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Objectives: Plasma leakage, a hallmark of disease in Dengue virus (DENV) infection, is an important clinical manifestation and is often associated with numerous factors such as viral factors. The aim of this study is to investigate the association of virus serotype, viral load kinetics, history of infection, and NS1 protein with plasma leakage., Methods: Subjects with fever ≤ 48 hours and positive DENV infection were included. Serial laboratory tests, viral load measurements, and ultrasonography examination to assess plasma leakage were performed., Results: DENV-3 was the most common serotype found in the plasma leakage group (35%). Patients with plasma leakage demonstrated a trend of higher viral load and a longer duration of viremia compared to those without. This was significantly observed on the fourth day of fever (p = 0.037). We found higher viral loads on specific days in patients with plasma leakage in both primary and secondary infections compared to those without. In addition, we also observed more rapid viral clearance in patients with secondary infection. NS1 protein, especially after 4 days of fever, was associated with higher peak viral load level, even though it was not statistically significant (p = 0.470). However, pairwise comparison demonstrated that peak viral load level in the group of patients with circulating NS1 detected for 7 days was significantly higher than the 5-day group (p = 0.037)., Conclusion: DENV-3 was the most common serotype to cause plasma leakage. Patients with plasma leakage showed a trend of higher viral load and a longer duration of viremia. Higher level of viral load was observed significantly on day 5 in patients with primary infection and more rapid viral clearance was observed in patients with secondary infection. Longer duration of circulating NS1 protein was also seen to be positively correlated with higher peak viral load level although not statistically significant., Competing Interests: The authors affirm no conflict of interest in this study., (Copyright: © 2023 Nainggolan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Dengue virus (DENV) infection remains to be a serious health problem in Indonesia. Community-based dengue studies to determine circulating DENV serotypes based on the geography and season are limited owing to the expensive cost and significant effort required. Many patients with DENV infection are not hospitalized and many visit the hospital in the later phase of the disease. In this study, we performed active DENV surveillance in a community in Jakarta to study the circulating dengue serotypes; adult febrile patients with fever less than 48 hours were recruited. Disease severity was defined using the World Health Organization (WHO) 1997 guidelines. Rapid NS1 dengue antigen detection was used to screen patients with DENV in the community. Viral culture using the C6/36 cell line, an increased antibody titer on hemagglutination inhibition test and enzyme linked immunosorbent assay, or detection of the viral genome on reverse transcription-polymerase chain reaction was used to confirm DENV infection. Of the 102 patients, 68 (66.7%) were confirmed to have DENV infection, with DENV-2 being the most dominant serotype, followed by DENV-3, DENV-1, and DENV-4, in concordance with several reports of mixed DENV infection. Interestingly, in terms of disease severity, although DENV-3 infection was not the predominant circulating serotype, infection with it tended to cause a more severe disease than infection with DENV-2.

The development of a dengue virus vaccine is a major priority in efforts to control the diseases. Several researchers are currently using the Asian 1 and Asian 2 genotypes as vaccine candidates for dengue type 2 virus (DENV-2). However, in this study, we constructed a recombinant plasmid-based prM/E gene, from a DENV-2 Cosmopolitan genotype strain as a dengue DNA vaccine candidate. The protein expression of the recombinant plasmid in CHO cells was analyzed using an enzyme-linked immunosorbent assay, western blotting, and sucrose gradient sedimentation. After being used to immunize ddY mice three times at doses of 25 or 100 μg, the DNA vaccine induced humoral immune responses. There was no difference in the neutralizing antibody titer (focus reduction neutralization test 50% value) of mice immunized with 25 and 100 μg DNA vaccine doses. When challenged with 3 × 10(5) FFU DENV-2, immunized mice could raise anamnestic neutralizing antibody responses, which were observed at day 4 and day 8 post-challenge. Analysis of immunogenicity using BALB/c mice showed that their antibody neutralization titers were lower than those of ddY mice. In addition, the antibodies produced after immunization and challenge could also neutralize a DENV-2 Asian 2 genotype (New Guinea C) strain. Therefore, the DENV-2 Cosmopolitan genotype may be a DENV-2 vaccine candidate.

Dengue hemorrhagic fever has become a worldwide health issue. Heterologous infection by different serotypes may lead to severe forms of dengue infection and even death. In a cohort study in Jakarta from 2009 to 2010 with inclusion criteria of adults with fever of less than 48 hours, 72% were confirmed dengue infection from a total of suspected 190 dengue patients. Using RT-PCR, 16 patients were infected with DENV-4 with or without co-infection with other serotype(s). Dengue fever (DF) patients (73%) were infected with DENV-4 alone, while mixed infections were present in more severe clinical manifestations of DHF grade I and II. The nucleotide sequences of envelope (E) protein gene of 3 isolates of DENV-4 showed that they belonged to genotype II. There were 50 nucleotide substitutions, but only 3 amino acid changes were found at positions 130, 233 and 455, present in E protein isolated from DHF grade II patient.

