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Background: Although child undernutrition and stunting has been decreasing worldwide while obesity rates increase, these extreme conditions might coexist in families from low- and middle-income countries. We examined the association between maternal and child anthropometric indicators using a population representative sample. Methods: 4,258 non-pregnant women and their children <60 months who participated in the 2006 Brazilian Demographic Health Survey. We compared the distributions of two nutritional indexes of children, height-for-age (HAZ) and body mass index-for age (BAZ) z-scores, by categories of maternal height, body mass index (BMI), and waist circumference (WC). Adjusted mean differences and 95% confidence intervals (95% CI) were estimated from linear regression, taking into account the complex survey design. We also examined the associations of maternal anthropometry with the prevalence of child stunting (HAZ<−2) and overweight/obesity (BAZ>2). Results: HAZ was positively associated with maternal height and WC in a linear fashion. After adjustment, for sociodemographic characteristics, children whose mothers' height was<145 cm had 1.2 lower HAZ than children whose mothers were ≥160 cm tall (p-trend<0.0001). After further adjustment for maternal height and maternal BMI, children of mothers with a waist circumference ≥88 cm had 0.3 higher HAZ than those of mothers with WC<80 cm (p-trend<0.01). Adjusted prevalence ratios and 95% CI for stunting by the categories of maternal height (<145, 145–149, 150–154, 155–159 and ≥160 cm) were, respectively, 2.95 (1.51;5.77), 2.29 (1.33;3.93), 1.09 (0.63;1.87), and 0.89 (0.45;1.77), (p-trend = 0.001). BAZ was positively associated with maternal BMI and WC. Conclusion: We observed a strong, positive association of maternal and child nutritional status. Mothers of low stature had children with lower stature, mothers with central obesity had taller children, and mothers with overall or abdominal obesity had children with higher BAZ. [ABSTRACT FROM AUTHOR]

Background: Road traffic injuries (RTI) are a major public health epidemic killing thousands of people daily. Low and middle-income countries, such as Brazil, have the highest annual rates of road traffic fatalities. In order to improve road safety, this study mapped road traffic fatalities on a Brazilian highway to determine the main environmental factors affecting road traffic fatalities. Methods and Findings: Four techniques were utilized to identify and analyze RTI hotspots. We used spatial analysis by points by applying kernel density estimator, and wavelet analysis to identify the main hot regions. Additionally, built environment analysis, and principal component analysis were conducted to verify patterns contributing to crash occurrence in the hotspots. Between 2007 and 2009, 379 crashes were notified, with 466 fatalities on BR277. Higher incidence of crashes occurred on sections of highway with double lanes (ratio 2∶1). The hotspot analysis demonstrated that both the eastern and western regions had higher incidences of crashes when compared to the central region. Through the built environment analysis, we have identified five different patterns, demonstrating that specific environmental characteristics are associated with different types of fatal crashes. Patterns 2 and 4 are constituted mainly by predominantly urban characteristics and have frequent fatal pedestrian crashes. Patterns 1, 3 and 5 display mainly rural characteristics and have higher prevalence of vehicular collisions. In the built environment analysis, the variables length of road in urban area, limited lighting, double lanes roadways, and less auxiliary lanes were associated with a higher incidence of fatal crashes. Conclusions: By combining different techniques of analyses, we have identified numerous hotspots and environmental characteristics, which governmental or regulatory agencies could make use to plan strategies to reduce RTI and support life-saving policies. [ABSTRACT FROM AUTHOR]

Malaria is a significant public health threat in the Brazilian Amazon. Previous research has shown that deforestation creates breeding sites for the main malaria vector in Brazil, Anopheles darlingi, but the influence of selective logging, forest fires, and road construction on malaria risk has not been assessed. To understand these impacts, we constructed a negative binomial model of malaria counts at the municipality level controlling for human population and social and environmental risk factors. Both paved and unpaved roadways and fire zones in a municipality increased malaria risk. Within the timber production states where 90% of deforestation has occurred, compared with areas without selective logging, municipalities where 0–7% of the remaining forests were selectively logged had the highest malaria risk (1.72, 95% CI 1.18–2.51), and areas with higher rates of selective logging had the lowest risk (0.39, 95% CI 0.23–0.67). We show that roads, forest fires, and selective logging are previously unrecognized risk factors for malaria in the Brazilian Amazon and highlight the need for regulation and monitoring of sub-canopy forest disturbance. [ABSTRACT FROM AUTHOR]

