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PubMed1997

Impact of dengue virus infection on feeding behavior of Aedes aegypti.

In addition to heavily infecting the salivary glands of Aedes aegypti (L.) mosquitoes, dengue viruses produce a significant infection of the nervous system, involving the brain, Johnston's organ, compound eye, and thoracic and abdominal ganglion. To determine if dengue infection affects feeding behavior of Ae. aegypti we measured feeding times, counted the number of feeding delays or interruptions, and by in situ immunocytochemistry techniques determined the spatial and temporal distribution of dengue infections in females parenterally infected with dengue 3 virus. The mean of the total time required for feeding by infected mosquitoes was significantly longer than the time required by uninfected mosquitoes. Similarly, the mean of the time spent probing was significantly longer in infected mosquitoes than in uninfected mosquitoes when day after inoculation was considered. Significant increases in the length of feeding activity in infected mosquitoes corresponded to virus infection in organs that are known to control or influence activities associated with blood feeding. Sequential infections of the salivary glands (five days postinoculation [PI]), brain and compound eye (eight days PI), and Johnston's organ and midgut and abdominal ganglion (11 days PI) of most mosquitoes were observed. The increased time required by infected Ae. aegypti mosquitoes to acquire a blood meal may contribute to the efficiency of Ae. aegypti as a vector of dengue virus. Longer feeding periods are more likely to be interrupted by the host, which increases the chance that an infected mosquito will probe or feed on additional hosts.

Aedes↗
PubMed1999

Increase of leptospirosis in dengue-negative patients after a hurricane in Puerto Rico in 1996 [correction of 1966].

Leptospirosis has rarely been reported in Puerto Rico, although in the period from 1948 to 1952, 208 cases of leptospirosis and an island-wide seroprevalence of antibody to Leptospira of 14% were documented. In Puerto Rico in October 1996, following rainfall and a period of flooding generated by Hurricane Hortense, serum specimens of 4 patients with suspected dengue fever that were negative for dengue tested positive for Leptospira-specific IgM antibodies in a dipstick assay. Subsequently, we used an island-wide dengue laboratory-based surveillance system to determine the increase in leptospirosis after hurricane-generated floods. All anti-dengue IgM-negative patients (n = 142) with disease onset from August 8 to October 6, 1996 from prehurricane and posthurricane groups were investigated for leptospirosis. Laboratory-confirmed leptospirosis cases were defined as microscopic agglutination test titers > or = 1 :400 to 1 or more serovars, or positive immunohistochemistry in autopsy tissues. Four (6%) of 72 prehurricane and 17 (24%) of 70 posthurricane patients had laboratory-confirmed cases of leptospirosis (relative risk [RR] = 4.4, 95% confidence interval [CI] = 1.6-12.4). The mean age of case-patients was 34 years (range = 13-64). Eighteen (86%) of 21 confirmed case-patients were males, including one patient who died (31 years old). Patients were located in 18 (38%) of 48 municipalities that submitted serum samples. Clinical features significantly associated with leptospirosis were eye pain (RR = 1.5, 95% CI = 1.3-1.9), joint pain (RR = 1.4, 95% CI = 1.1-1.6), diarrhea (RR = 1.7, 95% CI = 1.2-2.5), and jaundice (RR = 3.3, 95% CI = 1.5-7.2). This study demonstrates the utility of a dengue laboratory-based surveillance system for the detection of an increase of leptospirosis, which most likely would have gone unrecognized. Leptospirosis is treatable with antibacterial agents; knowledge of this diagnosis may significantly reduce morbidity and mortality.

Adolescent↗
PubMed1999

Detection of dengue virus RNA by reverse transcription-polymerase chain reaction in the liver and lymphoid organs but not in the brain in fatal human infection.

