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Identifying mixed infection and laboratory cross-contamination during Mycobacterial sequencing programs.

Wyllie, David H and Robinson, Esther R 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 infection and laboratory cross-contamination during Mycobacterial sequencing programs. Journal of clinical microbiology. ISSN 1098-660X.

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Abstract

Detecting laboratory cross-contamination and mixed tuberculosis infection are important goals of clinical Mycobacteriology laboratories. To develop a method detecting mixtures of different lineages in laboratories performing Mycobacterial next generation sequencing (NGS). Public Health England National Mycobacteriology Laboratory Birmingham, which performs Illumina sequencing on DNA extracted from positive Mycobacterial Growth Indicator tubes. We analysed 4,156 samples yielding from 663 MiSeq runs, obtained during development and production use of a diagnostic process using NGS. Counts of the most common (major) variant, and all other variants (non-major variants) were determined from reads mapping to positions defining lineages. Expected variation was estimated during process development. For each sample we determined the non-major variant proportions at 55 sets of lineage defining positions. The non-major variant proportion in the two most mixed lineage defining sets (F2 metric) was compared with that in the 47 least mixed lineage defining sets (F47 metric). 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 elevation 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 each pattern were 97%, 2.8%, and 0.001%, respectively. The F2 and F47 metrics described could be used for laboratory process control in laboratories sequencing .

Item Type: Article
Subjects: QW Microbiology. Immunology
Divisions: Clinical Support > Pathology
Related URLs:
Depositing User: Miss Emily Johnson
Date Deposited: 14 Sep 2018 12:43
Last Modified: 14 Sep 2018 12:43
URI: http://www.repository.heartofengland.nhs.uk/id/eprint/1736

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