Distribution of surface-exposed antigenic glycolipids in recent clinical isolates of Mycobacterium tuberculosis.

The distribution of surface-exposed antigenic glycolipids in seven recent clinical isolates of Mycobacterium tuberculosis was established. Thin-layer and liquid chromatographies revealed a uniformity in the glycolipid pattern. Chemical analysis of the individual glycolipids of a selected strain enabled the identification of glycolipids of serological interest in all the other clinical isolates. Phenolic glycolipid-Tb1 (PGL-Tb1) was lacking in all strains, but appreciable amounts of a partially deglycosylated version (PGL-Tb1D) were present in the seven isolates. Diacyltrehaloses (DATs) were detected in all strains, showing themselves to be major glycolipids. Lipooligosaccharides (LOS-II) were present in the seven strains studied though only in trace amounts. These results shed new light on the open debate on the distribution of these interesting glycolipids in typical clinical isolates of M. tuberculosis. In the search for a serological test for tuberculosis, and in accordance with our observations, we believe that PGL-Tb1 and LOS-II should not be the target molecules for serology and that it is worthwhile to continue investigating the value of DATs as antigens. We also believe that it would be of interest to undertake research to assess the usefulness of PGL-Tb1D as an antigen.

Lipid composition of leprosy-derived corynebacteria, a distinct group of corynebacteria, and of a reference Corynebacterium.

Leprosy-derived corynebacteria (LDC) are diphtheroid organisms isolated from leprosy patients and previously characterized by DNA and cell wall analysis. Three groups of LDC components of taxonomic value, glycolipids, and phospholipids and cell-wall-bound lipids were analyzed in comparison with those of a reference strain C. hoffmannii (CH). The main CH glycolipid, "cord factor" (trehalose dimycolate), was missing from LDC. Among phospholipids, phosphatidylinositol and phosphatidylglycerol had lowered proportions in LDC, as compared to CH, whereas phosphatidylethanolamine and cardiolipin were absent from both microorganisms. Bound lipids in acidic extracts of delipidated LDC yielded arabinose corynomycolate in lesser quantity with respect to CH. Alkaline hydrolysis of whole cells released fatty acids and mycolic acids, which were analyzed by gas chromatography/mass spectrometry. Reference CH, grown in the absence of serum, yielded C16:0 and C18:1 (major) and C18:0 (minor) fatty acids, as well as C32, C34, and C36 corynomycolic acids. All these components, particularly mycolates, had lowered proportions when this organism was grown in the presence of serum. Dominant LDC components were, in addition to C16:0, C18:0, and CI8:u fatty acids, cholesterol from serum. Very low concentrations of corynomycolic acids with a high degree of unsaturation were found in these organisms, suggesting a dependence of lipid metabolism on growth conditions. The presence in LDC of tuberculostearic acid (C19r:0), a mycobacterial component found in some pathogenic corynebacteria, was carefully explored: Traces of C19r:0 were found in LDC 19 grown in the presence of delipidated serum, but not in LDC 15 nor in C. hoffmannii. Present data, in conjunction with previous studies on DNA and mycolic acids, disclose basic differences in the composition of LDC and conventional corynebacteria.

Diglycosyl phenol phthiocerol diester of Mycobacterium leprae.

A diglycosyl phenol phthiocerol diester that had not been previously detected was isolated from M. leprae-infected armadillo tissues. Spectroscopy methods allowed the elucidation of its structure. The diglycoside was a 2,3-di-O-methylrhamnopyranosyl (alpha 1----2)3-O-methylrhamnopyranosyl (alpha 1-linked to the phenolic hydroxyl of phthiocerol dimycocerosates). It differs from the major phenolic glycolipid (PGL I) only by the absence of the terminal 3,6-di-O-methylglucopyranosyl unit. The diglycoside could be an intermediate in the synthesis of the latter antigen or a degradative product in the host detoxification process.

Distribution of phthiocerol diester, phenolic mycosides and related compounds in mycobacteria.

