This preprint evaluates the use of nanopore whole-genome sequencing (WGS) for routine clinical surveillance of carbapenem-resistant Enterobacterales (CRE). The authors compared nanopore WGS results with established routine diagnostics using isolates collected during standard hospital diagnostics at a tertiary-care centre. The study explores how nanopore WGS resolves genomic context, carbapenemase subtypes, strain identity, and plasmid carriage—information that routine assays typically do not provide—and defines recommended sequencing depths for different surveillance objectives.
The authors analysed a total of 110 clinical CRE isolates. This included 100 carbapenemase-producing CRE isolates identified through routine diagnostics, and an additional set of 10 carbapenem-non-susceptible CRE isolates for which routine diagnostics did not identify a carbapenemase gene despite phenotypic non-susceptibility. The isolates were used to directly compare nanopore WGS outputs with results from established diagnostic methods and to assess the sequencing depth required for various genomic interrogations.
Across the examined collection, nanopore WGS reproduced routine laboratory detections of carbapenemase families and pathogen identification. Beyond concordance with standard diagnostics, nanopore WGS provided higher-resolution data: it resolved carbapenemase subtypes, determined the genomic context of resistance genes (for example, whether genes were chromosomal or plasmid-borne), and delivered species- and strain-level typing. The authors also report that nanopore WGS identified resistance mechanisms that were missed by established routine diagnostics in some carbapenem-non-susceptible isolates.
These added capabilities are relevant because routine diagnostic assays—while able to detect common carbapenemases—do not reconstruct the surrounding genomic sequences and therefore cannot confirm plasmid carriage, cross-species dissemination, clonal transmission, or atypical mechanisms of resistance.
The study provides explicit recommendations for minimal sequencing depth depending on the intended clinical or surveillance objective. According to the authors:
These thresholds are presented as depth targets tied to progressively demanding analytical goals: from basic species call to the highest-resolution plasmid epidemiology needed for outbreak investigation.
Within the sampled isolates, the authors observed that the detected carbapenemases were mostly carried on plasmids. Two plasmid families are specifically noted: relatively conserved IncN plasmids and more heterogeneous IncL/M plasmids. Nanopore WGS enabled the authors to resolve these plasmids' sequences and place carbapenemase genes into their genomic context, which is essential for distinguishing between spread by clonal expansion and spread via mobile genetic elements.
The paper argues that strain typing and plasmid-level resolution provided by nanopore WGS are essential for infection control responses because they allow differentiation among three distinct transmission scenarios: clonal spread of a single strain, dissemination of a shared plasmid across different strains or species, and unrelated infection events. The authors conclude that cost-efficient nanopore WGS has strong potential for integration into CRE diagnostics, surveillance, and outbreak investigation workflows, provided sequencing depth is matched to the clinical question.
This work is presented as a preprint and has not been peer reviewed. The manuscript discloses a competing interest: one author (LU) received travel expenses from Oxford Nanopore Technologies to present preliminary results of this study. Funding sources declared include the University of Zurich and Helmholtz Zentrum München. The preprint format and declared competing interest should be considered when interpreting the findings.