@article{Hall2022,
abstract = {BACKGROUND Mycobacterium tuberculosis whole-genome sequencing (WGS) has been widely used for genotypic drug susceptibility testing (DST) and outbreak investigation. For both applications, Illumina technology is used by most public health laboratories; however, Nanopore technology developed by Oxford Nanopore Technologies has not been thoroughly evaluated. The aim of this study was to determine whether Nanopore sequencing data can provide equivalent information to Illumina for transmission clustering and genotypic DST for M tuberculosis. METHODS In this genomic analysis, we analysed 151 M tuberculosis isolates from Madagascar, South Africa, and England, which were collected between 2011 and 2018, using phenotypic DST and matched Illumina and Nanopore data. Illumina sequencing was done with the MiSeq, HiSeq 2500, or NextSeq500 platforms and Nanopore sequencing was done on the MinION or GridION platforms. Using highly reliable PacBio sequencing assemblies and pairwise distance correlation between Nanopore and Illumina data, we optimise Nanopore variant filters for detecting single-nucleotide polymorphisms (SNPs; using BCFtools software). We then used those SNPs to compare transmission clusters identified by Nanopore with the currently used UK Health Security Agency Illumina pipeline (COMPASS). We compared Illumina and Nanopore WGS-based DST predictions using the Mykrobe software and mutation catalogue. FINDINGS The Nanopore BCFtools pipeline identified SNPs with a median precision of 99{\textperiodcentered}3{\%} (IQR 99{\textperiodcentered}1-99{\textperiodcentered}6) and recall of 90{\textperiodcentered}2{\%} (88{\textperiodcentered}1-94{\textperiodcentered}2) compared with a precision of 99{\textperiodcentered}6{\%} (99{\textperiodcentered}4-99{\textperiodcentered}7) and recall of 91{\textperiodcentered}9{\%} (87{\textperiodcentered}6-98{\textperiodcentered}6) using the Illumina COMPASS pipeline. Using a threshold of 12 SNPs for putative transmission clusters, Illumina identified 98 isolates as unrelated and 53 as belonging to 19 distinct clusters (size range 2-7). Nanopore reproduced 15 out of 19 clusters perfectly; two clusters were merged into one cluster, one cluster had a single sample missing, and one cluster had an additional sample adjoined. Illumina-based clusters were also closely replicated using a five SNP threshold and clustering accuracy was maintained using mixed Illumina and Nanopore datasets. Genotyping resistance variants with Nanopore was highly concordant with Illumina, having zero discordant SNPs across more than 3000 SNPs and four insertions or deletions (indels), across 60 000 indels. INTERPRETATION Illumina and Nanopore technologies can be used independently or together by public health laboratories performing M tuberculosis genotypic DST and outbreak investigations. As a result, clinical and public health institutions making decisions on which sequencing technology to adopt for tuberculosis can base the choice on cost (which varies by country), batching, and turnaround time. FUNDING Academy for Medical Sciences, Oxford Wellcome Institutional Strategic Support Fund, and the Swiss South Africa Joint Research Award (Swiss National Science Foundation and South African National Research Foundation).},
author = {Hall, Michael B and Rabodoarivelo, Marie Sylvianne and Koch, Anastasia S and Dippenaar, Anzaan and George, Sophie and Grobbelaar, Melanie and Warren, Robin and Walker, Timothy M and Cox, Helen and Gagneux, Sebastien and Crook, Derrick and Peto, Tim and Rakotosamimanana, Niaina and {Grandjean Lapierre}, Simon and Iqbal, Zamin},
doi = {10.1016/s2666-5247(22)00301-9},
file = {:C$\backslash$:/Users/01462563/AppData/Local/Mendeley Ltd./Mendeley Desktop/Downloaded/Hall et al. - 2022 - Evaluation of Nanopore sequencing for iMycobacterium tuberculosisi drug susceptibility testing and outbreak investi.pdf:pdf},
issn = {26665247},
journal = {The Lancet Microbe},
keywords = {OA,fund{\_}not{\_}ack,original},
mendeley-tags = {OA,fund{\_}not{\_}ack,original},
month = {feb},
number = {2},
pages = {e84--e92},
pmid = {36549315},
publisher = {Elsevier BV},
title = {{Evaluation of Nanopore sequencing for \textit{Mycobacterium tuberculosis} drug susceptibility testing and outbreak investigation: a genomic analysis}},
url = {http://www.thelancet.com/article/S2666524722003019/fulltext http://www.thelancet.com/article/S2666524722003019/abstract https://www.thelancet.com/journals/lanmic/article/PIIS2666-5247(22)00301-9/abstract},
