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Review
. 2017 Sep 5;24(9):e00142-17.
doi: 10.1128/CVI.00142-17. Print 2017 Sep.

The Cross-Species Mycobacterial Growth Inhibition Assay (MGIA) Project, 2010-2014

Affiliations
Review

The Cross-Species Mycobacterial Growth Inhibition Assay (MGIA) Project, 2010-2014

Michael J Brennan et al. Clin Vaccine Immunol. .

Abstract

The development of a functional biomarker assay in the tuberculosis (TB) field would be widely recognized as a major advance in efforts to develop and to test novel TB vaccine candidates efficiently. We present preliminary studies using mycobacterial growth inhibition assays (MGIAs) to detect Mycobacterium bovis BCG vaccine responses across species, and we extend this work to determine whether a standardized MGIA can be applied in characterizing new TB vaccines. The comparative MGIA studies reviewed here aimed to evaluate robustness, reproducibility, and ability to reflect in vivo responses. In doing so, they have laid the foundation for the development of a MGIA that can be standardized and potentially qualified. A major challenge ahead lies in better understanding the relationships between in vivo protection, in vitro growth inhibition, and the immune mechanisms involved. The final outcome would be a MGIA that could be used with confidence in TB vaccine trials. We summarize data arising from this project, present a strategy to meet the goals of developing a functional assay for TB vaccine testing, and describe some of the challenges encountered in performing and transferring such assays.

Keywords: MGIA; correlates of immunity; mycobacterial growth inhibition assay; tuberculosis; vaccines.

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Figures

FIG 1
FIG 1
Schematic of the MGIT mycobacterial growth inhibition assay method. Whole blood or cell culture medium containing the appropriate concentration of PBMCs (or mouse splenocytes in mouse studies) is inoculated with an equal volume of mycobacteria at a low MOI (∼1 CFU/10,000 PBMCs). Cultures are incubated in 2-ml tubes at 37°C for 96 h, with 360° rotation. Cells are then lysed to remove red blood cells and/or to release intracellular mycobacteria. The lysate is inoculated into Bactec MGITs. The tubes are placed in the Bactec 960 system and incubated at 37°C until positivity is detected by fluorescence. On day 0, direct-to-MGIT viability controls are set up by directly inoculating MGIT tubes with the same volume of mycobacteria as the samples. The TTP data are converted to log10 CFU values using a standard curve, and the final values are expressed as absolute CFU values or values relative to the control value, indicating the amount of growth inhibition that occurred during the culture period.
FIG 2
FIG 2
Direct whole-blood MGIA for South African infants. (A) Whole-blood samples from 10 South African infants were assessed using the direct whole-blood MGIA. There was a significant correlation between growth of BCG SSI and M. tuberculosis (M.tb) H37Rv (Spearman's r = 0.73; P = 0.02). (B) Whole-blood samples from 10 South African infants were compared with those from 53 South African adults using the direct whole-blood MGIA. There was a significant reduction in growth of BCG SSI in infants, compared with adults. *, P < 0.05, Mann-Whitney U test between groups.
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