Stage Of Dengue Virus Infection Research Articles

Identification of the phytobioactive Polygonum cuspidatum as an antiviral source for restricting dengue virus entry

Dengue virus (DENV) is a mosquito-borne pathogen that is becoming a serious global threat, owing to its rising incidence in inter-tropical regions that yield over 50 million annual infections. There are currently no approved antiviral agents for the management of dengue, and recent shortcomings in its immunization called for immediate action to develop effective drugs with prophylactic ability to better manage its infection. In an attempt to discover novel antiviral sources, we identified the medicinal herb Polygonum cuspidatum (PC) as a bioactive botanical material against DENV infectivity. Specifically, the methanolic extract from PC rhizomes (PCME) potently inhibited DENV infection without causing significant cytotoxicity. Further examination on the viral life cycle demonstrated that PCME particularly targeted the initial stages of DENV infection, while pre- and post-infection treatments had no effect. More importantly, the PCME could efficiently inactivate DENV free virus particles and block the viral attachment and entry/fusion events without apparently influencing viral replication, egress, and cell-to-cell spread. The antiviral effect of PCME was also recapitulated in infection analysis using DENV pseudoparticles displaying viral structural proteins that mediate DENV particle entry. Besides, PCME treatment also inhibited direct DENV entry into several cell types relevant to its infection and reduced viral infectivity of other members of the Flaviviridae family, including the hepatitis C virus (HCV) and Zika virus (ZIKV). Due to its potency against DENV entry, we suggest that the phytobioactive extract from PC is an excellent starting point as an antiviral source material for further development of therapeutic strategies in the prophylactic management of DENV infection.

Upregulation of SCUBE1 in Dengue Virus Infection.

BACKGROUND:Dengue is a major communicable disease in tropical areas, with an increasing prevalence every year. Thrombocytopenia is one of the commonly used laboratory parameters for predicting the severity of the disease. It is detected on day 6 or day 7 after the febrile stage, and its presence indicates that the disease has become potentially fatal. Therefore, it is necessary to identify a marker for the early recognition of dengue virus infection during the febrile stage before the detection of thrombocytopenia on day 6 to prevent severe disease outcomes. Signal peptide-CUB- (complement C1r/C1s)-EGF (epidermal growth factor)-like domain-containing protein 1 (SCUBE1) is secreted in activated platelets under inflammatory conditions and enhances platelet-platelet adhesion and agglutination. This gene was first identified in human vascular endothelium, but its biological role in platelets remains unknown.AIM:This study aims to identify SCUBE1 expression during the febrile stage of dengue virus infection and examine the correlation of its expression with thrombocytopenia occurrence on day 6.MATERIAL AND METHODS:Blood samples were collected from 17 patients infected with dengue virus on day-3 fever and from 16 healthy controls who met the inclusion and exclusion criteria for dengue virus infection according to the World Health Organization (WHO) classification for dengue virus infection. All samples were subjected to SCUBE1 gene analysis using real-time reverse transcription quantitative PCR (RT-PCR).RESULTS:The results showed that upregulation of SCUBE1 gene in infected patients (8.9 ± 3.1-fold) compared to that in healthy controls, indicating SCUBE1 involvement in dengue virus infection. Furthermore, we analysed the laboratory parameters of infected patients on day 3 and day 6, when thrombocytopenia is usually detected. Platelet count was found to be significantly decreased from day 3 until day 6 in the infected patients. Unfortunately, our results showed no correlation between SCUBE1 expression in the febrile stage and the occurrence of thrombocytopenia on day 6.CONCLUSION:The conclusion of this study is SCUBE1 might play a role in dengue virus infection but does not correlate with thrombocytopenia on day-6 fever.

Raman spectroscopy based investigation of molecular changes associated with an early stage of dengue virus infection

Raman spectroscopy based investigations of the molecular changes associated with an early stage of dengue virus infection (DENV) using a partial least squares (PLS) regression model is presented. This study is based on non-structural protein 1 (NS1) which appears after three days of DENV infection. In total, 39 blood sera samples were collected and divided into two groups. The control group contained samples which were the negative for NS1 and antibodies and the positive group contained those samples in which NS1 is positive and antibodies were negative. Out of 39 samples, 29 Raman spectra were used for the model development while the remaining 10 were kept hidden for blind testing of the model. PLS regression yielded a vector of regression coefficients as a function of Raman shift, which were analyzed. Cytokines in the region 775–875 cm−1, lectins at 1003, 1238, 1340, 1449 and 1672 cm−1, DNA in the region 1040–1140 cm−1 and alpha and beta structures of proteins in the region 933–967 cm−1 have been identified in the regression vector for their role in an early stage of DENV infection. Validity of the model was established by its R-square value of 0.891. Sensitivity, specificity and accuracy were 100% each and the area under the receiver operator characteristic curve was found to be 1.