Among 28 mycobacterial species studied, only Mycobacterium tuberculosis, M. bovis, M. africanum, M. marinum, M. kansasii, M. gastri and M. ulcerans produced waxes yielding long-chain beta-diol components (called phthiocerol and companions) and polymethyl-branched fatty acids on saponification. The same mycobacterial species also produced diesters of phenol phthiocerol and companions. Fatty acids esterifying these fatty alcohols in M. marinum and M. ulcerans were found to belong to the phthioceranic series (dextrorotatory fatty acids), in contrast to those of the other species (laevorotatory fatty acids called mycocerosic acids), both groups having the same chain length and methyl-branched positions. M. kansasii and M. gastri contained the same waxes with identical structures, as did M. tuberculosis, M. bovis and M. africanum. Neither the type strain of M. tuberculosis, nor that of M. bovis or M. marinum accumulated the strain-specific phenolic glycolipids.

Structure of the major triglycosyl phenol-phthiocerol of Mycobacterium tuberculosis (strain Canetti).

Phenol-phthiocerol glycolipids have been found previously in Mycobacterium leprae, M. kansasii, M. bovis and M. marinum, but not in M. tuberculosis. A search for glycolipids in this latter species showed that the Canetti strains of M. tuberculosis synthesize a major triglycosyl phenol-phthiocerol, accompanied by minor amounts of other glycolipids with a similar aglycone moiety. The triglycoside moiety has the following structure: 2,3,4-tri-O-methyl L-fucopyranosyl(alpha 1----3)L-rhamnopyranosyl(alpha 1----3)2-O-methyl L-rhamnopyranosyl(alpha 1-. The aglycone moiety consists in phenol-phthiocerol (two homologs). Its two secondary alcohol functions are esterified by mycocerosic acids (homologs with 26-32 carbon atoms and with 2-4 methyl branches). The proposed structure differs on several points from the M. leprae glycolipids, but presents some analogy with the major glycolipid of M. kansasii. A minor monoglycosyl phenol-phthiocerol was also studied. Its overall structure is very similar to that of M. bovis, with 2-O-methyl rhamnose as sugar moiety.

Description of Corynebacterium tuberculostearicum sp. nov., a leprosy-derived Corynebacterium.

Leprosy-derived corynebacteria (LDC) have been extensively studied over the past decade. A composite of their biological properties (cell morphology, staining reactions, cellular inclusions and guanine-plus-cytosine content of their deoxyribonucleic acid; 16 strains studied) and their chemical structures (peptidoglycan type, major cell wall polysaccharide, major glycolipid as well as characteristic mycolic acids) appears to define them as members of the genus Corynebacterium. In relation to other corynebacteria found in humans, including "JK corynebacteria", they seem to be distinct. They are here named Corynebacterium tuberculostearicum sp. nov. because they produce a 10-methyloctadecanoic (tuberculostearic) acid (8 strains studied). This and some of their other attributes are considered in relation to properties of leprosy bacilli and Mycobacterium leprae.

Lipids as taxonomic markers for bacteria derived from leprosy infections.

Lipid analysis allows the specific detection of M. leprae among various other bacteria isolated from leprosy lesions. In this report mycolates and glycolipid compositions were used for such a discrimination. Comparative studies of the lipid composition of tissue fragments from different organs of experimentally infected armadillos, and of cultivable strains isolated from these tissues showed that the last ones did not multiply extensively in the tissues of the animals.

[Lipid composition of cultivable Mycobacteria isolated from livers of armadillos infected by Mycobacterium leprae]. / Etude de la composition lipidique de mycobactéries cultivables isolées de foies de tatous infectés par Mycobacterium leprae.

Four bacterial strains isolated from two livers of armadillos experimentally infected with Mycobacterium leprae were studied. Lipids obtained after saponification and methylation and complex lipids obtained by solvent extraction were examined. The presence of mycolates showed that the four strains belonged to the genus Mycobacterium, but the mycolate patterns, identical for the four strains, were different from those of all strains studied so far. Three of these strains contained phthioceranic acids, which were not found in the fourth one. Only the last strain contained mycosides of the C type, while the three others contained a new type of glycolipid. Their content in mycolates and in glycolipids demonstrated a clear-cut difference between these strains, on the one hand, and M. leprae on the other.

