[show abstract][hide abstract] ABSTRACT: Dengue is an acute illness caused by the positive-strand RNA dengue virus (DENV). There are four genetically distinct DENVs (DENV-1-4) that cause disease in tropical and subtropical countries. Most patients are viremic when they present with symptoms; therefore, RT-PCR has been increasingly used in dengue diagnosis. The CDC DENV-1-4 RT-PCR Assay has been developed as an in-vitro diagnostic platform and was recently approved by the US Food and Drug Administration (FDA) for detection of dengue in patients with signs or symptoms of mild or severe dengue. The primers and probes of this test have been designed to detect currently circulating strains of DENV-1-4 from around the world at comparable sensitivity. In a retrospective study with 102 dengue cases confirmed by IgM anti-DENV seroconversion in the convalescent sample, the RT-PCR Assay detected DENV RNA in 98.04% of the paired acute samples. Using sequencing as a positive indicator, the RT-PCR Assay had a 97.92% positive agreement in 86 suspected dengue patients with a single acute serum sample. After extensive validations, the RT-PCR Assay performance was highly reproducible when evaluated across three independent testing sites, did not produce false positive results for etiologic agents of other febrile illnesses, and was not affected by pathological levels of potentially interfering biomolecules. These results indicate that the CDC DENV-1-4 RT-PCR Assay provides a reliable diagnostic platform capable for confirming dengue in suspected cases.
[show abstract][hide abstract] ABSTRACT: Dengue is a potentially fatal acute febrile illness (AFI) caused by four mosquito-transmitted dengue viruses (DENV-1-4) that are endemic in Puerto Rico. In January 2010, the number of suspected dengue cases reported to the passive dengue surveillance system exceeded the epidemic threshold and an epidemic was declared soon after.
To characterize the epidemic, surveillance and laboratory diagnostic data were compiled. A suspected case was a dengue-like AFI in a person reported by a health care provider with or without a specimen submitted for diagnostic testing. Laboratory-positive cases had: (i) DENV nucleic acid detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in an acute serum specimen; (ii) anti-DENV IgM antibody detected by ELISA in any serum specimen; or (iii) DENV antigen or nucleic acid detected in an autopsy-tissue specimen. In 2010, a total of 26,766 suspected dengue cases (7.2 per 1,000 residents) were identified, of which 46.6% were laboratory-positive. Of 7,426 RT-PCR-positive specimens, DENV-1 (69.0%) and DENV-4 (23.6%) were detected more frequently than DENV-2 (7.3%) and DENV-3 (<0.1%). Nearly half (47.1%) of all laboratory-positive cases were adults, 49.7% had dengue with warning signs, 11.1% had severe dengue, and 40 died. Approximately 21% of cases were primary DENV infections, and 1-4 year olds were the only age group for which primary infection was more common than secondary. Individuals infected with DENV-1 were 4.2 (95% confidence interval [CI]: 1.7-9.8) and 4.0 (95% CI: 2.4-6.5) times more likely to have primary infection than those infected with DENV-2 or -4, respectively.
This epidemic was long in duration and yielded the highest incidence of reported dengue cases and deaths since surveillance began in Puerto Rico in the late 1960's. This epidemic re-emphasizes the need for more effective primary prevention interventions to reduce the morbidity and mortality of dengue.
[show abstract][hide abstract] ABSTRACT: Sequencing of dengue virus type 1 (DENV-1) strains isolated in Key West/Monroe County, Florida, indicate en-demic transmission for >2 years of a distinct and predomi-nant sublineage of the American–African genotype. DENV-1 strains isolated elsewhere in Florida grouped within a sepa-rate Central American lineage. Findings indicate endemic transmission of DENV into the continental United States. D engue is the most common mosquito-borne viral dis-ease; cases have been reported from ≈100 countries, and there are indications of increased incidence and sever-ity worldwide (1). The United States has reported year-round transmission of dengue virus (DENV) in Puerto Rico, the US Virgin Islands, and American Samoa and oc-casional transmission along the Texas–Mexico border. In the continental United States, DENV is the most frequent cause of febrile illness among travelers returning from the Caribbean, South America, and Asia (2,3). These frequent introductions of dengue infections and the increased pres-ence of vectors (i.e., Aedes aegypti and Ae. albopictus mosquitoes) in many US regions may portend the reintro-duction and extended transmission of DENV into the con-tinental United States. In September 2009, the Florida Department of Health (FDOH) and the Centers for Disease Control and Preven-tion (San Juan, Puerto Rico) investigated a case of DENV type 1 (DENV-1) infection in a person (index patient) who, as confirmed by reverse transcription PCR (RT-PCR), ac-quired the virus while traveling to Key West in Monroe County, Florida, USA. DENV-1 infections were subse-quently confirmed in 2 Monroe County residents without histories of recent travel. In addition, among 13 other cases in the county that were identified by serologic methods, 2 were confirmed as DENV-1 infections (4). Thus, a total of 5 DENV-1 cases were confirmed in Key West during 2009, and ≈5% of the serosurveyed population in Key West had evidence of recent DENV infection (4,5). In 2010, addi-tional dengue cases from Monroe County were reported, and DENV-1 was isolated from a mosquito pool (6) and a blood donor from Key West (7); isolates from the mosquito pool and blood donor appeared to be phylogenetically re-lated (7). This study determined the genetic relatedness of the DENV-1 isolates from dengue patients in Key West and 4 other Florida counties during 2009–2010, including the blood donor and mosquito isolates.
