scRNAseq analysis of mouse draining lymph nodes during Ross River virus or LCMV infection. scRNAseq analysis of mouse draining lymph nodes during Ross River virus or LCMV infection

The aim of this study was to compare murine immune responses, particularly CD8+ T cell activation, in acute infection of either Ross River virus (RRV) or lymphocytic choriomeningitis virus (LCMV). LCMV induces a strong CD8+ T cell response that is required for viral clearance. RRV induces CD8+ T cells that only affect viral clearance in certain tissue and do not contribute to prevention of viral RNA persistence in lymphoid and joint-associated tissue. We sequenced cells from the draining lymph node at 5 days post infection to assess activation states of CD8+ T cells at this timepoint. Overall design: To study immune responses in the draining lymph node after viral infection, with a focus on CD8+ T cell activation, three samples were generated: naive, RRV, and LCMV. Wild type mice each received adoptive transfer of 1 x 10^6 CD8+ T cells from a P14 transgenic mouse (specific for the gp33 epitope naturally found in LCMV and genetically expressed by RRV-gp33) and were then inoculated in the footpad with 10^3 infectious units of RRV-gp33 or LCMV. Five days post infection, draining lymph nodes (popliteal) were harvested from the infected mice and also uninfected wild type and P14 transgenic mice as a naive control. Cells from 7 mice were pooled for each group: naive, RRV, and LCMV. A portion of each sample was enriched for transgenic CD8+ T cells by magnetic bead negative selection and fluorescence-activated cell sorting and added back to the bulk sample at a 1:4 ratio. Libraries were prepared from 20,000 cells from each sample using a 10x Genomics Chromium Single Cell 5' Reagent Kit v2 with TCR enrichment and sequenced.