Journal: bioRxiv

Article Title: Chikungunya virus glycoproteins transform macrophages into productive viral dissemination vessels

doi: 10.1101/2023.05.29.542714

Figure Lengend Snippet: a, Human peripheral monocyte derived macrophages were infected with EGFP-labeled alphaviruses (SINV TE/5’2J/GFP, RRV strain T48, ONNV strain SG650, and CHIKV vaccine strain 181/clone 25) at MOI of 5 for 24 h. Levels of infection with different alphaviruses were determined by percent EGFP-positive cells evaluated by flow cytometry. Data are representative of 2 independent experiments performed in biological duplicates. b, THP-1 derived macrophages were infected with CHIKV 181/clone 25 or ONNV SG650 at MOI 5. Titration of supernatant virus samples was performed at 0, 6, 14, 24, 48, and 72 h.p.i by plaque assay on BHK-21 cells. Data are representative of 2 independent experiments. Mean values of biological duplicates were plotted with SD. Asterisks indicate statistically significant differences as compared to ONNV (Two-way ANOVA and Šidák’s multiple comparisons test: *, p≤0.05; ****, p≤0.0001). c, Levels of intracellular (-) vRNAs, the viral replication intermediate, at 4, 8, 14, and 24 h post transfection of THP-1 derived macrophages with CHIKV 181/clone 25 or ONNV SG650 were quantified through RT-qPCR with specific TaqMan probes. Data are representative of 2 independent experiments. Mean values of biological duplicates measured in technical duplicates were plotted with SD (Two-way ANOVA and Šidák’s multiple comparisons test: *, p≤0.05; **, p≤0.01; ***, p≤0.001). d, CHIKV and ONNV titers of supernatant samples collected from transfected THP-1 derived macrophages in were determined by plaque assay. Representative plaques of CHIKV and ONNV from 2 independent experiments (1:100 dilution) are shown.

Article Snippet: The appropriate reverse nsP1 primer, tag-specific forward primer, and FAM-labeled TaqMan probe were used in viral negative strand quantification with Luna Universal Probe qPCR Master Mix (New England Biolabs).

Techniques: Derivative Assay, Infection, Labeling, Flow Cytometry, Titration, Plaque Assay, Transfection, Quantitative RT-PCR