Comparison of Mycobacterium 23S rRNA Sequences by High-Temperature Reverse Transcription and PCR

Stone, Benjamin B.; Nietupski, Raymond M.; Breton, Gary L.; Weisburg, William G.

doi:10.1099/00207713-45-4-811

Cited by 18 publications

(13 citation statements)

References 26 publications

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“…However, when previously tested in the manual format, M. africanum gave positive results similar to those obtained with other M. tuberculosis complex organisms (39). These results are consistent with the observation that the target regions of both M. africanum and M. microti rRNA to which the capture probe binds differ from the M. tuberculosis target sequence by only one nucleotide (43).…”

Section: Discussionsupporting

confidence: 85%

“…Previously, inclusivity testing of all five M. tuberculosis complex members showed that M. bovis and M. tuberculosis strains were detected by hybridization and Q-Beta replicase amplification, as predicted by sequence data (43). When multiple samples of 10-fold dilutions of nine M. tuberculosis strains were tested, the lower limit of detection was determined to be Ͻ10 to 300 CFU.…”

Section: Discussionmentioning

confidence: 87%

“…Previously, we have shown that negative results were obtained when 10 7 CFU of M. avium was tested in the automated assay (40). Additionally, 23S rRNA sequence information (43) indicates that M. kansasii, M. celatum, M. xenopi, M. gastri, and M. gordonae have very similar sequences in the target region and none of these were positive by the Galileo assay with a high level of target input. During analysis of discrepant results, we found that one sample reported as MOTT came from a patient with clinical tuberculosis.…”

Section: Discussionmentioning

confidence: 96%

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Performance of an automated Q-beta replicase amplification assay for Mycobacterium tuberculosis in a clinical trial

Smith¹,

Radcliffe²,

Rigby³

et al. 1997

J Clin Microbiol

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We present data from a clinical trial study in which an automated version (Galileo) of a previously described Q-Beta replicase-amplified probe assay (J. S. Shah et al., J. Clin. Microbiol. 33:1435-1441, 1995) was used for the direct detection of Mycobacterium tuberculosis complex in sputum. The assay was designed to target specific regions of 23S rRNA found in M. tuberculosis, Mycobacterium bovis, Mycobacterium africanum, and Mycobacterium microti and had a sensitivity ranging from approximately <10 to 300 CFU. The assay was tested for cross-hybridization by using large numbers (e.g., 10 5 to 10 10 CFU/assay) of 133 other organisms commonly found in respiratory tract samples, including non-M. tuberculosis Mycobacterium spp., other bacteria, fungi, and viruses. All of these competitors tested negative by the assay. Automated assay results for 780 respiratory tract samples (sputum or bronchoalveolar lavage specimens) collected and tested at three trial sites in the United States) were compared with the results of culture and acid-fast microscopy. Aliquots of conventionally digested and decontaminated sputum pellets were heated at 100°C and mechanically disrupted prior to hybridization and background reduction, amplification, and detection in a closed disposable test pack. Pertinent elements of individual patient histories relating to tuberculosis exposure, previous active disease, antituberculosis therapy status, etc., were considered in the resolution of discrepant results for 48 (assay false-positive) samples. Seventy-one of 90 (78.9%) culture-positive samples were positive when tested in the Galileo assay, while 7% of culture-negative samples were assay positive, corresponding to a sensitivity of 79% and a specificity of 93%. Following resolution of discrepant results by chart review, the sensitivity and specificity for the Q-Beta replicase amplification assay with the Galileo analyzer were 84 and 97%, respectively. A total of 69.2% of smear-negative (culture positive) samples were detected by the assay. Ten test packs at a time were automatically processed by the Galileo analyzer without operator intervention following loading of samples. The first result was reported in approximately 3 h, and the last result was available in 6.5 h. To our knowledge, this is the first report of a clinical study with a fully automated amplification probe hybridization assay for the detection of pathogens directly from a clinical specimen.

