Amplification of a species-specific DNA fragment of Mycobacterium tuberculosis and its possible use in diagnosis

Portillo, Patricia Del; Murillo, Luis A.; Patarroyo, Manuel E.

doi:10.1128/jcm.29.10.2163-2168.1991

Cited by 181 publications

(72 citation statements)

References 21 publications

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“…The specificity of the PCR amplification process lies in the choice of primers used. Numerous PCR-based assays for the detection and identification of individual mycobacterium species, such as M. tuberculosis, M. leprae, and M. avium, have been described (2,3,(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46)(47). Many target sequences have been used, but the most thoroughly evaluated assays target the M. tuberculosisspecific repeated DNA element IS 6110 (40).…”

Section: Approaches Based On Molecular Biologymentioning

confidence: 99%

Diagnosis of tuberculosis: Available technologies, limitations, and possibilities

Garg

Tiwari

et al. 2003

Clinical Laboratory Analysis

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Rapid diagnosis and treatment are important for preventing transmission of Mycobacterium tuberculosis. However, the diagnosis of tuberculosis continues to pose serious problems, mainly because of difficulties in differentiating between patients with active tuberculosis and those with healed lesions, normal mycobacterium boris BCG (Bacillus Calmette Guerin) vaccinated individuals, and unvaccinated Manteux positives. Physicians still rely on conventional methods such as Ziehl-Neelsen (ZN) staining, fluorochrome staining, sputum culture, gastric lavage, and other non-traditional methods. Although the tuberculin test has aided in the diagnosis of tuberculosis for more than 85 years, its interpretation is difficult because sensitization with nontuberculous mycobacteria leads to false-positive tests. There have been numerous unsuccessful attempts to develop clinically useful serodiagnostic kits for tuberculosis. A number of proteinaceous and nonprotein antigens (such as acyltrehaloses and phenolglycolipids) have been explored from time to time for the development of such assays but they have not proved to be clinically useful. It has been difficult to develop an ELISA utilizing a suitable antigen because M. tuberculosis shares a large number of antigenic proteins with other microorganisms that may or may not be pathogenic. With the advent of molecular biology techniques, there have been significant advances in nucleic acid-based amplification and hybridization, which are helping to rectify existing flaws in the diagnosis of tuberculosis. The detection of mycobacterial DNA in clinical samples by polymerase chain reaction (PCR) is a promising approach for the rapid diagnosis of tuberculous infection. However, the PCR results must be corrected for the presence of inhibitors as well as for DNA contamination. In the modern era of genetics, marked by proteomics and genomics, the day is not far off when DNA chip-based hybridization assays will instantly reveal mycobacterial infections.

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Section: Approaches Based On Molecular Biologymentioning

confidence: 99%

Diagnosis of tuberculosis: Available technologies, limitations, and possibilities

Garg

Tiwari

et al. 2003

Clinical Laboratory Analysis

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“…The available PCR techniques are based either on the amplification of common sequences able to identify mycobacterial DNA (12,30,33) or on the amplification of sequences present in the strains of the so-called M. tuberculosis complex, which includes the humanpathogenic species M. tuberculosis, Mycobacterium bovis, and Mycobacterium africanum (25,28). On the basis of the nucleotide sequence of the MTP40 species-specific protein of the tuberculosis bacillus (24), we have developed a PCR technique which specifically amplifies a 396-bp fragment from the M. tuberculosis genome (7). However, although these assays have been shown to be useful in the diagnosis of tuberculosis, some limitations are now apparent.…”

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confidence: 99%

Multiprimer PCR system for differential identification of mycobacteria in clinical samples

et al. 1996

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A novel multiprimer PCR method with the potential to identify mycobacteria in clinical samples is presented. The assay relies on the simultaneous amplification of three bacterial DNA genomic fragments by using different sets of oligonucleotide primers. The first set of primers amplifies a 506-bp fragment from the gene for the 32-kDa antigen of Mycobacterium tuberculosis, which is present in most of the species belonging to the genus Mycobacterium. The second set of primers amplifies a 984-bp fragment from the IS6110 insertion sequence of the bacteria belonging to the M. tuberculosis complex. The third set of primers, derived from an M. tuberculosis species-specific sequence named MTP40, amplifies a 396-bp genomic fragment. Thus, while the multiprimer system would render three amplification fragments from the M. tuberculosis genome and two fragments from the Mycobacterium bovis genome, a unique amplification fragment would be obtained from nontuberculous mycobacteria. The results obtained, using reference mycobacterial strains and typed clinical isolates, show that the multiprimer PCR method may be a rapid, sensitive, and specific tool for the differential identification of various mycobacterial strains in a single-step assay. This paper is dedicated to the memory of Francisco Martín Luengo. 324 on July 6, 2020 by guest http://jcm.asm.org/ Downloaded from 326 DEL PORTILLO ET AL.

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“…The genomic DNA from the mycobacterial species was isolated by the method of Portillo et al [14]. Plasmid DNA extraction from E. coli followed the alkaline lysis procedure [15].…”

Section: Dna Isolationmentioning

confidence: 99%

Identification and characterization of a gene encoding a 35-kDa protein from Mycobacterium avium subspecies paratuberculosis

Banasure

2001

FEMS Microbiology Letters

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Mycobacterium avium subspecies paratuberculosis is the causative agent of Johne's disease, a chronic enteritis in ruminants. A gene homologous to that of 35-kDa antigen of Mycobacterium leprae was cloned and sequenced from Mycobacterium paratuberculosis. The database searches revealed 82.79% and 95.67% similarities of its nucleotide sequence, with those of immunodominant 35-kDa protein of M. leprae and M. avium, respectively. ß

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Amplification of a species-specific DNA fragment of Mycobacterium tuberculosis and its possible use in diagnosis

Cited by 181 publications

References 21 publications

Diagnosis of tuberculosis: Available technologies, limitations, and possibilities

Diagnosis of tuberculosis: Available technologies, limitations, and possibilities

Multiprimer PCR system for differential identification of mycobacteria in clinical samples

Identification and characterization of a gene encoding a 35-kDa protein from Mycobacterium avium subspecies paratuberculosis

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