Elizabeth Hunsperger

Dengue: a continuing global threat

Dengue fever and dengue haemorrhagic fever are important arthropod-borne viral diseases. Each year, there are ∼50 million dengue infections and ∼500,000 individuals are hospitalized with dengue haemorrhagic fever, mainly in Southeast Asia, the Pacific and the Americas. Illness is produced by any of the four dengue virus serotypes. A global strategy aimed at increasing the capacity for surveillance and outbreak response, changing behaviours and reducing the disease burden using integrated vector management in conjunction with early and accurate diagnosis has been advocated. Antiviral drugs and vaccines that are currently under development could also make an important contribution to dengue control in the future.

Evaluation of diagnostic tests: dengue

Dengue is an arthropod-borne flavivirus that comprises four distinct serotypes (DEN-1, DEN-2, DEN-3 and DEN-4) that constitute an antigenic complex of the genus flavivirus, family Flaviviridae. Infection by one serotype induces life-long immunity against reinfection by the same serotype, but only transient and partial protection against infection with the other serotypes1,2. Dengue virus infections can result in a range of clinical manifestations from asymp tomatic infection to dengue fever (DF) and the severe disease dengue haemorrhagic fever/dengue shock syndrome (DHF/ DSS). Most dengue infections are asymptomatic or cause mild symptoms, which are characterized by undifferentiated fever with or without rash. Typical DF is characterized by high fever, severe headache, myalgia, arthralgia, retro-orbital pain and maculopapular rash. Some patients show petechiae, bruising or thrombocytopenia. The clinical presentation of acute dengue infection is non-specific but 5–10% of patients progress to severe DHF/DSS, which can result in death if it is not managed appropriately. Plasma extravasation is the main pathophysiological finding of DHF/ DSS, which differentiates it from DF. DHF/ DSS is characterized by high fever, bleeding, thrombocytopenia and haemoconcentration (an increase in the concentration of blood cells as a result of fluid loss). Approximately 3–4 days after the onset of fever, patients can present with petechiae, rash, epistaxis, and gingival and gastrointestinal bleeding. Pleural effusion and ascites are common. Some patients develop circulatory failure (DSS), presenting with a weak and fast pulse, narrowing of pulse pressure or hypotension, cold and moist skin and altered mental state. Although there are no specific antiviral treatments for dengue infection, patients usually recover when the need for fluid management is identified early and electrolytes are administered3. It has been proposed that the classification of dengue disease should be simplified as severe and non-severe dengue. This simplified classification would make patient management and surveillance easier4. There is a need for specific, inexpensive dengue diagnostic tests that can be used for clinical management, surveillance and outbreak investigations and would permit early intervention to treat patients and prevent or control epidemics. Progress is being made in primary prevention, with several candidate dengue vaccines in late phases of development as well as improved vector control measures. Additionally, new techniques for the early detection of severe disease such as the use of biomarkers have the potential to decrease morbidity and

Evaluation of Commercially Available Anti–Dengue Virus Immunoglobulin M Tests

Anti–dengue virus immunoglobulin M kits were evaluated. Test sensitivities were 21%–99% and specificities were 77%–98% compared with reference ELISAs. False-positive results were found for patients with malaria or past dengue infections. Three ELISAs showing strong agreement with reference ELISAs will be included in the World Health Organization Bulk Procurement Scheme.

Dengue viremia in blood donors identified by RNA and detection of dengue transfusion transmission during the 2007 dengue outbreak in Puerto Rico

BACKGROUND: 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.

Comparison of Two Commercially Available Dengue Virus (DENV) NS1 Capture Enzyme-Linked Immunosorbent Assays Using a Single Clinical Sample for Diagnosis of Acute DENV Infection