Autopsy tissues from 18 children believed to have died of dengue hemorrhagic fever were tested for the presence of dengue virus RNA by reverse transcription-polymerase chain reaction (RT-PCR). Such RNA was found in 14 of 18 liver specimens, 13 of 18 spleen specimens and 7 of 16 mesenteric lymph node specimens. No dengue virus RNA was detected in 44 samples of brain tissue from 15 individuals, 1 or more of whose other tissues yielded such RNA. All tissues had been tested previously for dengue virus by mosquito inoculation. In those tests, virus was recovered from 5 of 18 liver and 2 of 18 spleen specimens. Thus, the RT-PCR is more sensitive than the most sensitive virus isolation technique for detecting dengue virus or its components in human tissue. Failure to isolate virus from most of spleen and all mesenteric lymph node specimens may indicate that those tissues contained primarily degraded virus undergoing inactivation.

Brain↗
PubMedDate not supplied

Dengue virus infects human endothelial cells and induces IL-6 and IL-8 production.

In this study dengue virus (DV) was found to infect primary endothelial cells derived from human umbilical cord veins (HUVEC) and alter their cytokine production. Dengue virus infection of HUVEC was confirmed by an increase in plaque-forming units in the culture supernatant and by immunofluorescence assay. HUVEC produced large amounts of interleukin (IL)-6 and IL-8 but not IL-1beta after DV infection. Both the replication of DV and the production of IL-6 and IL-8 by HUVEC after DV infection were inhibited by ribavirin, an antiviral synthetic guanosine analogue. Additionally, increased serum levels of IL-6 and IL-8 were observed in patients with dengue hemorrhagic fever but not dengue fever. Therefore, our results suggest that endothelial cells can be a target for DV infection, and that DV-induced IL-6 and IL-8 production by endothelial cells may contribute to the pathogenesis of dengue hemorrhagic fever.

Adolescent↗
PubMed2001

Prospective case-control study of encephalopathy in children with dengue hemorrhagic fever.

We present a prospective case-control study of 27 serologically confirmed dengue hemorrhagic fever (DHF) patients with severe central nervous system symptoms. Dengue associated encephalopathy accounted for 0.5% of 5,400 patients admitted with DHF. The mortality rate among children with encephalopathy was 22%, with the survivors experiencing a complete recovery. Liver enzymes and bilirubin were significantly elevated in the study group. In analysis of the cerebrospinal fluid (CSF), reverse transcriptase-polymerase chain reaction revealed dengue-3-specific RNA in one evaluated case. Dengue-specific immunoglobulin M was detected in CSF in 14 of 22 assessable patients, indicating a localized infection. Magnetic resonance imaging scans showed cerebral edema in the majority of patients, although encephalitis-like changes were less common. There was an equal distribution of primary and secondary infections. On the basis of previous reports and of the findings of our study, DHF probably encompasses an expanding clinical spectrum that infrequently involves encephalitis due to a direct neurotropic effect of dengue virus.

Adolescent↗
PubMedDate not supplied

Serotyping of dengue viruses by an enzyme-linked immunosorbent assay.

We have developed a sandwich enzyme-linked immunosorbent assay for serotyping dengue viruses. In this assay, we used antibody from dengue hemorrhagic fever patients for detection of flavivirus common antigens to confirm virus isolation in C6/36 cells and that from hyperimmune mouse ascitic fluids for serotyping. The anti-dengue antibody was immobilized on microplate wells for capturing of dengue antigens, which were then sandwiched with the same biotinylated antibody. Then the biotin in the solid phase was detected with peroxidase-conjugated streptavidin. We found that all the dengue strains tested were unequivocally identified by this method.

Antibodies, Viral↗
PubMed1998

[Serotypes and genotypes of dengue virus circulating in Venezuela, 1990-1997].

Due to the increasing severity of hemorrhagic dengue epidemics during the last years in Venezuela, a retrospective analysis was conducted to identify the behaviour of the dengue virus serotypes circulating in the country and the molecular evolution of dengue virus serotype 2. The data presented here indicates that dengue virus serotypes 1, 2 and 4 are endemic in Venezuela, they circulate simultaneously around the year in the biggest urban cities, however, one particular serotype is predominant during an epidemic period and replaces the virus serotype dominant during the previous epidemic period. The increased severity of dengue fever since 1989 in Venezuela might be associated to the introduction of the Asiatic genotype of virus which replaced the autochthonous Caribbean genotype. The Asiatic genotype is recognised as a more virulent virus.

