The Univeristy of Melbourne The Royal Melbourne Hopspital

A joint venture between The University of Melbourne and The Royal Melbourne Hospital

12 Dec 2017

MDU PHL transition to Whole Genome Sequencing serotyping for Shiga-toxin producing Escherichia coli

Escherichia coli is a commensal bacteria that is ubiquitous in the environment and is usually harmless. However, some strains have evolved the capability to cause disease in humans and/or animals by specific pathogenic mechanisms. Shiga toxin-producing E. coli (STEC) are a diverse group of organisms, which as per their name, produce shiga toxins which can cause severe gastrointestinal disease and haemolytic uremic syndrome (HUS). STEC infection in humans is generally via the consumption of contaminated food or water, but is also frequently associated with close contact with farm animals [1].

Serotyping is a method for characterising E. coli that has existed since the 1940s.  E. coli isolates are serologically differentiated based on three major surface antigens: the O (somatic), H (flagella), and K (capsule). The current serotyping scheme comprises 188 O groups designated O1 to O188 and 53 H antigens designated H1 to H56. PCR for the detection of virulence genes stx1, stx2, eaeA and ehxA has assisted in identifying E. coli virotypes [2].

The leading O serogroup associated STEC is O157, and is a significant food-borne pathogen worldwide. Australia’s largest outbreak of STEC, caused by STEC O157:H-, was found to be associated with feeding lambs, goats or animals and/or having hands licked by animals at the Royal Queensland show in 2013 [3]. Other notable outbreaks include a 1995 outbreak of HUS in children in Australia, which was found to be caused by a locally produced dry fermented sausage (mettwurst) contaminated with STEC O111:H- [4], and a large outbreak in Europe in 2011 caused by fenugreek sprouts contaminated with STEC O104:H4 [5].

As STEC is a notifiable disease in Victoria [6], the Microbiological Diagnostic Unit Public Health Laboratory (MDU PHL) receives E. coli isolates to determine the O:H serotype and virulence mechanisms. This facilitates the epidemiological surveillance of STEC conducted by the Victorian Government Department of Health and Human Services (DHHS) and the detection of STEC outbreaks. Although conventional serotyping of E. coli has been useful, performance of the serotyping method requires a high level of expertise, is laborious, time-consuming, expensive, and relies on the purchase or production, maintenance, and quality control of a large range of specific antisera. Whole-genome sequencing (WGS) has become an affordable, highly discriminatory tool that can be employed to determine the in silico (IS) serotype from sequencing reads.

The serotype prediction of E. coli from WGS data is based on the O-antigen processing genes wzx, wzy, wzm, and wzt and the flagellin genes fliC, flkA, fllA, flmA, and flnA [2] and is comparable to that obtained with conventional serotyping.  A significant advantage of IS serotyping over conventional serotyping is apparent where an O or H antigen cannot be determined by serotyping because a strain is phenotypically non-typable, rough (autoagglutination), or non-motile. As such, MDU PHL has now adopted WGS-based typing for STEC.  This will allow timely and discriminatory typing of STEC, which in turn, will facilitate more effective surveillance and outbreak detection of STEC in Victoria.