Impact of wet-lab protocols on quality of whole-genome short-read sequences from foodborne microbial pathogens.

Autor:	Forth LF; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Brinks E; Institute of Microbiology and Biotechnology, Max Rubner-Institut, Kiel, Germany., Denay G; Chemical and Veterinary Analytical Institute Rhein-Ruhr-Wupper (CVUA-RRW), Krefeld, Germany., Fawzy A; Department of Veterinary Medicine, Hessian State Laboratory, Giessen, Germany.; Department of Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Cairo University, Giza, Egypt., Fiedler S; Method Standardisation, Reference Laboratories, Resistance to Antibiotics, Federal Office of Consumer Protection and Food Safety, Berlin, Germany., Fuchs J; Chemical and Veterinary Analysis Agency Karlsruhe, Karlsruhe, Germany., Geuthner AC; Department of Food Safety, State Office for Consumer Protection Saxony-Anhalt, Halle, Germany., Hankeln T; Institute of Organismic and Molecular Evolution, Johannes Gutenberg University Mainz, Mainz, Germany.; StarSEQ GmbH, Mainz, Germany., Hiller E; Chemical and Veterinary Analysis Agency Stuttgart, Fellbach, Germany., Murr L; Bavarian Health and Food Safety Authority, Oberschleißheim, Germany., Petersen H; Chemical and Veterinary Analytical Institute Ostwestfalen-Lippe, Detmold, Germany., Reiting R; Hessian State Laboratory, Kassel, Germany., Schäfers C; Institute for Hygiene and Environment, Hamburg, Germany., Schwab C; Labor Kneißler GmbH & Co. KG, Burglengenfeld, Germany., Szabo K; Method Standardisation, Reference Laboratories, Resistance to Antibiotics, Federal Office of Consumer Protection and Food Safety, Berlin, Germany., Thürmer A; Methods Development and Research Infrastructure, Robert Koch-Institut, Berlin, Germany., Wöhlke A; Food and Veterinary Institute Braunschweig/Hannover, Lower Saxony State Office for Consumer Protection and Food Safety, Braunschweig, Germany., Fischer J; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Lüth S; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Projahn M; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Stingl K; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Borowiak M; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Deneke C; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Malorny B; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany., Uelze L; Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany.
Jazyk:	angličtina
Zdroj:	Frontiers in microbiology [Front Microbiol] 2023 Nov 29; Vol. 14, pp. 1253362. Date of Electronic Publication: 2023 Nov 29 (Print Publication: 2023).
DOI:	10.3389/fmicb.2023.1253362
Abstrakt:	For successful elucidation of a food-borne infection chain, the availability of high-quality sequencing data from suspected microbial contaminants is a prerequisite. Commonly, those investigations are a joint effort undertaken by different laboratories and institutes. To analyze the extent of variability introduced by differing wet-lab procedures on the quality of the sequence data we conducted an interlaboratory study, involving four bacterial pathogens, which account for the majority of food-related bacterial infections: Campylobacter spp., Shiga toxin-producing Escherichia coli , Listeria monocytogenes , and Salmonella enterica . The participants, ranging from German federal research institutes, federal state laboratories to universities and companies, were asked to follow their routine in-house protocols for short-read sequencing of 10 cultures and one isolated bacterial DNA per species. Sequence and assembly quality were then analyzed centrally. Variations within isolate samples were detected with SNP and cgMLST calling. Overall, we found that the quality of Illumina raw sequence data was high with little overall variability, with one exception, attributed to a specific library preparation kit. The variability of Ion Torrent data was higher, independent of the investigated species. For cgMLST and SNP analysis results, we found that technological sequencing artefacts could be reduced by the use of filters, and that SNP analysis was more suited than cgMLST to compare data of different contributors. Regarding the four species, a minority of Campylobacter isolate data showed the in comparison highest divergence with regard to sequence type and cgMLST analysis. We additionally compared the assembler SPAdes and SKESA for their performance on the Illumina data sets of the different species and library preparation methods and found overall similar assembly quality metrics and cgMLST statistics. Competing Interests: ClS was employed by the company Labor Kneißler GmbH & Co. KG. TH was emplyoed by StarSEQ GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. (Copyright © 2023 Forth, Brinks, Denay, Fawzy, Fiedler, Fuchs, Geuthner, Hankeln, Hiller, Murr, Petersen, Reiting, Schäfers, Schwab, Szabo, Thürmer, Wöhlke, Fischer, Lüth, Projahn, Stingl, Borowiak, Deneke, Malorny and Uelze.)
Databáze:	MEDLINE
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