volume = {4},
year = {2022}
}

{"_id":"niQg9pPfvPQDDpNZP","bibbaseid":"hall-rabodoarivelo-koch-dippenaar-george-grobbelaar-warren-walker-etal-evaluationofnanoporesequencingforimycobacteriumtuberculosisidrugsusceptibilitytestingandoutbreakinvestigationagenomicanalysis-2022","author_short":["Hall, M. B","Rabodoarivelo, M. S.","Koch, A. S","Dippenaar, A.","George, S.","Grobbelaar, M.","Warren, R.","Walker, T. M","Cox, H.","Gagneux, S.","Crook, D.","Peto, T.","Rakotosamimanana, N.","Grandjean Lapierre, S.","Iqbal, Z."],"bibdata":{"bibtype":"article","type":"article","abstract":"BACKGROUND Mycobacterium tuberculosis whole-genome sequencing (WGS) has been widely used for genotypic drug susceptibility testing (DST) and outbreak investigation. For both applications, Illumina technology is used by most public health laboratories; however, Nanopore technology developed by Oxford Nanopore Technologies has not been thoroughly evaluated. The aim of this study was to determine whether Nanopore sequencing data can provide equivalent information to Illumina for transmission clustering and genotypic DST for M tuberculosis. METHODS In this genomic analysis, we analysed 151 M tuberculosis isolates from Madagascar, South Africa, and England, which were collected between 2011 and 2018, using phenotypic DST and matched Illumina and Nanopore data. Illumina sequencing was done with the MiSeq, HiSeq 2500, or NextSeq500 platforms and Nanopore sequencing was done on the MinION or GridION platforms. Using highly reliable PacBio sequencing assemblies and pairwise distance correlation between Nanopore and Illumina data, we optimise Nanopore variant filters for detecting single-nucleotide polymorphisms (SNPs; using BCFtools software). We then used those SNPs to compare transmission clusters identified by Nanopore with the currently used UK Health Security Agency Illumina pipeline (COMPASS). We compared Illumina and Nanopore WGS-based DST predictions using the Mykrobe software and mutation catalogue. FINDINGS The Nanopore BCFtools pipeline identified SNPs with a median precision of 99˙3% (IQR 99˙1-99˙6) and recall of 90˙2% (88˙1-94˙2) compared with a precision of 99˙6% (99˙4-99˙7) and recall of 91˙9% (87˙6-98˙6) using the Illumina COMPASS pipeline. Using a threshold of 12 SNPs for putative transmission clusters, Illumina identified 98 isolates as unrelated and 53 as belonging to 19 distinct clusters (size range 2-7). Nanopore reproduced 15 out of 19 clusters perfectly; two clusters were merged into one cluster, one cluster had a single sample missing, and one cluster had an additional sample adjoined. Illumina-based clusters were also closely replicated using a five SNP threshold and clustering accuracy was maintained using mixed Illumina and Nanopore datasets. Genotyping resistance variants with Nanopore was highly concordant with Illumina, having zero discordant SNPs across more than 3000 SNPs and four insertions or deletions (indels), across 60 000 indels. INTERPRETATION Illumina and Nanopore technologies can be used independently or together by public health laboratories performing M tuberculosis genotypic DST and outbreak investigations. As a result, clinical and public health institutions making decisions on which sequencing technology to adopt for tuberculosis can base the choice on cost (which varies by country), batching, and turnaround time. FUNDING Academy for Medical Sciences, Oxford Wellcome Institutional Strategic Support Fund, and the Swiss South Africa Joint Research Award (Swiss National Science Foundation and South African National Research Foundation).","author":[{"propositions":[],"lastnames":["Hall"],"firstnames":["Michael","B"],"suffixes":[]},{"propositions":[],"lastnames":["Rabodoarivelo"],"firstnames":["Marie","Sylvianne"],"suffixes":[]},{"propositions":[],"lastnames":["Koch"],"firstnames":["Anastasia","S"],"suffixes":[]},{"propositions":[],"lastnames":["Dippenaar"],"firstnames":["Anzaan"],"suffixes":[]},{"propositions":[],"lastnames":["George"],"firstnames":["Sophie"],"suffixes":[]},{"propositions":[],"lastnames":["Grobbelaar"],"firstnames":["Melanie"],"suffixes":[]},{"propositions":[],"lastnames":["Warren"],"firstnames":["Robin"],"suffixes":[]},{"propositions":[],"lastnames":["Walker"],"firstnames":["Timothy","M"],"suffixes":[]},{"propositions":[],"lastnames":["Cox"],"firstnames":["Helen"],"suffixes":[]},{"propositions":[],"lastnames":["Gagneux"],"firstnames":["Sebastien"],"suffixes":[]},{"propositions":[],"lastnames":["Crook"],"firstnames":["Derrick"],"suffixes":[]},{"propositions":[],"lastnames":["Peto"],"firstnames":["Tim"],"suffixes":[]},{"propositions":[],"lastnames":["Rakotosamimanana"],"firstnames":["Niaina"],"suffixes":[]},{"propositions":[],"lastnames":["Grandjean