Biology 216 - Lect 27 Tuberculosis and Leprosy I. TB - M. tuberculosis and M. bovis - tubercle bacillus Consumption; Egyptian mummies (2400 BC); 1700s healthy climate, sanatorium (isolation, rest, diet) Koch's Postulates; Vaccine 1920s; Streptomycin 1940s; 1999 Genome sequence A. Bacteriology - same as MOTT M. tuberculosis - man only M. bovis - catttle and man; invasion through GI Obligate aerobes - tendency to colonize apex of lung Very slow growers - 4 - 6 weeks (generation of 24 hr) No exotoxins; Lipids make up 10 % of cell weight (60 % of cell wall) Sulfatides - sulfur lipids which inhibit phago-lysosome fusion Muramyl dipeptide active adjuvant; Freund's, wax D and MDP B. Pathogenesis and disease 1. Epidemiology - A disease of civilization Perspective - 1800s 1% of city population die from TB/yr (Ex. New York 100,000) 10 million/yr worldwide (3 million deaths); Estimated 2 billion infected. US - 28,000/yr epidemic (reverse trend downward) (15 million infected) - Plan for elimination by 2010 Incidence highest in crowded states (CA, NY, FL,TX). Control: antibiotics, living conditions, inexpensive screening (skin test and x-ray), case workers. Increase: immigration (1/4 of new cases, crowded, homeless shelters, prisons. Abandoned control in 1970s: not much time in med school, no new drugs, less research Currently #2 reportable infectious killer in US (AIDS is #1) 2. Transmission - M. bovis no longer major due to cattle control in developed countries Elimination of cattle, pasteurization. Airborne - coughed, dried secretions, small droplets from active or cavitation TB. Outbreaks - teacher, bus driver w/active TB, health care workers 3. Infection vs disease Man is highly susceptible to infection (skin test +), but resist disease. (less than 5 % symptomatic in one year),10 % chance of TB in lifetime (highest first 2 years) Disease depends on dose and host ability to respond and wall off (Crowding, nutritional inadequacy, alcoholism, young and very old, AIDS) 35 % of AIDS patients have TB -> death in 1 month from TB symptoms Genetics - Native Americans, Inbred mouse strains. 4. Primary TB - primary focus in alveoli (mild or asymptomatic) Macs phagocytize, TB multiply within, drain to hilar nodes, blood, miliary, bones... CMI by 4th week stops spread, tubercle walls off, active Macs kill (TB reduced numbers) T-cells -> IFN alpha important (stimulate Macs) TB more virulent in IFN negative mice, + skin test and + x-ray (both may be + for life) Healthy patient - flu-like. Unhealthy - miliary TB, impair function of lung, fatigue, fever, wasting (By Mac -> TNF alpha) Fever, cough, bloody sputum, wasting (consumption) 5. Secondary (Reactivation/reinfection) TB TB survive in tubercle (90 % latent) Decreased CMI (alcoholism, cancer, age, AIDS) years later Tubercle liquifies - spread, lung dysfunction, chest pain, fever, weight loss Cavitation susceptible to Nocardia New foci controlled, but massive pathophysiology - 50 % fatal w/o and 6 % w/ treatment. MDRTB - 50 % fatal C. Diagnosis:Early diagnosis important to treat and stop spread. History (socioeconomic, HIV..), x-ray, 1. Skin test - for DTH to TB antigen after 4 weeks of infection, BCG, specific anegy in active TB. - Intradermal injection of TB Ag (Mantoux)(tuberculin syringe) or multiple puncture (less reliable antigen delivery) - High population of sensitized T-cells in circulation, immobilized, cytokines -> Macs attracted. - 48 - 72 hr induration/red (raised and hard) and erythema (red) - measure diameter. - Antigen - OT (old tuberculin) - dialyzed TB PPD - AmSO4 ppt of OT (less Ag, more specific) - Interpretation - Greater than 10 mm = + (present or past infection) 5 - 9 mm = atypical Mycobacteria 0 = negative, injection too deep, anergy (compromised, severe TB, measles (Test for general anergy w/mumps, tetanus skin test) - Skin test dose is not immunizing dose - Suggested treatment if skin positive, even if not active TB 2. Direct exam of specimen (sputum) - Solubilize (2 % N-acetyl-L-cys) and decontaminate (2 % NaOH), then neutralize in 15 min. - Acid fast stain - Scan 100 fields under oil (3 sweeps through slide) picks up all Mycobacteria
|
Finding |
Report |
|
0 |
No AFB |
|
3 - 9/slide |
+ rare (repeat specimen) |
|
>10/slide |
++ Few |
|
1/field |
+++ Numerous |
3. Culture and ID - work in hood
Inoculate media with glycerol (C-source
for building triglycerides) and egg (slants)
Ex. 7H11, Lowenstein Jensen - Support variety
of Mycobacteria; Satisfactory for niacin
test; Malachite green inhibit others;
May add antibiotics.
