Dengue virus (DENV) is the most prevalent mosquito-borne virus causing human disease. Of the 4 DENV serotypes, epidemiological data suggest that DENV-2 secondary infections are associated with more severe disease than DENV-4 infections. Mass cytometry by time-of-flight (CyTOF) was used to dissect immune changes induced by DENV-2 and DENV-4 in human DCs, the initial targets of primary infections that likely affect infection outcomes. Strikingly, DENV-4 replication peaked earlier and promoted stronger innate immune responses, with increased expression of DC activation and migration markers and increased cytokine production, compared with DENV-2. In addition, infected DCs produced higher levels of inflammatory cytokines compared with bystander DCs, which mainly produced IFN-induced cytokines. These high-dimensional analyses during DENV-2 and DENV-4 infections revealed distinct viral signatures marked by different replication strategies and antiviral innate immune induction in DCs, which may result in different viral fitness, transmission, and pathogenesis.
Rebecca E. Hamlin, Adeeb Rahman, Theodore R. Pak, Kevin Maringer, Ignacio Mena, Dabeiba Bernal-Rubio, Uma Potla, Ana M. Maestre, Anthony C. Fredericks, El-ad D. Amir, Andrew Kasarskis, Irene Ramos, Miriam Merad, Ana Fernandez-Sesma
Comparing cellular marker positivity during mock, DENV-2, and DENV-4 infections by autogating.
Heat maps comparing cellular marker positivity by denoting cells as either positive or negative for each marker during mock, dengue virus 2 (DENV-2), and DENV-4 infections. For all plots, markers are ordered vertically by hierarchical clustering of the rows, which places markers with similar expression profiles closer together. Statistical significance was determined using a 2-sided Fisher’s exact test at a threshold of α = 10–5 with Bonferroni correction for multiple testing. Cellular markers not representing a significant difference between infection conditions at this threshold are labeled with “NS”; all others are significant. Calculations were made using the event counts from mass cytometry by time-of-flight (CyTOF) data, as determined by autogating of cells positive for each marker based on the 99th percentile threshold, which was set for mock treatment at 8 hours after infection (hpi). The comparative effects on each marker were calculated as the log10 of the relative risk for positive marker frequency between the indicated infection conditions. The histograms within the heat map color scales show the total number of cells matching that intensity range, revealing the overall distribution of the values. One representative DC donor of seven is shown. (