[show abstract][hide abstract] ABSTRACT: In June of 2007, West Nile virus (WNV) was detected in sentinel chickens and blood donors in Puerto Rico, where dengue virus (DENV) is hyperendemic. Enhanced human surveillance for acute febrile illness (AFI) began in eastern Puerto Rico on July 1, 2007. Healthcare providers submitted specimens from AFI cases for WNV and DENV virology and serology testing. Over 6 months, 385 specimens were received from 282 cases; 115 (41%) specimens were DENV laboratory-positive, 86 (31%) specimens were laboratory-indeterminate, and 32 (11%) specimens were laboratory-negative for WNV and DENV. One WNV infection was detected by anti-WNV immunoglobulin M (IgM) antibody and confirmed by a plaque reduction neutralization test. DENV and WNV infections could not be differentiated in 27 cases (10%). During a period of active WNV transmission, enhanced human surveillance identified one case of symptomatic WNV infection. Improved diagnostic methods are needed to allow differentiation of WNV and DENV in dengue-endemic regions.
The American journal of tropical medicine and hygiene 03/2013; · 2.53 Impact Factor
[show abstract][hide abstract] ABSTRACT: Dengue is endemic in the U.S. Virgin Islands, but no outbreaks have been reported since 2005. In November 2012, a school nurse in St. Croix reported suspected dengue in 27 (7%) of 369 students and staff members to the Virgin Islands Department of Health (VIDOH) and the CDC Dengue Branch in Puerto Rico. Four of 12 patient specimens sent to the CDC Dengue Branch for diagnostic testing were confirmed as dengue. Although VIDOH had observed an increase in passive dengue reporting, reliable baseline case counts were unavailable for comparison. An investigation was begun to determine the incidence of recent dengue virus (DENV) infection in schools and islandwide.
MMWR. Morbidity and mortality weekly report 03/2013; 62(09):172.
[show abstract][hide abstract] ABSTRACT: Background. The dengue virus serotype 3 (DENV-3) Indian subcontinent strain emerged in Puerto Rico in 1998 after a 21-year absence. The rapid expansion of DENV-3 on the island correlated with the withdrawal of the other serotypes for 7 years. The DENV-3 prevalence declined in 2008 and remains undetected. Methods. We sequenced complete genomes of 92 DENV-3 clinical isolates to characterize the molecular evolution and phylogeography throughout 10 years of continued sampling (1998-2007). Results. We documented 8 distinct lineages that emerged simultaneously and evolved independently. Two of the 8 lineages were highly associated with transient introductions of foreign viruses, and 2 of the 3 endemic lineages covered the entire study period. We found evidence of temporal-geographical clustering only within the 3 endemic lineages. The phylogeography analysis combined with serotype-specific incidence data showed that transmission of a DENV serotype in a given location and time is usually correlated with the absence of the other serotype. Conclusions. Our study shows the cotransmission of DENV-3 lineages through a complex dissemination pattern dissimilar to the evolutionary dynamics of the other serotypes in the island. High virus genetic diversity and a large naive population were underlying factors in the expansion and collapse of DENV-3 in Puerto Rico.
The Journal of Infectious Diseases 06/2012; 206(6):893-901. · 5.85 Impact Factor
[show abstract][hide abstract] ABSTRACT: TO THE EDITOR: Leptospirosis, caused by Leptospira spp. bacteria, and dengue, caused by dengue viruses (DENVs), are potentially fatal acute febrile illnesses (AFI) endemic to the tropics (1,2). Because their clinical manifestations are similar (3), leptospirosis may be misidentified as dengue (4). We report a fatal case of co-infection with Leptospira spp. and DENV-1 in a man in Puerto Rico.