The present study investigated the prevalence of HIV-1 multiple infections in a population composed by 47 patients under HAART failure and enrolled at the National DST/AIDS, Program, Ministry of Health, Brazil.Detection of multiple infections was done using a previously published RFLP assay for the HIV-1 protease gene, which is able of distinguishing between infections caused by a single or multiple HIV-1 subtypes. Samples with multiple infections were cloned, and sequence data submitted to phylogenetic analysis. We were able to identify 17 HIV-1 multiple infections out of 47 samples. Multiple infections were mostly composed by a mixture of recombinant viruses (94%), with only one case in which protease gene pure subtypes B and F were recovered. This is the first study that reports the prevalence of multiple infections and intersubtype recombinants in a population undergoing HAART in Brazil. Based on the data there was a steep increase of multiple infections after the introduction of the combined antiretroviral therapy in Brazil. Cases of multiple infections may be associated with HIV-1 genetic diversity through recombination allowing for the generation of viruses showing a combination of resistance mutations. [ABSTRACT FROM AUTHOR]

Background: American visceral leishmaniasis is caused by the protozoan Leishmania infantum. Dogs are the main reservoirs in the domestic transmission cycle. The limited accuracy of diagnostic tests for canine leishmaniasis may contribute to the lack of impact of control measures recommended by the Brazilian Ministry of Health. The objective of this study was to estimate the accuracy of two enzyme-linked immunosorbent assays employing L. major or L. infantum antigens and their reliability between three laboratories of different levels of complexity. Methods: A validation study of ELISA techniques using L. major or L. infantum antigens was conducted. Direct visualization of the parasite in hematoxylin/eosin-stained histopathological sections, immunohistochemistry, and isolation of the parasite in culture.were used as gold standard. An animal that was positive in at least one of the tests was defined as infected with L. infantum. Serum samples collected from 1,425 dogs were analyzed. Samples were separated in three aliquots and tested in three different laboratories. Sensitivity, specificity and the area under de ROC curve were calculated and the reliability was evaluated between the participant laboratories. Results: The sensitivity was 91.8% and 89.8% for the L. major and L. infantum assays, respectively. The specificity was 83.75% and 82.7% for the L. major and L. infantum assays, respectively. The area under de ROC curve was 0.920 and 0.898 for L. major and L. infantum, respectively. The mean intraclass correlation coefficients between laboratories ranged from 0.890 to 0.948 when L. major was used as antigen, and from 0.818 to 0.879 when L. infantum was used. Interpretation: ELISA tests using L. major or L. infantum antigens have similar accuracy and reliability. Our results do not support the substitution of the L. major antigen of the ELISA test currently used for the diagnosis of canine visceral leishmaniasis in Brazil. [ABSTRACT FROM AUTHOR]

Aedes aegypti has developed evolution-driven adaptations for surviving in the domestic human habitat. Several trap models have been designed considering these strategies and tested for monitoring this efficient vector of Dengue. Here, we report a real-scale evaluation of a system for monitoring and controlling mosquito populations based on egg sampling coupled with geographic information systems technology. The SMCP-Aedes, a system based on open technology and open data standards, was set up from March/2008 to October/2011 as a pilot trial in two sites of Pernambuco -Brazil: Ipojuca (10,000 residents) and Santa Cruz (83,000), in a joint effort of health authorities and staff, and a network of scientists providing scientific support. A widespread infestation by Aedes was found in both sites in 2008–2009, with 96.8%–100% trap positivity. Egg densities were markedly higher in SCC than in Ipojuca. A 90% decrease in egg density was recorded in SCC after two years of sustained control pressure imposed by suppression of >7,500,000 eggs and >3,200 adults, plus larval control by adding fishes to cisterns. In Ipojuca, 1.1 million mosquito eggs were suppressed and a 77% reduction in egg density was achieved. This study aimed at assessing the applicability of a system using GIS and spatial statistic analysis tools for quantitative assessment of mosquito populations. It also provided useful information on the requirements for reducing well-established mosquito populations. Results from two cities led us to conclude that the success in markedly reducing an Aedes population required the appropriate choice of control measures for sustained mass elimination guided by a user-friendly mosquito surveillance system. The system was able to support interventional decisions and to assess the program’s success. Additionally, it created a stimulating environment for health staff and residents, which had a positive impact on their commitment to the dengue control program. [ABSTRACT FROM AUTHOR]