[Specific lipids from Mycobacterium ulcerans]. / Lipides spécifiques de Mycobacterium ulcerans.

The main lipids synthesized by Mycobacterium ulcerans are specific for the species. Three products were isolated by chromatography. Their structures were determined by means of spectrographic methods performed on the natural substances or on their split products. The most abundant products were phthiodiolone diphthioceranate and phenolphthiodiolone diphthioceranate . These structures have some analogies with those of phthiocerol dimycocerosate synthesized by M. tuberculosis and M. bovis, and with those of phenolphthiocerol mycocerosate synthesized by M. bovis. The reverse configuration of the polymethyl-branched-chain fatty acids isolated from the substances, according to their origin, remains to be pointed out. Little attention has generally been paid to the stereochemistry of such molecules. We verified that the branched-chain fatty acids found in diacyl phthiocerol and in the mycoside of M. leprae have the same configuration as in the analogous molecules isolated from M. tuberculosis or M. bovis, contrary to M. ulcerans. Another peculiarity of phenolphthiodiolone isolated from M. ulcerans is the occurrence of the phenol group in free form.

Taxonomy of mycobacterial strains isolated from the tissues of leprosy patients.

Thirty-six slowly growing mycobacteria isolated from the tissues of leprosy patients were studied using 40 characteristics as well as susceptibility to 27 distinct mycobacteriophages. The composition in mycolic acids of selected strains was also studied. According to the data, the strains formed 5 clusters. Some of the clusters were possibly as yet undescribed species; however, comparison of the data with the known properties of Mycobacterium leprae leads to the conclusion that none of the strains were identical to the leprosy bacillus.

[Studies on the lipids of "Mycobacterium gordonae" in comparison with those of "M. leprae" and of some other scotochromogenic mycobacteria (author's transl)]. / Etude des lipides de Mycobacterium gordonae comparativement à ceux de M. leprae et de quelques mycobactéries scotochromogènes.

The mycolic acids were isolated from Mycobacterium gordonae (strain ATCC 14470), and purified by thin layer chromatography. Three species were studied by mass spectrometry. The analogy between M. gordonae and M. leprae, based on the lack of tuberculostearic acid, was supported by the comparison of the structures of their mycolic acids. Succinct analyses of the lipids of three other scotochromogenic strains of mycobacteria, using thin layer chromatography and gas-liquid chromatography, were performed. These strains were more remote from M. gordonae than is M. leprae, as far as the lipid content is concerned.

[Lipidic constituents of "Mycobacterium leprae" isolated from experimentally infected armadillo (author's transl)]. / Constituants lipidiques de Mycobacterium leprae isolé de tatou infecté expérimentalement.

Mycobacterium leprae (obtained from experimentally infected armadillo) was submitted to saponification. The liposoluble part was methylated and fractionated by chromatographic methods. Each fraction was studied by gas-liquid chromatography. Cholesterol (from the infected host) and the main fatty acids were identified. Mycolic acids were isolated, and their structures determined, using mass spectrometry. These structures are useful to make a comparison of M. leprae with some other mycobacteria. Some of these comparisons are discussed here. The absence-or, at least, the very low level-of tuberculostearate suggests comparative studies of M. leprae and M. gordonae.

Chemical characterization of organisms isolated from leprosy patients.

CHEMICAL ANALYSES OF THE CELL WALLS OF ORGANISMS ISOLATED IN VARIOUS PARTS OF THE WORLD FROM CASES OF LEPROMATOUS AND TUBERCULOID LEPROSY MAKE POSSIBLE THEIR ASSIGNMENT TO ONE OF THE THREE GENERA: Corynebacterium, Mycobacterium, or Propionibacterium. One, bacterium 22M, remains unassigned. The combined chemical and enzymatic properties attributed to leprosy bacilli freshly harvested from lepromata are found collectively, but not individually, in these three genera.