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Journal of Clinical Microbiology, October 2001, p. 3499-3504, Vol. 39, No. 10
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.10.3499-3504.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Phospholipase Region of Mycobacterium tuberculosis Is a Preferential Locus for IS6110 Transposition

Lucio Vera-Cabrera,1,* Marco A. Hernández-Vera,1 Oliverio Welsh,1 Wendy M. Johnson,2 and Jorge Castro-Garza3

Servicio de Dermatología, Hospital Universitario "José E. González,"1 and Centro de Investigación Biomédica del Noreste, IMSS,3 Monterrey, México, and Federal Laboratories for Health Canada, Winnipeg, Canada2

Received 10 November 2000/Returned for modification 17 February 2001/Accepted 27 July 2001

Enzymes with phospholipase C activity in Mycobacterium tuberculosis have been recently described. The three genes encoding these proteins, plcA, plcB, and plcC, are located at position 2351 of the genomic map of M. tuberculosis H37Rv and are arranged in tandem. We have previously described the presence of variations in the restriction fragment length polymorphism patterns of the plcA and plcB genes in M. tuberculosis clinical isolates. In the present work we investigated the origin of this polymorphism by sequence analysis of the phospholipase-encoding regions of 11 polymorphic M. tuberculosis clinical isolates. To do so, a long-PCR assay was used to amplify a 5,131-bp fragment that contains the plcA and plcB genes and part of the plcC gene. In the M. tuberculosis strains studied the production of an amplicon ~1,400 bp larger than anticipated was observed. Sequence analysis of the PCR products indicated the presence of a foreign sequence that corresponded to an IS6110 element. We observed insertion elements in the plcA, plcB, and plcC genes. One site in plcB had the highest incidence of transposition (5 out of 11 strains). In two strains the insertion element was found in plcA in the same nucleotide position. In all the cases, IS6110 was transposed in the same direction. The high level of transposition in the phospholipase region can lead to the excision of fragments of genomic DNA by recombination of neighboring IS6110 elements, as demonstrated by finding the deletion, in two strains, of a 2,837-bp fragment that included plcA and most of plcB. This can explain the negative results obtained by some authors when detecting the mtp40 sequence (plcA) by PCR. Given the high polymorphism in this region, the use of the mtp40 sequence as a genetic marker for M. tuberculosis sensu stricto is very restricted.

* Corresponding author. Mailing address: Servicio de Dermatología, Hospital Universitario "José E. González," Madero y Gonzalitos, Col. Mitras Centro, Monterrey, N.L., México. Phone: 011(528) 348-0383. Fax: 011(528) 348-4407. E-mail: luvera_99{at}yahoo.com.

Journal of Clinical Microbiology, October 2001, p. 3499-3504, Vol. 39, No. 10
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.10.3499-3504.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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