– From 1880 to 1900 are described in humans bacilli of leprosy and tuberculosis; other acid-fast bacilli are also observed in animals.
In 1885 is described in humans, the “smegma bacillus”, currently named Mycobacterium smegmatis.
– Between 1900 and 1950, Mycobacteria “nontuberculous” are observed in humans from various samples (tonsils, skin, pleural fluid, sputum, urine).
But it was difficult to clearly associate the presence of these bacteria in human disease.
– From 1950, emerges the concept of human infections with nontuberculous mycobacteria because laboratory tests become more efficient, as the frequency of tuberculosis begins to decrease with antibiotics and that bacterial-clinical correlations are performed.
– In 1954, a group led by Runyon studied hundreds of strains isolated from patients, leading in 1959 to the classification by the authors of mycobacteria.
These germs have received several names: paratuberculosis bacilli, pseudotuberculeux, unclassified, anonymous, non-tuberculous, atypical, opportunist, tuberculoid.
Currently, it is better to speak of so-called atypical mycobacteria or bacilli other than tuberculosis (MOTT = mycobacteria –other than tuberculous).
I – BACTERIOLOGICAL CLASSIFICATION RUNYON:
In 1959, Runyon offers bacteriological classification of nontuberculous mycobacteria. This classification therefore includes neither Mr. tuberculosis or M. bovis or M.africanum.
M. leprae, not cultivable, do not belong. This classification is based on the bacteria in vitro growth rate and on the conditions of any pigmentation of these organisms in culture.
– Growth rate:
Obtaining mature colonies and macroscopically visible more than 5 days is slow growing and in less than 5 days rapidly.
– Photochromogens Colonies:
These are non-pigmented colonies if their growth has occurred in the dark pigment after exposure to light in the presence of oxygen.
These are pigmented colonies that growth took place in the light or in the dark.
slowly growing mycobacteria and photochromogens
eg M. kansasii, M. marinum, M. simiae, M. asiaticum
slowly growing mycobacteria and scotochromogenic
eg M. scrofulaceum, M. gordonae, M. flavescens, M.xenopi Mr. szulgai
slowly growing mycobacteria and non-pigmented
eg M. avium-intracellulare, M. malmoense Mr. gastric M. haemophilum Mr. nonchromogenicum M. terrae, M. trivial M. ulcerans
rapidly growing mycobacteria, pigmented or not
eg M.fortuitum, M. chelonae, M. phlei, M. smegmatis, M. vaccae
This classification is not absolute because some pigmented mycobacteria may lose their pigment after subculture;Moreover, Mr. szulgai photochromogen is at 25 ° C and scotochromogène at 37 ° C. M. xenopi scotochromogène the only primary culture is often classified in group III because of its many similarities to avian mycobacteria.
II – CLASSIFICATION OF CLINICAL MYCOBACTERIA ACCORDING TO PATHOGENICITY HUMANS:
A – always considered pathogenic Mycobacteria:
Besides Mr. tuberculosis, M. bovis, M. africanum and M. Mycobacteria leprae there are rarely encountered:
1. M. ulcerans (group III):
Responsible painless skin ulcers, chronic and extensive without cellular reaction. This is the Bairnsdale ulcer or ulcer observed in Australia, Central Africa and Central America, which could be due to a toxin produced by the bacteria.
2. Mr. haemophilum (group III):
This bacterium has been described for the first time in Israel in 1978, and is responsible for disseminated skin lesions (cutaneous infiltration, abscesses, granulomas, fistulas) with cellular reaction.
3. Mr. szulgai (Group II):
Described in 1972, this bacterium is primarily responsible for lung conditions simulating tuberculosis but also of bursitis (olecranon). It was also found in lymph nodes and skin lesions.
Its natural habitat is unknown.
4. Mr. malmoense (group III):
Described in 1977 in Malmo, this bacterium has been associated with lung diseases (Sweden, Wales, Australia).
B – often pathogenic Mycobacteria:
1. M. marinum (group I):
This bacterium is responsible for chronic skin granulomas preferably sitting in the limbs. This is the “swimming pool granuloma” which resolves spontaneously within a few months more often or that causes deep ulcerations with cellular reaction (swellings stepped is sometimes observed on the lymph channels, simulating sporotrichosis).
The natural reservoir of the organism is represented by the cold-blooded animals, especially exotic fish in aquariums, swimming pools and the coastline of warm seas.
