patch sites included the tonsils (91.9%), the pharynx (55.9%), the larynx (27.8%) and others (24.4%). Complications included upper airway obstruction (42.3%), cardiac complications (10.0%) and neurological complications (4.7%). The mortality rate was 5.8%. There were significant associations between death and the presence of bull neck, laryngeal patch, airway obstruction and cardiac complications. Early recognition and prompt treatment will decrease complications and mortality in this group of patients.

Neonatal dengue infection: report of dengue fever in a 1-day-old infant.

A male infant was admitted because of fever. He was born at 37-weeks' gestation. His mother had experienced acute febrile illness with headache and myalgia. Her illness persisted with onset of active labor pain on day 5, which prompted cesarean section; postoperatively, the hematocrit decreased, requiring transfusion. The infant was well until fever developed at 16 hours after birth. There were petichiae on his face and trunk and the liver was enlarged. Fever subsided on day 5 without evidence of plasma leakage or severe hemorrhage. He made an uneventful recovery after 8 days of illness. Leukopenia and thrombocytopenia were present in the mother and infant. Both were diagnosed as dengue fever. Dengue type 1 was recovered from the infant by polymerase chain reaction. The dengue enzyme-linked immunoassay showed secondary infection in the mother and primary infection in the infant. In dengue-endemic areas, clinicians should be alert to dengue fever/dengue hemorrhagic fever in pregnant women presenting with acute febrile illness, and be prepared for proper management.

Clinical presentations of dengue hemorrhagic fever in infants compared to children.

BACKGROUND: Dengue infections are endemic in Thailand. Infants are at risk of developing dengue fever (DF) and dengue hemorrhagic fever (DHF). Many infants have severe DHF with unusual manifestations and may suffer from complications and even death because of difficulties in early diagnosis and improper management. OBJECTIVE: To study the incidence of infants DHF and to compare clinical and laboratory findings of DHF between infants and children in order to improve diagnosis and management of infants with DHF. MATERIAL AND METHOD: A retrospective review of hospital charts of DHF patients who were admitted to the Children's Hospital between 1995-1999 was carried out. Only confirmed DHF cases, either by serologic or virologic studies were included. Clinical and laboratory findings were compared using the standard software package SPSS. RESULTS: There were 4,595 confirmed dengue patients admitted to the Children's Hospital during this 5-year period and among these 245 infants were under one year old (5.3%). Unusual presentations of DHF including upper respiratory tract infection (URI) symptoms, diarrhea, convulsions and encephalopathy were found more in the infants (4.5%, 13.1%, 12.7% and 4.1%) than in the children (1.7%, 2.0%, 2.0% and 0.6%) but shock was found less in the infants (25%) than in the children (30.9%). The percentages of Tourniquet test positive and low WBC (< or = 5,000 cells/mm3) which help in the early diagnosis of dengue were less in the infants (50.2% and 26.8%) than in the children (92.2% and 71.9%). Liver involvement and/or dysfunction were found more in the infants as shown by mean aspartate aminotransferase/alanine aminotransferase (AST/ALT) elevation and prolonged prothrombin time. Duration of i.v. fluid was shorter in the infants than in the children (21.9 vs 38.2 hours). Complications of fluid overload were found more often in the infants than in the children (9% vs 3.6%). The case fatality rate was 1.2 in the infants and 0.3 in the children. CONCLUSION: Management of infants with DHF is critical because early diagnosis is rather difficult and they sometimes present with unusual manifestations (convulsions, encephalopathy and associated infections). Complications such as hepatic dysfunction and fluid overload are more commonly found in infants than in children and adults and also the case fatality rate is higher. Judicious i.v. fluid resuscitation and replacement is critical to the outcome of infants with DHF. It seems that the degree of plasma leakage is less in infants and the duration of plasma leakage is shorter in infants than in children.

Preparing for introduction of a dengue vaccine: recommendations from the 1st Dengue v2V Asia-Pacific Meeting.

