Antibiotic Resistance of Tuberculosis

Tuberculosis (TB) has affected human beings since Neolithic times (1).  In ancient Greece it was known as phthisis, which means “wasting.” During the 17th and 18th centuries in Europe it caused the “White Plague” and was known as consumption, accounting for 25 percent of all adult deaths during this period (2).  These two names reflect the slow deteriorative progression of the disease in the host.  It is the long time scale of the infection period that makes tuberculosis so dangerous.

Tuberculosis has become a threat again in the modern era of antimicrobial warfare, because its unique characteristics give it enormous potential for developing resistance to even the strongest antibiotics.  Tuberculosis combines one of the slowest division rates among bacteria with a hardy cell wall defense system (2).  Both of these factors stretch treatment into a multiple month process, creating a massive window for human error in the form of incorrect or missed dosages (1).  Similarly, this slow pace of infection increases the possibility of evolution-based antimicrobial resistance by giving tuberculosis bacteria time to mutate (2).  For these reasons, multi-drug resistant tuberculosis is one of the biggest health threats of this generation and the discovery of new treatment methods will be essential in the ongoing fight against this disease.

Tuberculosis
Tuberculosis is normally a chronic, widely variable disease that is usually caused by inhalation of the airborne causative agent (3).  The symptoms of TB include fever, cough, difficulty breathing, inflammatory infiltrations, formation of tubercles, caseation, pleural effusion and fibrosis (3).
TB is caused by a mycobacterium.  Mycobacterium tuberculosis (MTB) is the most common bacterial agent responsible for TB, however, M. bovis, M. microti canetti, and M. africanum can also result in TB.
MTB is an obligate aerobic bacterium, therefore it needs oxygen to survive (4).  Because of MTB’s large metabolic oxygen requirement, it is typically found in oxygen rich areas such as the respiratory system.  Although it generally starts in the lungs, it may spread to other regions of the body via the lymphatic system or blood vessels (3).

The Cell Wall
The cell wall of MTB is one of the major determining factors of its virulence (2). The structure of the cell wall has three major components: mycolic acids, cord factors, and Wax-D.  The mycolic acid molecules are of primary interest due to their deadly qualities.

Mycolic acids are unique to Mycobacterium and Corynebacterium (2).  They are alpha branched saturated fatty acids with chain lengths as long as 80 carbons (5).  Mycolic acids create a lipid shield, which protects against cationic proteins, lysozyme, and oxygen radicals of phagocytosis (2).

Gram staining is a means of categorizing bacteria based on the makeup of their cell walls.  The amount of the polymer peptidoglycan, which forms a mesh-like structure, determines whether or not the bacterium can hold onto the stain administered in gram staining. MTB cannot be classified as truly gram positive or negative, because its cell wall is impervious to gram staining due to the high content of lipids, especially mycolic acid.  This characteristic places MTB in a category of bacterium known as acid-fast bacteria, whose acid-rich cell walls retain a red dye used for staining, despite attempts at de-colorization (1).

The full genome of MTB was sequenced in 1998.  One of the major discoveries was that a large amount of coding is dedicated to the genesis and lysis of lipids, compared to other bacteria. This explained why over 60 percent of the MTB cell wall is composed of lipids (2).  This propensity for lipid production creates the unique mycobacterium cell wall, which is one of the most important characteristics of MTB as it is highly conducive to antibacterial resistance formation.

Pathology
The cell wall of MTB gives it many pathogenic properties.  For instance, MTB is a facultative intracellular parasite, which means that it can reproduce inside or outside of host cells.  As a result, MTB is able to survive within immune cells known as macrophages without being destroyed by phagocytosis (6).  When infectious particles reach the alveoli sacs in the lungs, macrophages phagocytose (engulf) the bacteria and clump together into granulomas in order to contain the infection.  Although this process keeps 95 percent of TB infections from becoming activated upon bacterial entry, MTB is able to remain dormant for many years, thanks to its cell wall which provides resistance to lethal oxidation (1, 2).

Once captive within immune cells, MTB is transferred to the lymph system and bloodstream (1).  Through this process MTB is able to spread to other organs and multiply in oxygen rich regions (1).  Despite the possibility of extra-pulmonary TB, the majority of cases occur in the upper lungs following reactivation of dormant MTB (1).

When MTB becomes active, its cell wall plays a major role in replication and resistance to immune responses.  After the initial infection, MTB can reproduce within the macrophage until the cell bursts, which alerts macrophages from peripheral blood (2).  However, the process continues because the newly produced MTB cannot be fully destroyed by the macrophages.

As MTB replicates further, T-cell lymphocytes are activated.  These immune cells are activated by major histocompatibility complex molecules, which allow the T-cells to recognize MTB antigens (2, 7).  At this point cytokines are released, which activate the macrophages and allow them to destroy MTB (2).  This activation is a cell-mediated immune response, as opposed to the original antibody-mediated immune response (2).