[show abstract][hide abstract] ABSTRACT: The objective of the study was to evaluate if the antibodies elicited after immunization with a tetravalent dengue vaccine, based on chimeric yellow fever 17D/dengue viruses, can neutralize a large range of dengue viruses (DENV). A panel of 82 DENVs was developed from viruses collected primarily during the last decade in 30 countries and included the four serotypes and the majority of existing genotypes. Viruses were isolated and minimally amplified before evaluation against a tetravalent polyclonal serum generated during vaccine preclinical evaluation in monkey, a model in which protection efficacy of this vaccine has been previously demonstrated (Guirakhoo et al., 2004). Neutralization was observed across all the DENV serotypes, genotypes, geographical origins and isolation years. These data indicate that antibodies elicited after immunization with this dengue vaccine candidate should widely protect against infection with contemporary DENV lineages circulating in endemic countries.
[show abstract][hide abstract] ABSTRACT: The incidence and severity of dengue in Latin America has increased substantially in recent decades and data from Puerto Rico suggests an increase in severe cases. Successful clinical management of severe dengue requires early recognition and supportive care.
Fatal cases were identified among suspected dengue cases reported to two disease surveillance systems and from death certificates. To be included, fatal cases had to have specimen submitted for dengue diagnostic testing including nucleic acid amplification for dengue virus (DENV) in serum or tissue, immunohistochemical testing of tissue, and immunoassay detection of anti-DENV IgM from serum. Medical records from laboratory-positive dengue fatal case-patients were reviewed to identify possible determinants for death.
Among 10,576 reported dengue cases, 40 suspect fatal cases were identified, of which 11 were laboratory-positive, 14 were laboratory-negative, and 15 laboratory-indeterminate. The median age of laboratory-positive case-patients was 26 years (range 5 months to 78 years), including five children aged < 15 years; 7 sought medical care at least once prior to hospital admission, 9 were admitted to hospital and 2 died upon arrival. The nine hospitalized case-patients stayed a mean of 15 hours (range: 3-48 hours) in the emergency department (ED) before inpatient admission. Five of the nine case-patients received intravenous methylprednisolone and four received non-isotonic saline while in shock. Eight case-patients died in the hospital; five had their terminal event on the inpatient ward and six died during a weekend. Dengue was listed on the death certificate in only 5 instances.
During a dengue epidemic in an endemic area, none of the 11 laboratory-positive case-patients who died were managed according to current WHO Guidelines. Management issues identified in this case-series included failure to recognize warning signs for severe dengue and shock, prolonged ED stays, and infrequent patient monitoring.
[show abstract][hide abstract] ABSTRACT: In 2007, a total of 10,508 suspected dengue cases were reported in Puerto Rico. Blood donations were tested for dengue virus (DENV) RNA and recipients of RNA-positive donations traced to assess transfusion transmission.
Blood donation samples from 2007 were maintained in a repository and tested individually for DENV RNA by transcription-mediated amplification (TMA); a subset was further tested by an enhanced TMA (eTMA) assay. TMA-reactive samples were considered confirmed if TMA (including eTMA) was repeat reactive (RR). All TMA-RR samples were tested by quantitative, DENV type-specific reverse transcriptase-polymerase chain reaction (RT-PCR) and for anti-DENV immunoglobulin (Ig)M by enzyme-linked immunosorbent assay. Samples positive by RT-PCR were further tested for infectivity in mosquito cell culture. Patients receiving components from TMA-RR donations were followed.
Of 15,350 donation samples tested, 29 were TMA-RR for a prevalence of 1 per 529 (0.19%). DENV Types 1, 2, and 3 with viral titers of 10(5) to 10(9) copies/mL were detected by RT-PCR in 12 samples of which all were infectious in mosquito culture. Six TMA-RR samples were IgM positive. Three of the 29 recipients receiving TMA-RR donations were tested. One recipient in Puerto Rico transfused with red blood cells containing 10(8) copies/mL DENV-2 became febrile 3 days posttransfusion and developed dengue hemorrhagic fever. The recipient was DENV-2 RNA positive by RT-PCR; both the donor and the recipient viruses had identical envelope sequences.
High rates of viremia were detected in blood donors in Puerto Rico coupled with the first documented transfusion transmission of severe dengue disease, suggesting that further research on interventions is needed.