Infection with dengue virus is a major public health problem in the Asia-Pacific region and throughout tropical and sub-tropical regions of the world. Vaccination represents a major opportunity to control dengue and several candidate vaccines are in development. Experts in dengue and in vaccine introduction gathered for a two day meeting during which they examined the challenges inherent to the introduction of a dengue vaccine into the national immunisation programmes of countries of the Asia-Pacific. The aim was to develop a series of recommendations to reduce the delay between vaccine licensure and vaccine introduction. Major recommendations arising from the meeting included: ascertaining and publicising the full burden and cost of dengue; changing the perception of dengue in non-endemic countries to help generate global support for dengue vaccination; ensuring high quality active surveillance systems and diagnostics; and identifying sustainable sources of funding, both to support vaccine introduction and to maintain the vaccination programme. The attendees at the meeting were in agreement that with the introduction of an effective vaccine, dengue is a disease that could be controlled, and that in order to ensure a vaccine is introduced as rapidly as possible, there is a need to start preparing now.

Classification of dengue illness based on readily available laboratory data.

The aim of this study was to examine retrospective dengue-illness classification using only clinical laboratory data, without relying on X-ray, ultrasound, or percent hemoconcentration. We analyzed data from a study of children who presented with acute febrile illness to two hospitals in Thailand. Multivariable logistic regression models were used to distinguish: (1) dengue hemorrhagic fever (DHF) versus dengue fever (DF), (2) DHF versus DF + other febrile illness (OFI), (3) dengue versus OFI, and (4) severe dengue versus non-severe dengue + OFI. Data from the second hospital served as a validation set. There were 1,227 patients in the analysis. The sensitivity of the models ranged from 89.2% (dengue versus OFI) to 79.6% (DHF versus DF). The models showed high sensitivity in the validation dataset. These models could be used to calculate a probability and classify patients based on readily available clinical laboratory data, and they will need to be validated in other dengue-endemic regions.

Molecular genetic relationship of the 3' untranslated region among Thai dengue-3 virus, Bangkok isolates, during 1973-2000.

Dengue virus serotype 3 (DENV-3) was associated with severe dengue epidemics in Thailand during 1973-1999. We studied Thai DENV-3 viruses isolated from hospitalized children in Bangkok with differing disease severity during that period. Viruses were sequenced at their 5' and 3' untranslated regions (UTRs), which are regions that play a pivotal role in viral replication. Our results indicated that the primary sequences as well as the secondary structures at both ends of Thai DENV-3 viruses were highly conserved over almost three decades. We found nucleotide insertions and deletions at the variable region (VR) that is located just downstream of the nonstructural protein 5 (NS5) stop codon among these viruses. The phylogenetic tree derived from the size heterogeneity of VR in the 3' UTR divided DENV-3 into four genotypes, and Thai DENV-3 viruses in this study belonged to genotype II. The replication efficiency of the candidate viruses with different lengths at the VR were assessed in the mosquito (C6/36) and human (HepG2) cell lines. Our results show that the viruses with nucleotide insertions at VR replicated better than the virus that contained deletions. Our findings indicate that Thai DENV-3 demonstrated a remarkable conservation of nucleotides over 28 years. Correlation with disease severity suggests that both primary sequences and secondary structures of the 3' UTR do not appear correlated with disease severity in humans.

Comparative analysis reveals no consistent association between the secondary structure of the 3'-untranslated region of dengue viruses and disease syndrome.

A comparative analysis was performed of the 3'-untranslated region (UTR) of Dengue virus (DENV) sampled from Bangkok, Thailand, over a 30 year period and representing all four serotypes. Considerable genetic variation was observed both within and among serotypes. Notably, a full-length version of the critical 3'-long stable hairpin structure was absent from some isolates, suggesting the occurrence of complex structural interactions within the 3'-UTR, including the influence of upstream mutations. The Thai sequences were then combined with 61 globally sampled isolates of DENV taken from patients with either dengue fever or severe dengue disease. No consistent association was found between 3'-UTR secondary structure and the clinical outcome of DENV infection, although some evidence for a trend in this direction was observed in DENV-2. It was concluded that the 3'-UTR is not the sole determinant of DENV virulence in nature, although variation in secondary structure may greatly influence viral fitness.

Structure and age of genetic diversity of dengue virus type 2 in Thailand.

