MICROBIOLOGY Fig 1. Colorex STEC agar (E&O Labs) showing differentiation of STEC colonies.


2023, surveillance data suggested a 10- fold increase of non-O157 STEC to 9,378 isolates infecting 9,311 individuals, from 338 different serotypes, with the most common being the O26, O146, O91, O128, O145 and O103 strains.22 One significant non-O157 STEC


outbreak in Germany in 2011 led to 3,816 cases, with HUS involved in 845 cases resulting in 54 fatalities.23


The


strain associated was O104:H4 and contaminated fenugreek sprouts were identified as the infection source. This outbreak highlighted the importance of non-O157 STEC and led to international collaboration, better diagnostics, enhanced surveillance, and stricter food safety regulations.


Laboratory diagnosis of STEC Rapid, accurate, and cost-effective detection of STEC is essential for effective patient care and outbreak control. Diagnostic O157 culture and identification is routine within most laboratories, by utilising its detection as a non-sorbitol fermenter on Sorbitol-MacConkey (SMAC) agar showing colourless colonies. However, local identification past the genus level for non-O157 STEC is currently difficult and individual serotype strain identification assays are often sent to the reference laboratory for identification. The rise in non-O157 STEC cases indicate a requirement for local laboratory diagnostics for all STECs as recently indicated in the recent non-O157 NEQAS distribution in 2024. Recently Pro-Lab Diagnostics and the Scottish E.coli and STEC reference laboratory undertook a research study looking into the feasibility of deploying fast, cost-effective diagnostics for non-O157 STEC, using differential agar culture, and latex agglutination assays to enable local diagnostics. The evaluation


42


analysed 32 clinical non-O157 STEC isolates confirmed by WGS, 18 E.coli reference strains, and 14 other enteric bacteria for growth on Colorex-STEC chromogenic medium from E&O Labs. The agar supports and differentiates growth of relevant STEC strains showing them as mauve colonies, with the non- pathogenic E.coli and other enteric bacteria being blue, green, or colourless on the agar (Fig 1).24


The evaluation subsequently analysed the mauve colonies to identify the specific strain. The Prolex E.coli non-O157 latex identification assay from Pro-Lab Diagnostics successfully identified all the specific strains. There were no false positives or negatives and results showed strong agglutination. These latex agglutination assays provide results within two minutes and have been developed to identify the non-O157 STEC strains while removing the common cross reactions with similar ‘O’ and ‘H’ antigens found in E. hartmanii. The Prolex E.coli STEC O157 and Non-O157 STEC assays shown in Figure 2 provide local laboratories an affordable, reliable and quick diagnostic solution for the current STEC strains.25 The study confirmed that the agar and latex assay provides a reliable and efficient method for detecting non-O157 STEC serogroups, and along with the O157 agar and reagents, will be an aid for local laboratory STEC diagnosis. In addition to culture-based methods, toxin detection assays play an important role in STEC diagnosis. Enzyme immunoassays (EIAs) are used to directly detect Shiga toxins in stool samples, offering high sensitivity in both clinical and outbreak scenarios. Rapid tests, including lateral flow immunoassays (LFIAs), provide results within minutes, which is particularly valuable for point-of- care screening during large outbreaks.


Molecular diagnostic techniques have further advanced STEC detection. Multiplex polymerase chain reaction (PCR) assays targeting virulence genes such as stx1, stx2, and eae offer both high sensitivity and specificity, enabling rapid diagnosis directly from faecal samples. Whole-genome sequencing (WGS) provides a comprehensive genetic profile of STEC strains, revealing details on virulence factors and antimicrobial resistance, and has become an indispensable tool in outbreak investigations. However, routine screening for STEC via molecular methods may not be cost effective for individual patient management at the local laboratory level and currently should only be considered for outbreak investigations at the regional level.


Conclusions


STEC continues to pose substantial challenges in individual case management and global public health outbreaks due to its clinical severity, economic impact, and complex epidemiology. Outbreaks not only cause severe illness and lasting complications, particularly HUS among vulnerable groups, but also result in significant healthcare burdens and economic losses to the food industry, through recalls and diminished consumer confidence. The impact of STEC both O157 and in the rise in non-O157 serotypes underscores the need for efficient and effective diagnostics, enhanced surveillance, policy re-evaluation, and broader preventive measures. Encouragingly, diagnostic advances as seen above will enable fast, efficient, and affordable diagnostics to be available at the local laboratory. Moreover, the move of STEC screening away from the reference laboratory setting will enable such laboratories to concentrate on the difficult cases, allow further diagnostic development, and identify new and emerging strains. The future management of


STEC requires innovative solutions including targeted vaccines against key virulence factors, bacteriophage therapy, therapeutic antibodies, and improved environmental monitoring through metagenomics. Additionally, understanding and adapting to the impacts of climate change on STEC ecology through refined agricultural practices and water treatment are crucial. Ultimately, addressing STEC holistically demands enhanced diagnostics, sustained research, international cooperation, and adaptive, multidisciplinary approaches to effectively reduce its public health burden and economic impact.


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