Anon Srikiatkhachorn

Spatial and Temporal Clustering of Dengue Virus Transmission in Thai Villages

Background Transmission of dengue viruses (DENV), the leading cause of arboviral disease worldwide, is known to vary through time and space, likely owing to a combination of factors related to the human host, virus, mosquito vector, and environment. An improved understanding of variation in transmission patterns is fundamental to conducting surveillance and implementing disease prevention strategies. To test the hypothesis that DENV transmission is spatially and temporally focal, we compared geographic and temporal characteristics within Thai villages where DENV are and are not being actively transmitted. Methods and Findings Cluster investigations were conducted within 100 m of homes where febrile index children with (positive clusters) and without (negative clusters) acute dengue lived during two seasons of peak DENV transmission. Data on human infection and mosquito infection/density were examined to precisely (1) define the spatial and temporal dimensions of DENV transmission, (2) correlate these factors with variation in DENV transmission, and (3) determine the burden of inapparent and symptomatic infections. Among 556 village children enrolled as neighbors of 12 dengue-positive and 22 dengue-negative index cases, all 27 DENV infections (4.9% of enrollees) occurred in positive clusters (p < 0.01; attributable risk [AR] = 10.4 per 100; 95% confidence interval 1–19.8 per 100]. In positive clusters, 12.4% of enrollees became infected in a 15-d period and DENV infections were aggregated centrally near homes of index cases. As only 1 of 217 pairs of serologic specimens tested in positive clusters revealed a recent DENV infection that occurred prior to cluster initiation, we attribute the observed DENV transmission subsequent to cluster investigation to recent DENV transmission activity. Of the 1,022 female adult Ae. aegypti collected, all eight (0.8%) dengue-infected mosquitoes came from houses in positive clusters; none from control clusters or schools. Distinguishing features between positive and negative clusters were greater availability of piped water in negative clusters (p < 0.01) and greater number of Ae. aegypti pupae per person in positive clusters (p = 0.04). During primarily DENV-4 transmission seasons, the ratio of inapparent to symptomatic infections was nearly 1:1 among child enrollees. Study limitations included inability to sample all children and mosquitoes within each cluster and our reliance on serologic rather than virologic evidence of interval infections in enrollees given restrictions on the frequency of blood collections in children. Conclusions Our data reveal the remarkably focal nature of DENV transmission within a hyperendemic rural area of Thailand. These data suggest that active school-based dengue case detection prompting local spraying could contain recent virus introductions and reduce the longitudinal risk of virus spread within rural areas. Our results should prompt future cluster studies to explore how host immune and behavioral aspects may impact DENV transmission and prevention strategies. Cluster methodology could serve as a useful research tool for investigation of other temporally and spatially clustered infectious diseases.

Serotype-Specific Differences in the Risk of Dengue Hemorrhagic Fever: An Analysis of Data Collected in Bangkok, Thailand from 1994 to 2006

Background It is unclear whether dengue serotypes differ in their propensity to cause severe disease. We analyzed differences in serotype-specific disease severity in children presenting for medical attention in Bangkok, Thailand. Methodology/Principal Findings Prospective studies were conducted from 1994 to 2006. Univariate and multivariate logistic and multinomial logistic regressions were used to determine if dengue hemorrhagic fever (DHF) and signs of severe clinical disease (pleural effusion, ascites, thrombocytopenia, hemoconcentration) were associated with serotype. Crude and adjusted odds ratios were calculated. There were 162 (36%) cases with DENV-1, 102 (23%) with DENV-2, 123 (27%) with DENV-3, and 64 (14%) with DENV-4. There was no significant difference in the rates of DHF by serotype: DENV-2 (43%), DENV-3 (39%), DENV-1 (34%), DENV-4 (31%). DENV-2 was significantly associated with increased odds of DHF grade I compared to DF (OR 2.9 95% CI 1.1, 8.0), when using DENV-1 as the reference. Though not statistically significant, DENV-2 had an increased odds of total DHF and DHF grades II, III, and IV. Secondary serologic response was significantly associated with DHF (OR 6.2) and increased when considering more severe grades of DHF. DENV-2 (9%) and -4 (3%) were significantly less often associated with primary disease than DENV-1 (28%) and -3 (33%). Restricting analysis to secondary cases, we found DENV-2 and DENV-3 to be twice as likely to result in DHF as DEN-4 (p = 0.05). Comparing study years, we found the rate of DHF to be significantly less in 1999, 2000, 2004, and 2005 than in 1994, the study year with the highest percentage of DHF cases, even when controlling for other variables. Conclusions/Significance As in other studies, we find secondary disease to be strongly associated with DHF and with more severe grades of DHF. DENV-2 appears to be marginally associated with more severe dengue disease as evidenced by a significant association with DHF grade I when compared to DENV-1. In addition, we found non-significant trends with other grades of DHF. Restricting the analysis to secondary disease we found DENV-2 and -3 to be twice as likely to result in DHF as DEN-4. Differences in severity by study year may suggest that other factors besides serotype play a role in disease severity.

