Food Safety Magazine, April/May 2012

O157:H7 does not determine the status of non-O157 STEC. Knowing this, our new challenge is similar to our experience in the 1990s with O157:H7—we must undertake the effort necessary to detect and control these additional six pathogenic STEC.

At this point, I want to be clear that the remainder of this article addresses only testing for non-O157 STEC. The reason for this distinction is that there are subtle differences between methods for O157:H7 testing compared with non-O157 STEC testing.

Developing STEC Testing Methods

Now that we understand the characteristics of pathogenic STEC, it’s important to differentiate between pathogenic STEC and other STEC. Surprisingly, stx-

Challenges for Non-O157 STEC Testing

producing E. coli that do not have the eae component are not typically associated with severe illness. Classified as STEC, there are literally hundreds of stx-containing E. coli serotypes that actually will pass through the digestive system harmlessly, simply because they lack the adhesion factor—eae. It really is remarkable, but when the adherence factor eae is missing, the bacteria cannot attach to the intestinal wall and are removed from the body by natural processes. (One notable exception to this oversimplification, as punctuated

Food Safety Magazine asked various companies within the food industry, What are the major challenges involved in non-O157 STEC testing? Several responses are provided below:

“To understand the

“Developing reagents that specifically detect the gene targets in the adulterated strains named by the U.S. Department of Agriculture, while at the same time minimizing cross-reactivity with detection of these same gene targets from nonpathogenic background or near neighbor organisms that can also be found in the beef sample. We accomplished this by integrating immunomagnetic separation prior to qPCR and using highly specific chemistry qPCR detection assays. Our workflow can help to minimize the potential for these false positives, while also providing the concentration and sensitivity needed for confident clearance of negative 375-g beef samples in as little as 8 hours.”

challenges of testing
for pathogenic
STEC, we need to
understand the
mechanics of what

“The biggest concern right now is the development of a defined and
industry-consistent process regarding the next steps companies should take if
they get a STEC suspect on the available PCR screening methods.”
“Will the industry want to carry these through to cultural confirmation and
if so, what is the expectation of industry regarding turnaround time and the
effect this will have on product disposition?”
“The biggest challenge to developing tests for non-O157 STEC is insufficient
understanding of all of the genetics relating to virulence factors. A single
genetic marker that can separate virulent from avirulent strains has not
yet been identified. Methods available today require a stepwise series of
screenings looking for several genetic markers. All of these markers must be
present. This screening approach adds a significant amount of time to the
testing process and to getting a final test result.”
“A key challenge in testing for the ‘Top Six’ non-O157 STEC is the inability to
easily and efficiently distinguish them from harmless bacteria using currently
available diagnostic tools. This is based on the fact that nonpathogenic
bacteria can naturally carry the same diagnostic markers associated with
non-O157 STEC, yielding similar reactions that may implicate large amounts
of products that are not contaminated with non-O157 STEC. Accordingly,
additional analyses would be needed to determine if pathogenic bacteria are
present, which at the time is limited to technically demanding, inefficient and
expensive cultural confirmation techniques. The industry will benefit greatly
from the much anticipated advances that can more efficiently and effectively
differentiate non-O157 STEC from harmless bacteria that normally occur in
agricultural products.”

makes them deadly.”

by the summer 2011 sprouts outbreak in Germany and Europe, is E. coli 104:H4, which is characterized by stx combined with non-eae adherence mechanisms.)

One last point we should cover at this stage is the potential sources of eae that are not STEC. There are many types of eae and many potential bacterial sources of it. There are also several variants of stx. So our challenge of testing a sample to determine if it contains pathogenic STEC is complicated, because we now know that stx and eae could potentially come from two or more bacterial sources and the presence of both virulence factors alone in a composite sample is not necessarily cause for concern. A successful test must determine if both stx and eae are present and also that both originate from a single E. coli bacterial source, specifically E. coli O26, O45, O103, O111, O121 or O145.