2. Mr. kansasii (group I):
This species causes lung diseases may resemble clinically and histologically tuberculosis. But it was found in lymph nodes, skin conditions, bursitis and urogenital infections and meningitis. In the USA this mycobacterium is frequently encountered as a cause of lung infection.
The natural reservoir of this organism is unknown. Sometimes is isolated bacterium samples of water and milk and exceptionally in bovines and pigs.
Note that strains with high catalase activity seem more pathogenic than those with low catalase activity.
3. M. simiae (group I):
This germ is sometimes responsible for lung disease in humans; it has sometimes found associated with M.tuberculosis in some patients. The germ of the reservoir is unknown; this bacterium has been isolated in monkeys and humans.
4. M. scrofulaceum (Group II):
D is responsible for submandibular lymphadenopathy festering in children under 8 years. A few cases of lung disease have also been described, especially in patients who already have caves to tuberculosis. The germ of the tank is not well known and some strains were isolated from milk, oysters, water and soil.
5. Mr. avium and M. intracellulare (group III) (or Battey bacillus)
These two species were initially separated on the basis of their pathogenicity for chickens and rabbits that are sensitive only to the injection of M. avium by intravenous route. But biochemically these two species can be differentiated and are part of avium-intracellulare complex. This group is not pathogenic for guinea pigs. In humans this complex is responsible for lung diseases.
In addition, in children, this seed is sometimes responsible cervical lymphadenopathy.
Joint infections, genitourinary and meningitis have sometimes been described.
Since the outbreak of the AIDS epidemic, atypical mycobacteria, including group M avium-intracellulare, have an increased frequency isolation. Indeed, disseminated mycobacteriosis occupy after the Pneumocystis cariniipneumonia and sepsis Cytomegalovirus, 3rd place among the terminal opportunistic infections in AIDS patients.Mycobacteria in question can be isolated by blood culture, bone marrow or stool cultures. These mycobacteria of theavium-intracellulare group are found in the bodies of nearly half of the patients who died of AIDS in the US
M. avium-intracellulare usually resistant to most antituberculosis including rifampicin. The clinical efficacy of ansamycin and clofazimine is discussed.
The germ is also isolated in birds with avian tuberculosis in mammals and the environment; the germ appears to remain viable and virulent in the soil for several years.
6. M. xenopi (Group II):
This species is often responsible for chronic lung disease in Northern Europe and Australia. She comes to Paris in third place after Mr. tuberculosis and M. Sometimes she bovis colonizes old tuberculous lesions.; it has also been isolated from tonsils.
The germ was found in the hot or cold water tap in the external environment and rarely in animals (pigs).
C – very pathogenic Mycobacteria:
1. Mycobacterium asiaticum (group I):
Initially isolated in monkeys, this mycobacterium was found in humans in the United States and Australia.
2. Mycobacterium fortuitum and M. chelonae (Group IV):
These two bacteria are often grouped under the term “complex fortuitum.”
These germs can be found everywhere in the environment (water, soil, dust) and in cold-blooded animals (fish, frogs). This is a common commensal of man.
In humans they mostly cause abscesses at injection drug products.
They are found in infections after orthopedic surgery (slow evolution), as well as keratitis and arthritis. Rare lung abscesses were observed on pre-existing lung injury, or after inhalation of oil particles or foreign bodies.
However, there may be prolonged colonization of the respiratory tract, without obvious clinical signs.
This is most often responsible for serious post-surgical infections mycobacteria in patients subjected to immunosuppressive therapy. This is incubated and long evolution infections.
D – Mycobacteria considered non-pathogenic to humans:
1. pathogenic species of animals:
– M. paratuberculosis, the agent of diarrheal hypertrophic enteritis of cattle (or Johne’s bacillus) growing on media supplemented Mycobactin J (siderophore mycobacteria)
– Mr. microti, agent of tuberculosis vole;
– Mr. lepraemurium, the agent of murine leprosy (Bacille Stefansky), difficult to grow, and used by Merkien Cottenot for serodiagnosis of leprosy;
– Mr. farcinogenes, agent farcy beef Chad;
– Mr. Senegal, agent farcy beef Senegal.
2. saprophytic species found both in the environment and in humans:
– M. gordonae: it is a saprophytic mycobacterium widespread in nature;
– Mr. flavescens: This is a common saprophyte in the environment and in the laboratory;
. – Mycobacteria complex “radish” or “terrae” They are also quite common saprophytes Mr gastri. Mr. nonchromogenicum M. terrae, M. trivial.
