Identifying Mixed Mycobacterium tuberculosis Infection and Laboratory Cross-Contamination during Mycobacterial Sequencing Programs.

Wyllie, David H and Robinson, Esther and Peto, Tim and Crook, Derrick W and Ajileye, Adebisi and Rathod, Priti and Allen, Rosemarie and Jarrett, Lisa and Smith, E Grace and Walker, A Sarah (2018) Identifying Mixed Mycobacterium tuberculosis Infection and Laboratory Cross-Contamination during Mycobacterial Sequencing Programs. Journal of clinical microbiology, 56 (11). ISSN 1098-660X. This article is available to all UHB staff and students via ASK Discovery tool http://tinyurl.com/z795c8c by using their UHB Athens login IDs

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Official URL: https://jcm.asm.org/content/56/11/e00923-18.long

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 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 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 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 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 genomes.

Item Type: Article
Additional Information: This article is available to all UHB staff and students via ASK Discovery tool http://tinyurl.com/z795c8c by using their UHB Athens login IDs
Subjects: QW Microbiology. Immunology
Divisions: Clinical Support > Pathology
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Depositing User: Beth Connors
Date Deposited: 11 Sep 2019 15:50
Last Modified: 11 Sep 2019 15:50
URI: http://www.repository.uhblibrary.co.uk/id/eprint/2382

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