Clinical presentations of dengue fever (DF) are diverse and non-specific, causing unpredictable progression and outcomes. Its progression and severity have been associated with cytokine levels alteration. In this study, dengue patients were classified into groups following the 2009 WHO dengue classification scheme to investigate the cytokine signature at different severity of the disease: dengue without warning sign symptoms (A); dengue with warning signs (B); severe dengue (C); other fever (OF) and healthy (Healthy). We analyzed 23 different cytokines simultaneously, namely IL-1b, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-33, CD14, CD54, CD62E, CD62L, CD62p, CD106, CD121b, CD154, CD178, GM-CSF, IFN-g, MIF, ST2 and TNF from patients admitted to National Cheng Kung University Hospital during the 2015 Taiwan dengue outbreak. Cytokines TNF, CD54, CD62E, CD62L, CD62P, GM-CSF, IL-1b, IL-2, IL-6, IL-8, IL-10, IL-12p70, IL-17A, INF-g and MIF were elevated while CD106, CD154, IL-4 and L-33 were decreased when compared to the control. IL-10 demonstrated to be a potential diagnostic marker for DF (H and A group; AUC = 0.944, H and OF group; AUC = 0.969). CD121b demonstrated to be predictive of the SD (A and B group; AUC = 0.744, B and C group; AUC = 0.775). Our results demonstrate the cytokine profile changes during the progression of dengue and highlight possible biomarkers for optimizing effective intervention strategies.

Background Recent studies have shown that dengue virus (DENV) can efficiently infect bone marrow hematopoietic stem cells (HSCs) as well as the placenta of pregnant women. Although mother-to-infant vertical transmission of DENV through the placenta has been well documented, the evidence of cell-associated vertical transmission is still unknown. Whether DENV can infect umbilical cord blood (UCB) cells before reaching the fetus remains to be explored. Here, we proposed that human UCB cells were permissive to the DENV infection and DENV infected CD133+ and CD34+ HSCs are reservoir of the virus that could be reactivated upon re-culturing in suitable cells. Methods Human UCB cells were freshly obtained and subjected to DENV infection. Multicolor flow cytometry (MFCM) was used to demonstrate the phenotypes of the infected HSC populations. Immunofluorescence analysis (IFA) and T-distributed Stochastic Neighbor Embedding (t-SNE) were used to show the association of the DENV antigen, non-structural protein1 (NS1) with HSCs. Key findings UCB cells were highly permissive to DENV infection. DENV altered the phenotype of the infected HSC population, increased the expression of HSCs, and affected the balance of transcription factors (TFs, GATA1/2/3). IFA revealed the association of the DENV antigen, non-structural protein1 (NS1), with CD34 + and CD133 + cells. T-distributed Stochastic Neighbor Embedding (t-SNE) analysis revealed heterogeneity in the distribution of CD133 + NS1 + , and CD34 + NS1 + cells. DENV particles were recovered from CD133 + and CD34 + cells even when virus production in the supernatant was negligible. Significance We predict that infection of CD133+ and CD34+ cells in the UCB serve as reservoirs for the amplification of DENV in UCB prior to the virus reaching the fetus and facilitate vertical transmission.

Although dengue virus (DENV) can establish infections in hematopoietic stem progenitor cells (HSPCs), there is little information on dengue virus persistent infection in CD34+ and CD133+ cell surface glycoproteins of hematopoietic stem cells (HSCs). CD34 and CD133 also function as cell–cell adhesion factors, which are present in umbilical cord blood (UCB). In this study, we sought to establish a persistent infection model of DENV infection in UCB using a prolonged period of infection lasting 30 days. Post-infection, the results exhibited a productive and non-productive phase of DENV production. Using a plaque assay, Western blot, and confocal microscopy, we demonstrated that CD133 and CD34 cells are target cells for DENV infection. Moreover, we showed that DENV particles can be recovered from the non-productive phase of DENV-infected CD34 and CD133 cells after co-incubation with Vero cells. We concluded that CD133 and CD34 retain their capacity to produce the infectious virus due to proliferation and their ability to repopulate, as deduced from a BrdU proliferation assay and flow cytometry analysis using t-distributed stochastic neighbor embedding. In summary, the platform to co-culture infected primitive HSCs from their non-productive phase onto Vero cells will give new insights into understanding the DENV dynamics in cell-to-cell transmission and reactivation of the virus.

Although dengue virus (DENV) can establish infection in hematopoietic stem progenitor cells (HSPCs), there is little information on dengue virus persistent infection in CD34+ and CD133+ cell surface glycoprotein of hematopoietic stem cells (HSCs). CD34 and CD133 also function as cell-cell adhesion factors which are present in umbilical cord blood (UCB). In this study, we attempted to establish a persistent infection model of DENV infection in UCB by infecting for a prolonged period of 30 days. Post-infection of DENV exhibited a productive and non-productive phase of DENV production. Using plaque assay, western blot, and confocal microscopy, we Show that CD133 and CD34 cells are target cells for DENV infection. Moreover, we show that DENV particles can be recovered from the non-productive phase of DENV infected CD34 and CD133 cells after co-incubation with Vero cells. We concluded that CD133 and CD34 retain their capacity to produce the infectious virus due to proliferation and their ability to repopulate, as deduced from BrdU proliferation assay and flow cytometry analysis using t-distributed stochastic neighbor embedding. In summary, the platform to co-culture infected primitive HSCs from its non-productive phase onto Vero cells will give new insight into understanding the DENV dynamics in cell-to-cell transmission and reactivation of the virus.

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