Mycobacterium Tuberculosis

Sample for Mycobacterium Tuberculosis

• This test is done on the skin of the patient.

Indications for Mycobacterium Tuberculosis

1. To diagnose a suspected case of tuberculosis.
2. Screening of people at risk like:
   1. Health worker.
   2. Immigrant to a high-risk area.
   3. IV drug user.
   4. Patients at increased risk of developing active TB.
3. So this TT should be targeted to the high-risk group which is called targeted tuberculin testing rather than people at low risk which is called screening test.

Epidemiology of Mycobacterium Tuberculosis:

1. Tuberculosis is the second leading cause of od death in the world.
   1. Nearly 3 million people die each year.
   2. 7% of all deaths occur worldwide.
   3. The global epidemic may become worse because of drug-resistant organisms and HIV patients.
2. Mycobacteria are widely distributed in our world; this is referred to as acid-fast bacilli because they resist decolorization after mordanting with strong acid.
   1. The staining property is related to the cell wall composition with high lipid contents.
   2. The acid-fastness results from the formation of complexes between the dye and mycolic acid, one of the cell wall lipids.

Laboratory characteristics of Mycobacterium tuberculosis:

1. They are weakly gram-positive organisms.
2. M. tuberculosis measures 0.2 to x 5.0 µm bacillus.
3. These are slender, beaded bacilli and non-sporing organisms.
4. It is an obligate aerobe.
5. It grows in high O2 tension (pO₂) areas like the lung apex tissue.
6. It is a slow-growing organism that needs 1 to 2 weeks or more to grow.
7. It has high cell-walled lipid content like mycosides and is impermeable to gram stains.
   1. Smears are treated with concentrated carbol fuchsin for 10 to 20 minutes, which acts as a mordant by heating, and then decolorized with 20% sulphuric acid+ alcohol; now, bacilli will have a bright red color.

Culture media for Mycobacterium Tuberculosis:

1. These bacteria need highly enriched media, Lowenstein-Jensen media. These bacteria do not grow on ordinary media.
   1. Lowenstein-Jensen media contains whole eggs, aspargine, glycerol, and malachite green, which will inhibit the growth of contaminants.
2. The specimen is incubated at 37 °C for 2 to 3 weeks, but the culture should be kept for 6 to 8 weeks.
3. Nowadays, these bacteria are identified under fluorescent microscopy.
4. These are divided into two groups:
   1. Rapid grower, which forms the colonies in 2 to 3 days.
   2. A slow grower who takes 2 to 3 weeks.
5. The medically important mycobacteria are:
   1. M. tuberculosis.
   2. M. bovis.
   3. M. leprae.
   4. Atypical mycobacteria.
6. Mycobacteria tuberculosis causes a slowly progressive chronic infection, usually of the lungs.
   1. But other organs are not spared by this disease.
   2. The main source is the lung’s secretions, so-called open disease.

Principle of Mycobacterium Tuberculosis  Tuberculin sensitivity test (TT):

1. The Tuberculin test (TT) is an important test in diagnosing and preventing tuberculosis.
   1. This test is more sensitive than the X-ray chest.
2. This test is done on the patient where purified protein derivatives (PPD) are stabilized by Tween 80 (polysorbate 80-coated nanoparticles) injected into the skin (Intradermal).
   1. The standard test material is intermediate-strength, which is 5 tuberculin units, PPD.
3. Lymphocytes will recognize PPD as an antigen and give rise to type IV, cell-mediated reaction.

4. There will be a local inflammatory reaction in a person who has TB or has past exposure to TB.
5. This test will give a severe reaction in the case of active TB or patients with vaccination. There may be sloughing of the skin.

Reading of the Tuberculin test (TT):

1. Read the test area after 48 to 72 hours of the injection.
2. The negative case is when there is no induration.
3. In positive cases, there will be induration at the site of injection appearing in 48 to 72 hours.

The American thoracic society and CDC criteria:

1. They suggest the following criteria for different population groups:
2. Induration <5 mm D is seen in:
   1. HIV patients or immunodeficiency state.
   2. Or a patient with a CMI defect.
   3. Close contact with active TB case.
   4. The patient with X-Ray finding of old TB.
   5. Patients with fibrosis or calcification in the lung suggest previous pulmonary tuberculosis.
3. Induration >10 mm in D seen in:
   1. I/V drug users.
   2. Nursing home residents.
   3. The traveler from the endemic or high-risk area of Tuberculosis, prisoners, homeless,  I/V drug abusers, and other chronically ill patients.
   4. A worker in the home care facility.
   5. Malnutrition, postgastrectomy, steroid use, and Diabetes.
4. Induration >15 mm in diameter seen in:
   1. Patient with active TB.
   2. All patients who don’t fulfill the above criteria.
5. A booster reaction may be mistaken for conversion among hospital workers or other people where repeated testing is needed.
   3. In such cases where the primary reaction is negative or the result is in doubt, give the second dose after one week.
   4. If the size of the induration is increased by more than 6 mm next year, a new tuberculous infection may be suggested.

Contraindications for Tuberculin test, Mantoux skin test (T T):

1. A patient who has active TB will have a very severe reaction.
2. A patient who has a vaccination with Bacillus Calmette Guerin (BCG).
3. No use in a malnourished patient.

The positive Tuberculin test, Mantoux skin test (T T) is seen in:

1. An active case of TB.
2. Other types of Mycobacterial infection.

The negative Tuberculin test, Mantoux skin test (T T)  is seen in:

1. Immune incompetent chronically ill patient.
2. The patient was not exposed to TB.
3. It is found 20% of the HIV-negative and 40% of HIV-positive cases with tuberculosis.
4. Viral infections, including measles, and rubella, can suppress tuberculin reactivity. However, this effect is transient.

Because of the tuberculin skin test’s limited sensitivity and specificity, new methods to diagnose Mycobacterium tuberculous infection are now being developed, like a sandwich-capture enzyme-linked immunoabsorbent assay.

Questions and answers:

 

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