Diagnostic guidelines

Indications for laboratory investigation

The following situations would indicate a need for a microbiological investigation of a stool sample:

  1. Diarrhoea and visible blood in the stool OR
  2. Haemolytic uremic syndrome

In persons who have travelled to Germany in the period starting from May 1st, 2011 OR have been in close contact to a patient with HUS or STEC diarrhoea after recent travel to Germany

Recommendations for laboratory diagnosis

The most important diagnostic characteristic is the ability of the E. coli strain to produce the Shiga toxin Stx2. Therefore, the stools should be assayed with a sensitive and specific test that detects the toxin gene stx2 or the isolation of the pathogen with detection of Stx2 production or the gene encoding it. The detection of Stx using ELISA directly from stool has a low specificity.


Faecal culture screening and isolation media

No special media are required: e.g.,  MacConkey agar is suitable for isolating E.coli. Based on the particular resistance characteristics with the ability of STEC O104:H4 to produce ESBL, selective chromogenic agar media designed for screening for ESBL-positive Enterobacteriaceae can be used for the isolation of the STEC O104:H4 strain.

STEC O104:H4 on Sorbitol-MacConkey agar is Sorbitol fermenting

Shiga toxin detection

The detection of the stx2 gene should be carried out using PCR (conventional or Light-Cycler) from washed-off colonies or stool enrichment. Detection Shiga toxin Stx2 can be carried out from E. coli culture using ELISA assays according to the manufacturer’s instructions.


E.coli strain characterisation for confirmation of epidemic strain type

The further characterisation of the organism for epidemiological purposes should follow the national procedures for sending samples as appropriate to the designated national reference laboratory for EHEC/STEC/VTEC. 

An epidemic case can be microbiologically confirmed for epidemiological purposes if the STEC strain harbours the O104 serogroup or the combination of stx2 and aggR genes. Other useful strain characteristics include its multidrug resistance pattern associated with production of Extended Spectrum Beta-Lactamase (ESBL) CTX-M-15 and negative eae gene PCR result.

Rapid Antigen Screening Test for O104 serogroup

Investigations conducted at the WHO Collaborating Centre for Reference and Research on Escherichia and Klebsiella showed that the E.coli O104 antigen is identical to the K9 capsular antigen (1). Based on this property, a bacterial cell slide agglutination assay using K9 antiserum with pooled E.coli faecal isolates can be used as screening tool for the detection of E.coli O104:H4 strain from suspected outbreak cases in clinical laboratories, before sending the strains to the National Reference Laboratory for confirmation. 

1. Characteristics of the enteroaggregative Shiga toxin/verotoxin-producing Escherichia coli O104:H4 strain causing the outbreak of haemolytic uraemic syndrome in Germany, May to June 2011


Further questions

For further questions and support please contact Food and Waterborne Disease FWD-Net contact at  FWD@ECDC.europa.eu