Virus-Induced Decline in Soluble Vascular Endothelial Growth Receptor 2 Is Associated with Plasma Leakage in Dengue Hemorrhagic Fever

ABSTRACT Some individuals infected with dengue virus develop dengue hemorrhagic fever (DHF), a viral hemorrhagic disease characterized by a transient period of localized plasma leakage. To determine the importance of vascular endothelial growth factor A (VEGF-A) in this syndrome, we compared plasma levels of VEGF-A and the soluble forms of its receptors in patients with DHF to patients with dengue fever (DF), a milder form of dengue virus infection without plasma leakage. We observed a rise in the plasma levels of free, but not total VEGF-A in DHF patients at the time of plasma leakage. This was associated with a decline in the soluble form of VEGF receptor 2 (VEGFR2) and VEGF-soluble VEGFR2 complexes, but not the soluble form of VEGFR1. The severity of plasma leakage in patients inversely correlated with plasma levels of soluble VEGFR2. In vitro, dengue virus suppressed soluble VEGFR2 production by endothelial cells but up-regulated surface VEGFR2 expression and promoted response to VEGF stimulation. In vivo, plasma viral load correlated with the degree of decline in plasma soluble VEGFR2. These results suggest that VEGF regulates vascular permeability and its activity is controlled by binding to soluble VEGFR2. Dengue virus-induced changes in surface and soluble VEGFR2 expression may be an important mechanism of plasma leakage in DHF.

Markers of dengue disease severity.

Infection with one of the four serotypes of dengue virus (DENV) causes a wide spectrum of clinical disease ranging from asymptomatic infection, undifferentiated fever, dengue fever (DF) to dengue hemorrhagic fever (DHF). DHF occurs in a minority of patients and is characterized by bleeding and plasma leakage which may lead to shock. There are currently no reliable clinical or laboratory indicators that accurately predict the development of DHF. Human studies have shown that high viral load and intense activation of the immune system are associated with DHF. Recently, endothelial cells and factors regulating vascular permeability have been demonstrated to play a role. In the absence of animal models that closely mimic DHF, human studies are essential in identifying predictors of severe illness. Well planned prospective studies with samples collected at different time points of the illness in well characterized patients are crucial for this effort. Ideally, clinical and laboratory predictive tools should be suitable for resource poor countries where dengue is endemic.

Natural History of Plasma Leakage in Dengue Hemorrhagic Fever A Serial Ultrasonographic Study

