The ribosomal intergenic spacer region: a target for the PCR based diagnosis of tuberculosis

Glennon, M.; Smith, Terry J.; Cormican, Martin; Noone, David; Barry, T. N.; Maher, Majella; Dawson, M.; Gilmartin, J J; Gannon, Frank

doi:10.1016/0962-8479(94)90081-7

Cited by 26 publications

(14 citation statements)

References 18 publications

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“…However, other semi-conserved genes, like hsp65 (Devallois et al, 1997 ;Lea4 o et al, 1999 ;Swanson et al, 1997 ;Telenti et al, 1993) and rpoB (Kim et al, 2001), and the 16S-23S rDNA internally transcribed spacer (ITS) region (Abed et al, 1995 ;Barry et al, 1991 ;Glennon et al, 1994 ;Portaels et al, 1996 ;Roth et al, 1998) have been shown to be more discriminating. The 16S-23S rDNA ITS sequence can be used to distinguish between the species M. avium and M. intracellulare.…”

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confidence: 96%

Molecular evidence to support a proposal to reserve the designation Mycobacterium avium subsp. avium for bird-type isolates and 'M. avium subsp. hominissuis' for the human/porcine type of M. avium

Mijs¹

2002

International Journal of Systematic and Evolutionary Microbiology

193

180

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In an attempt to clarify the taxonomy of the Mycobacterium avium complex, the relationship between IS1245 RFLP, growth temperature, 16S rDNA signature sequences and the 16S-23S rDNA internally transcribed spacer (ITS) of 160 M. avium-complex isolates from different sources was investigated. All 70 isolates identified as M. avium by INNO-LiPA MYCOBACTERIA (Innogenetics, Belgium), a DNA probe test that targets the ITS, and by 16S rDNA analysis carried multiple copies of IS1245. Three isolates with multiple copies of IS1245 were identified by 16S rDNA analysis as Mycobacterium intracellulare and by LiPA as M. intracellulare (n = 1) and M. avium-intracellulare complex (n = 2). A dichotomy among the M. avium isolates was found on the basis of a C and a G signature nucleotide at position 228 of the 16S-23S rDNA spacer sequence, and this grouping was largely confirmed on the basis of similarities in IS1245 RFLPs. Strains with the characteristic three-band IS1245 'bird-type', as well as M. avium subsp. silvaticum or 'wood-pigeon' strains, invariably contained the C signature. A third characteristic that separated the M. avium bird-type isolates from M. avium isolates from humans and other mammals was growth-temperature tolerance: in contrast to bird isolates, human/porcine isolates grew at 24 and 45 degrees C. Based on differences in IS1245 RFLP, 16S-23S rDNA ITS and growth temperature, M. avium isolates originating from birds should be considered as a separate, evolutionarily conserved taxon. Because all M. avium isolates from birds are invariably of this type, the designation M. avium subsp. avium should be reserved for these bird-type strains. For clarity in the epidemiology of M. avium-related disease, isolates from humans and pigs with multibanded IS1245 RFLPs merit a separate designation. The designation 'M. avium subsp. hominissuis' is suggested for this group of bacteria.

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confidence: 96%

Molecular evidence to support a proposal to reserve the designation Mycobacterium avium subsp. avium for bird-type isolates and 'M. avium subsp. hominissuis' for the human/porcine type of M. avium

Mijs¹

2002

International Journal of Systematic and Evolutionary Microbiology

193

180

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“…hominissuis sequavars (Turenne et al, 2006). Intergenic spacer (ITS) sequencing (Abed et al, 1995;Barry et al, 1991;Glennon et al, 1994;Roth et al, 1998) further subdivided MAC into 32 sequevars and recently delineated M. chimaera (Tortoli et al, 2004) and M. colombiense (Murcia et al, 2006). Herein, we studied partial rpoB gene sequencing as a new method for MAC species identification.…”

