Mycobacteria

Mycobacteria are a genus of Actinobacteria and belong to the family of Mycobacteriacae.
They are immobile, obligate aerobic, acid-fast Gram-positive bacilli with a high genomic G+C content (59-66%). Due to the rich content of mycolic acids/mycolates the cell wall is hydrophobic and waxy – attributes, which cause acid-fastness and makes substantial contribution to the hardiness of this genus.

The Mycobacterium genus comprises more than 120 different species. Their pathogens can cause serious diseases in humans and animals, for example tuberculosis and leprosy. Concerning their pathogenicity three groups can be subdivided:

• M. tuberculosis complex
• NTM (Non-Tuberculous Mycobacteria)
• M. leprae

Mycobacterium tuberculosis complex
The M. tuberculosis complex includes the closely related organisms

• M. tuberculosis
• M. africanum 
• M. bovis (subspecies bovis  and caprae)
• M. bovis BCG (Bacille Calmette-Guérin)
• M. microti
• M. canettii
• M. pinnipedii

These, with the exception of M. bovis BCG, are considered to cause TB in humans and animals. Despite their close genetic similarity, these organisms differ considerably with regard to epidemiology, pathogenicity, and their host spectrum.

M. tuberculosis is considered to be the principal cause of TB in humans. The TB germs are spread through droplet infection, and crowded and poor hygienic conditions aid the spread of the disease. The pathogen settles in the lung where it may induce TB or persist for many years in the defence cells of the body, the macrophage as latent tuberculosis. When a latent TB emerges as an active disease is not well predictable. Certainly patients with immune deficiency are at high risk. Other organs can be seized with the pathogens, if it comes to reactivation of the primary infection (post-primary tuberculosis) with dissemination of the bacilli (extrapulmonary tuberculosis). 

M. africanum occurs mainly in Africa and affects TB in humans, too.

M. bovis ssp. bovis is the major cause of TB in cattle and other animals and can be transmitted to humans by droplet infection or by food intake of unpasteurized milk. In the latter case an intestinal TB will be elicit. Bovine TB is also transferable back from humans to cattle. M. bovis ssp. bovis is resistance to pyrazinamide (PZA) unlike M. bovis ssp. caprae, which is PZA-sensitive. The resistance pattern serves as a criterion to distinguish the subspecies.

BCG vaccine strain, as a live vaccine for TB prophylactic immunization, was developed from M. bovis as an apathogenic variant. But up to now the most widespread form of disease, pulmonary tuberculosis in adults cannot be prevented. Due to consistently low TB incidences and a corresponding risk-benefit analysis the vaccination exists no longer for example in some European countries. Because of the high risk of infection in high-burden countries the vaccination is still recommended by the World Health Organization (WHO) for children under the age of 1 year as a protection against TB in these countries. The BCG strain, however, also  is used in the treatment of malignant tumors (e.g. bladder cancer).

M. microti mainly infects rodents; only few cases are reported as infections among humans. The human pathogen M. canettii has so far only rarely been isolated. M. pinnipedii infects primarily seals (pinnipeds), infection of cattle is also possible.

The M. tuberculosis complex can be cultivated on common liquid and solid culture media. Due to its slow growth a statement of negative culture cannot be made after an incubation time of 6 weeks for liquid culture and of 8 weeks for solid culture. A positive culture result is available after 2-4 weeks. Drug susceptibility testing (DST) takes additional 12-42 days. Molecular genetic systems can be an alternative to reduce time of identification to 1 day and additionally DST for first- and second-line drugs in case of acid-fast positive microscopy.

Non-tuberculous Mycobacteria
In addition to the representatives of the M. tuberculosis complex the non-tuberculous Mycobacteria can also cause mycobacterioses in humans which are mostly chronic. Synonyms are “Non-Tuberculous Mycobacteria” (NTM),  ”Mycobacteria Other Than Tuberculosis“ (MOTT) or “Atypical Mycobacteria”.
This heterogenic group is spread worldwide. The majority of NTM is living ubiquitous in nature. Only few are as parasites rely on host. They can be detected in soil and ground water, in sewage and dust as well as in fresh- and sea water. Being classic opportunists, NTM predominantly infect patients already suffering from pulmonary diseases or immunodeficiency; however, the number of mycobacterioses is also increasing among immunocompetent persons.
Therefore the M. avium complex is one of the most common pathogens in immunosuppressed patients, while M. kansasii, M. malmoense and M. xenopi are predominant in immunocompetent people. The clinical symptoms of mycobacterioses are various, ranging from pulmonary infections through granulomatous infections of the skin to lymphangitides.
Just as M. tuberculosis complex NTM can be cultivated on common liquid and solid culture media. Due to their growth behavior the classification is carried out in 4 groups according to RUNYON:

A triple isolation from direct patient material and a reliable species differentiation in combination with clinical symptoms are consequently of great importance to decide if it is a mycobacteroises which requires treatment.
Conventional biochemical methods are time-consuming and demand trained staff. Molecular genetic mycobacterial detection is therefore the method of choice. The GenoType^® Mycobacterium CM/AS allows the identification of M. tuberculosis complex and 30 of the most common NTM species from cultivated samples.
 

Mycobacterium leprae
Mycobacterium leprae, also known as Hansen`s bacillus, was discovered in 1873 by Gerhard Armauer Hansen in the skin nodules of patients with leprosy. It was the first bacterium to be identified as causes of human diseases. The Mycobacterium leprae occurs in the lesions of lepromatous leprosy in intra – and extracellular masses, known as globi.
The bacilli have never been cultivated on artificial cell culture. For this reason the microscopy is the common way of detection leprosy. Smears from skin or nasal smears are recommended.
This fast-acid bacillus closely resembles in size and shape to the M. tuberculosis complex. The rod shaped organism with parallel sides and rounded ends often grouped together like bundles of cigars or arranged in a palisade. It is assumed that only viable leprosy bacilli are stainable with carbol-fuchsin as solid acid-fast rods. Bacilli which show an irregular stain are supposedly dead and degenerating.
The morphological index together with the bacteriological index, calculated by counting six to eight stained smears and results are expressed on a logarithmic scale, help to assess the infection state of patients before and during treatment.
The GenoType LepraeDR enables the identification of M. leprae and its resistance to rifampicin, ofloxacin and dapson within one day.

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