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Section: Discussionsupporting

confidence: 85%

Section: Discussionmentioning

confidence: 87%

Section: Discussionmentioning

confidence: 96%

See 1 more Smart Citation

Performance of an automated Q-beta replicase amplification assay for Mycobacterium tuberculosis in a clinical trial

Smith¹,

Radcliffe²,

Rigby³

et al. 1997

J Clin Microbiol

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show abstract

“…Primer 3,5'-CCG TGA GGG AAA GGT GAA AAG TAC C-3', which corresponds to the sequence from 461 to 485 bp in the Escherichia coli nomenclature, was designed during this work. It is a slight modification of primer 3415 (26). Primer 4,5'-CCC GCT TAG ATG CTT TCA GC-3', which corresponds to sequence from 2744 to 2763 bp in the E. coli nomenclature, was described by Van Camp et al (7) as 97ar.…”

Section: Methodsmentioning

confidence: 99%

Phylogeny of Rhizobium galegae with respect to other rhizobia and agrobacteria

Terefework

Nick

Suomalainen

et al. 1998

International Journal of Systematic Bacteriology

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PCR-RFLP with nine restriction enzymes was applied to the 165 and 235 rRNA genes of 42 rhizobial and agrobacterial strains to determine the phylogenetic position of Rhizobium galegae and increase the understanding of the evolution of ribosomal operons. The strains were selected based on previous phylogenetic studies. PCR primers were designed so that they amplified a 2.3 kb fragment of the 23s rRNA gene (excluding the B8 loop). Universal primers rD1 and fD1 were used to amplify the full-length 165 rRNA. The RFLP analysis resulted in 27 and 32 different restriction patterns for 165 and 235, respectively. The RFLP patterns were transformed to genetic distances and dendrograms were constructed from the data using the unweighted pair group method with averages. The shapes of the dendrograms derived from the analysis of the 165 and 235 rRNA genes correlated well, with only a f e w strains having different positions. The 235 tree generally had deeper branching than the 165 tree, allowing better discrimination between species and strains. The combined data from the two analyses described 36 genotypes. The eight R. galegae strains formed a homogeneous cluster in all dendrograms. The RFLP analysis was confirmed by partial sequence analysis of the 165 rRNA gene (the first 800 bp), which correlated well with full-length 165 rRNA sequence analysis. The 165 data placed R. galegae near the genus Agrobacterium with Agrobacterium witis as its nearest neighbour, whereas in the 235 and the combined dendrograms it showed closer affinity to the genus Rhizobium.

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“…One M. smegmatis isolate was misidentified as M. fortuitum II by the GenoType. A possible explanation for this identification mistake is the high degree of homology in the 23S rRNA sequence for these two mycobacterial species (20). Interestingly, one other M. smegmatis isolate was correctly identified by GenoType as nontuberculous mycobacteria.…”

Section: Discussionmentioning

confidence: 98%

Comparative Evaluation of the New Version of the INNO-LiPA Mycobacteria and GenoType Mycobacterium Assays for Identification of Mycobacterium Species from MB/BacT Liquid Cultures Artificially Inoculated with Mycobacterial Strains

et al. 2004

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Comparison of Mycobacterium 23S rRNA Sequences by High-Temperature Reverse Transcription and PCR

Cited by 18 publications

References 26 publications

Performance of an automated Q-beta replicase amplification assay for Mycobacterium tuberculosis in a clinical trial

Performance of an automated Q-beta replicase amplification assay for Mycobacterium tuberculosis in a clinical trial

Phylogeny of Rhizobium galegae with respect to other rhizobia and agrobacteria

Comparative Evaluation of the New Version of the INNO-LiPA Mycobacteria and GenoType Mycobacterium Assays for Identification of Mycobacterium Species from MB/BacT Liquid Cultures Artificially Inoculated with Mycobacterial Strains

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