TREM-1 modulation during early stages of dengue virus infection

Uncontrolled and intricate production of inflammatory factors is the characteristic feature of dengue infection. The triggering receptor expressed in myeloid cells-1 (TREM-1), expressed on the surface of monocytes and neutrophils, is capable of enhancing and regulating the inflammatory response via the production of different mediators in bacterial and viral infections. Here, both the expression of TREM-1 on human monocytes and neutrophils from peripheral blood of dengue infected individuals, as well as the levels of the soluble form of TREM-1 (sTREM-1) in the sera of these patients were compared against healthy controls. A significant reduction of TREM-1 expression was observed in neutrophils during the first days of infection, followed by a gradual recovery throughout the course of infection. Also, sera from DENV-infected patients exhibited significantly higher sTREM-1 levels than healthy individuals. The difference was more pronounced during the first 5 days after the onset of symptoms. These findings highlight the dynamic process of TREM-1 expression during DENV infection. We hypothesized that increment of free sTREM-1 could be a compensatory mechanism aiming to counteract the inflammatory process elicited during DENV infection.

Adaptor protein complexes-1 and 3 are involved at distinct stages of flavivirus life-cycle

Intracellular protein trafficking pathways are hijacked by viruses at various stages of viral life-cycle. Heterotetrameric adaptor protein complexes (APs) mediate vesicular trafficking at distinct intracellular sites and are essential for maintaining the organellar homeostasis. In the present study, we studied the effect of AP-1 and AP-3 deficiency on flavivirus infection in cells functionally lacking these proteins. We show that AP-1 and AP-3 participate in flavivirus life-cycle at distinct stages. AP-3-deficient cells showed delay in initiation of Japanese encephalitis virus and dengue virus RNA replication, which resulted in reduction of infectious virus production. AP-3 was found to colocalize with RNA replication compartments in infected wild-type cells. AP-1 deficiency affected later stages of dengue virus infection where increased intracellular accumulation of infectious virus was observed. Therefore, our results propose a novel role for AP-1 and AP-3 at distinct stages of infection of some of the RNA viruses.

Spectrum of cardiac rhythm abnormalities and heart rate variability during the convalescent stage of dengue virus infection: a Holter study

Various minor cardiac rhythm abnormalities have been reported in patients with dengue virus infection. Previous studies have used only random electrocardiograms (ECG) to assess the incidence of cardiac arrhythmias, and the time when the ECGs were undertaken was not systematically defined. To evaluate cardiac arrhythmias and heart rate variability in children with dengue virus infection during the convalescent stage using Holter monitoring. Overnight 18-24-hour Holter monitoring was performed in 35 children [mean (SD) age 11·7 (2·3) y] at least 24 hours after defervescence (on the last day of admission). In 17 patients, time- and frequency-domain short-term (5 minutes) heart rate variability (HRV) during the convalescent stage was also compared with the value obtained during the follow-up visit (at least 14 days after defervescence). During the convalescent stage, cardiac rhythm abnormalities were found in ten patients (29%), including sinus pause (1), first-degree (2) and Mobitz type I second-degree AV block (Wenckebach) (3) and atrial (4) and ventricular ectopic beats (5). There was no relationship between the clinical severity of dengue virus infection (DF, DHF without shock and DSS) and the incidence of cardiac arrhythmia. There was no significant difference in the averaged RR interval, the time-domain HRV (SDNN, RMSSD, pNN 50) or frequency-domain HRV (LF, HF, LF/HF ratio) between the convalescent stage and at follow-up. Various benign bradyarrhythmias and ectopic beats are detected in patients with dengue virus infection during the convalescent stage.

Matrix Metalloproteinase-9 (MMP-9) in Children with Dengue Virus Infection

The purpose of this study was to investigate the role of matrix metalloproteinase-9 (MMP-9) in the pathogenesis of vascular leakage in patients with dengue virus infection. Serum samples from 24 children with serologically confirmed dengue virus infection (dengue fever [DF], 16; dengue hemorrhagic fever [DHF], 8; age, 9.5+/-2.4 years; 67% male] were analyzed for MMP-9 during the febrile and toxic stages and at follow-up. Serum samples obtained from 7 healthy children were used as controls. Serum MMP-9 levels in patients with dengue virus infection were found to be lower at the febrile (227.0+/-186.9 ng/ml) and toxic stages (150.9+/-151.7 ng/ml) than at follow-up (424.5+/-227.8 ng/ml) or in the control group (393.3+/-125.9 ng/ml, P<0.001 by one-way ANOVA). There was no significant difference between MMP-9 levels in patients with DHF and those with DF at any stage of the disease. In conclusion, MMP-9 levels are reduced during the febrile and toxic stages of dengue virus infection.

