Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Dec;83(12):4731-9.
doi: 10.1128/IAI.01055-15. Epub 2015 Sep 28.

Differential influence of nutrient-starved Mycobacterium tuberculosis on adaptive immunity results in progressive tuberculosis disease and pathology

Jes Dietrich  1 , Sugata Roy  2 , Ida Rosenkrands  2 , Thomas Lindenstrøm  2 , Jonathan Filskov  2 , Erik Michael Rasmussen  3 , Joseph Cassidy  4 , Peter Andersen  2
Affiliations

Differential influence of nutrient-starved Mycobacterium tuberculosis on adaptive immunity results in progressive tuberculosis disease and pathology

Jes Dietrich et al. Infect Immun. 2015 Dec.

Erratum in

Abstract

When infected with Mycobacterium tuberculosis, most individuals will remain clinically healthy but latently infected. Latent infection has been proposed to partially involve M. tuberculosis in a nonreplicating stage, which therefore represents an M. tuberculosis phenotype that the immune system most likely will encounter during latency. It is therefore relevant to examine how this particular nonreplicating form of M. tuberculosis interacts with the host immune system. To study this, we first induced a state of nonreplication through prolonged nutrient starvation of M. tuberculosis in vitro. This resulted in nonreplicating persistence even after prolonged culture in phosphate-buffered saline. Infection with either exponentially growing M. tuberculosis or nutrient-starved M. tuberculosis resulted in similar lung CFU levels in the first phase of the infection. However, between week 3 and 6 postinfection, there was a very pronounced increase in bacterial levels and associated lung pathology in nutrient-starved-M. tuberculosis-infected mice. This was associated with a shift from CD4 T cells that coexpressed gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) or IFN-γ, TNF-α, and interleukin-2 to T cells that only expressed IFN-γ. Thus, nonreplicating M. tuberculosis induced through nutrient starvation promotes a bacterial form that is genetically identical to exponentially growing M. tuberculosis yet characterized by a differential impact on the immune system that may be involved in undermining host antimycobacterial immunity and facilitate increased pathology and transmission.

PubMed Disclaimer

Figures

FIG 1
FIG 1
Generating nutrient-starved mycobacteria. (A) Overview of the procedure to generate nutrient-starved mycobacteria. (B) StarvM.tb and LogM.tb bacteria were transferred to Sauton medium, and the numbers of CFU were determined at the time points indicated. (C) StarvM.tb and LogM.tb bacteria were counted either by plating on agar or by determining the MPN. (D) Growth measured by absorbance of StarvM.tb and LogM.tb in liquid culture over a period of 15 days. (E and F) LogM.tb (E) or StarvM.tb (F) bacteria were stained with auramine-rhodamine. Magnifications are indicated on the panels.
FIG 2
FIG 2
In vivo growth of StarvM.tb and LogM.tb bacteria. (A and B) Bacterial load (shown as log10 CFU) in the lungs of mice infected with StarvM.tb or LogM.tb via the aerosol route. Two experiments are shown in panels A and B, respectively. Data represent the mean ± SD for a minimum of four mice per group. *, P < 0.05.
FIG 3
FIG 3
Macroscopic analysis and immunopathology of infected lungs. Mice infected by the aerosol route with StarvM.tb or LogM.tb were sacrificed 3 (A to C) or 6 (D and E) weeks after challenge and were examined macroscopically (A, D, and E) or subjected to histopathological examination (B and C [which shows the week 3 time point]). For histopathology, the right lung was fixed by immersion in 10% neutral buffered formalin and processed for examination. Cut sections were stained with hematoxylin and eosin (HE). Lesion morphometry was carried out using the public-domain, Java-based image processing program ImageJ. In panel C, results for lesions in lungs are from three independent experiments (represented by unique symbols).
FIG 4
FIG 4
T cell-induced IFN-γ secretion. (A) Lung cells were stimulated with TB10.4 or ESAT6 in vitro for 72 h, and IFN-γ levels in supernatants were assessed by ELISA. Data represent the mean ± SEM of the results for a minimum of three mice per group. (B) Mice were sacrificed 3 or 6 weeks after the challenge, and lymphocytes from lungs were stimulated in vitro with TB10.4 peptides prior to staining with anti-CD4, -CD8, -CD44, -IFN-γ, -IL-2, and -TNF-α. Cytokine profiles of specific CD4 and CD8 T cells 3 weeks after an aerosol challenge are shown. Frequencies represent cytokine-producing T cells within the subset of T cells specific for TB10.4. Data represent the mean ± SEM of the results for a minimum of three mice per group (*, P < 0.05 [two-way analysis of variance with Bonferroni's posttest]). (C) At week 3 after aerosol infection, lung cells were stimulated with TB10.4 or ESAT6 in vitro for 72 h, and IL-10 and IL-5 levels in supernatants were assessed by ELISA. Data represent the mean ± SEM of the results for a minimum of three mice per group.
FIG 5
FIG 5
Vaccination against infection with StarvM.tb. (A) Animals were vaccinated with Ag85B–ESAT-6 in CAF01 adjuvant and infected with StarvM.tb or LogM.tb at week 6 after the last vaccination. CFU levels were determined in the lungs at week 6 postinfection. (B) Cytokine profiles of ESAT-6-, Ag85B-, and TB10.4-specific CD4 T cells 6 weeks after aerosol challenge.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Smeulders MJ, Keer J, Speight RA, Williams HD. 1999. Adaptation of Mycobacterium smegmatis to stationary phase. J Bacteriol 181:270–283. - PMC - PubMed
    1. Gengenbacher M, Kaufmann SHE. 2012. Mycobacterium tuberculosis: success through dormancy. FEMS Microbiol Rev 36:514–532. doi:10.1111/j.1574-6976.2012.00331.x. - DOI - PMC - PubMed
    1. Kochi A. 1994. Tuberculosis: distribution, risk factors, mortality. Immunobiology 191:325–336. doi:10.1016/S0171-2985(11)80437-7. - DOI - PubMed
    1. Tsai MC, Chakravarty S, Zhu G, Xu J, Tanaka K, Koch C, Tufariello J, Flynn J, Chan J. 2006. Characterization of the tuberculous granuloma in murine and human lungs: cellular composition and relative tissue oxygen tension. Cell Microbiol 8:218–232. doi:10.1111/j.1462-5822.2005.00612.x. - DOI - PubMed
    1. Nathan C, Shiloh MU. 2000. Reactive oxygen and nitrogen intermediates in the relationship between mammalian hosts and microbial pathogens. Proc Natl Acad Sci U S A 97:8841–8848. doi:10.1073/pnas.97.16.8841. - DOI - PMC - PubMed

Publication types

MeSH terms