Background: A growing literature suggests that low birth weight increases the risk of poor health outcomes in adulthood. We tested this hypothesis among young adults living in São Paulo State, Brazil. Methods and Findings: To identify the effects of low birth weight on young adulthood outcomes, a medical assessment of 297 individuals born between 1977 and 1989 was conducted at a primary care unit in São Paulo State, Brazil. We analyzed body mass index (BMI), waist-hip ratio, blood pressure, fasting glucose and total cholesterol levels using linear and logistic regressions. Low birth was negatively associated with BMI (β = −2.0, 95% CI: −3.69, −0.27, p = 0.02), fasting glucose levels (β = −1.9, 95% CI: −3.9, −0.07, p = 0.05), waist-hip ratio (β = −0.03, 95% CI: −0.07, −0.01, p = 0.10), systolic blood pressure (β = −3.32, 95% CI: −7.60, 0.96, p = 0.12), and total cholesterol levels (β = −3.19, 95% CI: −16.43, 10.05, p = 0.636). Low birth weight was also associated with lower odds of young adults being overweight and obese, but neither association was statistically significant. Weight gain in the first 12 months of life was associated with higher adult BMI (β = 0.79, 95% CI: −0.0455, 1.623, p = 0.064) and blood pressure (β = 2.79, 95% CI: 0.22, 5.35, p = 0.034). No associations were found between low birth weight and early life (catch-up) growth. Conclusions: Low birth weight was not associated with poor health outcomes among young adults in Brazil. These results appear inconsistent with the original Barker hypothesis, but will need to be corroborated in larger samples with longer follow-ups to allow a more general evaluation of the validity of the hypothesis in low and middle income countries. [ABSTRACT FROM AUTHOR]

Background: Previous studies showed higher sepsis mortality rates in Brazil compared to other developed or developing countries. Moreover, another trial demonstrated an increased mortality rate in public hospitals compared to private hospitals in Brazil. The reasons for these findings may include delayed recognition and inadequate treatment of sepsis in public facilities. We designed this study to evaluate the factors associated with mortality in septic patients admitted to intensive care units in a network of public and private institutions. Materials and Methods: This study is a retrospective analysis of a prospective cohort of sepsis patients in 19 private and public institutions in Brazil. We analyzed data from the original database and collected additional data to assess compliance to the treatment guidelines and to determine the time from the onset of organ dysfunction and the sepsis diagnosis by the healthcare team. Results: A total of 396 patients were analyzed. Patients in public hospitals were younger, had a greater number of dysfunctional organs at baseline and a lower chance to have sepsis diagnosed within two hours of the onset of organ dysfunction. Private hospitals had a better compliance to lactate and blood culture sampling and maintenance of glycemic control. The multivariate analysis showed that age, disease severity at baseline and being treated at a public hospital were independent risk factors for mortality. A delay in the sepsis diagnosis of longer than two hours was associated with mortality only in the public setting. Conclusions: We confirmed a lower sepsis mortality rate in the private hospitals of this network. Being treated in a public hospital was an independent factor for mortality. Delayed recognition of sepsis was more frequent in public institutions and this might have been associated with a higher mortality. Improving sepsis recognition and early diagnosis may be important targets in public institutions. [ABSTRACT FROM AUTHOR]

