title: Clonal relationship and differentiation among Mycobacterium abscessus isolates as determined using the semiautomated repetitive extragenic palindromic sequence PCR-based DiversiLab system.
abstract: Mycobacterium abscessus is a rapidly growing mycobacterium that causes respiratory tract infections in predisposed patients, such as those with cystic fibrosis and nosocomial skin and soft tissue infections. In order to investigate the clonal relationships between the strains causing epidemic episodes, we evaluated the discriminatory power of the semiautomated DiversiLab (DL) repetitive extragenic palindromic sequence PCR (REP-PCR) test for M. abscessus genotyping. Since M. abscessus was shown to be composed of subspecies (M. abscessus subsp. massiliense, M. abscessus subsp. bolletii, and M. abscessus subsp. abscessus), we also evaluated the ability of this technique to differentiate subspecies. The technique was applied to two collections of clinical isolates, (i) 83 M. abscessus original isolates (43 M. abscessus subsp. abscessus, 12 M. abscessus subsp. bolletii, and 28 M. abscessus subsp. massiliense) from infected patients and (ii) 35 repeated isolates obtained over 1 year from four cystic fibrosis patients. The DL REP-PCR test was standardized for DNA extraction, DNA amplification, and electrophoresis pattern comparisons. Among the isolates from distinct patients, 53/83 (62%) isolates showed a specific pattern, and 30 were distributed in 11 clusters and 6 patterns, with 2 to 4 isolates per pattern. The clusters and patterns did not fully correlate with multilocus sequence typing (MLST) analysis results. This revealed a high genomic diversity between patients, with a discriminatory power of 98% (Simpson's diversity index). However, since some isolates shared identical patterns, this raises the question of whether it is due to transmission between patients or a common reservoir. Multiple isolates from the same patient showed identical patterns, except for one patient infected by two strains. Between the M. abscessus subspecies, the indexes were <70%, indicating that the DL REP-PCR test is not an accurate tool for identifying organisms to the subspecies level. REP-PCR appears to be a rapid genotyping method that is useful for investigating epidemics of M. abscessus infections.
authors.affiliation: APHP, Hôpitaux Universitaires Lariboisière-Saint Louis, Service de Bactériologie, Paris, France Université Paris Diderot, EA3964, Paris, France Centre National de Référence des Mycobactéries et Résistance des Mycobactéries aux Antituberculeux (CNR-MyRMA), Paris, France.
authors.forename: Faiza
authors.initials: F
authors.lastname: Mougari
authors.affiliation: APHP, Hôpitaux Universitaires Lariboisière-Saint Louis, Service de Bactériologie, Paris, France Centre National de Référence des Mycobactéries et Résistance des Mycobactéries aux Antituberculeux (CNR-MyRMA), Paris, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Raskine
authors.affiliation: APHP, Groupe Hospitalier Necker-Enfants Malades, Service de Bactériologie-Virologie-Parasitologie et Hygiène, Paris, France.
authors.forename: Agnes
authors.initials: A
authors.lastname: Ferroni
authors.affiliation: APHP, Hôpitaux Universitaires Paris Nord Val de Seine, Beaujon, Service de Microbiologie, Clichy, France.
authors.forename: Estelle
authors.initials: E
authors.lastname: Marcon
authors.affiliation: APHP, Groupe Hospitalier Necker-Enfants Malades, Centre de Ressources et de Compétences pour la Mucoviscidose (CRCM) et Centre de Formation de Traitement à Domicile Chez l'Enfant (CFTDE), Paris, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Sermet-Gaudelus
authors.affiliation: Centre National de Référence des Mycobactéries et Résistance des Mycobactéries aux Antituberculeux (CNR-MyRMA), Paris, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Veziris
authors.affiliation: APHP, Hôpital Ambroise Paré, Service de Microbiologie et Hygiène, Boulogne-Billancourt, France.
authors.forename: Beate
authors.initials: B
authors.lastname: Heym
authors.affiliation: APHP, Hôpital Ambroise Paré, Service de Microbiologie et Hygiène, Boulogne-Billancourt, France.
authors.forename: Jean-Louis
authors.initials: JL
authors.lastname: Gaillard
authors.affiliation: APHP, Groupe Hospitalier Necker-Enfants Malades, Service de Bactériologie-Virologie-Parasitologie et Hygiène, Paris, France.
authors.forename: Xavier
authors.initials: X
authors.lastname: Nassif
authors.affiliation: APHP, Hôpitaux Universitaires Lariboisière-Saint Louis, Service de Bactériologie, Paris, France Université Paris Diderot, EA3964, Paris, France Centre National de Référence des Mycobactéries et Résistance des Mycobactéries aux Antituberculeux (CNR-MyRMA), Paris, France emmanuelle.cambau@lrb.aphp.fr.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Cambau
mesh_terms: D057205:Automation, Laboratory / Q000379:methods; D015373:Bacterial Typing Techniques / Q000379:methods*; D016000:Cluster Analysis; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D017720:Molecular Epidemiology / Q000379:methods; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D009165:Mycobacterium Infections, Nontuberculous / Q000453:epidemiology / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*