1. MICR 454L Emerging and Re-Emerging Infectious Diseases Lecture 7:
M. tuberculosis
Dr. Nancy McQueen & Dr. Edith Porter

2. Overview M. tuberculosis Morphology
Growth and metabolic characteristics
Virulence factors
Diseases
Diagnosis
Culture
PCR
Immune response
Therapy
Threats

3. M. tuberculosis Acid fast rods Lipid-rich cell wall
Mycolic acids
Lowenstein Jensen agar
Eggs
Potatoes
Malachite green
Slow growth
Up to 6 weeks

4. M. tuberculosis: Virulence Factors
Lipid-rich cell wall
Mycolic acids
Resistant to host defense
Intracellular survival in macrophages
Iron capturing ability
Sulfolipids prevent phagsome-lysosome fusion
Eventually escape into the cytoplasm
Requires a T-cell mediated immune response for infection control/eradication
Granuloma formation

5. Removal of M. tuberculosis Depends on the Cell-Mediated Arm of Host Defense

8. The Course of TB Infection and Disease
Airborne Infection
90 %
10 %
Latent TB
TB Disease
No symptoms
Not sick
Cannot spread disease
Chest X Ray and sputum are normal
Symptoms
Can sp[read infection
Positive skin test
Possible abnormal chest X ray
Positive sputum smear or culture
Dissemination
AIDS increases susceptibility
Untreated:
Severe illness,
Death
Reactivation
(secondary) TB

9. M. tuberculosis: Diseases
General Symptoms
Feelings of sickness or weakness
Weight loss
Fever
Night sweats
Lung tuberculosis
Coughing
Chest pain
Hemoptysis
Extrapulmonary
Depends on localization

10. Lung Tuberculosis

11. Extrapulmonary Tuberculosis

12. M. tuberculosis: Diagnosis
History
Physical exam
Mantoux Skin test (tuberculin test with purified protein derivative)
QuantiFERON®-TB Gold Test
Chest radiograph
Sputum smear
Culture

13. Transmission of Tuberculosis
Inhalation of aerosols containing mycobacteria
Bacteria can float in the air for several hours
Ingestions (M. bovis)
According to CDC, TB is not spread by
Handshaking
Sharing food or a drink\touching bed lines or toilet seats
Sharing tooth brushes
Kissing

14. Principle of the Tuberculin Test

15. QuantiFERON®-TB Gold Test
Measure IFNg production by patient peripheral blood leukocytes in response to M. tuberculosis antigens (protein antigens ESAT-6 and CFP-10)
Rapid T cell response only in primed individuals

16. Interpretation of Tuberculin and QuantiFERON
Positive = previous contact with M. tuberculosis
Positive DOES NOT mean TB disease

17. M. tuberculosis: Therapy
Isoniazid (INH)
Rifampin (RIF)
Ethambutol
Pyrazinamide
DOTS
Direct observational therapy short course
At least 2 in combination (INH + RIF)
Prolonged time (at least 6 months)

18. Anti-Tuberculosis Drug Targets
Mycolic acid
INH inhibits mycolic acid synthesis
Ethambutol inhibits mycolic acid incorporation into the cell wall
Fatty acid synthetase I (FASI)
Pyrazinamide inhibits fatty acid synthesis
RNA synthesis
Inhibited by rifampin

19. Resistance of M. tuberculosis
Mutations in codon 306 of the embB gene (ethambutol) are discussed as marker and predictor of resistance development to multiple antibiotics
Not all mutated strains are resistant but resistant strains are mutated.
Alterations in RNA polymerase (Rifampicin)

20. Worldwide Threats by M. tuberculosis
Large fraction of the world population is infected
1/3 of world population is infected ~ (1.7 billion people)
Each year ~ 9 million new cases world wide
5 – 10 % will develop active tuberculosis (TB)
Worldwide almost 2 million deaths from TB
TB is the leading killer of people who are HIV infected
Even though TB has is declining in the US, the decline of the average annual percentage rate is slowing down
From 1993 – 2000 ~ 6.6% decline /year
From 2003 – 2008 only ~ 3.4% decline/year In the US decline of cases
In ,904 TB cases reported

21. Worldwide Threats by M. tuberculosis
Multidrug resistant TB
Case rates have also declined (407 cases in 1993 and 86 cases in 2008)
Result from sequential mutations
Extremely drug-resistant TB
First reported in 2005
3 cases reported in the US in 2006
Increase in Europe observed
7.3% of all MDR strains

22. Extremely Drug-Resistant M. tuberculosis
XDR TB
Resistant to almost all drugs used to treat TB, including the two best first-line drugs: isoniazid and rifampin
Resistant to the best second-line medications: fluoroquinolones (DNA gyrase mutations)
And at least one of three injectable drugs (i.e., amikacin, kanamycin, or capreomycin; mutations in 16sRNA and ribosomal protein genes).
Possibly involvement of drug efflux pumps.

23. Reported Cases of XDR in the US
MMWR Weekly, March 23rd, 2007

24. New Drugs are Needed Immune modulators
IL-2, IFN-gamma, GM-CSF, IL-12
New chemicals targeting essential genes of M. tuberculosis

25. Take Home Message
One third of the world population is infected with M. tuberculosis but only 10% develop active disease.
The lipid rich cell wall and slow growth contribute to resistance to host defense and difficulties in antibiotic treatment.
The emergence of extremely drug resistant tuberculosis strains poses a great threat to the public.

26. Additional Resources
The Microbial Challenge, by Krasner, ASM Press, Washington DC, 2002.
Brock Biology of Microorganisms, by Madigan and Martinko, Pearson Prentice Hall, Upper Saddle River, NJ, 11th ed, 2006.
Immunobiology, by Janeway,, Travers, Walport, and Shlomchik, Garland Science, 6th ed, 2005.
Malak Kotb Genetics of Susceptibility to Infectious Diseases Volume 70, Number 10, 2004 / ASM News Y
htttp://
Zager and McNerney (2008) Multidrug-resistant tuberculosis. BMC Infectious Disease. 8: 10.
Safi H, Sayers B, Hazbón MH, Alland D. (2008) Antimicrob Agents Chemother. Mar 31 [Epub ahead of print] Transfer of embB306 mutations into clinical Mycobacterium tuberculosis alters susceptibility to ethambutol, isoniazid and rifampin.
Zimhony O et al. (2000) Pyrazinamide inhibits the eukaryotic-like fatty acid synthetase I (FASI) of Mycobacterium tuberculosis. Nat Med. Sep;6(9):
(accessed )