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.