RT Journal
A1 Lohde, Mara
A1 Wagner, Gabriel E.
A1 Dabernig-Heinz, Johanna
A1 Viehweger, Adrian
A1 Braun, Sascha D.
A1 Monecke, Stefan
A1 Diezel, Celia
A1 Stein, Claudia
A1 Marquet, Mike
A1 Ehricht, Ralf
A1 Pletz, Mathias W.
A1 Brandt, Christian
T1 Accurate bacterial outbreak tracing with Oxford Nanopore sequencing and reduction of methylation-induced errors
JF Genome Research 
JO Genome Research 
YR 2024 
FD November 01 
VO 34 
IS 11 
SP 2039 
OP 2047 
DO 10.1101/gr.278848.123 
UL http://genome.cshlp.org/content/34/11/2039.abstract 
AB Our study investigates the effectiveness of Oxford Nanopore Technologies for accurate outbreak tracing by resequencing 33 isolates of a 3-year-long Klebsiella pneumoniae outbreak with Illumina short-read sequencing data as the point of reference. We detect considerable base errors through cgMLST and phylogenetic analysis of genomes sequenced with Oxford Nanopore Technologies, leading to the false exclusion of some outbreak-related strains from the outbreak cluster. Nearby methylation sites cause these errors and can also be found in other species besides K. pneumoniae. Based on these data, we explore PCR-based sequencing and a masking strategy, which both successfully address these inaccuracies and ensure accurate outbreak tracing. We offer our masking strategy as a bioinformatic workflow (MPOA) to identify and mask problematic genome positions in a reference-free manner. Our research highlights limitations in using Oxford Nanopore Technologies for sequencing prokaryotic organisms, especially for investigating outbreaks. For time-critical projects that cannot wait for further technological developments by Oxford Nanopore Technologies, our study recommends either using PCR-based sequencing or using our provided bioinformatic workflow. We advise that read mapping–based quality control of genomes should be provided when publishing results.