**9. Low iron**

Normally iron taken up by intestinal epithelial cells and bound to transferrin circulates within the body. This complex binds to cell surface receptors, and is internalized where it releases its iron to be bound by the host cellular factor ferritin. Infection and inflammation are natural signals to the host to limit availability of iron. Proinflammatory cytokines stimulate hepcidin production, decrease iron uptake from the gut, and inhibits the iron efflux protein ferroportin (Johnson and Wessingling-Resnick, 2012). Inflammation thus inhibits iron uptake by the intestinal epithelium thus preventing iron from being loaded onto transferrin. Interfering with uptake limits iron availability in the host, and *M. tuberculosis* has been shown to be severely growth restricted in a low iron environment. It has been demonstrated in African studies that iron supplementation increases incidence of tuberculosis. Thus being anemic may be protec‐ tive against infectious processes. Within human macrophages, Nramp1 (natural resistance associated macrophage protein) is produced and localizes to the phagosomal compartment where it reduces iron within this site possibly by extrusion. This function confers resistance to *M. tuberculosis* infections and mutations in the *nramp1* gene can result in increased suscepti‐ bility to active disease due to *M. tuberculosis* infection (Johnson and Wessingling-Resnick, 2012).

gulated by oxidative stress indicating this is an important stress *in vivo* (Wu et al, 2007). In addition nutrients are limited in the phagosome which may cause *M. tuberculosis* to enter a stationary phase of growth, which has been shown to induce internal oxidative damage. The gene *whiB1* is more active during stationary phase, and the protein produced by this gene has been shown to reduce cellular disulphide bridges that may predominate during this adapta‐

Mycobacteria contain a unique substance, mycothiol, which combats oxidative stress. Other bacterial species utilize glutathione which can also neutralize oxidative stress. Mycothiol contains cysteine residues which are oxidized when that condition predominates thus forming disulfide bonds, creating mycothione, and preventing other molecules in the mycobacterial cell from becoming oxidized (Table 1.). Human cells produce glutathione to combat oxidative damage, and glutathione is toxic to mycobacterial cells perhaps due to a redox imbalance generated by this substance in the mycobacteria (Venketaraman et al, 2008; Connell et al, 2008)). Mycobacteria also contain other molecules to detoxify oxidative damage including superoxide dismutase (SOD) and catalase (KatG) which can inactivate superoxide (Table 1.) (Shi et al, 2008). SOD and KatG are upregulated early in infection indicating an increase in oxidative damage due to superoxide. Oxidative damage is capable of harming DNA, and histone like proteins (LSR2) can protect against damage by compacting DNA and acting as a physical barrier. UvrB which repairs mycobacterial DNA damage also protects against

One of the hallmarks of tuberculosis is fever and night sweats in which body temperature increases and is suboptimal for *Mycobacterium tuberculosis* replication and survival. This allows the immune system a competitive edge over the invading microbes. Heat stress can cause damage to *M. tuberculosis* by causing proteins to unfold which may then be degraded. In response, *M. tuberculosis* can upregulate chaperonins which complex with unfolded proteins and help them refold (Table 1.). The α-crystalline protein, or Acr-2, is activated by heat shock,

Many proteins that are upregulated in *M. tuberculosis in vivo* are heat shock proteins that have chaperonine activity. While these proteins may benefit the organism by complexing with and refolding heat damaged proteins, they are also recognized by the immune system. Both the 65Kd heat shock protein and the HSP70 protein can be found extracellularly to *M. tuberculo‐*

Normally iron taken up by intestinal epithelial cells and bound to transferrin circulates within the body. This complex binds to cell surface receptors, and is internalized where it releases its

oxidative damage (Darwin and Nathan, 2005; Colangeli et al, 2009).

and has demonstrated chaperonin activity (Pang and Howard, 2007).

*sis,* and are potent stimulators of an inflammatory response (Anand et al, 2010).

tional phase (Garge et al, 2009).

