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In the absence of a correlate(s) of protection against human tuberculosis and a validated animal model of the disease, tools to facilitate vaccine development must be identified. We present an optimised  ex vivo  mycobacterial growth inhibition assay (MGIA) to assess the ability of host cells within the lung to inhibit mycobacterial growth, including Bacille Calmette–Guérin (BCG) and  Mycobacterium tuberculosis  (MTB) Erdman. Growth of BCG was reduced by 0.39, 0.96 and 0.73 log 10  CFU following subcutaneous (s.c.) BCG, intranasal (i.n.) BCG, or BCG s.c. + mucosal boost, respectively, versus naïve mice. Comparatively, a 0.49 (s.c.), 0.60 (i.n.) and 0.81 (s.c. + mucosal boost) log 10  reduction in MTB CFU was found. A BCG growth inhibitor, 2-thiophenecarboxylic acid hydrazide (TCH), was used to prevent quantification of residual BCG from i.n. immunisation and allow accurate MTB quantification. Using TCH, a further 0.58 log 10  reduction in MTB CFU was revealed in the i.n. group. In combination with existing methods, the  ex vivo  lung MGIA may represent an important tool for analysis of vaccine efficacy and the immune mechanisms associated with vaccination in the organ primarily affected by MTB disease.

Adaption of the ex vivo mycobacterial growth inhibition assay for use with murine lung cells