Background: Bacterial meningitis is associated with significant burden in Brazil. In 2010, both 10-valent pneumococcal conjugate vaccine and meningococcal capsular group C conjugate vaccine were introduced into the routine vaccination schedule. Haemophilus influenzae type b vaccine was previously introduced in 1999. This study presents trends in demographics, microbiological characteristics and seasonality patterns of bacterial meningitis cases in Brazil from 2000 to 2010. Methods and Findings: All meningitis cases confirmed by clinical and/or laboratory criteria notified to the national information system for notifiable diseases between 2000 and 2010 were analyzed. Proportions of bacterial meningitis cases by demographic characteristics, criteria used for confirmation and etiology were calculated. We estimated disease rates per 100,000 population and trends for the study period, with emphasis on H. influenzae, N. meningitidis and S. pneumoniae cases. In the decade, 341,805 cases of meningitis were notified in Brazil. Of the 251,853 cases with defined etiology, 110,264 (43.8%) were due to bacterial meningitis (excluding tuberculosis). Of these, 34,997 (31.7%) were due to meningococcal disease. The incidence of bacterial meningitis significantly decreased from 3.1/100,000 population in 2000–2002 to 2.14/100,000 in 2009–2010 (p<0.01). Among cases of meningococcal disease, the proportion of those associated with group C increased from 41% in 2007 to 61.7% in 2010, while the proportion of group B disease progressively declined. Throughout the study period, an increased number of cases occurred during winter. Conclusions: Despite the reduction in bacterial meningitis incidence during the last decade, it remains a significant healthcare issue in Brazil. Meningococcal disease is responsible for the majority of the cases with group C the most common capsular type. Our study demonstrates the appropriateness of introduction of meningococcal vaccination in Brazil. Furthermore, this study provides a baseline for future evaluation of the impact of the vaccines introduction in Brazil and changes in disease epidemiology. [ABSTRACT FROM AUTHOR]

A total of 179 individuals with acute Chagas disease mainly transmitted by oral source, from Pará and Amapá State, Amazonian, Brazil were included during the period from 1988 to 2005. Blood samples were used to survey peripheral blood for T. cruzi hemoparasites by quantitative buffy coat (QBC), indirect xenodiagnosis, blood culture and serology to detection of total IgM and anti-T. cruzi IgG antibodies by indirect immunofluorescence assay (IFA) and indirect hemagglutination assay (HA). All assays were performed pre-treatment (0 days) and repeated 35 (±7) and 68 (±6) days after the initiation of treatment with benznidazol and every 6 months while remained seropositive. The endpoint of collection was performed in 2005. Total medium period of follow-up per person was 5.6 years. Also, a blood sample was collected from 72 randomly chosen treated patients to perform polimerase chain reaction (PCR) method. Proportions of subjects with negative or positive serology according to the number of years after treatment were compared. In the endpoint of follow-up we found 47 patients (26.7%) serologically negative, therefore considered cured and 5 (2.7%) exhibited mild cardiac Chagas disease. Other 132 patients had persistent positive serologic tests. The PCR carried out in 72 individuals was positive in 9.8%. Added, there was evidence of therapeutic failure immediately following treatment, as demonstrated by xenodiagnosis and blood culture methods in 2.3% and 3.5% of cases, respectively. There was a strong evidence of antibody clearing in the fourth year after treatment and continuous decrease of antibody titers. Authors suggest that control programs should apply operational researches with new drug interventions four years after the acute phase for those treated patients with persistently positive serology. [ABSTRACT FROM AUTHOR]

The American/Asian genotype of Dengue virus type 2 (DENV-2) was introduced into the Americas in the 80′s. Although there is no data showing when this genotype was first introduced into Brazil, it was first detected in Brazil in 1990. After which the virus spread throughout the country and major epidemics occurred in 1998, 2007/08 and 2010. In this study we sequenced 12 DENV-2 genomes obtained from serum samples of patients with dengue fever residing in São José do Rio Preto, São Paulo (SJRP/SP), Brazil, in 2008. The whole open reading frame or envelope sequences were used to perform phylogenetic, phylogeographic and evolutionary analyses. Isolates from SJRP/SP were grouped within one lineage (BR3) close to isolates from Rio de Janeiro, Brazil. Isolates from SJRP were probably introduced there at least in 2007, prior to its detection in the 2008 outbreak. DENV-2 circulation in Brazil is characterized by the introduction, displacement and circulation of three well-defined lineages in different times, most probably from the Caribbean. Thirty-seven unique amino acid substitutions were observed among the lineages, including seven amino acid differences in domains I to III of the envelope protein. Moreover, we dated here, for the first time, the introduction of American/Asian genotype into Brazil (lineage BR1) to 1988/89, followed by the introduction of lineages BR2 (1998–2000) and BR3 (2003–05). Our results show a delay between the introduction and detection of DENV-2 lineages in Brazil, reinforcing the importance and need for surveillance programs to detect and trace the evolution of these viruses. Additionally, Brazilian DENV-2 differed in genetic diversity, date of introduction and geographic origin and distribution in Brazil, and these are important factors for the evolution, dynamics and control of dengue. [ABSTRACT FROM AUTHOR]

