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Publication Details
AFRICAN RESEARCH NEXUS
SHINING A SPOTLIGHT ON AFRICAN RESEARCH
Two new rapid SNP-typing methods for classifying mycobacterium tuberculosis complex into the main phylogenetic lineages
PLoS ONE, Volume 7, No. 7, Article e41253, Year 2012
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Description
There is increasing evidence that strain variation in Mycobacterium tuberculosis complex (MTBC) might influence the outcome of tuberculosis infection and disease. To assess genotype-phenotype associations, phylogenetically robust molecular markers and appropriate genotyping tools are required. Most current genotyping methods for MTBC are based on mobile or repetitive DNA elements. Because these elements are prone to convergent evolution, the corresponding genotyping techniques are suboptimal for phylogenetic studies and strain classification. By contrast, single nucleotide polymorphisms (SNP) are ideal markers for classifying MTBC into phylogenetic lineages, as they exhibit very low degrees of homoplasy. In this study, we developed two complementary SNP-based genotyping methods to classify strains into the six main human-associated lineages of MTBC, the "Beijing" sublineage, and the clade comprising Mycobacterium bovis and Mycobacterium caprae. Phylogenetically informative SNPs were obtained from 22 MTBC whole-genome sequences. The first assay, referred to as MOL-PCR, is a ligation-dependent PCR with signal detection by fluorescent microspheres and a Luminex flow cytometer, which simultaneously interrogates eight SNPs. The second assay is based on six individual TaqMan real-time PCR assays for singleplex SNP-typing. We compared MOL-PCR and TaqMan results in two panels of clinical MTBC isolates. Both methods agreed fully when assigning 36 well-characterized strains into the main phylogenetic lineages. The sensitivity in allele-calling was 98.6% and 98.8% for MOL-PCR and TaqMan, respectively. Typing of an additional panel of 78 unknown clinical isolates revealed 99.2% and 100% sensitivity in allele-calling, respectively, and 100% agreement in lineage assignment between both methods. While MOL-PCR and TaqMan are both highly sensitive and specific, MOL-PCR is ideal for classification of isolates with no previous information, whereas TaqMan is faster for confirmation. Furthermore, both methods are rapid, flexible and comparably inexpensive. © 2012 Stucki et al.
Authors & Co-Authors
Stucki, David
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Malla, Bijaya
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Hostettler, Simon
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Huna, Thembela
United Kingdom, London
Mrc National Institute for Medical Research
Feldmann, Julia
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Yeboah-Manu, Dorothy
Ghana, Accra
Noguchi Memorial Institute for Medical Research
Borrell, Sònia
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Fenner, Lukas
Switzerland, Bern
University of Bern
Comas, Iñaki T.
Spain, Valencia
Centre for Public Health Research
Spain, Madrid
Centro de Investigación Biomédica en Red de Epidemiología y Salud Pública
Coscollá, Mireia
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Gagneux, Sébastien P.
Switzerland, Allschwil
Swiss Tropical and Public Health Institute Swiss Tph
Switzerland, Basel
Universitat Basel
Statistics
Citations: 138
Authors: 11
Affiliations: 7
Identifiers
Doi:
10.1371/journal.pone.0041253
e-ISSN:
19326203
Research Areas
Genetics And Genomics