Specificity of IS6110-based amplification assays for Mycobacterium tuberculosis complex

Hellyer, T.J.; DesJardin, Lucy E.; Assaf, Melissa; Bates, Joseph H.; Cave, M. Donald; Eisenach, Kathleen D.

doi:10.1128/jcm.34.11.2843-2846.1996

Cited by 68 publications

(22 citation statements)

References 25 publications

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“…Our previous study demonstrated the presence of a homologous sequence by PCR, and this was confirmed by dot blot hybridization of genomic DNA (10). Despite the hybridization data, other investigators have suggested that our results may have been due to PCR contamination (8). In this study PCR was only used to prepare probes, homology was demonstrated by Southern hybridization of genomic DNA, and a higher stringency than that of the international standard method was used, implying that the sequences have a high degree of homology for IS6110.…”

Section: Discussionsupporting

confidence: 81%

“…No positive results were obtained, although the amount of DNA used in the experiments was not quantified. These data suggest either that there is a lower degree of homology or that the homology that we have demonstrated does not extend to the region covered by their primers (4,8). We believe that it is more likely that mycobacteria may possess a family of IS3 sequences which are closely re-lated.…”

Section: Discussionmentioning

confidence: 64%

“…If the multiple bands are due to the presence of an IS, then the distribution that we have shown in the genus could have arisen by movement of IS6110 between species. Hellyer et al (8) reported the use of a PCR amplifying the region of IS6110 from positions 762 to 865 on DNA derived from MOTT provided by our laboratory. No positive results were obtained, although the amount of DNA used in the experiments was not quantified.…”

Section: Discussionmentioning

confidence: 99%

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IS6110 homologs are present in multiple copies in mycobacteria other than tuberculosis-causing mycobacteria

1997

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We have previously demonstrated homology between a 181-bp fragment of IS6110 and DNA from mycobacteria other than tuberculosis-causing mycobacteria (MOTT). Genomic DNA from 14 strains of MOTT was digested with PvuII and was hybridized with a probe derived from the 181-bp fragment and the INS1/INS2 international standard probe at high stringency. Multiple banding patterns were obtained from isolates of M. avium-M. intracellulare, M. fortuitum, M. kansasii, and M. malmoense. Differences in the banding patterns between and within species were obtained. This suggests that mycobacteria possess a family of IS3-like elements. The species of isolates suspected of being M. tuberculosis must be carefully determined before IS6110 restriction fragment length polymorphism analysis, and caution must be used in designing and evaluating diagnostic PCR tests based on this element.

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Section: Discussionsupporting

confidence: 81%

Section: Discussionmentioning

confidence: 64%

Section: Discussionmentioning

confidence: 99%

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IS6110 homologs are present in multiple copies in mycobacteria other than tuberculosis-causing mycobacteria

1997

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“…However, this homology lies in segments outside the region amplified by the T4 and T5 primers that we use (17). Therefore, PCR using the T4 and T5 primers for the IS6110 repeat sequence is useful in differentiating Mycobacterium tuberculosis from atypical mycobacterial infection because it is not found in myobacteria other than tuberculosis (18). It makes a more rapid diagnosis of rare or atypical manifestation of tuberculosis infection than tissue culture.…”

Section: Discussionmentioning

confidence: 99%

Perianal Ulceration in a “Healthy” Chinese Man with Disseminated Tuberculosis

Lee

Chong

2002

The Journal of Dermatology

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Orificial tuberculosis (OTB) is a rare form of cutaneous Mycobacterium tuberculosis infection affecting the mucosa and skin around orifices in patients with advanced internal tuberculosis and poor general health. We report a 72-year-old Chinese man who had a 10-year history of OTB with disseminated tuberculosis infection of the lungs and urinary tract. He appeared surprisingly healthy and had been free from systemic symptoms all along despite widespread tuberculosis. The diagnosis of OTB was established by the microscopic presence of acid-fast bacilli (AFB) in the tissue section and was rapidly confirmed by polymerase chain reaction (PCR) to be Mycobacterium tuberculosis. PCR shortens the time of diagnosing rare presentations of cutaneous tuberculosis and prevents delays in treatment. Conventional culture is still important in confirming the diagnosis and screening for drug resistance.

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“…Various PCR methods with different DNA targets are specific and sensitive for the rapid detection of MTB in sputum and other body fluids (Eisenach et al 1991;Salian et al 1998;Haldar et al 2007;Gopinath & Singh 2009). As the insertion sequence IS6110 is normally present in multiple copies (1-25 copies) in the MTB genome and as it is specific for the MTB complex (Hellyer et al 1996), we considered it an excellent target for the detection of MTB.…”

Section: Introductionmentioning

confidence: 99%

Pulmonary tuberculosis in the West Bank, Palestinian Authority: molecular diagnostic approach

Ereqat

Bar‐Gal

Nasereddin

et al. 2010

Tropical Med Int Health

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Summaryobjective To compare the effectiveness and feasibility of an insertion sequence (IS6110)-based polymerase chain reaction (PCR) assay with conventional methods of detecting Mycobacterium tuberculosis and to analyse mutations present in the hot spot region of the RNA polymerase B subunit (rpoB) gene associated with rifampin resistance by DNA sequencing.methods Ninety-five sputum samples from 84 clinically suspected cases of tuberculosis were tested for mycobacterial infections by Ziehl Neelsen smear examination, Lowenstein-Jensen culture and IS6110-based PCR assay.results Sensitivity and specificity of the PCR were 94%; the sensitivity of culture was 65%, and of smear tests, 59%. Both smear microscopy and culture had 100% specificity. DNA sequencing data of the 305-bp fragment of the rpoB gene for nine clinical isolates revealed one point mutation at position I572F and double mutations at position S531F in two isolates obtained from two patients who did not respond to the anti-tuberculosis therapy.conclusion IS6110-based PCR can be used routinely in clinical laboratories for rapid detection of Mycobacterium tuberculosis and thus allow early diagnosis and treatment of any contacts by the cheapest method currently available in the Palestinian Authority region. Rapid detection of rifampin resistance isolates will enable efficient treatment of patients and assist in eradication of the disease in the Palestinian territories.

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Specificity of IS6110-based amplification assays for Mycobacterium tuberculosis complex

Cited by 68 publications

References 25 publications

IS6110 homologs are present in multiple copies in mycobacteria other than tuberculosis-causing mycobacteria

IS6110 homologs are present in multiple copies in mycobacteria other than tuberculosis-causing mycobacteria

Perianal Ulceration in a “Healthy” Chinese Man with Disseminated Tuberculosis

Pulmonary tuberculosis in the West Bank, Palestinian Authority: molecular diagnostic approach

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