Dataset Information

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

ABSTRACT:

Background

Different combinations of variable number of tandem repeat (VNTR) loci have been proposed for genotyping Mycobacterium tuberculosis complex (MTBC). Existing VNTR schemes show different discriminatory capacity among the six human MTBC lineages. Here, we evaluated the discriminatory power of a "customized MIRU12" loci format proposed previously by Comas et al. based on the standard 24 loci defined by Supply et al. for VNTR-typing of MTBC in Ghana.

Method

One hundred and fifty-eight MTBC isolates classified into Lineage 4 and Lineage 5 were used to compare a customized lineage-specific panel of 12 MIRU-VNTR loci ("customized MIRU-12") to the standard MIRU-15 genotyping scheme. The resolution power of each typing method was determined based on the Hunter-Gaston- Discriminatory Index (HGDI). A minimal set of customized MIRU-VNTR loci for typing Lineages 4 (Euro-American) and 5 (M. africanum West African 1) strains from Ghana was defined based on the cumulative HGDI.

Results and conclusion

Among the 106 Lineage 4 strains, the customized MIRU-12 identified a total of 104 distinct genotypes consisting of 2 clusters of 2 isolates each (clustering rate 1.8%), and 102 unique strains while standard MIRU-15 yielded a total of 105 different genotypes, including 1 cluster of 2 isolates (clustering rate: 0.9%) and 104 singletons. Among, 52 Lineage 5 isolates, customized MIRU-12 genotyping defined 51 patterns with 1 cluster of 2 isolates (clustering rate: 0.9%) and 50 unique strains whereas MIRU-15 classified all 52 strains as unique. Cumulative HGDI values for customized MIRU-12 for Lineages 4 and 5 were 0.98 respectively whilst that of standard MIRU-15 was 0.99. A union of loci from the customised MIRU-12 and standard MIRU-15 revealed a set of customized eight highly discriminatory loci: 4052, 2163B, 40, 4165, 2165, 10,16 and 26 with a cumulative HGDI of 0.99 for genotyping Lineage 4 and 5 strains from Ghana.

SUBMITTER: Asante-Poku A

PROVIDER: S-EPMC3965448 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Publications

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

Asante-Poku Adwoa A Nyaho Michael Selasi MS Borrell Sonia S Comas Iñaki I Gagneux Sebastien S Yeboah-Manu Dorothy D

PloS one 20140325 3

<h4>Background</h4>Different combinations of variable number of tandem repeat (VNTR) loci have been proposed for genotyping Mycobacterium tuberculosis complex (MTBC). Existing VNTR schemes show different discriminatory capacity among the six human MTBC lineages. Here, we evaluated the discriminatory power of a "customized MIRU12" loci format proposed previously by Comas et al. based on the standard 24 loci defined by Supply et al. for VNTR-typing of MTBC in Ghana.<h4>Method</h4>One hundred and f ...[more]

PMID: 24667333

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Project description:BackgroundCombining different molecular typing methods for Mycobacterium tuberculosis complex (MTBC) can be a powerful tool for molecular epidemiology-based investigation of TB. However, the current standard method that provides high discriminatory power for such a combination, mycobacterial interspersed repetitive units-variable numbers of tandem repeats typing (MIRU-VNTR), is laborious, time-consuming and often too costly for many resource-limited laboratories. We aimed to evaluate a reduced set of loci for MIRU-VNTR typing in combination with spoligotyping and SNP-typing for routine molecular epidemiology of TB.MethodSpoligotyping and SNP-typing, in combination with the 15 loci MIRU-VNTR typing, were first used to type clinical MTBC isolates (n = 158) from Madagascar. A step by step reduction of MIRU-VNTR loci number was then performed according to the Hunter and Gaston Discriminatory Index (HGDI) and to the Principal component analysis (PCA) correlation with the spoligotype profiles to evaluate the discrimination power inside the generated spoligotype clusters. The 15 MIRU-VNTR was used as reference and SNP-typing was used to determine the main MTBC lineages.ResultsOf the 158 clinical isolates studied, the SNP-typing classified 23 into Lineage 1 (14.6%), 31 into Lineage 2 (19.6%), 23 into Lineage 3 (14.6%) and 81 into Lineage 4 strains (51.3%). 37 different spoligotypes profiles were obtained, 15 of which were unique and 20 in clusters. 15-loci MIRU-VNTR typing revealed 144 different genotypes: 132 isolates had a unique MIRU-VNTR profile and 27 isolates were grouped into 12 clusters. After a stepwise reduction of the MIRU-VNTR loci number within each main spoligotype families, three different sets composed of 5 customised MIRU-VNTR loci had a similar discrimination level to the reference 15 loci MIRU-VNTR in lineage 1, lineage 2 and lineage 3. For lineage 4, a set of 4 and 3 MIRU-VNTR loci were proposed to subtype the Harleem and LAM spoligotype families, respectively. For the T spoligotype family, a set of 5 MIRU-VNTR loci was proposed.ConclusionAccording to the lineages and the spoligotype families, the number of MIRU-VNTR loci can be reduced to get an optimal classification of MTBC. These customized sets of MIRU-VNTR loci reduce workload and save resources while maintaining optimal discriminatory power.

