Background: The host immune response is crucial in determining the outcome after exposure
to Mtb. A substantial proportion of exposed individuals (40-70%) never develop infection
indicating an efficient cell mediated immune response against Mtb and early clearance by
the host immune defense (Verall et al, Immunology 2014). Thus, the outcome of exposure is
highly dependent on host immunity. In this study, our aim was to investigate whether the
previously developed mycobacterium groth inhibition assay (MGIA) method (Andersson et al,
Tuberculosis 2020) could be used to quantify the host immune response to Mtb.

Study design: In this study we will include healthy donors (n=80), patient with active TB
(n=40), subjects recently exposed to active TB (n=80) and patients with latent TB (LTBI)
(n=80). Exclusion criteria for all groups are known HIV infection or other
immunosuppression from treatment or disease. Patients and healthy blood donors will be
recruited at the departments of infectious diseases in Region Kalmar and Region
Östergötland. From patients giving oral and written consent, 40 ml of blood will be
collected for MGIA analysis.

Primary aim: To determine the difference in MGIA between healthy blood donors and
patients with latent and active TB as well as the variation in host control of Mtb within
each group.

Method: The Mycobacterial growth inhibition assay (MGIA) using human PBMCs isolated by
density gradient by Lymhoprep (Axis-Shield) and SepMate tubes (StemCell Technologies)
will be performed as described in study I with minor modifications. The PBMCs are
infected with GFP expressing M. tuberculosis H37Rv and intracellular growth of Mtb and
viability of PBMCs are assessed by live-cell imaging (Incucyte©) and measured as relative
fluorescence units (RFU) during 5 days with and without stimulation with the purified
protein derivate (PPD) is included. The cytokine response following exposure to
gamma-irradiated H37Rv will be quantified by Olink proteomics.