Identification of 54 Mycobacterial Species by PCR-Restriction Fragment Length Polymorphism Analysis of the hsp65 Gene

PCR-restriction fragment length polymorphism analysis (PRA) of thehsp65 gene present in all mycobacteria was used in the present investigation to characterize Mycobacterium leprae. Bacilli were extracted and purified from different organs from experimentally infected armadillos and nude mice (Swiss mice ofnu/nu origin). A total of 15 samples were assayed in duplicate, and the results were compared with those obtained for a total of 147 cultivable mycobacteria representing 34 species. Irrespective of its origin or viability, M. leprae strains from all the samples were uniformly characterized by two fragments of 315 and 135 bp upon BstEII digestion and two fragments of 265 and 130 bp upon HaeIII digestion. PRA is a relatively simple method and permits the conclusive identification of M. leprae to the species level.

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Rapid identification of mycobacteria to species level by PCR-restriction fragment length polymorphism analysis of the hsp65 gene and proposition of an algorithm to differentiate 34 mycobacterial species.

Journal of Clinical Microbiology ◽

10.1128/jcm.35.11.2969-2973.1997 ◽

1997 ◽

Vol 35 (11) ◽

pp. 2969-2973 ◽

Cited By ~ 119

Author(s):

A Devallois ◽

K S Goh ◽

N Rastogi

Keyword(s):

Restriction Fragment Length Polymorphism ◽

Length Polymorphism ◽

Fragment Length ◽

Fragment Length Polymorphism ◽

Rapid Identification ◽

Species Level ◽

Polymorphism Analysis ◽

Mycobacterial Species ◽

Hsp65 Gene ◽

Restriction Fragment Length

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Species Identification of Mycobacteria by PCR-Restriction Fragment Length Polymorphism of therpoB Gene

Journal of Clinical Microbiology ◽

10.1128/jcm.38.8.2966-2971.2000 ◽

2000 ◽

Vol 38 (8) ◽

pp. 2966-2971 ◽

Cited By ~ 140

Author(s):

Hyeyoung Lee ◽

Hee-Jung Park ◽

Sang-Nae Cho ◽

Gill-Han Bai ◽

Sang-Jae Kim

Keyword(s):

Restriction Fragment Length Polymorphism ◽

Restriction Fragment ◽

Length Polymorphism ◽

Fragment Length ◽

Fragment Length Polymorphism ◽

Species Level ◽

Polymorphism Analysis ◽

Mycobacterial Species ◽

Bacterial Strains ◽

Restriction Fragment Length

PCR-restriction fragment length polymorphism analysis (PRA) using the novel region of the rpoB gene was developed for rapid and precise identification of mycobacteria to the species level. A total of 50 mycobacterial reference strains and 3 related bacterial strains were used to amplify the 360-bp region of rpoB, and the amplified DNAs were subsequently digested with restriction enzymes such as MspI and HaeIII. The results from this study clearly show that most of the mycobacterial species were easily differentiated at the species level by this PRA method. In addition, species with several subtypes, such as Mycobacterium gordonae, M. kansasii, M. celatum, andM. fortuitum, were also differentiated by this PRA method. Subsequently, an algorithm was constructed based on the results, and a blinded test was carried out with more than 260 clinical isolates that had been identified on the basis of conventional tests. Comparison of these two sets of results clearly indicates that this new PRA method based on the rpoB gene is more simple, more rapid, and more accurate than conventional procedures for differentiating mycobacterial species.

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