Wyllie, DH;
Robinson, E;
Peto, TEA;
Crook, DW;
Ajileye, A;
Rathod, P;
Allen, R;
... Walker, AS; + view all
(2018)
Identifying mixed Mycobacterium tuberculosis infection and laboratory cross-contamination during Mycobacterial sequencing programs.
Journal of Clinical Microbiology
, 56
(11)
, Article e00923-18. 10.1128/JCM.00923-18.
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Abstract
The detection of laboratory cross-contamination and mixed tuberculosis infections is an important goal of clinical mycobacteriology laboratories. The objective of this study was to develop a method to detect mixtures of different Mycobacterium tuberculosis lineages in laboratories performing mycobacterial next-generation sequencing (NGS). The setting was the Public Health England National Mycobacteriology Laboratory Birmingham, which performs Illumina sequencing on DNA extracted from positive mycobacterial growth indicator tubes. We analyzed 4,156 samples yielding M. tuberculosis from 663 MiSeq runs, which were obtained during development and production use of a diagnostic process using NGS. The counts of the most common (major) variant and all other variants (nonmajor variants) were determined from reads mapping to positions defining M. tuberculosis lineages. Expected variation was estimated during process development. For each sample, we determined the nonmajor variant proportions at 55 sets of lineage-defining positions. The nonmajor variant proportion in the two most mixed lineage-defining sets (F2 metric) was compared with that of the 47 least-mixed lineage-defining sets (F47 metric). The following three patterns were observed: (i) not mixed by either metric; (ii) high F47 metric, suggesting mixtures of multiple lineages; and (iii) samples compatible with mixtures of two lineages, detected by differential F2 metric elevations relative to F47. Pattern ii was observed in batches, with similar patterns in the M. tuberculosis H37Rv control present in each run, and is likely to reflect cross-contamination. During production, the proportions of samples in the patterns were 97%, 2.8%, and 0.001%, respectively. The F2 and F47 metrics described could be used for laboratory process control in laboratories sequencing M. tuberculosis genomes.
| Type: | Article |
|---|---|
| Title: | Identifying mixed Mycobacterium tuberculosis infection and laboratory cross-contamination during Mycobacterial sequencing programs |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1128/JCM.00923-18 |
| Publisher version: | https://doi.org/10.1128/JCM.00923-18 |
| Language: | English |
| Additional information: | Copyright © 2018 Wyllie et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) |
| Keywords: | Mycobacterium tuberculosis, next-generation sequencing, quality control, mixed infection, cross-contamination |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > Inst of Clinical Trials and Methodology UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > Inst of Clinical Trials and Methodology > MRC Clinical Trials Unit at UCL |
| URI: | https://discovery-pp.ucl.ac.uk/id/eprint/10054956 |
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