ABSTRACT Dengue virus (DENV) nonstructural protein 1 (NS1) has shown promise as a novel diagnostic marker of acute DENV infection. Current techniques used to diagnose acute DENV infection, including virus isolation and reverse transcription-PCR (RT-PCR), are costly and difficult to perform, while traditional serological assays have low sensitivities during the acute stage of infection. Two commercially available NS1 antigen capture enzyme-linked immunosorbent assays (ELISAs), the Platelia dengue NS1Ag test (Bio-Rad Laboratories, Marnes La Coquette, France) and the Pan-E dengue early ELISA test (Panbio Diagnostics, Brisbane, Australia), were evaluated against a well-characterized panel of 208 real-time RT-PCR- and virus isolation-positive sera, as well as 45 real-time RT-PCR- and serologically negative sera from patients with other acute febrile illnesses. The overall sensitivities were 64.9% (95% confidence interval [CI95], 58.2 to 71.1%) for the Panbio test and 83.2% (CI95, 77.5 to 87.7%) for the Bio-Rad test, with interserotype variation, especially for DENV serotype 4. Predictive models were constructed to identify factors that had a significant influence on a tests outcome with respect to this panel of samples in order to identify the conditions in which the test will be most effective as a diagnostic tool. The immunoglobulin G titer was found to be the only covariate that significantly influenced results in the Bio-Rad test, while serotype and the day postonset were found to significantly influence results in the Panbio test. We concluded that the NS1 capture ELISA is a useful tool that can improve testing algorithms to diagnose DENV infection in single samples from acute and early convalescent cases.

Dengue Fever Seroprevalence and Risk Factors, Texas-Mexico Border, 2004

High seroprevalence of dengue was found on both sides of the border.

Dengue outbreak in Key West, Florida, USA, 2009.

After 3 dengue cases were acquired in Key West, Florida, we conducted a serosurvey to determine the scope of the outbreak. Thirteen residents showed recent infection (infection rate 5%; 90% CI 2%–8%), demonstrating the reemergence of dengue in Florida. Increased awareness of dengue among health care providers is needed.

Temporal analyses of the neuropathogenesis of a West Nile virus infection in mice

A West Nile virus (WNV) infection in humans can produce neurological symptoms including acute flaccid paralysis, encephalitis, meningitis and myelitis. To investigate the pathogenesis of WNV in the peripheral and the central nervous system (PNS and CNS), the authors used a murine footpad inoculation model of WNV infection. Survival curves of virus-infected animals of ages 4- to 6-weeks-old demonstrated age-dependent mortality where older animals (6-weeks-old) had a higher mortality rate compared to younger animals (4- and 5-weeks-old). The mice that survived the virus infection formed WNV-reactive antibodies, confirming viral infection and clearance. The localization of viral RNA (vRNA) and antigen in infected murine tissues was investigated using TaqMan and immunohistochemistry (IHC) respectively. During a nine day infection, vRNA levels in the spinal cord and brainstem fluctuated, suggesting early viral clearance from these tissues by days 3–4 p.i. with later re-introduction. Viral antigens detected using IHC were primarily observed in three main regions of the brain: cortex, hippocampus and brainstem. Additionally, the dorsal root ganglion neurons of the PNS stained positive for viral antigens. These data are consistent with multiple routes of neuroinvasion following a peripheral inoculation of virus and do not preclude the previous observation that virus-infected peripheral neurons can introduce virus into the CNS by a retrograde transport mechanism.

Structural Optimization and De Novo Design of Dengue Virus Entry Inhibitory Peptides

Viral fusogenic envelope proteins are important targets for the development of inhibitors of viral entry. We report an approach for the computational design of peptide inhibitors of the dengue 2 virus (DENV-2) envelope (E) protein using high-resolution structural data from a pre-entry dimeric form of the protein. By using predictive strategies together with computational optimization of binding “pseudoenergies”, we were able to design multiple peptide sequences that showed low micromolar viral entry inhibitory activity. The two most active peptides, DN57opt and 1OAN1, were designed to displace regions in the domain II hinge, and the first domain I/domain II beta sheet connection, respectively, and show fifty percent inhibitory concentrations of 8 and 7 µM respectively in a focus forming unit assay. The antiviral peptides were shown to interfere with virus:cell binding, interact directly with the E proteins and also cause changes to the viral surface using biolayer interferometry and cryo-electron microscopy, respectively. These peptides may be useful for characterization of intermediate states in the membrane fusion process, investigation of DENV receptor molecules, and as lead compounds for drug discovery.

Virus-Specific Differences in Rates of Disease during the 2010 Dengue Epidemic in Puerto Rico

Background 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. Methodology/Principal Findings 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. Conclusions/Significance 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 1960s. This epidemic re-emphasizes the need for more effective primary prevention interventions to reduce the morbidity and mortality of dengue.