Lapierre"],"firstnames":["Simon"],"suffixes":[]},{"propositions":[],"lastnames":["Iqbal"],"firstnames":["Zamin"],"suffixes":[]}],"doi":"10.1016/s2666-5247(22)00301-9","file":":C$\\$:/Users/01462563/AppData/Local/Mendeley Ltd./Mendeley Desktop/Downloaded/Hall et al. - 2022 - Evaluation of Nanopore sequencing for iMycobacterium tuberculosisi drug susceptibility testing and outbreak investi.pdf:pdf","issn":"26665247","journal":"The Lancet Microbe","keywords":"OA,fund_not_ack,original","mendeley-tags":"OA,fund_not_ack,original","month":"feb","number":"2","pages":"e84–e92","pmid":"36549315","publisher":"Elsevier BV","title":"Evaluation of Nanopore sequencing for <i>Mycobacterium tuberculosis</i> drug susceptibility testing and outbreak investigation: a genomic analysis","url":"http://www.thelancet.com/article/S2666524722003019/fulltext http://www.thelancet.com/article/S2666524722003019/abstract https://www.thelancet.com/journals/lanmic/article/PIIS2666-5247(22)00301-9/abstract","volume":"4","year":"2022","bibtex":"@article{Hall2022,\r\nabstract = {BACKGROUND Mycobacterium tuberculosis whole-genome sequencing (WGS) has been widely used for genotypic drug susceptibility testing (DST) and outbreak investigation. For both applications, Illumina technology is used by most public health laboratories; however, Nanopore technology developed by Oxford Nanopore Technologies has not been thoroughly evaluated. The aim of this study was to determine whether Nanopore sequencing data can provide equivalent information to Illumina for transmission clustering and genotypic DST for M tuberculosis. METHODS In this genomic analysis, we analysed 151 M tuberculosis isolates from Madagascar, South Africa, and England, which were collected between 2011 and 2018, using phenotypic DST and matched Illumina and Nanopore data. Illumina sequencing was done with the MiSeq, HiSeq 2500, or NextSeq500 platforms and Nanopore sequencing was done on the MinION or GridION platforms. Using highly reliable PacBio sequencing assemblies and pairwise distance correlation between Nanopore and Illumina data, we optimise Nanopore variant filters for detecting single-nucleotide polymorphisms (SNPs; using BCFtools software). We then used those SNPs to compare transmission clusters identified by Nanopore with the currently used UK Health Security Agency Illumina pipeline (COMPASS). We compared Illumina and Nanopore WGS-based DST predictions using the Mykrobe software and mutation catalogue. FINDINGS The Nanopore BCFtools pipeline identified SNPs with a median precision of 99{\\textperiodcentered}3{\\%} (IQR 99{\\textperiodcentered}1-99{\\textperiodcentered}6) and recall of 90{\\textperiodcentered}2{\\%} (88{\\textperiodcentered}1-94{\\textperiodcentered}2) compared with a precision of 99{\\textperiodcentered}6{\\%} (99{\\textperiodcentered}4-99{\\textperiodcentered}7) and recall of 91{\\textperiodcentered}9{\\%} (87{\\textperiodcentered}6-98{\\textperiodcentered}6) using the Illumina COMPASS pipeline. Using a threshold of 12 SNPs for putative transmission clusters, Illumina identified 98 isolates as unrelated and 53 as belonging to 19 distinct clusters (size range 2-7). Nanopore reproduced 15 out of 19 clusters perfectly; two clusters were merged into one cluster, one cluster had a single sample missing, and one cluster had an additional sample adjoined. Illumina-based clusters were also closely replicated using a five SNP threshold and clustering accuracy was maintained using mixed Illumina and Nanopore datasets. Genotyping resistance variants with Nanopore was highly concordant with Illumina, having zero discordant SNPs across more than 3000 SNPs and four insertions or deletions (indels), across 60 000 indels. INTERPRETATION Illumina and Nanopore technologies can be used independently or together by public health laboratories performing M tuberculosis genotypic DST and outbreak investigations. As a result, clinical and public health institutions making decisions on which sequencing technology to adopt for tuberculosis can base the choice on cost (which varies by country), batching, and turnaround time. 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