Rapid and simple approach for identification of Mycobacterium tuberculosis complex isolates by PCR-based genomic deletion analysis

Stewart Cole
2002
Article

Résumé

Although the virulences and host ranges differ among members of the Mycobacterium tuberculosis complex (TBC; M. tuberculosis, M. africanum, M. canettii, M. microti, M. bovis, and M. bovis BCG), commercially available molecular assays cannot differentiate these organisms because of the genetic identities of their 16S rRNA gene sequences. Comparative genomic analyses with the complete DNA sequence of M. tuberculosis H37Rv has provided information on regions of difference (RD 1 to RD 16) deleted in members of the TBC other than M. tuberculosis. To determine whether deletion analysis could accurately differentiate members of TBC, we used PCR to assess the presence or absence of specific regions of the genome in 88 well-characterized isolates of M. tuberculosis, M. africanum, M. microti, M. bovis, and M. bovis BCG. The identifications obtained by use of the specific deletion profiles correlated 100% with the original identifications for all TBC members except M. africanum, but further characterization resulted in profiles specific for all members. Although six RD regions were used in the analyses with the original 88 isolates, it was found that the use of RD 1, RD 9, and RD 10 was sufficient for initial screenings, followed by the use of RD 3, RD 5, and RD 11 if the results for any of the first three regions were negative. When 605 sequential clinical isolates were screened, 578 (96%) were identified as M. tuberculosis, 6 (1%) were identified as M. africanum, 8 (1%) were identified as M. bovis, and 13 (2%) were identified as M. bovis BCG. Since PCR-based assays can be implemented in most clinical mycobacteriology laboratories, this approach provides a rapid and simple means for the differentiation of members of TBC, especially M. bovis and M. tuberculosis, when it is important to distinguish between zoonotic sources (i.e., cattle and unpasteurized dairy products) and human sources of tuberculosis disease.

Official source

https://infoscience.epfl.ch/record/151252?ln=fr

À propos de ce résultat

Cette page est générée automatiquement et peut contenir des informations qui ne sont pas correctes, complètes, à jour ou pertinentes par rapport à votre recherche. Il en va de même pour toutes les autres pages de ce site. Veillez à vérifier les informations auprès des sources officielles de l'EPFL.

Concepts associés

Chargement

Publications associées

Chargement

Stewart Cole
2002
Article

Résumé

Official source

https://infoscience.epfl.ch/record/151252?ln=fr

À propos de ce résultat

Concepts associés (10)

Concepts associés

Chargement

Publications associées (104)

Publications associées

Chargement

Mycobacterial genome structure

Stewart Cole

Genome maps have been constructed for the mycobacterial pathogens Mycobacterium leprae and Mycobacterium tuberculosis, as well as for the attenuated vaccine strain Mycobacterium bovis BCG Pasteur. While the chromosomes of M. tuberculosis and M. bovis BCG Pasteur show extensive conservation at the gross level, comparison with M. leprae revealed a high degree of diversification, with a mosaic-like pattern apparent. The ordered libraries of M. tuberculosis and M. leprae produced during the course of these studies played a central role in the genome sequencing projects of these two bacilli, showing the utility of this approach for systematic sequencing of bacterial genomes.

1998

Chargement

In tuberculosis, it is often important to establish the source of infection and to determine whether disease is due to a new strain of Mycobacterium tuberculosis or to relapse. To cope with the resurgence of tuberculosis and atypical mycobacterioses in AIDS patients, on the one hand, and to overcome the limitations of classical bacteriological procedures on the other, the development of rapid, sensitive, and reliable diagnostic and epidemiologic tools is highly desirable. Molecular typing methods are often based on repeated genes such as those for rRNA. Ribotyping is of limited use with pathogenic mycobacteria, as the slow-growers possess a single rRNA operon, while the fast-growers have two. This problem has been overcome by the discovery and study of repeated DNA elements in mycobacterial genomes, as these provide an alternative pathway for diagnostic and epidemiological investigations.

1995

Chargement

Comparative genomics of Esx genes from clinical isolates of Mycobacterium tuberculosis provides evidence for gene conversion and epitope variation

Stewart Cole, Jacques Rougemont, Swapna Uplekar

The 23-membered Esx protein family is involved in the host-pathogen interactions of Mycobacterium tuberculosis. These secreted proteins are among the most immunodominant antigens recognized by the human immune system and have thus been used to develop vaccines and immunodiagnostic tests for tuberculosis (TB). Gene pairs for 10 Esx proteins are contained in the ESX-1 to ESX-5 loci, encoding type VII secretion systems. A subset of Esx proteins can be further classified into the Mtb9.9, QILSS, and TB10.4 subfamilies. To survey genetic diversity in the Esx family and its potential for antigenic variation, we sequenced all esx genes from 108 clinical isolates of M. tuberculosis from different clades by using a targeted approach. A total of 109 unique single nucleotide polymorphisms (SNPs) were observed, and 59 of these were nonsynonymous. Some of the resultant amino acid substitutions affect known Esx epitopes, including two in the EsxB (CFP-10) and EsxH (TB10.4) antigens. Assessment of the SNP distribution across the Esx proteins revealed high genetic variability, especially in the Mtb9.9 and QILSS subfamilies, and more conservation in the ESX-1 to ESX-4 loci. Comparison of the DNA sequences of variable esx genes provided clear evidence for recombination events between different genes in the same strain, some of which are predicted to truncate the corresponding protein. Many of these polymorphisms escape detection by ultrahigh-throughput sequencing using short sequence reads, as such approaches cannot distinguish between closely related genes. The esx gene family is dynamic, and sequence changes likely lead to immune variation.

2011

Chargement