Distinct gene expression signatures comparing latent tuberculosis infection with different routes of Bacillus Calmette-Guérin vaccination [blood]

RNA-Seq was utilized to provide an unbiased assessment of systemic and mucosal immunity using blood cells from LTBI subjects and recipients of ID and oral (PO) BCG. Blood CD4+ T cells were studied from groups of Mtb/BCG-naïve, BCG-vaccinated and LTBI subjects. We combined the pre-vaccination studies of individuals who subsequently received ID and/or PO BCG. Blood was drawn into CPT tubes and PBMC isolated, aliquoted and frozen. Following thawing, CD4+ T cells were isolated using Miltenyi positive selection kits. Autologous monocyte-derived dendritic cells (MDDC) were prepared (Hoft et al, Mucosal immunology. 2018;11(2):486-95). MDDC were cultured overnight with autologous CD4+ T cells (20:1, T cell:MDDC ratio), in presence of either Danish BCG (Statens Serum Institut, Copenhagen, MOI 20:1), or medium alone. Cell pellets were lysed for RNA extraction.