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PubMed1985

Isolation, identification, and structural analysis of the mycobactins of Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum, and Mycobacterium paratuberculosis.

Methods were devised to purify the cell-associated, iron-binding compounds known as mycobactins from the closely related species Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum (i.e., the MAIS complex of organisms). The mycobactins from these three species showed a structure that is common to the mycobactins from all the mycobacteria examined to date. However, these mycobactins were unique in that they had more than one alkyl chain. The M. scrofulaceum mycobactins differed from other MAIS mycobactins by a shift in the position of the double bond in the R1 alkyl chain. Traces of other mycobactin types were observed in ethanol extracts of the three species, and examination of the chromatographic properties of these mycobactins showed that each species produced five mycobactin types. Each mycobactin could be subdivided further by the length of its R1 alkyl chain. No differences in the production of these novel mycobactin were observed among species. Mycobactins from three strains of Mycobacterium paratuberculosis and two wood pigeon strains of Mycobacterium avium which had lost their original growth requirements for mycobactin after repeated subculturing in laboratory growth media were examined by thin-layer chromatography and high-pressure liquid chromatography. Each organism produced a mycobactin with similar chromatographic properties to those synthesized by MAIS organisms. M. paratuberculosis NADC 18 produced at least two components in our laboratory, and nuclear magnetic resonance analysis of the major component showed this mycobactin to be identical to that produced by M. intracellulare M12. However, a sample of mycobactin J isolated by Merkal and McCullough (Curr. Microbiol. 7:333-335, 1982) from M. paratuberculosis NADC 18 was different from our isolates and appeared to correspond to a minor mycobactin component we had seen by thin-layer chromatography. No reason for this difference could be evinced. Our findings indicate that there is a close taxonomic relationship between M. paratuberculosis and the MAIS complex.

Chemical Phenomena↗

PubMed2001

Fluorescent acid-fast microscopy for measuring phagocytosis of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum by Tetrahymena pyriformis and their intracellular growth.

Fluorescent acid-fast microscopy (FAM) was used to enumerate intracellular Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum in the ciliated phagocytic protozoan Tetrahymena pyriformis. There was a linear relationship between FAM and colony counts of M. avium cells both from cultures and within protozoa. The Ziehl-Neelsen acid-fast stain could not be used to enumerate intracellular mycobacteria because uninfected protozoa contained acid-fast, bacterium-like particles. Starved, 7-day-old cultures of T. pyriformis transferred into fresh medium readily phagocytized M. avium, M. intracellulare, and M. scrofulaceum. Phagocytosis was rapid and reached a maximum in 30 min. M. avium, M. intracellulare, and M. scrofulaceum grew within T. pyriformis, increasing by factors of 4- to 40-fold after 5 days at 30 degrees C. Intracellular M. avium numbers remained constant over a 25-day period of growth (by transfer) of T. pyriformis. Intracellular M. avium cells also survived protozoan encystment and germination. The growth and viability of T. pyriformis were not affected by mycobacterial infection. The results suggest that free-living phagocytic protozoa may be natural hosts and reservoirs for M. avium, M. intracellulare, and M. scrofulaceum.

Animals↗

PubMed2003

Factors influencing the chlorine susceptibility of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum.

The susceptibility of representative strains of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum (the MAIS group) to chlorine was studied to identify factors related to culture conditions and growth phase that influenced susceptibility. M. avium and M. intracellulare strains were more resistant to chlorine than were strains of M. scrofulaceum. Transparent and unpigmented colony variants were more resistant to chlorine than were their isogenic opaque and pigmented variants (respectively). Depending on growth stage and growth rate, MAIS strains differed in their chlorine susceptibilities. Cells from strains of all three species growing in early log phase at the highest growth rates were more susceptible than cells in log and stationary phase. Rapidly growing cells were more susceptible to chlorine than slowly growing cells. The chlorine susceptibility of M. avium cells grown at 30 degrees C was increased when cells were exposed to chlorine at 40 degrees C compared to susceptibility after exposure at 30 degrees C. Cells of M. avium grown in 6% oxygen were significantly more chlorine susceptible than cells grown in air. Chlorine-resistant MAIS strains were more hydrophobic and resistant to Tween 80, para-nitrobenzoate, hydroxylamine, and nitrite than were the chlorine-sensitive strains.

