title: High-throughput mycobacterial interspersed repetitive-unit-variable-number tandem-repeat genotyping for Mycobacterium tuberculosis epidemiological studies.
abstract: The emergence of drug-resistant forms of tuberculosis (TB) represents a major public health concern. Understanding the transmission routes of the disease is a key factor for its control and for the implementation of efficient interventions. Mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) marker typing is a well-described method for lineage identification and transmission tracking. However, the conventional manual genotyping technique is cumbersome and time-consuming and entails many risks for errors, thus hindering its implementation and dissemination. We describe here a new approach using the QIAxcel system, an automated high-throughput capillary electrophoresis system that also carries out allele calling. This automated method was assessed on 1,824 amplicons from 82 TB isolates and tested with sets of markers of 15 or 24 loci. Overall allele-calling concordance between the methods from 140 to 1,317 bp was 98.9%. DNA concentrations and repeatability and reproducibility performances showed no biases in allele calling. Furthermore, turnaround time using this automated system was reduced by 81% compared to the conventional manual agarose gel method. In sum, this new automated method facilitates MIRU-VNTR genotyping and provides reliable results. Therefore, it is well suited for field genotyping. The implementation of this method will help to achieve accurate and cost-effective epidemiological studies, especially in countries with a high prevalence of TB, where the high number of strains complicates the surveillance of circulating lineages and requires efficient interventions to be carried out in an urgent manner.
authors.affiliation: Laboratoire des Pathogènes Emergents (LPE), Fondation Mérieux-Centre International de Recherche en Infectiologie (CIRI), Lyon, France marie.gauthier@fondation-merieux.org.
authors.forename: Marie
authors.initials: M
authors.lastname: Gauthier
authors.affiliation: Laboratoire des Pathogènes Emergents (LPE), Fondation Mérieux-Centre International de Recherche en Infectiologie (CIRI), Lyon, France.
authors.forename: Floriane
authors.initials: F
authors.lastname: Bidault
authors.affiliation: Laboratoire des Pathogènes Emergents (LPE), Fondation Mérieux-Centre International de Recherche en Infectiologie (CIRI), Lyon, France.
authors.forename: Amandine
authors.initials: A
authors.lastname: Mosnier
authors.affiliation: National Center for Tuberculosis and Lung Disease (NCTLD), Tbilisi, Georgia.
authors.forename: Nino
authors.initials: N
authors.lastname: Bablishvili
authors.affiliation: National Center for Tuberculosis and Lung Disease (NCTLD), Tbilisi, Georgia.
authors.forename: Nestani
authors.initials: N
authors.lastname: Tukvadze
authors.affiliation: Centre d'Infectiologie Christophe Mérieux du Laos (CICML), Vientiane, Lao People's Democratic Republic.
authors.forename: Silaphet
authors.initials: S
authors.lastname: Somphavong
authors.affiliation: Centre d'Infectiologie Christophe Mérieux du Laos (CICML), Vientiane, Lao People's Democratic Republic.
authors.forename: Phimpha
authors.initials: P
authors.lastname: Paboriboune
authors.affiliation: Les Centres GHESKIO, Port-au-Prince, Haiti.
authors.forename: Oksana
authors.initials: O
authors.lastname: Ocheretina
authors.affiliation: Les Centres GHESKIO, Port-au-Prince, Haiti.
authors.forename: Jean William
authors.initials: JW
authors.lastname: Pape
authors.affiliation: Laboratoire des Pathogènes Emergents (LPE), Fondation Mérieux-Centre International de Recherche en Infectiologie (CIRI), Lyon, France.
authors.forename: Glaucia
authors.initials: G
authors.lastname: Paranhos-Baccala
authors.affiliation: Laboratoire des Pathogènes Emergents (LPE), Fondation Mérieux-Centre International de Recherche en Infectiologie (CIRI), Lyon, France.
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Berland
mesh_terms: D057205:Automation, Laboratory / Q000379:methods; D060005:Genotyping Techniques; D057166:High-Throughput Screening Assays; D006801:Humans; D018598:Minisatellite Repeats; D017720:Molecular Epidemiology / Q000379:methods; D058889:Molecular Typing / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000145:classification* / Q000235:genetics*; D015203:Reproducibility of Results; D013997:Time Factors; D014376:Tuberculosis / Q000453:epidemiology / Q000382:microbiology*