Towards a Phylogeny and Definition of Species at the Molecular Level within the Genus Mycobacterium

Abstract

were determined and compared. The sequence data were used to infer a phylogenetic tree, which provided the basis for a systematic phylogenetic analysis of the genus Mycobacterium. The groups of slow-and fast-growing mycobacteria could be differentiated as distinct entities. We found that M. simiae occupies phylogenetically an intermediate position between these two groups. The phylogenetic relatedness within the slow-growing species did not reflect the Runyon classification of photochromogenic, scotochromogenic, and nonchromogenic mycobacteria. In general, the phylogenetic units identified by using rRNA sequences confirmed the validity of phenotypically defined species; an exception was M. gastri, which was indistinguishable from M. kunsasii when this kind of analysis was used.Mycobacteria are aerobic, nonmobile bacteria that are characteristically acid fast. The property of acid fastness, which is due to waxy materials in cell walls, is particularly important for recognizing mycobacteria. Members of the genus Mycobacterium are widespread in nature and range from soil-dwelling saprophytes to pathogens of humans and animals (22, 34). A major descriptive division of mycobacteria is related to growth rate and pigmentation. On the basis of these criteria, the genus Mycobacterium has been divided into four groups. Group I consists of the photochromogenic (pigmented) species of slow growers; members of group I1 are scotochromogenic slow growers; group I11 contains the nonchromogenic slow growers; and group IV consists of rapid growers (defined as maturing in less than 1 week) (22,34).Taxonomic analysis of the genus Mycobacterium is complicated by the fact that a variety of specialized and complex tests must be used. Numerical taxonomic analysis, which requires that some dozens of characters be tested (e.g., enzymatic activity, growth, morphology, and drug susceptibility), is now being applied to circumscription of clusters and description of strains. Early on, the problems and difficulties of traditional taxonomy with respect to mycobacteria were recognized, prompting a number of investigators in the field to organize themselves into the International Working Group on Mycobacterial Taxonomy and to undertake a number of cooperative taxonomic studies (18, 29, Attempts to subdivide mycobacterial species by using immunological approaches, DNA composition, and similar characteristics (1-3,9,10) proved to be taxonomically useful but gave little phylogenetic information. The use of macromolecular comparisons to infer phylogenetic relationships is generally accepted and well established. Of the macromolecules used for phylogenetic analysis, the rRNAs, in particular 16s rRNA, have proven to be the most useful for establishing phylogenetic relationships because of their high information content, conservative nature, and universal dis-31-33).* Corresponding author. tribution (7,35). We recently developed a general procedure for the isolation and direct complete nucleotide determination of entire genes coding for 16s rRNA in w...

Comparison of Mycobacterium 23S rRNA Sequences by High-Temperature Reverse Transcription and PCR

“…Several of these organisms, including M. haemophilum, Mycobacterium shimoidei, and Mycobacterium ulcerans, are considered pathogenic for humans. We compared our results with the comprehensive overview provided by the 16s rRNA phylogenetic tree of the RDP, as well as with the results of specific analyses described previously (13,29,31,36). For the most part, our results corroborate the 16s rRNA tree data, although there are some exceptions.…”

“…Assuming that the extracted and reanalyzed positions of M. chelonae and its fellow fast-growing species in Fig. 2B are correct, this is the only example of a contradiction with the slow-fast phylogenetic split (31,36). The results of several independent approaches (16s rRNA, 23s rRNA, DNA hybridization) have placed M. farcinogenes with the fast growers, and slow growth appears to be a recently acquired trait (29).…”

“…Several important human pathogens, including Mycobacterium tuberculosis and Mycobacterium leprue, are members of the slowly growing group, as are a number of opportunistic pathogens (Table 1) (15). In recent reports workers have described methods for using nucleic acid sequences to identify various specific members of the genus Mycobacterium (4,5,20,21,29,31,37,38). However, not all known members of the genus have been examined; the organisms that have not been examined include some strains which have recently been shown to be problematic in probe-based assays (7,12,14).…”

Mycobacterium celatum sp. nov.

“…The M. celatum strains were more closely related to M. avium (Dice the slowly growing mycobacteria included in this study were found to be highly related, with similarity values of 94.7% or greater (Table 3). This degree of similarity has been noted previously (24,30). When they were compared with the previously described sequences of other Mycobacterium species, the rRNA sequences of the M. celatum strains were most similar to the Mycobacterium asiaticum sequence (average level of similarity, 97.5%).…”

Discrimination of members of the Mycobacterium avium complex by polymerase chain reaction

“…The 16S rRNA sequencing has been particularly helpful in clarifying the complex taxonomical status of the members of the M.avium-M.intracellulare complex. (12,13). Unfortunately, this methodology remains relatively cumbersome, with application limited to reference laboratories.…”