1. A 32-year-old woman presented with a cough for several weeks and a 15-lb weight loss. She also had night sweats and fevers and felt fatigued. Despite erythromycin treatment for suspected pneumonia given by her family physician, her fever and cough got progressively worse. She complained about coughing blood-tinged sputum. She had emigrated from Venezuela to the United States 3 years before her illness, but she frequently returned to Venezuela to visit relatives.

2. Physical examination Temperature: 38.6˚C Pulse: 96/min Respiration: 18/min Blood pressure: 112/60 mm Hg Examination was remarkable for bilateral rales and lymphadenopathy Chest X-ray revealed right upper lobe infiltrates

3. Likely causes of illness Actinomyces spp. Anaerobes (aspiration pneumonia) Endemic fungi (Histoplasma capsulatum) Legionella pneumophila Mycobacterium tuberculosis Mycoplasma pneumoniae Nocardia spp.

4. MYCOBACTERIA The captain of all the men of death that came against him to take him away, was the Consumption, for it was that that brought him down to his grave John Bunyan (1628-1688) English author

5. MYCOBACTERIA Died of tuberculosis: Chopin Paganini Thoreau Keats Elizabeth Barrett Browning Emily and Charlotte Bronte

6. MYCOBACTERIA In the 19th Century, one quarter of the population of Europe is thought to have died of tuberculosis

9. MYCOBACTERIA 1.9 billion people infected with Mycobacterium tuberculosis Tuberculosis kills about 2 million people and infects about 8 million others every year Risk factors: poverty, malnutrition and poor housing

11. MYCOBACTERIA May proceed to a generalized infection (“miliary” Tuberculosis) Transmission electron micrograph of M. tuberculosis Primarily a disease of the lungs

12. Mycobacteria stained with the Ziehl-Neelsen stain Acid-fast staining “Acid-fast bacteria”

13. M. tuberculosis colonies on Lowenstein-Jensen agar

14. MYCOBACTERIA Physiology and structure Complex cell wall Peptidoglycan layer linked with arabinose- galactose-mycolic acid (arabinogalactan mycolate) Mycolic acids Major lipids Long chain (C 78 -C 90 )

15. Mycobacterial cell wall structure

16. MYCOBACTERIA Physiology and structure Free lipids Waxes: wax D (Freund's adjuvant) Helps withstand drying and thus increases survival Mycosides: complex saturated glycolipids Cord factor: 6,6'-dimycolate of trehalose Inhibits neutrophil migration Thought to mediate granuloma

17. Trehalose 6,6’-dimycolate RCO: mycolyl

18. MYCOBACTERIA Physiology and structure Polypeptides Act as antigens Used in skin testing (PPD)

20. MYCOBACTERIA Physiology and structure Runyon classification according to rate of growth and pigmentation Photochromogens: Organisms that produce pigments after exposure to light Scotochromogens: Produce pigments both in the light and dark

21. Photochromogenic M. kansasi colonies on Middlebrook agar

22. Pathogenesis Infection via inhalation of infectious aerosols Replicates in alveolar macrophages and destroys the cells Infected macrophages migrate to the local (tracheobronchial) lymph nodes, the bloodstream and other tissues Mycobacterium tuberculosis

23. Tubercle bacilli enter through the respiratory tract

24. Stealth invader: MTB in macrophages

25. Tubercle bacilli multiply in phagocytes and spread to lymph nodes and the circulation

26. Pathogenesis No exotoxin or endotoxin ”Exported repetitive protein" prevents the phagosome from fusing with the lysosome Mycobacterium tuberculosis

27. Pathogenesis Two types of lesions Exudative: Acute inflammatory response at the initial site of infection (usually lungs) Granulomatous: A central area of infected giant cells (Langhans’cells) surrounded by epithelioid cells (tubercles) Mycobacterium tuberculosis

28. Pathogenesis Tubercles May heal spontaneously May become fibrotic or calcified Caseous (cheesy) necrosis and cavitation in the center of the granuloma, due to: Cellular immune response Enzymes and reactive oxygen intermediates from dying macrophages Mycobacterium tuberculosis

29. Pathogenesis The exudative lesion and the draining lymph nodes fibrose, and sometimes calcify, to produce the Ghon complex seen in X-rays Latent bacilli can be re-activated when the patient's immune system is weakened (malnutrition, alcoholism, diabetes, old age, emotional stress) Mycobacterium tuberculosis

30. Pulmonary tuberculosis

32. MTB can cause devastating damage to the lungs

33. Pathogenesis and immunity Intracellular replication stimulates helper and cytotoxic lymphocytes T-cells release interferon-  and other cytokines -> Macrophage activation -> MTB killing Tuberculin skin test: purified protein derivative (PPD) Positive 4-6 weeks after infection Mycobacterium tuberculosis

