Hybrid strains of enterotoxigenic/Shiga toxin-producing , United Kingdom, 2014-2023.

Diarrhoeagenic (DEC) pathotypes are defined by genes located on mobile genetic elements, and more than one definitive pathogenicity gene may be present in the same strain. In August 2022, UK Health Security Agency (UKHSA) surveillance systems detected an outbreak of hybrid Shiga toxin-producing /enterotoxigenic (STEC-ETEC) serotype O101:H33 harbouring both Shiga toxin () and heat-stable toxin (). These hybrid strains of DEC are a public health concern, as they are often associated with enhanced pathogenicity. However, little is known about their epidemiology, clinical significance and associated public health burden. The aim of this study was to describe the microbiology, epidemiology and genomic analysis of this novel hybrid serotype in the context of the STEC-ETEC strains in the UKHSA archive. From 2014 to 2023, STEC isolated from faecal specimens testing positive for STEC by PCR were sequenced on the NextSeq 1000 short read platform and a subset were selected for long read nanopore sequencing. Genomes were analysed to determine serotype, subtype, DEC pathogenicity genes and antimicrobial resistance determinants. There were 162 STEC-ETEC strains isolated between 2014 and 2023, of which 117/162 were human clinical isolates and 45 were of food or animal origin. An average of 16 STEC-ETEC strains were identified each year, exhibiting a range of different subtypes, the most common profiles being (=65, 40%) and (=48, 30%). The most common sequence types were ST329 and ST200 (=24 each), and the most frequently detected serotype was O187:H28 (=25). Nine cases of genetically linked STEC-ETEC O101:H33, were detected between 8 August and 21 September 2022. Although the temporal and geographical distribution of the cases was characteristic of a foodborne outbreak, the contaminated vehicle was not identified. Phylogenetic analysis and long-read sequencing of the outbreak strain provided insight into the stepwise acquisition of and and the evolutionary history of STEC-ETEC pathotypes. The integration of epidemiological data and whole-genome sequencing for routine surveillance of gastrointestinal pathogens is key to understanding the emergence of zoonotic hybrid DEC pathotypes and monitoring foodborne threats to public health.