Chlorine↗

PubMed1991

[Identification of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium tuberculosis complex by Gen-Probe Rapid Diagnostic System].

DNA probe testing for Mycobacterium avium, Mycobacterium intracellulare and Mycobacterium tuberculosis complex (MTC) was performed using Gen-Probe Rapid Diagnostic System (Gen-Probe Inc., San Diego, Calif., U.S.A.). By DNA probe test carried out blindfold for 48 mycobacterial strains with code numbers obtained from Kyoto University (Prof. F. Kuze), 13, 7, and 5 strains were identified as to be M. avium, M. intracellulare, and MTC, respectively. The diagnostic specificity and sensitivity of this testing were 100%. In this experiment, % hybridization of M. avium complex (MAC) and MTC were 25-55% and 45-52%, respectively. DNA probe test for 54 MTC strains including M. tuberculosis, M. bovis, M. africanum and M. microti revealed that 53 strains, except for one strain donated as a niacin-negative M. tuberculosis, reacted with MTC probe but not with MAC-probes. The one exceptional strain reacted with both the MTC- and M. avium-probes. However, when ten colonies randomly isolated from this strain on 7H11 agar plate were subjected to the DNA probe test again, all of these colonies reacted with M. avium probe, but not with MTC probe. Moreover, one representative colony was found to have alpha-antigen specific for the MAC.

DNA Probes↗

PubMed1993

Serovar determination and molecular taxonomic correlation in Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum: a cooperative study of the International Working Group on Mycobacterial Taxonomy.

A cooperative study was conducted by the International Working Group on Mycobacterial Taxonomy to correlate the agglutination serovar designations of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum strains with the species ascriptions of these organisms according to molecular criteria and cultural properties and to assess the reproducibility of serovar determinations for a set of 63 reference strains of these species. Among the molecular criteria, the level of agreement between results obtained with nucleic acid probes and T-catalase serology results was 94% for strains of M. avium and M. intracellulare. Nucleic acid probes were not available for M. scrofulaceum, but none of the 10 strains ascribed to this species on the basis of catalase serology data reacted with a nucleic acid probe for M. avium or M. intracellulare. Ascription to a species on the basis of mycolic acid high-performance liquid chromatography patterns was in agreement with catalase serology results in 86% of the cases examined. Most strains belonging to serovars 1 through 6 and 8 through 11 were identified by molecular criteria as M. avium, most strains belonging to serovars 7, 12 through 20, 23, and 25 were identified as M. intracellulare, and most strains belonging to serovars 41 through 43 were identified as M. scrofulaceum, in agreement with common current practice. Evidence for assigning serovar 27 to M. scrofulaceum was obtained. However, two strains of a given serovar may, on occasion, be placed in different species. The dominant species assignments for strains belonging to serovars 21, 24, 26, and 28 remain unresolved.(ABSTRACT TRUNCATED AT 250 WORDS)

Agglutination Tests↗

PubMed1990

Analysis of cellular fatty acids and proteins by capillary gas chromatography and sodium dodecyl sulphate polyacrylamide gel electrophoresis to differentiate Mycobacterium avium, Mycobacterium intracellulare and Mycobacterium scrofulaceum (MAIS) complex species.

Infections due to atypical mycobacteria have increased during the past 30 years. Species of Mycobacterium avium, Mycobacterium intracellulare and Mycobacterium scrofulaceum are among the most common non-tuberculous mycobacteria isolated from patients with AIDS or immunosuppressed. These three organisms are taxonomically closely related and identification, according to cultural characteristics and biochemical tests, is not always evident, so some of these related strains are grouped in a "MAIS" complex. Analysis of cellular constituents is an aid to identification. Gas chromatography was used to study mycolic acids and a secondary alcohol was found which is a discriminating constituent between M. scrofulaceum and the other two species. The lipidic analysis was not able to separate M. avium and M. intracellulare, so cell proteins were considered. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of proteins reflects genetic relatedness between strains; the different patterns obtained from these three species are described and it is shown that this method is very useful in classification and epidemiology.

Bacterial Proteins↗

PubMed1992

Epidemiology of infection by nontuberculous mycobacteria. Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum in acid, brown-water swamps of the southeastern United States and their association with environmental variables.

Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum (MAIS) organisms were isolated and identified from waters, soils, aerosols, and droplets ejected from water collected from four geographically separate aquatic environments (Okefenokee Swamp, GA; Dismal Swamp, VA; Claytor Lake, VA; and Cranberry Glades, WV) during several seasons. Recovery of MAIS was significantly higher from waters, soils, and aerosols collected from the two acid, brown-water swamps located in the southeastern coastal plain. High MAIS numbers correlated with warmer temperature, low pH, low dissolved oxygen, high soluble zinc, high humic acid, and high fulvic acid. This research, in relation to previous findings for the geographic distribution and physiologic ecology of MAIS, supports the conclusion that waters, soils, and aerosols of the acid, brown-water swamps of the southeastern United States coastal plain represent major environmental sources likely connected with the higher incidence of human infection in this region.

Georgia↗

PubMed1999

Purification and inhibition by quinolones of DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum.

The DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum, which are species naturally resistant, moderately susceptible and susceptible to fluoroquinolones, respectively, were purified by affinity chromatography on novobiocin-Sepharose columns. The DNA gyrase inhibiting activities (IC50 values) of classical quinolones and fluoroquinolones were determined from the purified enzymes and were compared to the corresponding antibacterial activities (MICs). Regarding M. fortuitum bv. peregrinum, which is nearly as susceptible as Escherichia coli, the corresponding MIC and IC50 values of quinolones were significantly lower than those found for M. avium and M. smegmatis (e.g. for ofloxacin, MICs of 0.25 versus 32 and 1 microg ml(-1), and IC50 values of 1 versus 8 and 6 microg ml(-1), respectively). Such a result could be related to the presence of Ser-83 in the quinolone-resistance-determining region of the gyrase A subunit of M. fortuitum bv. peregrinum, as found in wild-type E. coli, instead of Ala-83 in M. avium and M. smegmatis, as found in fluoroquinolone-resistant E. coli mutants. The IC50 values of quinolones against the M. avium and M. smegmatis DNA gyrases were similar, while the corresponding MICs were 32-fold higher for M. avium when compared to M. smegmatis, suggesting that an additional mechanism, such as a low cell wall permeability or a drug efflux, could contribute to the low antibacterial potency of quinolones against M. avium.

4-Quinolones↗

PubMed1987

In vitro susceptibility of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium avium, Mycobacterium fortuitum, and Mycobacterium chelonae to ticarcillin in combination with clavulanic acid.

The in vitro susceptibility of Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium africanum, Mycobacterium avium, Mycobacterium fortuitum, and Mycobacterium chelonae (M. chelonei) to ticarcillin in combination with calvulanic acid (CA) was studied by the agar dilution method. All the M. tuberculosis, M. bovis, and M. africanum strains were inhibited at a ticarcillin concentration of 32 micrograms/ml or lower in combination with 5 micrograms of CA. M. chelonae and M. avium strains proved resistant to more than 128 micrograms of ticarcillin plus 5 micrograms of CA per ml. M. fortuitum strains needed 128 micrograms of ticarcillin plus 5 micrograms of CA to inhibit approximately 30% of the isolates.

Clavulanic Acid↗

PubMed1992

A new glycolipid from Mycobacterium avium--Mycobacterium intracellulare complex.

From a nonpolar lipid fraction of Mycobacterium avium--Mycobacterium intracellulare complex cell mass, a new glycolipid was obtained, which was shown to be 5-mycoloyl-beta-arabinofuranosyl-(1-->2)-5-mycoloyl-alpha-ar abinofuranosyl- (1-->1')-glycerol. When examined by TLC, all the 12 strains of this species tested, including clinical isolates, were found to contain this glycolipid. But the glycolipid was not detected in Mycobacterium bovis BCG or Mycobacterium tubrculosis H37Rv.

Chromatography, Thin Layer↗

PubMed1986

Plasmid DNA profiles as epidemiological markers for clinical and environmental isolates of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum.

Plasmid DNA was isolated, and profiles of a variety of clinical and environmental isolates of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum (MAIS) were compared to determine whether plasmid DNA content would be useful as an epidemiological marker for these environmental pathogens. Since plasmids are common in clinical isolates and are stable during culture and exposure to NaOH, plasmid DNA analysis appears to be a suitable epidemiological tool. Based on the high frequency of plasmids in only clinical (56%) and aerosol (75%) isolates and low frequencies in soil (5%), dust (7%), sediment (less than 6%), and water (25%) isolates, the data suggest that MAIS-laden aerosols generated over waters of the southeastern United States are a likely source of human infection.