[show abstract][hide abstract] ABSTRACT: Dengue is an acute febrile illness caused by four mosquito-borne dengue viruses (DENV-1 to -4) that are endemic throughout the tropics. After returning from a 1-week missionary trip to Haiti in October of 2010, 5 of 28 (18%) travelers were hospitalized for dengue-like illness. All travelers were invited to submit serum specimens and complete questionnaires on pre-travel preparations, mosquito avoidance practices, and activities during travel. DENV infection was confirmed in seven (25%) travelers, including all travelers that were hospitalized. Viral sequencing revealed closest homology to a 2007 DENV-1 isolate from the Dominican Republic. Although most (88%) travelers had a pre-travel healthcare visit, only one-quarter knew that dengue is a risk in Haiti, and one-quarter regularly used insect repellent. This report confirms recent DENV transmission in Haiti. Travelers to DENV-endemic areas should receive dengue education during pre-travel health consultations, follow mosquito avoidance recommendations, and seek medical care for febrile illness during or after travel.
The American journal of tropical medicine and hygiene 01/2012; 86(1):16-22. · 2.53 Impact Factor
[show abstract][hide abstract] ABSTRACT: Dengue virus currently causes 50-100 million infections annually. Comprehensive knowledge about the evolution of Dengue in response to selection pressure is currently unavailable, but would greatly enhance vaccine design efforts. In the current study, we sequenced 187 new dengue virus serotype 3 (DENV-3) genotype III whole genomes isolated from Asia and the Americas. We analyzed them together with previously-sequenced isolates to gain a more detailed understanding of the evolutionary adaptations existing in this prevalent American serotype. In order to analyze the phylogenetic dynamics of DENV-3 during outbreak periods; we incorporated datasets of 48 and 11 sequences spanning two major outbreaks in Venezuela during 2001 and 2007-2008, respectively. Our phylogenetic analysis of newly sequenced viruses shows that subsets of genomes cluster primarily by geographic location, and secondarily by time of virus isolation. DENV-3 genotype III sequences from Asia are significantly divergent from those from the Americas due to their geographical separation and subsequent speciation. We measured amino acid variation for the E protein by calculating the Shannon entropy at each position between Asian and American genomes. We found a cluster of seven amino acid substitutions having high variability within E protein domain III, which has previously been implicated in serotype-specific neutralization escape mutants. No novel mutations were found in the E protein of sequences isolated during either Venezuelan outbreak. Shannon entropy analysis of the NS5 polymerase mature protein revealed that a G374E mutation, in a region that contributes to interferon resistance in other flaviviruses by interfering with JAK-STAT signaling was present in both the Asian and American sequences from the 2007-2008 Venezuelan outbreak, but was absent in the sequences from the 2001 Venezuelan outbreak. In addition to E, several NS5 amino acid changes were unique to the 2007-2008 epidemic in Venezuela and may give additional insight into the adaptive response of DENV-3 at the population level.
Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases 09/2011; 11(8):2011-9. · 3.22 Impact Factor
[show abstract][hide abstract] ABSTRACT: To study the evolution of dengue virus (DENV) serotype 2 in Puerto Rico, we examined the genetic composition and diversity of 160 DENV-2 genomes obtained through 22 consecutive years of sampling. A clade replacement took place in 1994-1997 during a period of high incidence of autochthonous DENV-2 and frequent, short-lived reintroductions of foreign DENV-2. This unique clade replacement was complete just before DENV-3 emerged. By temporally and geographically defining DENV-2 lineages, we describe a refuge of this virus through 4 years of low genome diversity. Our analyses may explain the long-term endurance of DENV-2 despite great epidemiologic changes in disease incidence and serotype distribution.