III – GERM TANK AND EPIDEMIOLOGY:
Atypical mycobacteria are bacteria widespread in nature. Ubiquitous, they were isolated from water, soil, plants and many animals both domestic and wild, but there is not really duly proven natural reservoir as is the case for Mr.tuberculosis or bovis. These germs are frequently isolated from water distribution networks, including in hospitals.
The most exposed subjects to mycobacterial diseases are those with an overall deficit of cellular immunity (cancer, transplant, AIDS), or a local deficit of immunity by decreasing the activity of alveolar macrophages (silicosis, pneumoconiosis). These infections can produce lymphadenopathy in children.
They are usually benign and often affect a lymph node. The distribution of these infected nodes suggests localized nasopharyngeal colonization of mucous membranes, bronchial and intestinal also by waterborne bacteria. Humans become infected by drinking contaminated water or being exposed to aerosols of tap water (showers). There are also infections related to the accidental introduction of germs in the tissues (syringes, implantation of foreign material contamination).
IV – BODIES MOST FREQUENTLY AFFECTED:
A – Lung:
II is the most frequent target reached (80-90%) by mycobacteria, particularly M. kansasii, M. avium-intracellulare andM. xenopi Rarely are criminalized. M. scrofulaceum Mr. szulgai, M.fortuitum-chelonae, M. malmoense, M. simiae, M. asiaticum. Different
clinical forms exist, ranging from asymptomatic lesions discovered during a routine X-rays, to cavitation accompanied by hemoptysis.
The typical patient is a human male, aged over 40 years with chronic lung injury. Miners, arc welders are particularly vulnerable.
Factors favoring the emergence of a pulmonary nontuberculous mycobacterial infection are:
– Factors related to the nature of the infecting germ,
– Factors related to the resistance of the infected organism,
– The level of local resistance associated with bronchopulmonary underlying conditions (pneumoconiosis, chronic bronchitis, bronchiectasis, tuberculosis sequelae, lung cancer)
– The level of general resistance.
However, cases are sometimes observed among women, young men without preexisting lung lesions or without deficit of cellular immunity.
The isolation of saprophytic mycobacteria from sputum is common.
This is why the pulmonary mycobacteriosis diagnosis will be focused on the convergence of the following arguments:
a / absence of mycobacterium tuberculosis (mixed infections are common);
b / isolation of germ SEVERAL times to days or weeks apart with a significant number of colonies in culture and / or positive direct microscopy (Ziehl)
c / clinical and radiological signs more or less suggestive of tuberculosis;
d / possibly isolation of mycobacteria directly in lesions after surgery or autopsy.
Unlike tuberculosis, pulmonary mycobacteriosis these are usually not contagious from person to person: mycobacteria in question come from the environment.
B – Lymph nodes:
Most cases occur in children under 5-8 years (range, 7 months to 12 years). These cold cervical lymphadenopathy are mostly but can sometimes sit in the limbs or in the parotid region. There is little or no painful unilateral lymphadenopathy in a child with a good conservation condition. Usually these lesions tend to fistuliser.
Sometimes lymph nodes remain stable or regress spontaneously. Recovery is usually spontaneous, although relapses can be observed.
Three species are mainly involved: Mr. scrofulaceum, M. avium-intracellulare, M. kansasii and sometimes Mr. szulgaiand M.fortuitum.
C – The soft tissue, the skin:
It is skin ulcers (M. ulcerans), chronic granulomas (M. marinum), abscess secondary to localized drug injections(M.fortuitum-chelonae). And Mr. M. haemophilum szulgai were also found in these locations.
D – Bones and Joints:
Synovial, tendon sheaths, bursae can be infected with M. avium-intracellulare, M. kansasii, M. fortuitum, M. szulgaiand bones by Mr. fortuitum-chelonae.
E – widespread disease:
It can be observed in immunocompromised patients, especially in patients with AIDS. The germs involved are especially M. avium-intracellulare, M. sometimes kansasii, M. and M. szulgai xenopi.
V – IDENTIFICATION MYCOBACTERIA LABORATORY:
A – Identification of the seed:
The techniques used are those used for tuberculosis bacilli.
However, some mycobacteria (M. fortuitum and M. chelonae) are very sensitive to decontaminants usually used.
Moreover, the culture of M. haemophilum requires special environments: or chocolate agar media supplemented with lysed red blood cells or hemoglobin 0.4% or 60 microM hemin; or Löwenstein-Jensen with 1% of ferric ammonium citrate.