Aerosols↗

PubMed1984

Induction of bacteriophage from members of the Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum serocomplex.

Bacteriophages have been induced from strains in the Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum serocomplex by exposure of cultures to UV light or treatment with mitomycin C. One-sixth of the strains examined, representing all but one of the 31 authenticated serotypes, were found to possess phages lytic for a Mycobacterium smegmatis indicator strain. Four single-plaque isolated phages, TM4, TM9, TM10 and TM20, were purified and shown to have a similar morphology on electron micrographs. They had an isometric head of diameter 50-58 nm and a flexible non-contractile tail about 170 nm in length with a terminal bulb. All had an identical buoyant density in CsCl of 1.521 g cm-3 and extreme sensitivity to chloroform. The induced phages differed in host range and possessed the unique ability to lyse other members of the serocomplex. Interest in these phages centres on a possible role in mediating genetic interrelationships between members of the serocomplex.

Lysogeny↗

PubMed1987

[Clinical trial of forphenicinol for the lung infection with Mycobacterium avium, Mycobacterium intracellulare complex].

Clinical evaluation of forphenicinol, a low molecular weight immunomodulator, in patients with Mycobacterium avium, Mycobacterium intracellulare complex pulmonary infections has been conducted in a multicenter trial participated by 9 institutions in Kyushu during a period of 12 months from July 1982 to August 1983. Forphenicinol was administered for 6 months without changing the regimen of antituberculous drugs used previously. The following results were obtained. 1. Ten out of 33 eligible patients were evaluated as showing good responses; In 5 of them, elimination of M. avium, M. intracellulare complex from sputum was observed. In the other 5 patients, number of bacilli excreted was decreased significantly after treatment. 2. All of the cases with good responses were those which were simultaneously administered with antituberculous drugs. 3. Patients with thick-walled cavities, following cured tuberculosis, which was superimposed with M. avium, M. intracellulare complex, were poorly responded to forphenicinol. 4. Side effects were observed in 3 out of 41 patients; 1 case each with fever, abdominal distension and anorexia. No abnormalities in laboratory test values were observed.

Adult↗

PubMed2001

Factors influencing numbers of Mycobacterium avium, Mycobacterium intracellulare, and other Mycobacteria in drinking water distribution systems.

Eight water distribution systems were sampled over an 18-month period (528 water and 55 biofilm samples) to measure the frequency of recovery and number of mycobacteria, particularly Mycobacterium avium and Mycobacterium intracellulare, in raw source waters before and after treatment and within the distribution system. The systems were chosen to assess the influence of source water, treatment, and assimilable organic carbon levels on mycobacterial numbers. Overall, mycobacterial recovery from the systems was low (15% of samples). Numbers of mycobacteria ranged from 10 to 700,000 CFU liter(-1). The number of M. avium in raw waters was correlated with turbidity. Water treatment substantially reduced the number of mycobacteria in raw waters by 2 to 4 log units. Mycobacterial numbers were substantially higher in the distribution system samples (average, 25,000-fold) than in those collected immediately downstream from the treatment facilities, indicating that mycobacteria grow in the distribution system. The increase in mycobacterial numbers was correlated with assimilable organic carbon and biodegradable organic carbon levels (r(2) = 0.65, P = 0.03). Although M. intracellulare was seldom recovered from water samples, it was frequently recovered (six of eight systems) in high numbers from biofilms (average, 600 CFU/cm(2)). Evidently, the ecological niches of M. avium and M. intracellulare are distinct.

Biofilms↗

PubMed2005

Elemental analysis of Mycobacterium avium-, Mycobacterium tuberculosis-, and Mycobacterium smegmatis-containing phagosomes indicates pathogen-induced microenvironments within the host cell's endosomal system.