[show abstract][hide abstract] ABSTRACT: The purpose of this investigation was to identify the mosquito (Diptera: Culicidae) vectors of West Nile virus (WNV; family Flaviviridae, genus Flavivirus) during an epizootic WNV outbreak in eastern Puerto Rico in 2007. In June 2006, 12 sentinel chicken pens with five chickens per pen were deployed in six types of habitats: herbaceous wetlands, mangrove forests, deciduous forests, evergreen forests, rural areas, and urban areas. Once WNV seroconversion in chickens was detected in June 2007, we began trapping mosquitoes using Centers for Disease Control and Prevention (CDC) miniature (light/CO2-baited) traps, CMT-20 collapsible mosquito (CO2- and ISCA SkinLure-baited) traps, and CDC gravid (hay infusion-baited) traps. We placed the CDC miniature traps both 2-4 m and >30 m from the chicken pens, the collapsible traps 2-4 m from the pens, and the gravid traps in backyards of houses with sentinel chicken pens and in a wetland adjacent to an urban area. We found numerous blood-engorged mosquitoes in the traps nearest to the sentinel chickens and reasoned that any such mosquitoes with a disseminated WNV infection likely served as vectors for the transmission of WNV to the sentinels. We used reverse transcriptase-polymerase chain reaction and isolation (C636) on pools of heads, thoraxes/ abdomens, and legs of collected blood-engorged mosquitoes to determine whether the mosquitoes carried WNV. We detected WNV-disseminated infections in and obtained WNV isolates from Culex nigripalpus Theo (minimum infection rate [MIR] 1.1-9.7/1,000), Culex bahamensis Dyar and Knab (MIR 1.8-6.0/1,000), and Aedes taeniorhynchus (Wied.) (MIR 0.34-0.36/1,000). WNV was also identified in and isolated from the pool of thoraxes and abdomens of Culex quinquefasciatus Say (4.17/1,000) and identified in one pool of thoraxes and abdomens of Culex habilitator Dyar and Knab (13.39/1,000). Accumulated evidence since 2002 suggests that WNV has not become endemic in Puerto Rico.
Journal of Medical Entomology 11/2010; 47(6):1185-95. · 1.86 Impact Factor
[show abstract][hide abstract] ABSTRACT: The flavivirus genus includes viruses with a remarkable ability to produce disease on a large scale. The expansion and increased endemicity of dengue and West Nile viruses in the Americas exemplifies their medical and epidemiological importance. The rapid detection of viral infection and induction of the innate antiviral response are crucial to determining the outcome of infection. The intracellular pathogen receptors RIG-I and MDA5 play a central role in detecting flavivirus infections and initiating a robust antiviral response. Yet, these viruses are still capable of producing acute illness in humans. It is now clear that flaviviruses utilize a variety of mechanisms to modulate the interferon response. The non-structural proteins of the various flaviviruses reduce expression of interferon dependent genes by blocking phosphorylation, enhancing degradation or down-regulating expression of major components of the JAK/STAT pathway. Recent studies indicate that interferon modulation is an important factor in the development of severe flaviviral illness. This suggests that an increased understanding of viral-host interactions will facilitate the development of novel therapeutics to treat these viral infections and improved biological models to study flavivirus pathogenesis.
[show abstract][hide abstract] ABSTRACT: Periodic outbreaks of dengue fever occur in the United States Virgin Islands. In June 2005, an outbreak of dengue virus (DENV) serotype-2 with cases of dengue hemorrhagic fever (DHF) was detected in St. Croix, US Virgin Islands. The objective of this report is to describe this outbreak of DENV-2 and the findings of a case-control study examining risk factors for DHF.
This is the largest dengue outbreak ever recorded in St. Croix, with 331 suspected dengue cases reported island-wide during 2005 (62.2 cases/10,000 population); 54% were hospitalized, 21% had at least one hemorrhagic manifestation, 28% had thrombocytopenia, 5% had DHF and 1 patient died. Eighty-nine laboratory-positive hospitalized patients were identified. Of these, there were 15 (17%) who met the WHO criteria for DHF (cases) and 74 (83%) who did not (controls). The only variable significantly associated with DHF on bivariate or multivariable analysis was age, with an adjusted odds ratio (95% confidence interval) of 1.033 (1.003,1.064).
During this outbreak of DENV-2, a high proportion of cases developed DHF and increasing age was significantly associated with DHF.
PLoS ONE 01/2010; 5(10):e13729. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: Dengue virus is sensed in mammalian cells by Toll-like receptors and DExD/H box RNA helicases, triggering a Type 1 interferon response. Interferon acts upon infected and noninfected cells by stimulating the JAK/STAT signaling pathway resulting in the activation of interferon stimulated genes that lead cells toward the establishment of an antiviral response. The recognition of the importance of this rapid protective response should come with the realization that dengue virus would circumvent the interferon response to propagate in the host. There is recent, mounting evidence for mechanisms encoded by the dengue virus that weaken interferon signaling. Nonstructural proteins expressed separately or in replicon vectors block phosphorylation and down-regulate expression of major components of the JAK/STAT pathway, causing reduced activation of gene expression in response to IFNalpha/beta interferon. As our understanding of viral-host interaction increases, opportunities for improved biological models and therapeutics discovery arise.
Current topics in microbiology and immunology 01/2010; 338:35-44. · 4.86 Impact Factor