Low induction of proinflammatory cytokines parallels evolutionary success of modern strains within the Mycobacterium tuberculosis Beijing genotype

Abstract

One of the most widespread clades of Mycobacterium tuberculosis worldwide, the Beijing genotype family, consists of ancient (atypical) and modern (typical) strains. Modern Beijing strains outcompete ancient strains in terms of prevalence, while reserving a higher degree of genetic conservation. We hypothesize that their selective advantage lies in eliciting a different host immune response. Bead-disrupted lysates of a collection of different M. tuberculosis strains of the modern (n = 7) or ancient (n = 7) Beijing genotype, as well as the Euro-American lineage (n = 6), were used for induction of ex vivo cytokine production in peripheral blood mononuclear cells (PBMCs) from 10 healthy individuals. Hierarchical clustering and multivariate regression analyses were used to study possible differences in production of nine cytokines. Modern and ancient M. tuberculosis Beijing genotypes induced different cytokine signatures. Overall induction of interleukin-1β (IL-1β), gamma interferon (IFN-γ), and IL-22 was 38 to 40% lower after stimulation with modern Beijing strains (corrected P values of <0.0001, 0.0288, and 0.0002, respectively). Euro-American reactivation strains induced 2-fold more TNF-α production than both types of Beijing strains. The observed differences in cytokine induction point to a reduction in proinflammatory cytokine response as a possible contributing factor to the evolutionary success of modern Beijing strains.

Fig 1

Mycobacterium tuberculosis strains used for this study, with genetic markers and countries of origin. IS6110 RFLP profiles and clustering are shown, as well as typing by spoligotyping and determination of the presence of IS6110 in the NTF region. Note that all reactivation strains were isolated from Dutch patients over 70 years of age. n/a, not available.

Fig 2

Hierarchical clustering of modern and ancient Beijing strains and Euro-American reactivation strains. The figure shows clustering of the virtual distances between the different M. tuberculosis strains based on the cytokines they induce in PBMCs. The distance matrix shows similarity (red) and dissimilarity (green) between cytokine signatures of the respective strains. Analysis was performed on ln-transformed and normalized data with the open source software J Express (17). See the text for further details.

Fig 3

(A) Differences in cytokine response after stimulation with modern (red) compared to ancient (blue) Beijing strains. (B) Differences in cytokine response after stimulation with modern (red) and ancient (blue) Beijing strains compared to Euro-American reactivation strains (green). Relative cytokine responses were calculated by multivariate regression. The shaded areas show the confidence intervals, which were adjusted by the Bonferroni correction to take into account the multiple comparisons made. The axis is on the ln scale; corresponding percentages are shown in the legend. A lower response is indicated by projection toward the center of the figure. Asterisks indicate the significance of the differences, as follows: *, P < 0.05; ***, P < 0.001; ****, P < 0.0001.

Fig 4

IL-1β responses of 10 healthy donors to modern and ancient Beijing strains and Euro-American reactivation strains. (A) Strain-associated differences in cytokine production. (B) Donor-associated differences in cytokine production for modern and ancient Beijing strains, for all 10 donors individually.