Increased level of hepatocyte growth factor in children with dengue virus infection

Evidence of hepatocellular damage is common in dengue-infected individuals. Hepatocyte growth factor (HGF), a key cytokine responsible for liver regeneration, may play a prognostic role in dengue virus infection. To determine the relationship between serum HGF level and disease severity in patients with dengue virus infection. Serum samples from 27 children [17 dengue fever (DF), ten dengue haemorrhagic fever (DHF)] with serologically confirmed dengue virus infection during the febrile, toxic stages and at follow-up were analysed for HGF. Serum samples obtained from nine healthy children served as the control group. In dengue-infected patients, serum HGF was significantly higher at the febrile and toxic stages than at follow-up (p<0.05). In comparison with DF, patients with DHF had a greater level of HGF at the febrile stage (p<0.05). A cut-off HGF level of 1220 pg/mL obtained during the febrile stage showed a sensitivity of 90% and a specificity of 53% for predicting clinical progression to DHF (area under the ROC curve 0.75). Serum HGF level at the early stage of dengue virus infection is elevated and may be a useful predictor for clinical progression to DHF.

Effect of dengue virus infection on the host signal transduction pathways

Background: Dengue virus (DENV) is a single, positive-stranded RNA virus from the family Flaviviridae. Dengue infection is a medically important infectious disease affecting many tropical and subtropical countries. To date, vaccine and anti-viral drugs for DENV infection is still not available. This is mainly due to the incomplete understanding of the pathogenesis and the underlying molecular mechanism involved during infection. This project aims to study the virus-host interplay by investigating the signal transduction pathways that are activated during DENV infection. Methods: RT2 Profiler PCR array from SABiosciences was employed to screen for the pathways that were affected during DENV infection on Human Embryonic Kidney (HEK)-293 cells. A panel of 84 genes from 18 biological pathways were analyzed and quantified simultaneously. The HEK293 cells were infected with DENV at multiplicity of infection of 1 and the total RNA were extracted after 12hr, 24hr, and 48hr post infection (p.i.). The RNA of infected cells and mock infected cells were reverse-transcribed into cDNA which were then served as templates for real-time polymerase chain reaction in a 96-well plate containing predispensed gene specific primer sets. The gene expression profiling of DENV-infected cells were compared to that of mock-infected cells. Results: The PCR array data showed that there was no significant change in the gene expression profile of infected cells at 12hr p.i. as compared to mock infected cells. Nevertheless, the number of activated genes increased from 24hr p.i. to 48hr p.i. Among 18 signal transduction pathways, the most affected pathways are NFκB, CREB, Jak-Stat, phospholipase C, insulin and low density lipoprotein pathways. Besides, there are also novel genes identified to be activated in this study. All the activated genes were analyzed and an interactome map was generated. It provides a better understanding of the virus-host interplay during different stages of DENV infection. Conclusion: PCR array allows fast and convenient analysis of many gene expression profiles simultaneously for different samples. Therefore, the comparison of the gene expression profiles of all four different DENV serotypes can be carried out to study the similarities and differences in virus-host interaction. Abstracts for SupplementInternational Journal of Infectious DiseasesVol. 14Preview Full-Text PDF Open Archive

The role of vascular endothelial growth factor leading to vascular leakage in children with dengue virus infection

Increased vascular permeability is the main aetiology for hypovolaemic shock and circulatory failure in dengue haemorrhagic fever (DHF). To investigate the role of vascular endothelial growth factor (VEGF) in the pathogenesis of DHF. Serum samples from 41 patients [15 dengue fever (DF), 26 DHF] with serologically confirmed dengue virus infection during the febrile, toxic, convalescent stages and at follow-up were analysed for VEGF. Plasma samples from an additional 27 children (16 DF and 11 DHF) during the febrile, toxic stages and at 4-week follow-up and from eight healthy controls were analysed for VEGF. Serum and plasma VEGF levels were not elevated during the febrile or toxic stages of dengue virus infection and did not differ between patients with DF and DHF. Plasma leakage in patients with DHF cannot be explained by elevation of VEGF during the toxic stage of the illness.

Activation of coagulation and fibrinolysis during dengue virus infection.

Dengue virus infection can induce mild dengue fever (DF) or severe dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS) in human. The pathogenesis of hemorrhage in dengue virus infection is not fully understood. Since hemostasis depends on the balance between coagulation and fibrinolysis, alternation of some coagulation parameters (platelet count and activated partial thromoboplastin time, APTT) as well as fibrinolytic parameters (tissue plasminogen activator, tPA and plasminogen activator inhibitor-1, PAI-1) were compared in 8 DHF/DSS and 17 DF patients. Patients showed thrombocytopenia, APTT prolongation, and tPA increase in the acute stage of disease, indicating activation of coagulation and fibrinolysis. The activation of coagulation and fibrinolysis in DHF/DSS patients was much more severe than DF patients. In the convalescent stage, a rise of PAI-1 level and platelet count with concomitant decline of tPA level and APTT returned to normal in both DHF/DSS and DF patients. Therefore, the activation of coagulation and fibrinolysis during the acute stage of dengue virus infection is offset by the increase of platelet and PAI-1 during convalescent stage. Taken together, these results suggest that the degree of coagulation and fibrinolysis activation induced by dengue virus infection is associated with the disease severity.