Project description:BackgroundTuberculosis (TB) resulted in an estimated 1.7 million deaths in the year 2016. The disease is caused by the members of Mycobacterium tuberculosis complex, which includes Mycobacterium tuberculosis, Mycobacterium bovis and other closely related TB causing organisms. In order to understand the epidemiological dynamics of TB, national TB control programs often conduct standardized genotyping at 24 Mycobacterial-Interspersed-Repetitive-Units (MIRU)-Variable-Number-of-Tandem-Repeats (VNTR) loci. With the advent of next generation sequencing technology, whole-genome sequencing (WGS) has been widely used for studying TB transmission. However, an open-source software that can connect WGS and MIRU-VNTR typing is currently unavailable, which hinders interlaboratory communication. In this manuscript, we introduce the MIRU-profiler program which could be used for prediction of MIRU-VNTR profile from WGS of M. tuberculosis.ImplementationThe MIRU-profiler is implemented in shell scripting language and depends on EMBOSS software. The in-silico workflow of MIRU-profiler is similar to those described in the laboratory manuals for genotyping M. tuberculosis. Given an input genome sequence, the MIRU-profiler computes alleles at the standard 24-loci based on in-silico PCR amplicon lengths. The final output is a tab-delimited text file detailing the 24-loci MIRU-VNTR pattern of the input sequence.ValidationThe MIRU-profiler was validated on four datasets: complete genomes from NCBI-GenBank (n = 11), complete genomes for locally isolated strains sequenced using PacBio (n = 4), complete genomes for BCG vaccine strains (n = 2) and draft genomes based on 250 bp paired-end Illumina reads (n = 106).ResultsThe digital MIRU-VNTR results were identical to the experimental genotyping results for complete genomes of locally isolated strains, BCG vaccine strains and five out of 11 genomes from the NCBI-GenBank. For draft genomes based on short Illumina reads, 21 out of 24 loci were inferred with a high accuracy, while a number of inaccuracies were recorded for three specific loci (ETRA, QUB11b and QUB26). One of the unique features of the MIRU-profiler was its ability to process multiple genomes in a batch. This feature was tested on all complete M. tuberculosis genome (n = 157), for which results were successfully obtained in approximately 14 min.ConclusionThe MIRU-profiler is a rapid tool for inference of digital MIRU-VNTR profile from the assembled genome sequences. The tool can accurately infer repeat numbers at the standard 24 or 21/24 MIRU-VNTR loci from the complete or draft genomes respectively. Thus, the tool is expected to bridge the communication gap between the laboratories using WGS and those using the conventional MIRU-VNTR typing.

Project description:INTRODUCTION:Recurrent tuberculosis (TB) is one of the main challenges in TB control. Genotyping based on Mycobacterial Interspersed Repetitive Units-Variable Tandem Repeats (MIRU-VNTR) has been widely used to differentiate between relapse and reinfection, which are the two main causes of recurrent TB. There is a lack of data regarding the causes of TB recurrence in Georgia, and while differentiating between relapse and reinfection plays a key role in defining appropriate interventions, the required genotyping methodologies have not been implemented. The objective of this study was to implement MIRU-VNTR genotyping at the National Center for Tuberculosis and Lung Diseases (NCTBLD) and differentiate between relapse and reinfection in multidrug resistant (MDR-) TB patients from Tbilisi, Georgia. METHODS:Recurrent MDR tuberculosis cases from 2014-2016 diagnosed at NCTLD were included in the study when bacterial samples from both episodes were available. Genotyping based on the MIRU-VNTR 24 loci was implemented and used for differentiating between relapse and reinfection. Paired samples showing the same MIRU-VNTR pattern or one locus difference were classified as relapse, while two and more loci differences were treated as reinfection. Exact logistic regression was used to identify predictors of recurrence. RESULTS:Thirty two MDR-TB patients (64 samples) were included and MIRU-VNTR 24 typing was performed on the corresponding paired samples. Of the 32 patients, 25 (83.3%) were identified as relapse while 5 (16.7%) were due to re-infection. Patients with a history of incarceration were significantly associated with TB reinfection (p< 0.05). CONCLUSION:Recurrent TB in MDR patients in Georgia are mainly caused by relapse, raising concerns on the efficacy of the TB control program. An association between incarceration and reinfection likely reflects high levels of ongoing TB transmission in prisons, indicating the need for better TB infection control measures in these settings. Our results add to the rationale for implementing genotypic surveillance of TB more broadly to support TB control in Georgia.

Dataset Information

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

Background

Method

Results and conclusion

Publications

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

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