CD4⁺ T cells from Ech_0660 mutant vaccinated and wild-type <i>E</i>. <i>chaffeensis</i> infected animals proliferate in response to <i>E</i>. <i>chaffeensis</i> antigen.

<p>PBMC from dogs vaccinated with Ech_0660 and challenged with wild-type <i>E</i>. <i>chaffeensis</i> via needle inoculation (group 1, left panels), vaccinated with Ech_0660 and challenged with wild-type <i>E</i>. <i>chaffeensis</i> via tick inoculation (group 2, middle panels), or unvaccinated and infected with wild-type <i>E</i>. <i>chaffeensis</i> or Ech_0480 via tick inoculation (group 3, right panels) were labeled with Cell Trace Violet, then cultured for 5 days at 4x10⁶ cells/mL in the presence or absence of 10 ug/mL <i>E</i>. <i>chaffeensis</i> host-cell free lysate grown in the tick ISE6 cell line. On day 5, CD4⁺ T cells were analyzed by flow cytometry for Cell Trace Violet dilution as a measure of proliferation. (A) Representative Cell Trace Violet dilution profiles, gated on total live cells and total CD3⁺CD4⁺ T cells. (B) The percentage of CD4⁺ T cells that have proliferated in response to <i>E</i>. <i>chaffeensis</i> antigens as measured over the course of the experiment. The background (mock stimulated) proliferation was subtracted, and results represent change over mock. Each line is representative of a single animal.