Background: High technology in the field of interventional cardiology applied in tertiary hospitals has brought enormous benefits in the treatment of ischemic heart disease (IHD). However, IHD mortality rates remain high. We analyzed the relationship between IHD mortality rate and the socioeconomic, demographic, and geographic conditions in 399 cities in Parana state, Brazil, from 2006 to 2010. Methods and Results: Data were obtained from the Mortality Information System and the Brazilian Institute of Geography and Statistics and evaluated through Exploratory Spatial Data Analysis. GeoDa™ was used to analyze 29.351 deaths across 399 cities. We found a positive spatial autocorrelation regarding IHD mortality (I = 0.5913, p = 0.001). There was a significant positive association between each of three socioeconomic and demographic indicators and IHD mortality rate: Population Elderly Index (I = 0.3436), Illiteracy Rate (I = 0.1873) and City Development Index (I = 0.0900). In addition, two indicators presented significant negative association with IHD mortality rate: Adjusted Population Size (I = −0.1216) and Gross Domestic Product (I = −0.0864). We also found a positive association between IHD mortality rates and the geographic distances between patients’ city of residence and their corresponding regional referral centers in interventional cardiology (I = 0.3368). Cities located within Regional Health Units with Reference Interventional Cardiology Center presented a significantly lower average specific mortality rate by IHD. The high mortality rate by IHD within the Regional Health Units was not restricted to socioeconomic and demographic variables, but dependent on the distance between each city and their reference interventional cardiology center. Conclusions: We conclude that geographic factors play a significant role in IHD mortality within cities. These findings have important policy implications regarding the geographic distribution of cardiac health care networks in Latin America and in other emerging countries. [ABSTRACT FROM AUTHOR]

Previous work suggests that Brazilian Plasmodium falciparum has limited genetic diversity and a history of bottlenecks, multiple reintroductions due to human migration, and clonal expansions. We hypothesized that Brazilian P. falciparum would exhibit clonal structure. We examined isolates collected across two decades from Amapá, Rondônia, and Pará state (n = 190). By examining more microsatellites markers on more chromosomes than previous studies, we hoped to define the extent of low diversity, linkage disequilibrium, bottlenecks, population structure, and parasite migration within Brazil. We used retrospective genotyping of samples from the 1980s and 1990s to explore the population genetics of SP resistant dhfr and dhps alleles. We tested an existing hypothesis that the triple mutant dhfr mutations 50R/51I/108N and 51I/108N/164L developed in southern Amazon from a single origin of common or similar parasites. We found that Brazilian P. falciparum had limited genetic diversity and isolation by distance was rejected, which suggests it underwent bottlenecks followed by migration between sites. Unlike Peru, there appeared to be gene flow across the Brazilian Amazon basin. We were unable to divide parasite populations by clonal lineages and pairwise FST were common. Most parasite diversity was found within sites in the Brazilian Amazon, according to AMOVA. Our results challenge the hypothesis that triple mutant alleles arose from a single lineage in the Southern Amazon. SP resistance, at both the double and triple mutant stages, developed twice and potentially in different regions of the Brazilian Amazon. We would have required samples from before the 1980s to describe how SP resistance spread across the basin or describe the complex internal migration of Brazilian parasites after the colonization efforts of past decades. The Brazilian Amazon basin may have sufficient internal migration for drug resistance reported in any particular region to rapidly spread to other parts of basin under similar drug pressure. [ABSTRACT FROM AUTHOR]

Background: Neuromyelitis optica (NMO) is considered relatively more common in non-Whites, whereas multiple sclerosis (MS) presents a high prevalence rate, particularly in Whites from Western countries populations. However, no study has used ancestry informative markers (AIMs) to estimate the genetic ancestry contribution to NMO patients. Methods: Twelve AIMs were selected based on the large allele frequency differences among European, African, and Amerindian populations, in order to investigate the genetic contribution of each ancestral group in 236 patients with MS and NMO, diagnosed using the McDonald and Wingerchuck criteria, respectively. All 128 MS patients were recruited at the Faculty of Medicine of Ribeirão Preto (MS-RP), Southeastern Brazil, as well as 108 healthy bone marrow donors considered as healthy controls. A total of 108 NMO patients were recruited from five Neurology centers from different Brazilian regions, including Ribeirão Preto (NMO-RP). Principal Findings: European ancestry contribution was higher in MS-RP than in NMO-RP (78.5% vs. 68.7%) patients. In contrast, African ancestry estimates were higher in NMO-RP than in MS-RP (20.5% vs. 12.5%) patients. Moreover, principal component analyses showed that groups of NMO patients from different Brazilian regions were clustered close to the European ancestral population. Conclusions: Our findings demonstrate that European genetic contribution predominates in NMO and MS patients from Brazil. [ABSTRACT FROM AUTHOR]