The incubation temperature is usually 37 ° C; but the products of the skin covering or surface origin, it must be both 30 ° C and 37 ° C.
B – Identification tests of mycobacteria:
1. Check that these are acid-fast bacilli.
2. Check the purity of the strain because mycobacteria associations are not uncommon.
3. Study the germ growth rate at various temperatures (28 ° C,
37 ° C, 42 ° C). The speed and the optimum growth temperature will be estimated on a subculture of the strain having single colonies.
4. Observe the appearance of colonies:
– Rough Colonies R (or eugoniques), as Mr. tuberculosis.
– Smooth Colonies S (smooth), as Mr. Hovis.
– Intermediate Aspect.
– Pigmentation: photochromogens colonies scotochromogenic not pigmented.
Looking for a photo-inducible pigmentation must be made as follows: a strain suspension to be tested is diluted to obtain isolated colonies on medium 3 tubes to the egg. Two of these tubes were wrapped in aluminum foil and the third remains unprotected from light. A few days after the onset of colonies on the unprotected tube growth is controlled to the mycobacterium in the wrapped tubes and one of them is only exposed for 5 hours at a 100 W lamp having uncapped tube or loosening the screw cap to promote oxygenation of the culture.
All cultures are returned to the oven, and the appearance of a possible pigmentation is observed after 24, 48 or 72 hours.
5. Biochemical Properties
a / synthesis and release of nicotinic acid in the medium (niacin-test)
B / Study of enzymatic activities: nitrate reductase, catalase at 22 ° C, catalase at 68 ° C and pH 7, arylsulfatase, hydrolysis of Tween 80, urease pyrazinamidase, amidases, P-glucosidase, growth in the presence of fructose …
6. Resistance to various agents
5% NaCl, NaNO2, TCH (hydrazide of thiophene-2-carboxylic acid), Tbi (thiosemicarbazone), PAS, ethambutol, MacConkey medium without crystal violet, para-nitrobenzoic acid, hydroxylamine, D-cycloserine.
C – Identification itself:
1. Bacteria slow growing photochromogens (Table I):
Photochromogénicité studies should be performed with care because they are important for the correct identification of these germs.
Cultures to confluent growth or aged colonies can not produce pigment after exposure to light.
These bacteria have a thermostable catalase.
If the catalase activity of M. kansasii is high, the germ is deemed more pathogenic than if it has a low activity. In Ziehl, it looks grainy.
2. Mycobacteria slow growing scotochromogenic (Table II):
These bacteria have a thermostable catalase.
M. szulgai photochromogen is at 24 ° C and scotochromogène at 37 ° C.
M. xenopi is often pigmented as the primary culture. These are small colonies long bacilli. M. xenopi grows faster at 42 ° C than at 37 ° C.
3. Bacteria slow-growing, non-pigmented (Table III):
M. tuberculosis, M. bovis, M. africanum, M. and M. malmoense gastric have catalasc thermolabile at 68 ° C.
M. ulcerans is isolated from skin or subcutaneous. It grows at 30 ° C in 6-9 weeks.
Mr. malmoense resistant to INH, streptomycin, PAS, and rifampin.
M. haemophilum grows at 30 ° C, requires hemin or pushes on Löwenstein-Jensen containing 1% ferric ammonium citrate.
Other bacteria without clinical interest: mycobacteria of “radish” complex or “terrae”: this group is sensitive to ethambutol.
4. Bacteria fast growth (Table IV):
They form colonies in 5-7 days and grow on common bacteriological media agar.
a / Mycobacteriumfortuitum-chelonae
It is non-pigmented bacteria growing on MacConkey medium without crystal violet.
M.fortuitum sometimes absorbs green and malachite green colonies present on egg-based medium.
b / pigmented species
II are many no medical value, but sometimes encountered in analytical laboratory: M. vaccae (photochromogen), Mr.auruin, M. parafortuitum Mr. neoaurum, etc.
c / thermophilic species more or less pigmented Mr phlei, M. thermoresistibile, M. smegmatis sometimes used as a nonspecific stimulant in cancer immunity.
5. Identification by DNA probes:
Specific nucleic probes allow the identification of certain so-called atypical mycobacterial species: M. aviumm, intracellulare, M. and M. gordonae kausasii. The results are very satisfactory, but the probes still have the disadvantage of being radiolabeled.
VI – PATHOGENICITY METHODS:
– The guinea pig is not sensitive to the so-called atypical mycobacteria injected subcutaneously.