At this phase in the disease, small rounded nodules known as tubercles  form and create an environment in which MTB is unable to multiply.  However, because of its cell wall, MTB can survive in the low pH and anoxic tubercles for long periods of time (2).  These tubercles are surrounded by many inactivated macrophages in which MTB is able to replicate (2). Through this process, although the cell-mediated immune response is capable of destroying individual bacterium, it is also responsible for the growth of tubercles, which occurs as MTB replicates within and subsequently ruptures inactivated macrophages (2).

In these ways, the cell wall of MTB allows it to evade or complicate each step of the immune process, creating a need for man-made antibiotics.

Antimicrobial Resistance

The waxy, hydrophobic cell wall of MTB gives it the ability to survive long exposure to substances such as acids, detergents, oxidative bursts, and antibiotics. In fact, the typical “short” treatment of MTB involves a four antibiotic treatment for two months and then a two antibiotic treatment for an additional four months. The antibiotics involved are isoniazid, rifampicin, pyrazinamide, and ethambutol.

The cell wall of MTB is so resistant to normal antibiotic measures that the above listed antibiotics, especially isoniazid, are targeted at the synthesis of mycolic acids (8). The inhibition of the gene InhA has been found to induce the lysis of MTB cells (8). However, some MTB strands have mutated and begun to augment the mycolic acids of their cell walls with cycloprophyl groups (8). Although these groups have been shown to hamper persistent infection, they also protect MTB against immune responses (8).

This is simply one example of the adaptive ability of MTB. The long period of treatment, combined with a laundry list of side effects, which can include hepatitis, optic neuritis, and seizures, compels many patients to stop taking medication after symptoms subside (1, 6). The standard therapy for active TB is a six-month program with two months dedicated to isoniazid, rifampin, and pyrazinamide and four months of isoniazid, rifamate, and rimactane (1, 6). Ethambutol or streptomycin is also added until the patient’s drug sensitivity is known (6).  This long period of treatment is a direct result of the slow reproductive time and resistant cell wall of MTB (1, 2).  Both of these factors give MTB ample time to capitalize on patient or doctor error by producing mutations like the addition of cycloprophyl mentioned above.

Two types of drug resistant MTB strains are currently recognized.  Multi-drug-resistant tuberculosis (MDR TB) is resistant to at least two of the four first-line drugs listed above (2).  Extensively drug resistant tuberculosis (XDR TB) is defined as resistant to isoniazid, rifampin and also to fluoroquinolone and at least one of three injectable second-line drugs (2).  XDR TB has an estimated cure rate of only 30 percent in patients with an uncompromised immune system compared to a 95 percent cure rate of normal tuberculosis (2, 9).

The four-drug regimen of tuberculosis treatment is a means of avoiding MDR TB and XDR TB.  In 2008, the World Health Organization (WHO) indicated that MDR TB was at a record high of 489 cases, compared to 139 cases in 2006, and that XDR TB had been reported in 45 countries (2).  These findings reflect a pressing need not only for greater adherence to prescribed drug treatments, but also for the discovery of new antibiotics or other means of combating this rapidly growing problem.

The WHO has implemented a new system in response to its drug-resistant tuberculosis findings, which is known as directly observed treatment, short-course (DOTS) (10).  This approach facilitates cooperation between doctors, health workers, and primary health care agencies in order to monitor tuberculosis patients and facilitate the complete eradication of infection (10).

Conclusion
As demonstrated, even in its non-resistant form, MTB provides a massive challenge for the immune system through its unique cellular properties.  It is capable of infiltrating the body’s own immune cells, of surviving for weeks outside of the body, and for resisting most standard antibiotics.  When coupled with human error, tuberculosis proves a deadly adversary.  As demonstrated by the findings of the WHO, MDR TB is a global concern, which needs to be closely monitored in the future.  Similarly, although still rare, XDR TB threatens the medical landscape of this generation and needs to be met with patient compliance and cooperative action on the part of doctors and health agencies.

References
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4. Aerobic (2005).  Available at http://www2.merriam-webster.com/cgi-bin/mwmednlm (19 November 2008).
5. J. Lackie, The Dictionary of Cell and Molecular Biology (2008). Available at http://cancerweb.ncl.ac.uk/cgi-bin/omd?mycolic+acid (19 November 2008).
6. S. Swierzewski, Tuberculosis (2007). Available at http://www.pulmonologychannel.com/tuberculosis/index.shtml (29 November 2008).
7. MeSH Descriptor Data (2008). Available at http://www.nlm.nih.gov/cgi/mesh/2008/MB_cgi?mode=&term=Major+Histocompatibility+Comple (19 November 2008).
8. W. Jacobs, Mycolic Acids of Mycobacterium tuberculosis: An Achilles Heel or a Neutralizing Weapon? (2001). Available at http://www.rockefeller.edu/lectures/jacobs011901.html (29 November 2008).
9. Extensively Drug Resistant Tuberculosis (2007). Available at http://www.medicinenet.com/extensively_drug-resistant_tuberculosis_xdr_tb/article.htm (29 November 2008).
10. Tuberculosis: An Airborne Disease (2004). Available at http://findarticles.com/p/articles/mi_m1309/is_/ai_54157859 (29 November 2008).