Characterization of a Mycobacterium tuberculosis insertion sequence, IS6110, and its application in diagnosis

Thierry, Dominique; Brisson-Noël, A; Vincent-Lévy-Frébault, Véronique; Nguyen, Sa Van; Guesdon, Jean‐Luc; Gicquel, Brigitte

doi:10.1128/jcm.28.12.2668-2673.1990

Cited by 409 publications

(167 citation statements)

References 28 publications

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“…DNA was extracted from the paraffin blocks according to the method described by Shibata et al 6 The oligonu- l of DNA solution 7 . Denaturation was at 948C for 40 s with annealing at 658C for 40 s and extension at 728C for 15 s. Amplification was done by 50 cycles.…”

Section: P O Ly M E R a S E C H A I N R E Ac T I O Nmentioning

confidence: 99%

Nodular granulomatous phlebitis of the skin: a fourth type of tuberculid

et al. 1997

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We present five cases of granulomatous phlebitis of the skin and compare them with a case of miliary tuberculosis with granulomatous phlebitis. All five patients were hypersensitive to purified protein derivative, but without active tuberculosis. Although anti-tuberculous drugs were effective, no tubercle bacilli were isolated from the skin. Clinically, subcutaneous nodules were felt along the course of the leg vein. Histologically, epithelioid cell granulomas with Langhans' giant cells were observed within the walls of the cutaneous veins. In a later stage, granulomatous panniculitis was often associated. Using the polymerase chain reaction method. Mycobacterium tuberculosis DNA was detected in four of the five cases of granulomatous phlebitis of the skin. Granulomatous phlebitis of the skin seems to represent a relatively early phase of delayed-type hypersensitivity reactions to Mycobacterium tuberculosis and may represent a distinct entity different from other types of tuberculid-a new tuberculid. Nevertheless, before making the diagnosis, the possibility of true tuberculosis must always be excluded. Nodular granulomatous phlebitis of the skin would be an appropriate name for the newly described condition.

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Section: P O Ly M E R a S E C H A I N R E Ac T I O Nmentioning

confidence: 99%

Nodular granulomatous phlebitis of the skin: a fourth type of tuberculid

et al. 1997

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“…Genomic DNA from Myco. tuberculosis was prepared by phenolchloroform extraction [13]. The gene for MPT64 was amplified from Myco.…”

Section: Cloning Of Mpt64 Gene Into Recombinant Myco Smegmatismentioning

confidence: 99%

Expression of Mycobacterium tuberculosis MPT64 in recombinant Myco. smegmatis: purification, immunogenicity and application to skin tests for tuberculosis

Roche

Winter

Triccas

et al. 1996

Clinical and Experimental Immunology

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SUMMARYProteins secreted across the cell wall of mycobacteria are important antigens recognized early in the host response to mycobacterial infection. MPT64 is a 23-kD secreted protein restricted to members of the Mycobacterium tuberculosis complex which elicits T cell responses and cutaneous DTH reactions in Myco. tuberculosis-infected animals. Patients with tuberculosis and their tuberculin-positive contacts respond to the protein, but recipients of bacille Calmette-Guérin (BCG) vaccine strains lacking the mpt64 gene do not. In the present study, we describe the development of a unique recombinant mycobacterial vector which secretes the encoded Myco. tuberculosis protein MPT64 at high levels into the culture filtrate, from which the protein is isolated by a single-step affinity chromatographic step. The purified protein was recognized by both polyclonal and monoclonal anti-MPT64 antibodies. The T cell reactivity of the protein was confirmed by its ability to stimulate human anti-rMPB64 T cell lines. The Myco. smegmatis recombinant MPT64 protein was superior to the Escherichia coli rMPB64 protein, which has identical amino acid sequence, in eliciting cutaneous DTH reactions in guinea pigs sensitized with Myco. tuberculosis. Animals sensitized with BCG strains lacking the mpb64 gene failed to respond to MPT64. Similarly, interferon-gamma (IFN-) responses in tuberculosis patients and their contacts were higher to the Myco. smegmatis form of the protein. The potential of this form of the Myco. tuberculosis MPT64 protein as a skin test reagent for tuberculosis is discussed.

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“…The mycobacterial 902-bp HindIII-BamHI DNA fragment of pMT02 described elsewhere [12] was labeled with (a-32p)dCTP by using the multiprime DNA labeling kit (Amersham International, Amersham, UK). Alternatively, the whole pMT02 plasmid was chemically labeled with acetoxy-acetylaminofluorene (AAF) as previously described [13,14].…”

Section: Labelling Of Dna Probesmentioning

confidence: 99%

“…The DNA fragments were electrophoretically separated through 0.8% agarose gels at 30 V for 18 h (20-cm gel) in 0.04 M Tris-acetate buffer and 0.002 M EDTA; the gels were then incubated once for 15 min in 0.25 M HC1, twice for 15 min each time in 1.5 M NaC1-0.5 M NaOH, and twice for 15 min each time in 1 M CH3COONH 4. The DNA fragments were transferred to a Hybond N filter (Amersham, UK) or nitrocellulose membrane (Schleicher and Schuell, FRG); hybridization was performed with the denaturated 32p_ or AAF-labeled probes as de-scribed by Thierry et al [12]. Hybridization and washings were done at 65°C.…”

Section: Southern Hybridization Analysismentioning

confidence: 99%

Use of the insertion element IS6110 for DNA fingerprinting of Mycobacterium tuberculosis isolates presenting various profiles of drug susceptibility

Thierry

1993

FEMS Immunology and Medical Microbiology

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IS6100 is an insertion sequence of the IS3 family and it is present in multiple copies in the chromosome of Mycobacterium tuberculosis. Four to 15 copies are present in various strains of M. tuberculosis. In this study, the value of IS6110 as an epidemiological marker of tuberculosis was examined. Unrelated clinical strains from Greek patients presented, in restriction fragment length polymorphism analysis, a high degree of polymorphism, whereas patterns of related clinical strains from familial outbreaks were identical. Since RFLP analysis with acetylaminofluorene labeled IS6110 as the probe gave satisfactory results, it is suggested that this non-radioactive probe can be used in hospitals and health centres for the epidemiological survey of M. tuberculosis infections.

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Characterization of a Mycobacterium tuberculosis insertion sequence, IS6110, and its application in diagnosis

Cited by 409 publications

References 28 publications

Nodular granulomatous phlebitis of the skin: a fourth type of tuberculid

Nodular granulomatous phlebitis of the skin: a fourth type of tuberculid

Expression of Mycobacterium tuberculosis MPT64 in recombinant Myco. smegmatis: purification, immunogenicity and application to skin tests for tuberculosis

Use of the insertion element IS6110 for DNA fingerprinting of Mycobacterium tuberculosis isolates presenting various profiles of drug susceptibility

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