34. Pathogenesis and immunity Localized activated macrophages can penetrate into small granulomas (< 3 mm) and kill all bacilli Larger necrotic or caseous granulomas become encapsulated with fibrin that protects the bacilli Mycobacterium tuberculosis

35. Pathogenesis and immunity The pathogen population does not increase, but the immune system cannot get rid of the bacteria It is not known how MTB can sit around in tissues for years or decades MTB eats carbon from lipids via the glyoxylate pathway Isocitrate lyase critical for this pathway Mycobacterium tuberculosis

36. Epidemiology Transmitted by aerosols More common among The urban poor Patients with suppressed immune systems Immigrants from high-incidence areas Health care workers are at risk for infection! Mycobacterium tuberculosis

37. Annual incidence of MTB in the US 1981-1995 In 2005: 14,093 cases 4.8 per 100,000 California: 2,900 cases 8 per 100,000

38. Epidemiology An estimated 10-15 million people in the US are infected with M. tuberculosis without displaying symptoms 1 in 10 of these individuals will develop active TB at some time in their lives Mycobacterium tuberculosis

39. Epidemiology What caused TB to return? HIV/AIDS epidemic Immigration from countries with many TB cases Increased poverty, injection drug use, homelessness Failure to take antibiotics as prescribed Increased number of people in long-term care Mycobacterium tuberculosis

40. Clinical syndromes Primary tuberculosis mild and asymptomatic in 90% of cases does not proceed further In the remaining 10% malaise, weight loss, productive cough, night sweats Bloody and purulent sputum Mycobacterium tuberculosis

41. Clinical syndromes Reactivation of latent bacilli years later when the patient's immunologic responsiveness wanes: Old age Immunosuppressive disease or therapy Malnutrition Alcoholism Diabetes Stress Mycobacterium tuberculosis

42. Clinical syndromes Necrosis may erode blood vessels -> hemorrhage Most infections in the lungs Can disseminate to the lymph nodes, pleura, the liver, genitourinary tract, bone marrow, spleen, kidneys, central nervous system M. tuberculosis can also cause meningitis Mycobacterium tuberculosis

43. Acute miliary tuberculosis leading to meningitis Latin: milium, millet seed Each tubercle resembles a millet seed

44. Laboratory diagnosis Skin test: "purified protein derivative" injected into the intradermal layer of the skin. Induration of 10 mm or more 48 hours later indicates exposure to MTB Microscopy: Clinical specimen stained with carbolfuchsin (Ziehl-Neelsen and Kinyoun stains) or fluorochrome dyes (Truant auramine-rhodamine), de- colorized with acid-alcohol solution, and counterstained Mycobacterium tuberculosis

45. Acid-fast stain (carbolfuchsin) of Mycobacterium tuberculosis M. tuberculosis stained with the fluorescent dyes auramine and rhodamine

46. Laboratory diagnosis Culture: After treatment of specimen with 2% NaOH (to kill rapidly growing bacteria) MTB is grown on Lowenstein-Jensen (egg-based) medium, or Middlebrook (agar-based) medium BACTEC measures the metabolism of 14 C-palmitic acid to 14 CO 2 Mycobacterium tuberculosis

47. Laboratory diagnosis Biochemical tests: The production of niacin, and nitrate reductase. Analysis of cell wall lipids Nucleic acid probes and nucleic acid sequencing PCR Mycobacterium tuberculosis

48. Treatment and control Isoniazid: inhibits mycolic acid biosynthesis (used for prophylaxis) Rifampin Pyrazinamide (active at low pH; mechanism unknown) Ethambutol (inhibits arabinogalactan synthesis) American Thoracic Society recommendations: Initial treatment with these 4 drugs for 2 months, followed by INH/RIF for 4-6 months Mycobacterium tuberculosis

49. Treatment and control Multiple-drug-resistant (MDR) strains Mutations in genes for mycolic acid synthesis catalase-peroxidase required to activate isoniazid Need in vitro susceptibility testing Mycobacterium tuberculosis

50. Treatment and control Noncompliance -> emergence of MDR strains Directly observed therapy (DOT) BCG: Bacille Calmette-Guérin (attenuated M. bovis) Used in most developing countries to reduce the severe consequences in infants and children Variable efficacy in adults Mycobacterium tuberculosis

51. Treatment and control PA-824 New drug in clinical trials (“nitroimidazopyran”) Active against both actively dividing and slow growing MTB Mycobacterium tuberculosis

52. Pathogenesis Asymptomatic colonization: immunocompetent patients Localized pulmonary disease: patients with chronic bronchitis Disseminated disease: immunocompromised individuals Organ dysfunction due to abundance of bacteria and host response to infection Mycobacterium avium-intracellulare

53. M. avium-intracellulare infected tissue from an AIDS patient

54. M. avium-intracellulare in fixed-tissue macrophages in a bone marrow biopsy from a patient with AIDS (anti-mycobacterial Ab immunoperoxidase stain)

