Gene transfer from NDM-5-producing and OXA-48-producing Enterobacter hormaechei ST79 on contaminated dicloxacillin capsules to other Enterobacterales in Europe, 2020-23: a retrospective, observational, molecular epidemiological study

BACKGROUND: In February, 2023, an outbreak of Enterobacter hormaechei ST79 carrying blaNDM-5 and blaOXA-48 was linked to contaminated dicloxacillin capsules administered to approximately 79 000 individuals in Denmark. Initial clonal outbreak investigations identified 11 patients with the E hormaechei ST79 outbreak strain, which carried blaNDM-5 on a distinct IncX3 plasmid, and in nine cases, blaOXA-48 was on a distinct IncL plasmid. Interspecies plasmid transfer was observed in one patient, suggesting a potential plasmid-mediated outbreak involving other Enterobacterales. However, no studies have characterised the progression of a clonal outbreak originating from a contaminated medicine into plasmid-mediated dissemination of carbapenemase genes. Hence, we aimed to characterise the clonal and plasmid-mediated spread of carbapenemase genes in this outbreak.

METHODS: We conducted a retrospective genomic and epidemiological investigation using existing short-read whole-genome sequencing data from all carbapenemase-producing Enterobacterales (CPE) from the Danish national surveillance, collected between Jan 1, 2014, and Oct 1, 2023. All confirmed CPE isolates were eligible for inclusion. Using in-silico screening for unique fragments of the two outbreak plasmids, we selected 160 isolates for long-read sequencing to obtain complete plasmid sequences for outbreak investigation. Analyses were descriptive and included comparison of sequence identity and coverage to define outbreak-associated plasmids and summary statistics of patient characteristics.

FINDINGS: Data from 1829 isolates were obtained. We detected 16 of 53 isolates involved in the outbreak using conventional outbreak detection methods. The remaining 37 isolates were detected using plasmid-specific screening and long-read sequencing. 15 patients carried the outbreak E hormaechei strain, including the 11 patients previously reported. Three of the 15 patients presented with at least one additional bacterial species carrying one or both outbreak plasmids (pDcap_OXA-48 and pDcap_NDM-5). A further 24 patients, sampled between July 1, 2020, and Oct 1, 2023, presented with other Enterobacterales carrying one or both outbreak-associated plasmids but not the original E hormaechei ST79 strain. In four cases, outbreak-associated plasmids differed structurally from the original outbreak plasmid.

INTERPRETATION: This study describes how a clonal CPE outbreak caused by a contaminated medicine evolved into a complex plasmid-mediated outbreak involving multiple Enterobacterales species. Most patients related to the outbreak did not present with the original E hormaechei ST79 outbreak strain and were therefore not identified using standard outbreak detection methods. These findings highlight the importance of using plasmid-focused approaches in outbreak investigations.

FUNDING: The Danish Ministry of Health, SSI-Seq (cofunded by EU4Health).