title: Mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) genotyping of mycobacterium intracellulare for strain comparison with establishment of a PCR-based database.
abstract: Strain comparison is important to population genetics and to evaluate relapses in patients with Mycobacterium avium complex (MAC) lung disease, but the "gold standard" of pulsed-field gel electrophoresis (PFGE) is time-consuming and complex. We used variable-number tandem repeats (VNTR) for fingerprinting of respiratory isolates of M. intracellulare from patients with underlying bronchiectasis, to establish a nonsequence-based database for population analysis. Different genotypes identified by PFGE underwent species identification using a 16S rRNA gene multiplex PCR. Genotypes of M. intracellulare were confirmed by internal transcribed spacer 1 (ITS1) sequencing and characterized using seven VNTR primers. The pattern of VNTR amplicon sizes and repeat number defined each specific VNTR type. Forty-two VNTR types were identified among 84 genotypes. PFGE revealed most isolates with the same VNTR type to be clonal or exhibit similar grouping of bands. Repetitive sequence-based PCR (rep-PCR) showed minimal pattern diversity between VNTR types compared to PFGE. Fingerprinting of relapse isolates from 31 treated patients using VNTR combined with 16S multiplex PCR unambiguously and reliably distinguished different genotypes from the same patient, with results comparable to those of PFGE. VNTR for strain comparison is easier and faster than PFGE, is as accurate as PFGE, and does not require sequencing. Starting with a collection of 167 M. intracellulare isolates, VNTR distinguished M. intracellulare into 42 clonal groups. Comparison of isolates from different geographic areas, habitats, and clinical settings is now possible.
authors.affiliation: Mycobacteria/Nocardia Laboratory, The University of Texas Health Science Center at Tyler, Tyler, Texas, USA.
authors.forename: Elena
authors.initials: E
authors.lastname: Iakhiaeva
authors.forename: Steven
authors.initials: S
authors.lastname: McNulty
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Brown Elliott
authors.forename: Joseph O
authors.initials: JO
authors.lastname: Falkinham
authors.forename: Myra D
authors.initials: MD
authors.lastname: Williams
authors.forename: Ravikiran
authors.initials: R
authors.lastname: Vasireddy
authors.forename: Rebecca W
authors.initials: RW
authors.lastname: Wilson
authors.forename: Christine
authors.initials: C
authors.lastname: Turenne
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Wallace
mesh_terms: D000483:Alleles; D015373:Bacterial Typing Techniques; D030561:Databases, Nucleic Acid; D005838:Genotype; D006801:Humans; D018598:Minisatellite Repeats; D008969:Molecular Sequence Data; D058885:Multilocus Sequence Typing; D015269:Mycobacterium avium Complex / Q000145:classification* / Q000235:genetics*; D015270:Mycobacterium avium-intracellulare Infection / Q000175:diagnosis*; D012336:RNA, Ribosomal, 16S
chemical_list: D012336:RNA, Ribosomal, 16S