3-4 weeks growth - dry, cream colored
ID - pigment, niacin, tween hydrolysis...,
DNA hybridization (probe for TB)
Antibiotic sensitivity
4. BACTEC - liquid culture with 14C-substrate ->
14C-CO2 -> fluorescence -> detector (14 days)
Can be used to test antibiotics
5. PCR - Primers for TB specific IS6110
6. Mycophage with luciferase:
Infect isolate, grow in antibiotic -> ?
D. Treat
1. Combination antibiotics, long term (up to 2 years);
Less than 20 % finish -> resistance
Possible subq implants being considered;
Single pill with all antibiotics.
Non-contagious after 1 - 2 weeks;
Lungs take months to improve.
Outpatient treatment when not communicable.
Used to have sanatoria; may return in the future;
in hospital - negative pressure isolation room.
MDR - 50 % mortality.
E. Prevention - Dosage and host immunity
most important considerations.
1. INH treat contacts and skin test+ converts.
2. Better housing, sanitation, nutrition, health
3. Animal control (M. bovis - treat, moniter, pasteurize)
4. Vaccine - BCG (bacillus of Calmette and Guerin) -
live attenuated (1921), enters Macs, then dies
One of safest vaccines known; 3 Billion doses given;
Given orally; 10 cents/dose
Efficacy in question - effective in children, 50 % in adults,
not effective in preventing reactivation TB.
Given to exposed and high risk in some countries.
(Controversy - health care workers)
Not given to immunocompromised
- disseminated disease in AIDS
Not given in US -> conversion to skin test +;
eliminates tool for diagnosis and epidemiology.
Work on cloning protective Ag into BCG
Experimental treatmentof tumors
- BCG attracts and immobilizes Macs
II. M. leprae - leprosy (Hansen's disease)
A. Bacteriology - Obligate intracellular parasite
(Never grown in lab)
Few generations in Mac tissue culture;
mouse foot pad; armadillo blood (endemic)
(2 week generation time)
In vivo growth very slow - incubation period of years.
Prefers low temp - colonize skin,
mucus membrane, peripheral nerves.
Sequenced: 1999
B. Pathogenesis - 10 - 15 million world total (mostly tropics).
1. Transmission - person -> person by nasal
secretion or skin (children especially susceptible)
Lepromatous shed 10(8)/day in nasal.
Survive dessication.
Infection through cuts, mucus membranes (ex URT)
Infection requires - high number, compromised
and prolonged exposure (not very contagious)
Predisposition - crowding, genetic, CMI, nutrition.
2. Very long incubation - ave 4 yrs
Proliferate in Mac and Schwann
(affect peripheral nerves and tissues)
- damage from CMI
Usually asymptomatic or minor skin lesion (blotch)
- CMI protective.
3. Clinical - 2 major forms with intermediate forms
a. Tuberculoid leprosy - early loss of skin pigmentation
Large granuloma formation, may destroy nerve cell ->
Paralysis of muscles, anaesthesia (numbness)
Lesions on face, trunk, skin -> deformity
Good CMI - walled off, few organisms shed,
slow progressive healing, scaring.
b. Lepromatous leprosy (high # and low CMI) - severe form
Anergy to leprosy (specifically) by glycolipid -> Ts
Evidence - Ts low in tuberculoid, high in lepromatous
No granuloma formation, no walling off,
lesions coalesce, large # in macs and schwann
(less nerve damage since no granuloma)
Lesions coalesce -> loss of nasal, lips,
earlobe, digits; Highly contagious.
Susceptible to secondary infection
Ab/Ag complex deposition -> renal failure,
pulmonary blockage
C. Diagnosis - symptomology, h istory of contact
1. AFB in scrapings of lesion (low # in tuberculoid)
2. Skin test - Lepromin (ground material) -
prognostic of patient CMI
3. Cannot grow in lab; Serology available.
D. Treatment
1. Long term antibiotics (1 - 5 years), combination,
relapse may occur
Some resistance to Dapsone
Prophylaxis of close contacts to lepromatous patient.
E. Prevention
1. Chemoprophylaxis for close contacts (lepromatous);
Isolation of patient if lepromatous.
2. BCG vaccine - cross reacts (questionable efficacy)
3. Cloned surface Ag into E. coli
Expression when subcloned behind lac promoter
Currently experimental work on this as vaccine
Other experimental vaccines:
TB/leprosy genes into vaccinia virus