Dengue↗
PubMed1999

Dengue fever. An Australian perspective.

BACKGROUND: An estimated 2.5 billion people live in areas where dengue fever is endemic with around 100 million cases per year. A more serious form of the disease called dengue haemorrhagic fever (DHF) is the most common cause of hospitalisation and death among children in some Southeast Asian countries. OBJECTIVE: This article reviews the ecology and the pathogenesis of the disease and its clinical features. The history of dengue fever in Australia will be presented and the prospects for the future discussed. DISCUSSION: Dengue fever was common in Australia in the late 19th century and the first clinical description of DHF were published by an Australian clinician in Charters Towers north Queensland. Since that time there have been sporadic outbreaks of dengue fever. In recent years there has been an increase in the number of epidemics in north Queensland. If Australia follows a pattern observed elsewhere in the world there might again be cases of DHF recorded on Australian soil.

Adolescent↗
PubMed1999

Infection of dengue 2 virus strains isolated from patients exhibiting different disease severities to human peripheral blood leukocytes and production of cytokines in the infected culture supernatant.

Three strains of type 2 dengue virus (DV-2), which had been isolated from patients exhibiting different disease severity, were inoculated to primary culture of human peripheral blood leukocytes from 3 healthy donors. The percentage of dengue antigen positive cells was highest for the strain isolated from a case of dengue shock syndrome, followed by the strain isolated from a case of dengue hemorrhagic fever, and lowest for the strain isolated from a mild case of dengue fever (DF). Generally, similar trend was observed for the amount of some cytokines released into infected culture supernatant, such as interleukin 6, and tumor necrosis factor-alpha. However, such a trend was not observed for interleukin 1 beta.

Animals↗
PubMed2001

Underdiagnosis of dengue--Laredo, Texas, 1999.

Dengue outbreaks have been reported in communities along the Mexico-U.S. border since 1980; however, during 1987-July 1999, no cases were reported from Laredo, Texas (1999 population: 162,000). During January-July 1999, approximately 300-325 dengue cases were reported from Nuevo Laredo, Tamaulipas, Mexico (1999 population: 274,000), a city across the Rio Grande from Laredo. To determine whether undiagnosed or unreported dengue cases had occurred in Laredo, the Texas Department of Health (TDH) reviewed medical records from five Laredo health facilities (the two city hospitals and the three largest of five community clinics). This report summarizes the findings of the review, which indicated that during July 23-August 20, 1999, 50% of suspected case-patients had undiagnosed dengue infection. Recognition of the diagnosis of dengue can be improved through heightened surveillance, professional and public education, and prompt reporting of cases by the health-care providers to local or state health departments.

Antibodies, Viral↗
PubMed2000

IgM-capture ELISA of serum samples collected from Filipino dengue patients.

Viral antigens for 4 dengue serotypes were produced in C6/36 Aedes albopictus cells. These were used as assay antigens for IgM-capture ELISA to detect IgM antibodies in sera of dengue patients from 3 hospitals in Metro Manila, Philippines. A total of 378 serum samples came from National Children's Hospital (NCH), San Lazaro Hospital (SLH), and St Luke's Medical Center (SLMC), from January to November 1995. Three hundred and four (304) out of 378 serum samples, or 80.42% showed positive IgM ELISA titer against at least one of the 4 assay antigens. Dengue type 4 (D4) antigen detected antibodies in 61.90% (234/378) of these serum samples, whereas type 1 (D1), type 3 (D3), and type 2 (D2) had detection rates of 60.05% (227/378), 50.79% (192/378) and 49.47% (187/378) respectively. Although the results show that both D1 and D4 are the most effective antigens in identifying dengue infections for this batch of samples, the use of a cocktail of antigens is still recommended. The results of this study are the basis for the IgM-capture ELISA protocol presently applied for the laboratory confirmation of dengue cases in the Philippines.

