# Isolation and Characterization Through Whole-Genome Sequencing of STEC Strains from Free-Ranging Red Deer

## TL;DR

This study identifies and genetically characterizes STEC strains in free-ranging red deer in the Italian Alps, highlighting their potential role in transmitting these food-borne pathogens to humans.

## Contribution

The study reports the isolation and whole-genome sequencing of STEC strains from wild red deer, revealing cross-pathotype virulence features and genetic diversity.

## Key findings

- STEC strains were isolated from 12% of red deer fecal samples, showing diverse genetic subtypes.
- One isolate belonged to serotype O26:H11 and carried the stx2a and eae genes, linked to severe human disease.
- Some isolates displayed shuffled virulence features of multiple E. coli pathotypes.

## Abstract

Shiga toxin-producing Escherichia coli (STEC) are important human food-borne pathogens and red deer have been recently identified as STEC carriers. This study aimed to isolate and characterize by whole-genome sequencing (WGS) STEC strains from free-ranging red deer in the central Italian Alps. Fecal samples from 92red deer hunted between September and December 2022 were analyzed by bacteriology and WGS analysis. STEC strains were isolated from 11 (12%) samples. Different genetic features were identified in STEC strains, which included one isolate belonging to serotype O26:H11 and possessing the stx2a subtype along with the eae gene, frequently associated with severe disease in humans. Our results show a wide range of strain types, including some with cross-pathotype features, present in a red deer population from the central Italian Alps. Results underscore red deer as STEC carriers and confirm the need of STEC monitoring in wild ruminants to understand the potential role for these animals in the transmission of infections to humans.

Whole-genome sequence (WGS) analysis was used in this study to characterize Shiga toxin-producing Escherichia coli (STEC) isolates in free-ranging red deer from the central Italian Alps. Fecal samples from 92 hunted red deer collected between September and December 2022 were analyzed for the presence of STEC. Single E. coli colonies positive by PCR for stx genes were analyzed by WGS. STEC were isolated from eleven (12%) samples, showing eight stx2b, one stx2a, two stx1c, and one stx1a subtypes. Different serotypes and sequence types were identified (n = 8 each). Three isolates of O27:H30 serotype and ST753 showed no correlation in the cgMLST analysis (AD range 44–98). All strains harbored additional virulence factors. The only isolate harboring stx2a also possessed the eae gene and belonged to serotype O26:H11. Some isolates displayed shuffled virulence features of more than one E. coli pathotype. The high genetic diversity of strains circulating in the red deer population living in the central Italian Alps, including the STEC O26:H11 strain associated with STEC from severe disease in humans, confirms red deer as STEC reservoirs and highlights the need for monitoring the presence of these pathogens in wild ruminants.