Background: Although plasma leakage is the major cause of mortality and morbidity in patients with dengue hemorrhagic fever (DHF), a detailed assessment of the natural course of this process is still lacking. We employed serial ultrasound examination to delineate the locations and the timing of plasma leakage and to evaluate the usefulness of ultrasound in detecting plasma leakage in DHF. Method: Daily ultrasound examinations of the abdomen and right thorax were performed in 158 suspected dengue cases to detect ascites, thickened gall bladder wall and pleural effusions. Cases were classified into dengue fever (DF), DHF or other febrile illness (OFI) based on serology and evidence of plasma leakage including hemoconcentration and pleural effusion detected by chest radiograph. Results: Ultrasonographic evidence of plasma leakage was detected in DHF cases starting from 2 days before defervescence and was detected in some cases within 3 days after fever onset. Pleural effusion was the most common ultrasonographic sign of plasma leakage (62% of DHF cases one day after defervescence). Thickening of the gallbladder wall and ascites were detected less frequently (43% and 52% of DHF cases respectively) and resolved more rapidly than pleural effusions. The size of pleural effusions, ascites and gall bladder wall thickness in DHF grade I and II were smaller than those of grade III patients. Ultrasound detected plasma leakage in 12 of 17 DHF cases who did not meet the criteria for significant hemoconcentration. Conclusions: Ultrasound examinations detected plasma leakage in multiple body compartments around the time of defervescence. Ultrasonographic signs of plasma leakage were detectable before changes in hematocrits. Ultrasound is a useful tool for detecting plasma leakage in dengue infection.

Protection From Arthritis and Myositis in a Mouse Model of Acute Chikungunya Virus Disease by Bindarit, an Inhibitor of Monocyte Chemotactic Protein-1 Synthesis

Chikungunya virus (CHIKV) is associated with outbreaks of infectious rheumatic disease in humans. Using a mouse model of CHIKV arthritis and myositis, we show that tumor necrosis factor-α, interferon-γ, and monocyte chemotactic protein 1 (MCP-1) were dramatically induced in tissues from infected mice. The same factors were detected in the serum of patients with CHIKV-induced polyarthralgia and polyarthritis, with MCP-1 levels being particularly elevated. Bindarit (MCP inhibitor) treatment ameliorated CHIKV disease in mice. Histological analysis of muscle and joint tissues showed a reduction in inflammatory infiltrate in infected mice treated with bindarit. These results suggest that bindarit may be useful in treating CHIKV-induced arthritides in humans.

A Shorter Time Interval Between First and Second Dengue Infections Is Associated With Protection From Clinical Illness in a School-based Cohort in Thailand

BACKGROUND Despite the strong association between secondary dengue virus (DENV) infections and dengue hemorrhagic fever (DHF), the majority of secondary infections are subclinical or mild. The determinants of clinical severity remain unclear, though studies indicate a titer-dependent and time-dependent role of cross-protective anti-DENV antibodies. METHODS Data from 2 sequential prospective cohort studies were analyzed for subclinical and symptomatic DENV infections in schoolchildren in Kamphaeng Phet, Thailand (1998-2002 and 2004-2007). Children experiencing ≥ 1 DENV infection were selected as the population for analysis (contributing 2169 person-years of follow-up). RESULTS In total, 1696 children had ≥ 1 DENV infection detected during their enrollment; 268 experienced 2 or more infections. A shorter time interval between infections was associated with subclinical infection in children seronegative for DENV at enrollment, for whom a second-detected DENV infection is more likely to reflect a true second infection (average of 2.6 years between infections for DHF, 1.9 for DF, and 1.6 for subclinical infections). CONCLUSIONS These findings support a pathogenesis model where cross-reactive antibodies wane from higher-titer, protective levels to lower-titer, detrimental levels. This is one of the first studies of human subjects to suggest a window of cross-protection following DENV infection since Sabins challenge studies in the 1940s.

Plasma leakage in dengue haemorrhagic fever

Dengue viruses (DENV), a group of four serologically distinct but related flaviviruses, are the cause of one of the most important emerging viral diseases. DENV infections result in a wide spectrum of clinical disease including dengue haemorrhagic fever (DHF), a viral haemorrhagic disease characterised by bleeding and plasma leakage. The characteristic feature of DHF is the transient period of plasma leakage and a haemorrhagic tendency. DHF occurs mostly during a secondary DENV infection. Serotype cross-reactive antibodies and mediators from serotype cross-reactive Dengue-specific T cells have been implicated in the pathogenesis. A complex interaction between virus, host immune response and endothelial cells likely impacts the barrier integrity and functions of endothelial cells leading to plasma leakage. Recently the role of angiogenic factors and the role of dengue virus on endothelial cell transcription and functions have been studied. Insights into the mechanisms that confer protection or cause disease are critical in the development of prophylactic and therapeutic modalities for this important disease.