Section: Introductionmentioning

confidence: 99%

rpoB sequence-based identification of Mycobacterium avium complex species

et al. 2008

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The Mycobacterium avium complex (MAC) comprises slowly growing mycobacteria responsible for opportunistic infections and zoonoses. The ability to speciate MAC isolates in the clinical microbiology laboratory is critical for determining the organism implicated in clinical disease and for epidemiological investigation of the source of infection. Investigation of a 711 bp variable fragment of rpoB flanked by the Myco-F/Myco-R primers found a 0.7-5.1 % divergence among MAC reference strains, with Mycobacterium chimaera and Mycobacterium intracellulare being the most closely related. Using a 0.7 % divergence cut-off, 83 % of 100 clinical isolates, which had been previously identified by phenotypic characteristics and 16S-23S rDNA intergenic spacer (ITS) probing, were identified as M. avium, 8 % as M. intracellulare and 2 % as M. chimaera. The uniqueness of seven isolates, exhibiting ,99.3 % rpoB sequence similarity with MAC reference strains, was confirmed by 16S rDNA, ITS and hsp65 sequencing and phylogenetic analyses. Partial rpoB gene sequencing using the Myco-F/Myco-R primers permits one-step identification of MAC isolates at the species level and the detection of potentially novel MAC species.

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“…Positive slides were confirmed by Ziehl-Neelsen staining. Bacterial colonies were simultaneously identified as M. tuberculosis or other different species of mycobacteria by 16SrRNA sequencing [18] and conventional methods, including growth characteristics, pigment production, colony morphology, and growth in the presence of PNB (p-nitrobenzoic acid) and TCH (thiophene-2-carboxylic acid hydrazide). The exact methods used in our laboratory were in accord with those described in Chinese Laboratory Science Procedure of Diagnostic Bacteriology in Tuberculosis.…”

Section: Methodsmentioning

confidence: 99%

The Conservation and Application of Three Hypothetical Protein Coding Gene for Direct Detection of Mycobacterium tuberculosis in Sputum Specimens

et al. 2013

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BackgroundAccurate and early diagnosis of tuberculosis (TB) is of major importance in the control of TB. One of the most important technical advances in diagnosis of tuberculosis is the development of nucleic acid amplification (NAA) tests. However, the choice of the target sequence remains controversial in NAA tests. Recently, interesting alternatives have been found in hypothetical protein coding sequences from mycobacterial genome.Methodology/Principal FindingsTo obtain rational biomarker for TB diagnosis, the conservation of three hypothetical genes was firstly evaluated in 714 mycobacterial strains. The results showed that SCAR1 (Sequenced Characterized Amplified Region) based on Rv0264c coding gene showed the highest conservation (99.8%) and SCAR2 based on Rv1508c gene showed the secondary high conservation (99.7%) in M. tuberculosis (MTB) strains. SCAR3 based on Rv2135c gene (3.2%) and IS6110 (8%) showed relatively high deletion rate in MTB strains. Secondly, three SCAR markers were evaluated in 307 clinical sputum from patients in whom TB was suspected or patients with diseases other than TB. The amplification of IS6110 and 16SrRNA sequences together with both clinical and bacteriological identification was as a protocol to evaluate the efficacy of SCAR markers. The sensitivities and specificities, positive predictive value (PPV) and negative predictive value (NPV) of all NAA tests were higher than those of bacteriological detection. In four NAA tests, IS6110 and SCAR3 showed the highest PPV (100%) and low NPV (70% and 68.8%, respectively), and SCAR1 and SCAR2 showed the relatively high PPV and NPV (97% and 82.6%, 95.6% and 88.8%, respectively).Conclusions/SignificanceOur result indicated that SCAR1 and SCAR2 with a high degree of sequence conservation represent efficient and promising alternatives as NAA test targets in identification of MTB. Moreover, the targets developed from this study may provide more alternative targets for the development of a multisite system to effectively detect MTB in samples.

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The ribosomal intergenic spacer region: a target for the PCR based diagnosis of tuberculosis

Cited by 26 publications

References 18 publications

Molecular evidence to support a proposal to reserve the designation Mycobacterium avium subsp. avium for bird-type isolates and 'M. avium subsp. hominissuis' for the human/porcine type of M. avium

Molecular evidence to support a proposal to reserve the designation Mycobacterium avium subsp. avium for bird-type isolates and 'M. avium subsp. hominissuis' for the human/porcine type of M. avium

rpoB sequence-based identification of Mycobacterium avium complex species

The Conservation and Application of Three Hypothetical Protein Coding Gene for Direct Detection of Mycobacterium tuberculosis in Sputum Specimens

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