Mycobacterium avium and Mycobacterium tuberculosis are human pathogens that infect and replicate within macrophages. Both organisms live in phagosomes that fail to fuse with lysosomes and have adapted their lifestyle to accommodate the changing environment within the endosomal system. Among the many environmental factors that could influence expression of bacterial genes are the concentrations of single elements within the phagosomes. We used a novel hard x-ray microprobe with suboptical spatial resolution to analyze characteristic x-ray fluorescence of 10 single elements inside phagosomes of macrophages infected with M. tuberculosis and M. avium or with avirulent M. smegmatis. The iron concentration decreased over time in phagosomes of macrophages infected with Mycobacterium smegmatis but increased in those infected with pathogenic mycobacteria. Autoradiography of infected macrophages incubated with (59)Fe-loaded transferrin demonstrated that the bacteria could acquire iron delivered via the endocytic route, confirming the results obtained in the x-ray microscopy. In addition, the concentrations of chlorine, calcium, potassium, manganese, copper, and zinc were shown to differ between the vacuole of pathogenic mycobacteria and M. smegmatis. Differences in the concentration of several elements between M. avium and M. tuberculosis vacuoles were also observed. Activation of macrophages with recombinant IFN-gamma or TNF-alpha before infection altered the concentrations of elements in the phagosome, which was not observed in cells activated following infection. Siderophore knockout M. tuberculosis vacuoles exhibited retarded acquisition of iron compared with phagosomes with wild-type M. tuberculosis. This is a unique approach to define the environmental conditions within the pathogen-containing compartment.

Animals↗

PubMed1988

Relationship between Mycobacterium avium, Mycobacterium paratuberculosis and "wood pigeon mycobacteria". Determinations by DNA-DNA hybridization.

The DNA-DNA homology percentages obtained in this study indicate that M. avium and M. paratuberculosis belong to one species. Consequently, M. paratuberculosis ought to be considered a variant of M. avium, and the following designations are proposed: Mycobacterium avium, subsp. avium. Mycobacterium avium, subsp. paratuberculosis. Identification and classification of "wood pigeon mycobacteria" occurring in wild animals have been problematic due to their dysgonic and mycobactin-dependent growth. DNA-DNA homology percentages indicate that these bacteria are closely related to reference strains both of M. avium and of M. paratuberculosis. "Wood pigeon mycobacteria" should therefore be classified as atypical strains of M. avium, and the following designation is proposed: Mycobacterium avium subsp. columbae.

Animals↗

PubMed1991

Antigens of the Mycobacterium avium, Mycobacterium intracellulare complex.

In the past decade, the clinical significance of the Mycobacterium avium, Myobacterium intracellulare complex (MAC) has increased dramatically primarily because of the association between the MAC and the acquired immunodeficiency syndrome (AIDS). Recent hospital reports have suggested that about one-half of AIDS patients in the United States are infected with the MAC. The resulting myobacteremia is a primary cause of mortality in 5-10% of these patients. This increased clinical importance of the MAC has generated renewed interest in MAC immunobiology. In this review, recent immunological and biochemical characterizations of four classes of dominant myobacterial antigens - glycopeptidolipids, arabinogalactan, lipoarabinomannan and MAC proteins - is examined. In addition, future prospects for improved diagnosis of MAC disease using defined monospecific antigens is discussed.

Acquired Immunodeficiency Syndrome↗

PubMed1990

Epidemiology of infection by nontuberculous mycobacteria IX. Evidence for two DNA homology groups among small plasmids in Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum.

A 12.9 kb plasmid, pVT2, from a clinical Mycobacterium avium isolate, MD1, was cloned and radiolabeled for use as a DNA probe to examine the relatedness of plasmids in M. avium complex. That probe hybridized with plasmids isolated from M. avium complex strains from the environment (7 of 16) and from non-acquired immunodeficiency syndrome (AIDS) (10 of 17) and AIDS (5 of 6) clinical isolates. The similarity of plasmids from the environment with those from patients supports the hypothesis that the environment is a source of human M. avium complex infection. More striking was the observation that pVT2 hybridized with every plasmid (13 of 13 clinical and 5 of 5 environmental isolates) of 13.5 kb or smaller. A second probe, consisting of a 15.3 kb plasmid (pLR7) from another clinical isolate of the M. avium complex, hybridized with plasmids of 15.3 to 25 kb from environmental and clinical (AIDS and non-AIDS) isolates. There was no hybridization between pVT2 and pLR7. Thus, these two probes define two different groups of small mycobacterial plasmids.

Acquired Immunodeficiency Syndrome↗