55. Epidemiology Ingestion of contaminated food or water Found in soil and water, and infected poultry and swine Inhalation of infectious aerosols: minor role Immunocompromised patients (esp. AIDS) and those with long-standing pulmonary disease at greatest risk Mycobacterium avium-intracellulare

56. Clinical syndromes The most common mycobacterial isolate in AIDS patients Disseminated disease: Tissues filled with MAC Bacteremia: 100-1000/ml in blood Mycobacterium avium-intracellulare

57. Treatment and control Resistant to most antimycobacterial drugs Combination therapy Clarithromycin (or azithromycin) + Ethambutol + Rifabutin For prophylaxis in AIDS patients with low CD4 counts: Clarithromycin or Azithromycin Mycobacterium avium-intracellulare

58. Pathogenesis Causes leprosy or Hansen's disease Tuberculoid leprosy Strong cellular immune reaction Lymphocytes and granulomas, but relatively few bacteria Production of interferon-  and IL-2 -> macrophage activation, phagocytosis and bacterial clearance Mycobacterium leprae

59. A. Tuberculoid leprosy Macules with hypo- pigmentation B. Lepromatous leprosy with extensive infiltration, edema and corrugation of the face

60. Pathogenesis Lepromatous leprosy Specific defect in the cellular immune response to M. leprae antigens Bacilli populate dermal macrophages and Schwann cells of peripheral nerves The most infectious form of leprosy Mycobacterium leprae

61. Tuberculoid leprosy Lepromatous leprosy

62. Ziehl-Neelsen staining of Mycobacterium leprae

63. Clinical syndromes Tuberculoid Lepromatous leprosy Most infectious, with large numbers of bacilli in infected tissues Causes disfigurations in the skin Mycobacterium leprae

64. Epidemiology > 12 million cases of leprosy worldwide 250 cases annually in the U.S. ( California, Texas, Louisiana and Hawaii), mostly in immigrants Spread by person-to-person contact Inhalation of infectious aerosols, skin contact with respiratory secretions or wound exudates. Arthropod vectors? Armadillos? Mycobacterium leprae

65. Armadillos and M. leprae?

66. Treatment and control Tuberculoid leprosy Dapsone (antifolate) + Rifampin (min 6 months) Lepromatous leprosy Clofazimine in addition to Dapsone and Rifampin (therapy extended to 12 months) Controlled by prompt recognition and treatment of infected people Mycobacterium leprae

67. Mycobacterium kansasii Causes chronic pulmonary disease Disseminated disease in immunocompromised patients

68. Mycobacterium marinum Causes "swimming pool granulomas” nodular lesions along the lymphatics -> ulceration

69. Fish tank granuloma caused by Mycobacterium marinum infection of a lesion acquired while cleaning out a fish tank

70. NOCARDIA

71. Strict aerobic, Gram-positive bacilli Form branched hyphae The cell wall contains mycolic acids "acid-fast” (in contrast to Actinomyces) Nocardia

72. Nocardia asteroides causes bronchopulmonary disease that can spread to the skin and CNS Nocardia brasiliensis causes cutaneous disease Nocardia

73. Acid-fast stain of Nocardia asteroides in expectorated sputum

74. Gram stain of Nocardia asteroides in expectorated sputum

75. Pathogenesis Found worldwide in organic-rich soil Colonizes the oropharynx Aspiration of oral secretions causes infection of the lower airways Nocardia

76. Pathogenesis Causes necrosis and abscess formation Can survive in phagocytes Traumatic introduction into subcutaneous tissue > Cutaneous nocardiosis Nocardia

77. Epidemiology Infections are exogenous (not caused by normal human flora) Nocardia asteroides infections occur in patients with impaired cell-mediated immunity Nocardia

78. Epidemiology Immunocompetent patients with chronic pulmonary disease (bronchitis, emphysema) Nocardia brasiliensis infections of the skin can occur in immunocompetent persons Nocardia

79. Mycetoma caused by Nocardia brasiliensis The foot is grossly enlarged and covered with multiple draining sinus tracts

80. Clinical syndromes Bronchopulmonary infections: Cough, dyspnea (difficulty in breathing), fever Cutaneous infections: Cellulitis (inflammation of the soft or connective tissue), pustula, pyoderma (purulent skin disease), chronic ulcerative lesions, subcutaneous abscesses Nocardia

81. Clinical syndromes Lymphocutaneous infections: Regional lymph nodes Mycetoma: Chronic granulomatous disease; may involve bone, connective tissue and muscle CNS: Single or multiple brain abscesses Nocardia

82. Treatment Proper wound care and trimethoprim- sulfamethoxazole for 6 weeks or more Prognosis is poor for immunocompromised patients with disseminated disease Nocardia