Aedes↗
PubMed2001

[Secondary serologic responses to dengue epidemic in 1998 in Salta, Argentina, where other flavivirus co-circulate].

In 1998, a dengue outbreak (serotype 2) occurred in Salta province in Northern Argentina, following the first detection of dengue in the same area in 1997. We classified the serologic response of cases from 1998 as primary or secondary, since the risk of severe disease is greater for secondary cases. We studied 154 cases by plaque reduction neutralization and hemagglutination inhibition tests. Thirty-eight cases (25%) were classified as primary serologic responses and 84 cases (54%) as secondary responses. Thirty-two cases (21%) with borderline IgG titers could not be classified. Previous exposure to potentially cross-reacting flaviviruses (Saint Louis Encephalitis [SLE] and Yellow Fever [YF] viruses) was analyzed, as a possible cause of the secondary response pattern. Our results indicated that among cases classified as dengue secondary response, 83% could be attributed to previous SLE or YF exposure or serologic cross-reactivity. Vaccination against YF virus was at most a minor contributor to the secondary response pattern. The finding of a positive YF serologic result among persons not vaccinated may indicate silent circulation of YF in a region that can support both urban and jungle cycles. Other cases showing dengue secondary responses remained unexplained, suggesting the unrecognized occurrence of a previous infection with other dengue serotypes or of flaviviruses other than SLE or YF.

Antibodies, Viral↗
PubMed2003

A glance at the von Willebrand factor in dengue virus infection.

To identify the level of von Willebrand factor (vWF) in dengue infection, especially severe DHF, and correlate the increase in vWF with thrombocytopenia, children admitted with dengue fever/DHF were examined for hemoglobin, hematocrit, platelet, and vWF for three consecutive days. Anti-dengue IgM and IgG were determined. Correlations between vWF and thrombocytopenia were analyzed using multivariate analysis. Forty-one patients were eligible for the study; of whom almost three fourths had a secondary infection, as proved serologically. At the beginning of the study, a high level of vWF along with a low platelet count were seen, which seemed to fit the hypothesis that an increase in vWF in the serum will be followed by a decrease in platelets, as a result of the platelet aggregation process in the peripheral blood vessels, predisposed by the immune complex events in DHF. Observations for three consecutive days revealed significant changes of vWF levels (p = 0.000) as well as platelet counts (p = 0.002). However in the context of dengue infections, these changes did not correlate well (p = 0.988). Could there have been a significant correlation if cases were followed for a longer period of time? Being a part of a more comprehensive study, it appeared that in patients with dengue infections, vWF and platelets were not the only factors involved in bleeding, indicating that activation of endothelium is one factor in a multifactorial process.

Adolescent↗
PubMed2004

[Demonstration of simultaneous infection with dengue type 2 and 3 in a Chinese patient].

IgG and IgM antibodies to dengue virus(DEN) in patient serum sample were detected with an indirect immunofluorescence assay. The patient's serum was inoculated into the culture of C6/36 cells to isolate dengue virus. Viral RNA was extracted from cultured supernatant and RT-PCR amplification and sequencing were carried out. The result showed that there were anti-dengue virus IgG and IgM antibodies in the serum. Both dengue type 2 and 3 virus specific sequences were confirmed by RT-PCR and sequence analysis. The results presented here demonstrated the simultaneous infection with dengue type 2 and 3 viruses in the Chinese patient.

Adult↗
PubMed2005

Short report: assessment of the World Health Organization scheme for classification of dengue severity in Nicaragua.

The World Health Organization (WHO) scheme for classification of dengue severity was evaluated in a three-year study of 1,671 confirmed dengue cases in three Nicaraguan hospitals. The WHO classification of dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) was compared with the presence of hemorrhagic manifestations, signs of vascular permeability, marked thrombocytopenia, and shock in 114 infants, 1,211 children, and 346 adults. We found that strict application of the WHO criteria fails to detect a significant number of patients with severe manifestations of dengue, especially in adults.

