German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing
We compared the consistency, accuracy and reproducibility of next-generation short read sequencing between ten laboratories involved in food safety (research institutes, state laboratories, universities and companies) from Germany and Austria. Participants were asked to sequence six DNA samples of t...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2020-09-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fmicb.2020.573972/full |
| _version_ | 1818538273588379648 |
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| author | Laura Uelze Maria Borowiak Markus Bönn Erik Brinks Carlus Deneke Thomas Hankeln Sylvia Kleta Larissa Murr Kerstin Stingl Kathrin Szabo Simon H. Tausch Anne Wöhlke Burkhard Malorny |
| author_facet | Laura Uelze Maria Borowiak Markus Bönn Erik Brinks Carlus Deneke Thomas Hankeln Sylvia Kleta Larissa Murr Kerstin Stingl Kathrin Szabo Simon H. Tausch Anne Wöhlke Burkhard Malorny |
| author_sort | Laura Uelze |
| collection | DOAJ |
| description | We compared the consistency, accuracy and reproducibility of next-generation short read sequencing between ten laboratories involved in food safety (research institutes, state laboratories, universities and companies) from Germany and Austria. Participants were asked to sequence six DNA samples of three bacterial species (Campylobacter jejuni, Listeria monocytogenes and Salmonella enterica) in duplicate, according to their routine in-house sequencing protocol. Four different types of Illumina sequencing platforms (MiSeq, NextSeq, iSeq, NovaSeq) and one Ion Torrent sequencing instrument (S5) were involved in the study. Sequence quality parameters were determined for all data sets and centrally compared between laboratories. SNP and cgMLST calling were performed to assess the reproducibility of sequence data collected for individual samples. Overall, we found Illumina short read data to be more accurate (higher base calling accuracy, fewer miss-assemblies) and consistent (little variability between independent sequencing runs within a laboratory) than Ion Torrent sequence data, with little variation between the different Illumina instruments. Two laboratories with Illumina instruments submitted sequence data with lower quality, probably due to the use of a library preparation kit, which shows difficulty in sequencing low GC genome regions. Differences in data quality were more evident after assembling short reads into genome assemblies, with Ion Torrent assemblies featuring a great number of allele differences to Illumina assemblies. Clonality of samples was confirmed through SNP calling, which proved to be a more suitable method for an integrated data analysis of Illumina and Ion Torrent data sets in this study. |
| first_indexed | 2024-12-11T21:26:55Z |
| format | Article |
| id | doaj.art-964142d9a1214b648ac9597a52bd335f |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-11T21:26:55Z |
| publishDate | 2020-09-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-964142d9a1214b648ac9597a52bd335f2022-12-22T00:50:19ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2020-09-011110.3389/fmicb.2020.573972573972German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read SequencingLaura Uelze0Maria Borowiak1Markus Bönn2Erik Brinks3Carlus Deneke4Thomas Hankeln5Sylvia Kleta6Larissa Murr7Kerstin Stingl8Kathrin Szabo9Simon H. Tausch10Anne Wöhlke11Burkhard Malorny12Department of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyLandesamt für Verbraucherschutz Sachsen-Anhalt (LAV), Halle (Saale), GermanyDepartment of Microbiology and Biotechnology, Max Rubner-Institut (MRI), Kiel, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyInstitute of Organismic and Molecular Evolution, AG Molecular Genetics and Genome Analysis, Johannes Gutenberg Universität Mainz, Mainz, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyBavarian Health and Food Safety Authority (LGL), Oberschleißheim, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyDepartment 5, Federal Office of Consumer Protection and Food Safety (BVL), Berlin, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyFood and Veterinary Institute, Lower Saxony State Office for Consumer Protection and Food Safety (LAVES), Braunschweig, GermanyDepartment of Biological Safety, German Federal Institute for Risk Assessment (BfR), Berlin, GermanyWe compared the consistency, accuracy and reproducibility of next-generation short read sequencing between ten laboratories involved in food safety (research institutes, state laboratories, universities and companies) from Germany and Austria. Participants were asked to sequence six DNA samples of three bacterial species (Campylobacter jejuni, Listeria monocytogenes and Salmonella enterica) in duplicate, according to their routine in-house sequencing protocol. Four different types of Illumina sequencing platforms (MiSeq, NextSeq, iSeq, NovaSeq) and one Ion Torrent sequencing instrument (S5) were involved in the study. Sequence quality parameters were determined for all data sets and centrally compared between laboratories. SNP and cgMLST calling were performed to assess the reproducibility of sequence data collected for individual samples. Overall, we found Illumina short read data to be more accurate (higher base calling accuracy, fewer miss-assemblies) and consistent (little variability between independent sequencing runs within a laboratory) than Ion Torrent sequence data, with little variation between the different Illumina instruments. Two laboratories with Illumina instruments submitted sequence data with lower quality, probably due to the use of a library preparation kit, which shows difficulty in sequencing low GC genome regions. Differences in data quality were more evident after assembling short reads into genome assemblies, with Ion Torrent assemblies featuring a great number of allele differences to Illumina assemblies. Clonality of samples was confirmed through SNP calling, which proved to be a more suitable method for an integrated data analysis of Illumina and Ion Torrent data sets in this study.https://www.frontiersin.org/article/10.3389/fmicb.2020.573972/fullinterlaboratory studywhole-genome sequencingfood safetyilluminaion torrent |
| spellingShingle | Laura Uelze Maria Borowiak Markus Bönn Erik Brinks Carlus Deneke Thomas Hankeln Sylvia Kleta Larissa Murr Kerstin Stingl Kathrin Szabo Simon H. Tausch Anne Wöhlke Burkhard Malorny German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing Frontiers in Microbiology interlaboratory study whole-genome sequencing food safety illumina ion torrent |
| title | German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing |
| title_full | German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing |
| title_fullStr | German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing |
| title_full_unstemmed | German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing |
| title_short | German-Wide Interlaboratory Study Compares Consistency, Accuracy and Reproducibility of Whole-Genome Short Read Sequencing |
| title_sort | german wide interlaboratory study compares consistency accuracy and reproducibility of whole genome short read sequencing |
| topic | interlaboratory study whole-genome sequencing food safety illumina ion torrent |
| url | https://www.frontiersin.org/article/10.3389/fmicb.2020.573972/full |
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