This ppt focusses on basics about mycobacterium tuberculosis, especially helpful for MBBS and PG medical students.

1. Mycobacterium tuberculosis
Dr Shilpi Gupta
MD (Microbiology)

2. MYCOBACTERIA 2/2
M. leprae & Atypical
Professor Sudheer Kher

4. • Mycobacterium tuberculosis
• One of the most serious infectious diseases in the
developing world
• Thirty million people have active disease
• Nine million new cases occur
• Three million people die of the disease, each year
• Problems
– association with AIDS
– Multiple drug-resistance
– Chronic disease
– Prolonged treatment
Tuberculosis (Koch’s Disease)

5. General characters
• Slender rods, long, filamentous, branching
• Resist staining but once stained, resist
decolorization by dilute mineral acids; hence
called ACID FAST BACILLI (AFB) due to
presence of mycolic acids in cell wall
• Aerobic, Non-motile, Non-sporing, Non-
capsulated.
• Growth generally slow

6. Resistance
• Not specifically resistant to heat, 60°C for 20 min
destroys.
• In sputum can survive 20-30 hrs and in droplet nuclei
up to 8-10 days
• Relatively resistant to disinfectants.
• Survives exposure to
– 5 % Phenol
– 15 % Sulphuric acid
– 3 % Nitric acid
– 5 % Oxalic acid
– 4 % NaOH
Killed by formaldehyde, glutaraldehyde and ethanol

7. Modes of infection
1. Droplet infection
Person to person by inhalation aerosols
Mycobacterium tuberculosis (Pulmonary tuberculosis)
2. Ingestion of milk
Infected cattle
Mycobacterium bovis (Intestinal tuberculosis)
3. Contamination of abrasion
Laboratory workers (Skin infection)

9. Pathogenesis
• Infects lung
• Distributed within macrophages
• Facultative intracellular pathogen
– inhibits phagosome-lysosome fusion
– resists lysosomal enzymes

10. Cell-mediated immunity -tuberculosis
• Infiltration
– macrophages
– lymphocytes
• Granulomas

11. Classical tubercular lesion – Granuloma with typical
Langhan’s giant cells, epithelioid cells, lymphocytes and
necrosis.

17. Clinical Features
* Low grade fever
* Weight loss
* Night sweats
* Fatigue
* Cough & haemoptysis

19. Diagnosis
• Demonstrating bacilli by microscopy
• Isolating the bacilli in culture
• Biochemical identification
• Demonstrating hypersensitivity to
tuberculoprotein (Tuberculin test)
• Molecular methods

20. There are multiple light areas (opacities) of varying size that run
together (coalesce). Arrows indicate the location of cavities within
these light areas. The appearance is typical for chronic pulmonary
tuberculosis.

21. Specimen
• Pulmonary
– Sputum
– Bronchial washings or
laryngeal
– In children gastric lavage
• Extra-pulmonary
– CSF
– Joint fluid
– Biopsy material
(endometrial)
– Urine
– Blood or any other body
fluid
* Specimens need appropriate processing
Liquefaction with N-acetyl-L- cysteine
Sputum Decontamination with NaOH
Centrifugation

22. 1. Microscopy
• Ziehl-Neelsen Stain (ZN stain)
– Primary stain – Carbol fuchsin
– Mordant – gentle intermittent heating
– Decolourisation – 20% H₂S0₄ or 3% acid 95% alcohol
– Secondary stain – Malachite green/ Methylene blue
• Kinyoun’s modification
– Modified cold method
• Auramine rhodamine

24. 2. Culture

25. Lowenstein Jensen Medium –
Selective. Always in screw capped bottle. Bluish Green.
Contains – Egg protein – Solidifying agent
Mineral salts – Mg sulphate, Mg citrate
Asparagine
Malachite Green – Selective agent
Sterilized by - Inspissation

29. 3. Biochemical methods
• M.tuberculosis – positive biochemical tests
– Niacin test
– Peroxidase test
– Nitrate reduction test
– Pyrazinamidase test

30. 4. Molecular methods
• Real Time PCR
• Polymerase chain reaction (PCR)
• Transcription mediated amplification (TMA)
• Line probe assay (LPA)
– Rapid 3-4 hour
– Expensive

37. 5. Tuberculin testing (Mantoux)
• Delayed hypersensitivity skin test to assay cell
mediated immunity to tubercle bacillus
• Material: A purified protein derivative (PPD)
• Dose : 0.1 ml of (PPD) is injected intradermal
• Reading : Positive test is defined as
- Induration equal or greater than 10 mm
- Develop 48-72 hours after injection

42. Treatment
Use multiple drug therapy to prevent emergence of resistant
mutants
* Long duration treatment (6-18 months)
* Four drugs are usually started in initial therapy due to:
- Intracellular location of bacilli
- Slow growth rate of bacilli
- Caseous material blocks penetration of drugs
- Some bacilli persist in a metabolically inactive state
* Sputum becomes non-infective 2-3 weeks after starting
therapy

43. Treatment
Drugs used :
1- First line drugs :
- Isoniazid - Rifampicin - Pyrazinamide
- Ethambutol - Streptomycin
2- Second line drugs (more toxic and less effective):
- Kanamycin - capreomycin - Cycloserin
- ethionamide - ciprofloxacin - Ofloxacin
* Noncompliance (failure to complete the course):
Directly observed therapy (DOT)
Health care workers observe the medication

44. Immuno-prophylaxis
• Intradermal injection of live attenuated
vaccine Bacille Calmette-Guerin (BCG)
• Given at birth
• Immunity lasts for 10-15 years. Immunity 60-
80%
• Protects against TB, the disease runs milder
course in protected, prevents skeletal,
meningeal & miliary forms

45. THANK YOU