Mycobacterium tubercolosis

Mycobacterium tuberculosis (also Koch's bacillus' - KB') is a medium-long, sometimes granular acid-resistant rod. It is an obligate pathogen of mycobacteria and is the causative agent of tuberculosis. It is closely related to Mycobacterium bovis.

Characteristics
Mycobacteria are strictly aerobic.

The 'high hydrophobicity of the bacterium makes processing by lysosomal macrophage enzymes difficult (see below).

They have a long generation time (20-30 hours, cf. Mycobacterium leprae). Therefore, long cultivation on so-called slanted soils is necessary for their capture. This can take 6-10 weeks.
 * They are intracellular parasites and cause chronic infections,
 * do not produce toxins.

Acid resistance
Acid-resistance is poor staining with organic dyes, resistance to decolourisation by acids, bases and alcohol. It is related to high wall lipid content (a typical feature of mycobacteria). They are based on mycolic acids - the longest fatty acids in nature (C60-C90, similar but shorter in nocardia and corynebacteria).

Antigeny
Old (Tuberculinum vetus) was first prepared by Koch. It was obtained by concentrating liquid soil after separation of the bacterial mass. It had no therapeutic effects.

Purified Protein Derivative (PPD), purified tuberculin. It is still used for Mantoux test (Charles Mantoux - French physician), i.e. detection of cellular hypersensitivity of late type against tuberculin:
 * PPD
 * 1) intradermal injection (so that it doesn't wash away)
 * 2) after 24-48 hours, if positive (in infected and vaccinated), an inflammatory infiltrate (pimple) formed by T-lymphocytes and macrophages
 * 3) induction (not erythema) is measured - positive> 5 mm

Cord-factor (trehalose dimycolate)

 * toxic glycolipid, part of the cell wall
 * it's a virulence factor
 * alters mitochondrial membrane → inhibition of respiration and phosphorylation

Other antigens

 * arabinogalactan
 * peptidoglycan
 * complex glycolipids

Direct evidence
Directly demonstrate Mycobacterium tuberculosis, or its components, by the following methods: It has, compared to microscopy, a higher sensitivity for detection. For TB gold standard. Result in 24-48 hours. PCR is used, but also non-variable bacteria are positive (proof of dead DNA), i.e. you cannot make a diagnosis based on PCR alone (e.g. if the clinic is not expressed)!
 * direct microscopy = basic method; high specificity, low sensitivity
 * Ziehl-Neelsen staining
 * hot carbolfuchsin
 * acid decolourisation (mycobacteria remain stained)
 * staining of other structures with methylene blue or malachite green
 * examination of 100 fields of view
 * fluorescence staining
 * hot auramine
 * acid decolourisation
 * fuchsin staining
 * examination under UV - lower magnification is sufficient (25-50 fields of view)
 * Culture card
 * 1) sample decontamination (destruction of faster growing microbes with 4% NaOH)
 * 2) centrifugation (→ thickening)
 * 3) inoculation on Löwenstein-Jensen soil (egg) / liquid soil - Šula
 * 4) reading of the result at 3, 6 and 9 weeks
 * Accelerated cultivation in systems derived from hemoculture (indicator is CO2); result in 5 days
 * Molecular genetics:

Indirect evidence
The QuantiFERON® method is available for indirect confirmation: There is also a risk of false negatives in immunodeficient individuals! The test scheme is as follows:
 * the patient's blood is put into three tubes
 * shaking to wash the antigen from the tube wall
 * the test is not positive in vaccinated patients
 * 1) the tube is empty (negative control)
 * 2) the tube contains T-lymphocyte activator (positive control) - active T-lymphocytes produce IFN-γ which is quantified and indicates the positivity of the reaction
 * 3) tube contains specific peptides from the bacterium

Prevention
Vaccination with BCG' (attenuated strain of Mycobacterium bovis) protects against the development of Mycobacterium tuberculosis infection:
 * induces the formation of an arteficial primary complex at the site of injection
 * develops a nodular granuloma after 2-4 weeks, which heals in 6-8 weeks
 * those already vaccinated or infected have an accelerated reaction in as little as 24-48 hours → they may develop necrosis' (so-called Koch phenomenon)
 * vaccination efficacy is 40-80%
 * previously, active search for infection was performed by chest X-ray, but this method is now obsolete

Treatment
Treatment of tuberculosis takes a long time (months). To avoid the risk of resistance, combinations of antituberculotics are used. In the case of monotherapy, there is a risk of developing a fall & rise phenomenon, i.e. selection of resistant mutants that trigger the next wave of infection. The proportional test is used to test antibiotic susceptibility:
 * Proportional test
 * bacteria are urinated on soil without antibiotic and on soil with a critical antibiotic concentration.
 * if the soil with antibiotic grows 0-1% of colonies relative to the soil without antibiotic, the strain is susceptible

antituberculosis' are used to treat tuberculosis:
 * isoniazid' - bactericidal, highly effective - inhibits mycobacteria for 5-6 days after one dose
 * rifampicin' - bactericidal, blocks RNA polymerase, stains body fluids orange ("blood sweat")
 * pyrazinamide - bactericidal, active at more acidic pH (suitable for intracellular environment); with rifampicin kills even semi-sleeping mycobacteria
 * streptomycin' - bactericidal aminoglycoside (binding to the 30S subunit of the ribosome), ototoxic
 * ethambutol' - bacteriostatic, inhibits synthesis of some metabolites

Related articles

 * Tuberculosis
 * Atypical mycobacteria

References used



 * BEDNÁŘ, M, et al. Lékařská mikrobiologie. 1. vydání. Marvil, s. r. o., 1996. s. 305–313.  ISBN 80-238-0297-6.