Adolescent↗
PubMed2006

Serum levels of sICAM-1 and sE-selectin in patients with dengue virus infection.

The purpose of this study was to measure the serum level of sICAM-1 and sE-selectin as markers for endothelial damage in patients with dengue fever (DF) and dengue hemorrhagic fever (DHF). Twenty-nine patients with serologically-proven dengue virus infection (age 10.6 +/- 2.4 years, 55% male, DF = 13 and DHF = 16) were enrolled. Serum samples were collected from 25 healthy children (age 10.6 +/- 2.3 years, 40% male) as the control group. A follow-up was done at a mean interval of 15.9 +/- 1.6 days. The level of sICAM-1 at the toxic stage was significantly elevated compared to its level at the follow-up (494.1 +/- 107.4 versus 358.2 +/- 67.6 ng/ml, P = 0.001), but no difference was found between patients with DF and patients with DHF (444.1 +/- 158.0 versus 465.1 +/- 154.6 ng/ml, P = 0.74). The sICAM-1 level at the follow-up was similar to that of the control group (396.9 +/- 113.0 ng/ml, P = 0.56). The level of sE-selectin at the toxic stage was not different from its level at the follow-up (75.9 +/- 33.0 versus 75.5 +/- 31.7 ng/ml, P = 0.96), and no difference was found between the DF group and the DHF group (64.1 +/- 25.7 versus 78.8 +/- 39.9 ng/ml, P = 0.30). These levels were not elevated compared to the sE-selectin level that was determined in 8 patients in the control group (94.7 +/- 20.5 ng/ml, P = 0.12). In conclusion, there is evidence of endothelial activation by an increased sICAM-1 level in patients with dengue virus infection. However, the degree of endothelial activation alone may be similar for patients with DF and patients with DHF, and this fact by itself cannot explain the difference between the two clinical syndromes of dengue virus infection. The sE-selectin level was not elevated for patients included in this study.

Child↗
PubMed2005

Dengue vaccine.

Dengue is an expanding health problem. About two-fifths of the world population are at risk for acquiring dengue with 50-100 million cases of acute febrile illness yearly including about 500,000 cases of DHF/DSS. No antiviral drugs active against the flavivirus exist. Attempts to control mosquito vector has been largely unsuccessful. Vaccination remains the most hopeful preventive measure. Dengue vaccine has been in development for more than 30 years, yet none has been licensed. The fact that enhancing antibody from previous infection and high level of T cell activation during secondary infection contribute to immunopathology of DHF, the vaccine must be able to induce protective response to four dengue serotypes simultaneously. Inactivated vaccine is safe but needs a repeated booster thus, development is delayed. Tetravalent live attenuated vaccine and chimeric vaccine using yellow fever or dengue viruses as a backbone are being carried out in human trials. DNA vaccine and subunit vaccine are being carried out in animal trials.

Dengue↗
PubMed2006

A simplified PCR methodology for semiquantitatively analyzing dengue viruses.

The standard methodology for titrating dengue viruses, the plaque assay, is slow, time consuming and relatively expensive. Other methods require machinery that may not be routinely accessible to all researchers, particularly those in developing nations. We therefore sought to develop a rapid, simplified semiquantitative polymerase chain reaction (PCR) methodology based on the use of a template mimic. In particular, it was desired that the mimic should be applicable for use a DNA template to avoid the requirement for producing an in vitro RNA transcript. A 511 base pair fragment of the capsid-PrM junction of dengue serotype 4 was cloned into pGEM-T Easy vector and subjected to splicing overlap extension-PCR to generate a 160 base pair deletion. The deleted plasmid mimic competed competitively against the parent plasmid as well as the first strand cDNA of all four dengue viruses. The primers used are specific for the dengue virus, and no product was seen with first strand cDNA from a closely related flavivirus, Japanese encephalitis virus. Under the conditions used, accurate quantitation of the dengue viruses in the range of 10(3) to 10(6) pfu can be achieved in a single day, as opposed to the 7 days required for conventional plaque assay.

Capsid↗