– The rabbit and hen are susceptible to M. avium administered intravenously.
– The mouse can be useful but is not necessary to differentiate M. kansasii, M. marinum and M. ulcerans. M. kansasiiinjected intravenously in this animal produces a disease of internal organs only. Tail and foot pads, the temperature is below 37 ° C, remain unharmed. The other two species after intravenous injection or footpads, produce lesions in the cooler parts of the body like the tail, foot pads, nose, scrotum etc.
M. ulcerans grows very slowly compared to M. marinum but causes more serious damage.
VII – sensitins:
The mechanisms of immunity and hypersensitivity caused by non-tuberculous mycobacteria are similar to those of tuberculosis. The sensitins are soluble antigens derived from cytoplasmic fractions after rupture of the wall of the bacilli.
Their use is the same as that of the tuberculin: detect delayed hypersensitivity to a nontuberculous mycobacterium.Unfortunately there are significant cross-reactivity between the various sensitins, that is to say that a given mycobacterium infection is detected by the counterpart sensitin and sensitins from other mycobacterial species.However homologous reaction is generally greater than the heterologous reaction, provided the use of well standardized sensitins. Currently, use of purified fractions of sensitins known PPD (“purified protein derivative”). For example, in the case of infection with M. avium-intracellulare will be conducted in parallel an intradermal PPD-bird to a patient’s arm and another PPD of tuberculosis in the other arm. We compare the surfaces of the two reactions: reading must be made by at least two observers; if the surface of the reaction to the PPD-bird is greater than that of the reaction to the PPD tuberculosis, we conclude to infection by M. avium-intracellulare. In practice, this technique is useful in young children who have not had contact with other mycobacterial antigens, particularly in the case of lymphadenopathy. In adult humans, the situation is more complex to consider.
PPD is provided by the Statens Serum Institute in Copenhagen.
VIII – serotyping MYCOBACTERIA TELL OF NON-STANDARD:
Some species of mycobacteria include various serotypes, which is of interest for epidemiological studies.
M. avium-intracellulare has 28 serotypes (1-28) (serotype 8 is most often encountered in humans)
M. scrofulaceum: 3 or 4 serotypes (41 to 43 or 44)
M. marinum: 2 serotypes (1-2)
M.fortuitum 2 serotypes (1-2)
M. gordonae: 7 serotypes (1-7)
M. simiae 1 serotypes.
M. chelonae, M. and M. szulgai kansasii are serologically homogeneous (single serotype).
IX – THERAPEUTIC:
The sensitivity of the so-called atypical mycobacteria is usually evaluated by the method of solid medium proportions.
The so-called atypical mycobacteria are almost always resistant to INH and PAS, and many are sensitive to D-cycloserine (except szulgai M. chelonae and M. fortuitum-). However, the observed in vitro sensitivity does not always result in vivo efficacy of the drug; the reasons are unclear.
Antimicrobial susceptibility by the radiometric method is not applicable to so-called atypical mycobacteria. Many alternative protocols are under study, aimed at standardizing the susceptibility of non-tuberculous mycobacteria.
M. kansasii usually responds well to appropriate treatment. It is especially sensitive to the thiosemicarbazone, ethambutol, ethionamide and rifampicin. However Mr. kansasii appears wrongly resistant to rifampicin on conventional susceptibility when rifampicin is replaced by rifamycin SV, more stable, and having the same vis-à-visMr. activity tuberculosis. Now Mr. kansasii is resistant vis-à-vis of rifamycin SV; it is necessary to measure the sensitivity of this organism to real rifampicin on a medium prepared extemporaneously.
M. marinum: infections usually recover completely but slowly (several months). Was successfully used in the treatment: trimethoprim-sulfamethoxazole, cyclins rifampicineéthambutol or association to which the germ is often sensitive in vitro.
M. avium-intracellulare, M. scrofulaceum, M. xenopi, M. and M. szulgai simiae are usually multiresistant to antibioticsin vitro and respond poorly to antibiotic treatment. For pulmonary infection with M. avium-intracellulare have been proposed including associations ansamycin (derivative rifampin) or D-cycloserine. Surgical treatment give satisfactory results after stabilization of lesions by antibiotic treatment.
The use of clofazimine (leprosy) of clarithromycin and ansamycin was advocated in generalized infection with M.avium-intracellulare.
For Me. fortuitum-chelonae, amikacin, erythromycin, tetracyclines, sulfonamides, ethionamide can be effective.