8 Tuberculosis - Current Issues in Diagnosis and Management

**8. Heat shock**

**9. Low iron**

Mycobacteria have a variety of systems which aid in the uptake of iron and the regulation of iron responsive genes. As mycobacteria have been shown to be somewhat novel among gram positive bacteria, they possess an outer mycolic acid based membrane, as well as an inner membrane and periplasmic space. Porins in the outer membrane appear to transport iron in the presence of high iron conditions (Jones and Niederweis, 2010). *M. tuberculosis* under low iron conditions can produce the siderophore carboxymycobactin as well as my‐ cobactin (Table 1.) (Banerjee et al, 2011). These molecules bind with a higher affinity to iron than the human host's storage proteins and steal iron from the host. Mycobactin is present within the inner membrane and thus can only bind iron imported into the periplasmic space. Interestingly lipid membranes with associated mycobactins may diffuse out, travel to lipid vesicles in the host cell, and sequester iron. These structures may recycle back to in‐ teract with the mycobacterium. Disruption of the genes responsible for production of my‐ cobactins can cause these mutant mycobacteria to replicate less well in macrophages (Banerjee et al, 2011). Carboxymycobactins are excreted possibly by the type VII secretion or ESX system. Externally the carboxymycobactins bind available iron from transferrin (Banerjee et al, 2011). Porins and also ABC transporters may allow import of these iron loaded carboxymycobactins (Banerjee et al, 2011). The host cell, in response to infection and inflammation, produces siderocalins such as lipocalin-2 that can bind to and inactivate my‐ cobactin from *M. tuberculosis* thus interfering with mycobacterial iron acquisition (Johnson and Wessingling-Resnick, 2012). In fact mice deleted for genes involved in production of siderocalin are much more susceptible to mortality due to *M. tuberculosis* infection (Johnson and Wessingling-Resnick, 2012). Inside the mycobacterial cell, iron is stored in bacterioferri‐ tin and a ferritin like protein. These proteins are required for replication in human macro‐ phages and guinea pigs, act to store iron, and also to limit excess iron in the cells that can lead to iron mediated oxidative damage due to the Fenton reaction (Reddy et al, 2011).

Iron responsive genes in *M. tuberculosis* are controlled in part by the iron dependent regulator IdeR. This protein can act both as an activator and a repressor depending on where it binds within a mycobacterial promoter region (Manabe et al, 1999; Banerjee et al, 2011). Within promoters of genes involved in mycobactin synthesis it acts as a repressor, inhibiting expres‐ sion of these genes at high iron concentrations. In promoters of iron storage proteins it acts as an activator, stimulating expression of these genes at high iron concentrations and thus avoiding iron stimulated oxidative damage.

In Vivo Condition or Location Mycobacterial Response

Two components systems are common in many bacteria. These systems are comprised of a sensor kinase which phosphorylates the response regulator as a result of an environmental signal, which is often a stress. The sensor kinases are trans membrane proteins which are embedded into membranes. They sense external stresses and transmit these signals internally into the bacterial cell by phosphorylating a response regulator that binds to its cognate promoter DNA, and regulates transcription. The mycobacterial genome contains 11 two component systems (Hett and Rubin, 2008). The large number of these systems in the myco‐ bacterial coding regions is likely the result of evolution to accommodate bacterial responses

macrophages, granulomas, liquified

macrophages, granulomas, low

all stress conditions, macrophages,

lesions and sputum

granulomas,sputum

liquified lesions,sputum, conditions leading to dormancy

low oxygen, macrophages, conditions leading to dormancy

low oxygen, macrophages, possibly acidity

In all conditions *in vivo*

Fever

hypoxia

**12. Two component systems**

to diverse stresses.

oxidative stress, macrophages

macrophages, phagocytosis,

**Table 1.** Mycobacterial responses to in vivo stressors and conditions.

iron

increased lipid metabolism in bacillus, or induction of same in host

*Mycobacterium tuberculosis* Adaptation to Survival in a Human Host

http://dx.doi.org/10.5772/54956

11

Siderophore production

lipid body production

differential sigma factor utilization

DosR two component system activity

PhoP two component system activity Constitutive thick waxy cell wall construction, may be upregulated Mycothiol, SOD, & KatG production Heat shock protein production toxin-antitoxin system function