Dengue virus type 2 (DENV-2) is a common viral infection and an important health concern in South-East Asia. To determine the molecular evolution of DENV-2 in Thailand, 105 isolates of the E (envelope) gene and 10 complete genomes sampled over a 27 year period were sequenced. Phylogenetic analysis of these data revealed that three genotypes of DENV-2 have circulated in Thailand, although, since 1991, only viruses assigned to Asian genotype I have been sampled from the population. A broader analysis of 35 complete genomes of DENV-2 revealed that most amino acids are subject to strong selective constraints, indicative of widespread purifying selection against deleterious mutations. This was further supported by an analysis of genome-wide substitution rates, which indicated that DENV-2 fixes approximately 10 mutations per genome per year, far lower than expected from its mutational dynamics. Finally, estimates of the age of DENV-2 were remarkably consistent among genes, indicating that the current genetic diversity in this virus probably arose within the last 120 years, concordant with the first determination of the aetiology of dengue disease.

Clade replacements in dengue virus serotypes 1 and 3 are associated with changing serotype prevalence.

The evolution of dengue virus (DENV) is characterized by phylogenetic trees that have a strong temporal structure punctuated by dramatic changes in clade frequency. To determine the cause of these large-scale phylogenetic patterns, we examined the evolutionary history of DENV serotype 1 (DENV-1) and DENV-3 in Thailand, where gene sequence and epidemiological data are relatively abundant over a 30-year period. We found evidence for the turnover of viral clades in both serotypes, most notably in DENV-1, where a major clade replacement event took place in genotype I during the mid-1990s. Further, when this clade replacement event was placed in the context of changes in serotype prevalence in Thailand, a striking pattern emerged; an increase in DENV-1 clade diversity was associated with an increase in the abundance of this serotype and a concomitant decrease in DENV-4 prevalence, while clade replacement was associated with a decline in DENV-1 prevalence and a rise of DENV-4. We postulate that intraserotypic genetic diversification proceeds at times of relative serotype abundance and that replacement events can result from differential susceptibility to cross-reactive immune responses.

Dengue hemorrhagic fever in infants: research opportunities ignored.

The age distribution of cases of dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS) in infants under the age of 1 year are reported from Bangkok, Thailand, and for the first time for Ho Chi Minh City, Vietnam; Yangon, Myanmar; and Surabaya, Indonesia. The four dengue viruses were isolated from Thai infants, all of whom were having a primary dengue infection. Progress studying the immunologically distinct infant DHF/DSS has been limited; most contemporary research has centered on DHF/DSS accompanying secondary dengue infections. In designing research results obtained in studies on a congruent animal model, feline infectious peritonitis virus (FIPV) infections of kittens born to FIPV-immune queens should be considered. Research challenges presented by infant DHF/DSS are discussed.

Clinical and laboratory presentations of dengue patients with different serotypes.

A five –year retrospective review of confirmed dengue patients admitted to the Children’s Hospital, Bangkok, during 1995-1999 was undertaken to compare the clinical and laboratory presentations between DEN-1,2,3 and 4. Dengue serotype could be identified in 50.6% (2398 cases) of all the confirmed cases (Total: 4743 cases) during this period. DEN-3 was the predominant serotype found in 50.6%, followed by DEN-1 (25.8%), DEN-2 (20.9%) and DEN-4 (2.7%). There was no gender difference. The mean age of patients was significantly higher in DEN-4 (9.8 years) while in DEN-1, DEN-2 and DEN-3 it was 8.2, 8.8 and 8 years, respectively. The peak incidence of DEN-1, DEN-2 and DEN-3 was among children between the age of 5-9 years while in DEN-4 it was between 10-14 years. From this study it was concluded that 91.8% of DEN-2 infections were secondary and it presented with more severe disease as demonstrated by more degree of plasma leakage, more shock and more complication of fluid overload cases. DEN-3 and DEN-4 tended to have more degree of liver involvement as demonstrated by more patients with abnormal elevation of liver enzyme AST and ALT and higher mean value of both AST and ALT. Because 29.6% of DEN -3 presented as primary infections and DEN-4 was the rare serotype, so that hepatic dysfunction / encephalopathy was not a big problem during this study period. DEN-1 seemed to have mildest clinical presentation with 29.6% of DEN-3 presented as primary infections. With this knowledge, if the circulating dengue serotype in the area is known, a better plan for case management can be established; preparation for more colloidal solutions and probably blood transfusion in DEN-2 and DEN-1 outbreaks and preparation for the management of DHF patients with hepatic dysfunction/encephalopathy if there is DEN-3 or DEN-4 outbreak.