Cell cycle protein expression plays an important role in the pathophysiology of cervical cancer. However, few studies have attempted to correlate the use of these biomarkers with the clinical progression of the tumor. Objectives: 1) To analyze the expression of Ki-67, p53 and p16INK4a in cervical cancer, 2) to correlate the relative expression of these proteins as well as clinical parameters with the stage of disease, and 3) to determine the HPV DNA prevalence and subtype distribution. Methods: Tissue Micro-Arrays (TMA) from patients with invasive cervical cancer (ICC) and controls were analyzed. HPV DNA detection was done by PCR and in situ hybridization. Ki-67, p53 and p16INK4a were analyzed by immunohistochemistry; clinical data was derived from the chart review. Results: Advanced tumor stage (III and IV) was strongly associated (p<0.005) with advanced age (>55 years old), with more than four pregnancies and with the lack of formal education. HPV DNA was found in 94.3% of cases with the most prevalent types being HPV16 (67.5%), followed by HPV33 (12.0%) and HPV35 (3.6%). High expression of Ki-67 and p16 was more common in the advanced FIGO stages (p = 0.023). Women with HPV16 tended to be younger (50.9 years; SE 1.9) compared to women with other types (59.9 years; SE 2.8). Conclusion: We found that Ki-67 and p16 expression were independently associated with the tumor stage. We also noted that about 1/3 of the cervical cancers in this Brazilian cohort were not associated with HPV types directly targeted by the current HPV vaccines. [ABSTRACT FROM AUTHOR]

Background: Although known to be highly endemic in the Amazon regions of Brazil, the presence of cutaneous leishmaniasis (CL) in the subtropical southern part of the country has largely been ignored. This study was conducted to demonstrate CL is emerging in the Brazilian state of Santa Catarina, as well as to characterize the epidemiological profile and Leishmania species involved. Methodology/Principal Findings: For this cross-sectional study, data from all CL cases from Santa Catarina, Brazil, reported to the Brazilian National Notifiable Diseases Information System from 2001 to 2009 were investigated. Amplification of the kDNA minicircle conserved region followed by restriction fragment length polymorphism (PCR-RFLP) was conducted to screen for Leishmania species present in patient biopsy. Overall, 542 CL cases were reported, with majority resulting from autochthonous transmission (n = 401, 73.99%) and occurring in urban zones (n = 422, 77.86%). Age, gender, zone of residence, origin of case, clinical form and case outcome were found to differ significantly by region. Imported cases were over seven times more likely to relapse (95% CI 2.56–21.09). Mapping of cases revealed new endemic areas in northeastern Santa Catarina with two species present. With the exception of three L. (Leishmania) amazonensis cases (1.20%), majority of PCR positive samples were found to be L. (Viannia) braziliensis (n = 248, 98.80%). Conclusions/Significance: CL is now endemic in the state of Santa Catarina, Brazil, with case profiles varying significantly by region. L. (V.) braziliensis has been identified as the predominant species in the region. [ABSTRACT FROM AUTHOR]

This manuscript summarizes the papers presented at the 19th International Papillomavirus Conference, held in Florianopolis, Brazil, September 1–7, 2001, divided in four main areas: Clinical diagnosis and screening, Epidemiology, Biology and Immunology. It provides an overview of what we know about the biology and life cycle of these viruses, their interaction with human and animal hosts, and the diseases that they cause. Highlights derive from the analysis of more than 500 papers presented at the Conference. [Copyright &y& Elsevier]

The article presents the author's views on the detection of various dengue serotypes which are increasing in the hyperendemicity in urban areas of Brazil. Brief information on terms including dengue fever and hyperendemicity are presented. Four serotypes of the fever that are discussed includes DENV-1, DENV-2, DENV-3 and DENV-4.