TY - JOUR
T1 - IS-seq
T2 - a novel high throughput survey of in vivo IS6110 transposition in multiple Mycobacterium tuberculosis genomes
AU - Reyes, Alejandro
AU - Sandoval, Andrea
AU - Cubillos-Ruiz, Andrés
AU - Varley, Katherine E.
AU - Hernández-Neuta, Ivan
AU - Samper, Sofía
AU - Martín, Carlos
AU - García, María J.
AU - Ritacco, Viviana
AU - López, Lucelly
AU - Robledo, Jaime
AU - Zambrano, María M.
AU - Mitra, Robi D.
AU - Del Portillo, Patricia
N1 - Funding Information:
We would like to thank Jessica Hoisington-Lopez from Washington University Genomic Technology Access Center (GTAC), and Cofactor Genomics for Illumina sequencing; Elida Gravina for isolating and genotyping Argentinean strains and providing clinical/demographic information; Isabel Millán for helping with the confirmation of the IS insertions in the strains from Spain; and Lucía Barrera and her team at the Instituto Malbrán for microbiological support. We also thank Elena del Corral, Wellman Ribon and Maria Lilia Diaz for comments and help with the preliminary data analysis, and Dr. Roberto Kolter for helpful discussions and analysis of the data. This work was supported by Colciencias project No. 657040820410, grant No. 431–2004 for the Colombian Center for Excellence in Tuberculosis Research, CCITB, the StopLATENT-TB network (Collaborative Project) supported by the EC under the Health Cooperation Work Programme of the 7th Framework Programme (G.A. no. 200999) (http://cordis.europa.eu/fp7/dc/index.cfm), the Children’s Discovery Institute grant MC-II-2006-1, and the NIH Epigenetics Roadmap grant (1R01DA025744-01 and 3R01DA025744-02 S1). Part of this work was developed in the context of the Network of multidrug resistant tuberculosis in Latin America supported by Latin-American Science & Technology Development Programme (CYTED). Reyes A. is the recipient of an International Fulbright Science and Technology Program award. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
PY - 2012/6/15
Y1 - 2012/6/15
N2 - Background: The insertion element IS6110 is one of the main sources of genomic variability in Mycobacterium tuberculosis, the etiological agent of human tuberculosis. Although IS 6110 has been used extensively as an epidemiological marker, the identification of the precise chromosomal insertion sites has been limited by technical challenges. Here, we present IS-seq, a novel method that combines high-throughput sequencing using Illumina technology with efficient combinatorial sample multiplexing to simultaneously probe 519 clinical isolates, identifying almost all the flanking regions of the element in a single experiment.Results: We identified a total of 6,976 IS6110 flanking regions on the different isolates. When validated using reference strains, the method had 100% specificity and 98% positive predictive value. The insertions mapped to both coding and non-coding regions, and in some cases interrupted genes thought to be essential for virulence or in vitro growth. Strains were classified into families using insertion sites, and high agreement with previous studies was observed.Conclusions: This high-throughput IS-seq method, which can also be used to map insertions in other organisms, extends previous surveys of in vivo interrupted loci and provides a baseline for probing the consequences of disruptions in M. tuberculosis strains.
AB - Background: The insertion element IS6110 is one of the main sources of genomic variability in Mycobacterium tuberculosis, the etiological agent of human tuberculosis. Although IS 6110 has been used extensively as an epidemiological marker, the identification of the precise chromosomal insertion sites has been limited by technical challenges. Here, we present IS-seq, a novel method that combines high-throughput sequencing using Illumina technology with efficient combinatorial sample multiplexing to simultaneously probe 519 clinical isolates, identifying almost all the flanking regions of the element in a single experiment.Results: We identified a total of 6,976 IS6110 flanking regions on the different isolates. When validated using reference strains, the method had 100% specificity and 98% positive predictive value. The insertions mapped to both coding and non-coding regions, and in some cases interrupted genes thought to be essential for virulence or in vitro growth. Strains were classified into families using insertion sites, and high agreement with previous studies was observed.Conclusions: This high-throughput IS-seq method, which can also be used to map insertions in other organisms, extends previous surveys of in vivo interrupted loci and provides a baseline for probing the consequences of disruptions in M. tuberculosis strains.
UR - http://www.scopus.com/inward/record.url?scp=84862209699&partnerID=8YFLogxK
U2 - 10.1186/1471-2164-13-249
DO - 10.1186/1471-2164-13-249
M3 - Article
C2 - 22703188
AN - SCOPUS:84862209699
SN - 1471-2164
VL - 13
JO - BMC genomics
JF - BMC genomics
IS - 1
M1 - 249
ER -