Dengue hemorrhagic fever: the sensitivity and specificity of the world health organization definition for identification of severe cases of dengue in Thailand, 1994-2005.

BACKGROUND Dengue virus infection causes a spectrum of clinical manifestations, usually classified according to the World Health Organization (WHO) guidelines into dengue fever (DF) and dengue hemorrhagic fever (DHF). The ability of these guidelines to categorize severe dengue illness has recently been questioned. METHODS We evaluated dengue case definitions in a prospective study at a pediatric hospital in Bangkok, Thailand, during 1994-2005. One thousand thirteen children were enrolled within the first 3 days after onset of fever and observed with standardized data collection. Cases were classified on the basis of application of the strict WHO criteria. All dengue virus infections were laboratory confirmed. We retrospectively grouped patients on the basis of whether they received significant intervention based on fluid replacement and/or requirements for blood transfusion. RESULTS Eighty-five (58%) of 150 persons with DHF, 40 (15%) of 264 with DF, and 73 (12%) of 599 with other febrile illnesses (OFIs) received significant intervention. Sixty-eight percent of dengue cases requiring intervention met strict WHO criteria for DHF. In contrast, only 1% of OFI cases met WHO criteria for DHF. Plasma leakage and thrombocytopenia were the 2 components contributing to the specificity of the WHO case definition and identified dengue cases that required intervention. Hemorrhagic tendency did not reliably differentiate DF and DHF. In DF cases, thrombocytopenia and bleeding were associated with severity. CONCLUSIONS Dengue illness is heterogeneous in severity, and severe clinical features occurred in patients whose cases were not characterized as DHF. The WHO case definition of DHF demonstrated sensitivity of 62% and specificity of 92% for identification of dengue illness requiring intervention, without the need for laboratory confirmation of dengue virus infection, in an area of endemicity.

Fine Scale Spatiotemporal Clustering of Dengue Virus Transmission in Children and Aedes aegypti in Rural Thai Villages

Background Based on spatiotemporal clustering of human dengue virus (DENV) infections, transmission is thought to occur at fine spatiotemporal scales by horizontal transfer of virus between humans and mosquito vectors. To define the dimensions of local transmission and quantify the factors that support it, we examined relationships between infected humans and Aedes aegypti in Thai villages. Methodology/Principal Findings Geographic cluster investigations of 100-meter radius were conducted around DENV-positive and DENV-negative febrile “index” cases (positive and negative clusters, respectively) from a longitudinal cohort study in rural Thailand. Child contacts and Ae. aegypti from cluster houses were assessed for DENV infection. Spatiotemporal, demographic, and entomological parameters were evaluated. In positive clusters, the DENV infection rate among child contacts was 35.3% in index houses, 29.9% in houses within 20 meters, and decreased with distance from the index house to 6.2% in houses 80–100 meters away (p<0.001). Significantly more Ae. aegypti were DENV-infectious (i.e., DENV-positive in head/thorax) in positive clusters (23/1755; 1.3%) than negative clusters (1/1548; 0.1%). In positive clusters, 8.2% of mosquitoes were DENV-infectious in index houses, 4.2% in other houses with DENV-infected children, and 0.4% in houses without infected children (p<0.001). The DENV infection rate in contacts was 47.4% in houses with infectious mosquitoes, 28.7% in other houses in the same cluster, and 10.8% in positive clusters without infectious mosquitoes (p<0.001). Ae. aegypti pupae and adult females were more numerous only in houses containing infectious mosquitoes. Conclusions/Significance Human and mosquito infections are positively associated at the level of individual houses and neighboring residences. Certain houses with high transmission risk contribute disproportionately to DENV spread to neighboring houses. Small groups of houses with elevated transmission risk are consistent with over-dispersion of transmission (i.e., at a given point in time, people/mosquitoes from a small portion of houses are responsible for the majority of transmission).