Dengue is a major public health threat worldwide, affecting approximately 3 billion people. More than 100 countries in the world located in tropical and subtropical areas, there are at least 100 to 400 million people infected with the dengue virus which causes dengue hemorrhagic fever (DHF). Soluble Non Structural Protein (sNS1) DENV is a soluble NS1 protein that is secreted and found in the serum of patients during acute infection. Because of its presence early in infection, sNS1 is used as a diagnostic indicator of acute dengue infection. NS1 can directly activate platelets through TLR4 and can further increase platelet aggregation, endothelial cell adhesion, and phagocytosis by macrophages that can cause thrombocytopenia so that high sNS1 levels are associated with disease severity. From the results of the study showed p <0.05. This indicates that there is a correlation between the presence of NS1 and the severity of dengue infection. [ABSTRACT FROM AUTHOR]

Dengue fever is widely prevalent in Indonesia and many other tropical countries. There has been no antiviral protection against DENV (dengue virus) due to the scarcity of animal models that replicate the symptoms of its infection in humans. Subsequently, Balb/ c mice were used to optimize the animal models that were infected with DENV, and measurements of DENV titer, platelet count and leukocytes counts were taken after six days of CA (Cassia alata leaves extract) treatment to examine the inhibitory activity of viral replication on dengue-infected mice. The results showed that CA significantly reduced DENV titer and increased platelet count compared to the control group (P < 0.05) but had no significant effect on leukocyte counts (P > 0.05). Therefore, it could be implied that Cassia alata could be further investigated for medication against DENV, prospectively. [ABSTRACT FROM AUTHOR]

BACKGROUND Early diagnosis of dengue virus (DENV) infection is essential for patient management and disease control. Detection of the antigen non-structural protein 1 (NS1) has been proven to provide early diagnosis of DENV infection. Thus, commercial NS1 antigen detection assays have been increasingly used and are becoming the tool of choice among clinicians to confirm DENV infection in Indonesia. METHODS To obtain anti-NS1 DENV antibody, NS1 protein (90 µg/ml) from the collection of the Department of Microbiology, Faculty of Medicine, Universitas Indonesia was injected into a rabbit. The anti-NS1 antibody from the rabbit was then labeled with horseradish peroxidase (HRP) using the periodate oxidation method. Sera were tested by enzyme-linked immunosorbent assay (ELISA) to detect NS1 from DENV-infected patients. RESULTS Serially diluted antibody labeled with HRP tested using the direct ELISA method showed the highest absorbance value at a 1:100 dilution (Mean [SD] = 1.35 [0.35]); even at a dilution as high as 1:3,200 (0.22 [0.15]), antibody labeled with HRP was able to detect the NS1 protein, although the absorbance value did not differ greatly from that of the negative control (0.13 [0.01]). CONCLUSIONS In an attempt to develop an NS1-based diagnostic test, polyclonal anti- NS1 DENV antibody was successfully produced as a diagnostic assay to determine the presence of DENV NS1 antigen in patients' sera. [ABSTRACT FROM AUTHOR]

Introduction: Dengue Virus (DENV) infection is a mosquito-borne disease, the most severe manifestation of the infection could lead to a lethal Dengue Haemorrhagic Fever (DHF) and/or Dengue Shock Syndrome (DSS). The currently licensed DENV vaccine has been launched in the market with some limitation applications in the field. Therefore, accurate diagnosis system is needed to identify effective treatment of DENV-infected patients and prevent the fatal outcomes caused by the severe complications. Non-Structural-1 (NS1) protein of DENV has been known as a biomarker for dengue diagnosis since the protein is abundantly circulating in the blood during the acute phase of the disease. Aim: To produce and characterise mouse monoclonal Antibodies (mAbs) anti-DENV NS1 antigen. Materials and Methods: This was an experimental study. Hybridoma mAb-producing cells were obtained by the fusion between PAI mouse myeloma cells and B cells from an immunized mice with lysed Chinese Hamster Ovary (CHO-K1) cells expressing NS1 protein. Monoclonal antibodies were tested against native NS1 antigen from DENV1, DENV2, DENV3, and DENV4 respectively, by ELISA, IFA and Western Blot analysis. Results: Hybridoma selection with indirect ELISA showed 16 clones that potentially produce anti-NS1 antibodies. Six out of 16 clones showed positive results by IFA analysis against NS1 transfected CHO-K1 cells. Three mAb clones 4-2D, 4-4F, and 2-7A were able to recognise the specific NS1 antigen for all DENV serotypes and did not react against other DENV proteins. Moreover, clone 4-4F showed dissociation constant (Kd) approximately 1.25+0.47 nM indicating the antibody has a strong binding affinity to NS1 antigen. Conclusion: The monoclonal antibodies anti-NS1 induced by DENV3 Indonesian clinical isolate are potentially used as the main reagent for development of the DENV NS1 diagnostic test. [ABSTRACT FROM AUTHOR]

BACKGROUND Dengue virus (DENV) can infect and replicate in monocytes, resulting in antibody-dependent enhancement. The liver is the main target of DENV, and the infection mechanisms of DENV include direct cytopathic effects (CPEs) of the virus, mitochondrial dysfunction, and effect of cellular and humoral immune factors in the liver. This study was aimed to explore the infectivity of DENV and viability of human hepatocytes using Huh 7it-1 cells cocultured with peripheral blood mononuclear cells (PBMCs). METHODS Huh 7it-1 cells were infected with dengue virus serotype-2 (DENV-2) New Guinea C strain at multiplicity of infection of 0.5 and 1 FFU/cell, and cocultured in vitro with and without adherent PBMCs. The infectivity of DENV was assessed by immunoperoxidase staining. The viability of Huh 7it-1 cells was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT, a tetrazole) assay and trypan blue staining. Data were statistically analyzed by Shapiro-Wilk and analysis of variance for normality significances. RESULTS The result showed that addition of PBMCs to DENV-2 infected Huh 7it-1 cells decreased the infectivity of DENV (15-37%). DENV-2 infection decreased the viability of Huh 7it-1 cells (15.5-20.8%). Despite the decrease in infectivity of DENV, the addition of PBMCs increased the Huh 7it-1 cells viability (4.50-10.2%). CONCLUSIONS Addition of PBMCs to Huh 7it-1 cells that are infected with DENV-2 decreased the infectivity of DENV and increased Huh 7it-1 cells viability. [ABSTRACT FROM AUTHOR]