Types and functions of heterogeneity in mycobacteria

Abstract

The remarkable ability of Mycobacterium tuberculosis to survive attacks from the host immune response and drug treatment is due to the resilience of a few bacilli rather than a result of survival of the entire population. Maintenance of mycobacterial subpopulations with distinct phenotypic characteristics is key for survival in the face of dynamic and variable stressors encountered during infection. Mycobacterial populations develop a wide range of phenotypes through an innate asymmetric growth pattern and adaptation to fluctuating microenvironments during infection that point to heterogeneity being a vital survival strategy. In this Review, we describe different types of mycobacterial heterogeneity and discuss how heterogeneity is generated and regulated in response to environmental cues. We discuss how this heterogeneity may have a key role in recording memory of their environment at both the single-cell level and the population level to give mycobacterial populations plasticity to withstand complex stressors.

Figure 1.. Mycobacterial asymmetry.

(a) In mycobacteria, growth is asymmetric: the old pole elongates more than the new pole. The sister that inherits the old pole from the mother called the accelerator cell) is born longer and elongates more than the sister that inherits the new pole (the alternator cell). (b) The cell elongation and division protein Wag31 and growth inhibitor LamA are regulators of asymmetric growth in mycobacteria. Inhibition of growth at the new pole by LamA is relieved after a time lag. MurG, GlfT2 and Pks13 are cell wall synthesis complexes in Mycobacterium smegmatis that are localized in the subpolar regions with their highest local concentration at the old pole. (c) In M. smegmatis, chromosome positioning is asymmetric with the terminus (ter) closer to the new pole than the origin of replication (ori) is to the old pole. The asymmetric positioning of the chromosome is proportional to cell size. The future sites of asymmetric division are determined in mother and grandmother cells and can be observed by surface wave troughs that are inherited from previous generations. (d) Asymmetric distribution of irreversibly oxidized proteins (IOPs) in Mycobacterium tuberculosis creates sister cells with different growth properties and sensitivities against antibiotic stresses depending on their IOP-burdens. Specifically, sister cells with a high amount of IOPs are more sensitive to drugs.

Figure 2.. Environmental heterogeneity.

Microenvironments in tuberculosis lesions (granulomas) vary among patients, within one patient, and even within a lesion as shown with examples in the first, second, and third columns, respectively. Lesion composition is dependent on the stage of disease progression and the network of immune cells present. Many types of cells respond to infection with Mycobacterium tuberculosis, including macrophages, neutrophils, T and B cells, epithelioid cells, and fibroblasts, and populate cellular granulomas. M. tuberculosis in infected macrophages encounter an environment characterized by low pH, oxidative stress and abundant lipids. As M. tuberculosis utilizes these host lipids, it secretes lipid vesicles into the macrophage cytoplasm, inducing some of the infected macrophages to become foamy with lipid droplets. Cytoplasmic debris from necrotic macrophages form the granuloma's caseous center, which may be acidic or neutral and vary in oxygen content.

Figure 3.. Population structure and complex Mycobacterium tuberculosis phenotypes.

A hypothetical population structure framework maps single-cell and population-level cellular states and phenotypes across multiple processes. The locations, dense regions and spread in this phase diagram change due to adaptation to different environmental conditions, including drug treatment. Each axis (property A and property B) can represent cellular states, such as cell size, gene expression or virulence. Abstracted axes, as being used for illustrative examples here, may be made quantitative using weighted sums of specific cellular features through dimensionality reduction techniques such as principal component analysis. Condition 1 is an example wherein cells are evenly distributed throughout the range of properties A and B. During adaptation to new environments and stressors, the population structure can be shifted and/or concentrated to other regions of the phase diagram. The degree of bacterial heterogeneity can become amplified (condition 2) or attenuated (condition 3). Following antibiotic treatment, the size of the tolerant subpopulation may vary depending on which condition Mycobacterium tuberculosis has adapted to (for example, 21% in condition 1, 55% in condition 2, 0.04% in condition 3 following treatment with drug A). The region that encompasses drug-tolerant cell states will be dependent on the activity and mechanism of action of the drug (compare drug A versus drug B), highlighting that there are different kinds of persister cells.