E coli O157:H7 PCR test for horses
equine assay data sheet
B0059 - Ultrasensitive qualitative detection of
E. coli strain O157:H7 by
real time polymerase chain reaction. This assay is specific for
E. coli strain O157:H7 and
does not detect other unrelated E.
Test B0059 is
- waterborne pathogens screening panel
serotype O157:H7 is a gram-negative, rod-shaped bacterium and is one
of hundreds of serotypes of the bacterium
E. coli. While most
strains are harmless and are found normally in the intestines of
mammals, strain O157:H7 produces shiga-like toxins, causes severe
illness, and is a member of a class of pathogenic E. coli known as
enterohemorrhagic Escherichia coli
(EHEC). They are sometimes also referred to by their toxin producing
capabilities, eg “verocytotoxin producing
E. coli” (VTEC) or
“shiga-like toxin producing E.
from other pathogenic E. coli
in that it is not invasive, elaborates no colonization
factors (CFA/I nor CFA/II), does not produce heat stable or heat
labile toxins and is non-hemolytic. In addition,
E. coli O157:H7 is
sorbitol negative whereas 93% of all
E. coli ferment sorbitol.
E. coli O157:H7 also does not hydrolyze
4-methylumbelliferyl-ß-D-glucuronide (MUG), nor does it grow at 45ºC
in the presence of 0.15% bile salts. Because of the latter
characteristic this serotype cannot be isolated by using standard
fecal coliform methods that include incubation at 45ºC.
O157:H7 can be transmitted in food or drinking water, and outbreaks of
E. coli O157:H7
infection have been attributed to the presence of this bacterium in
groundwater and surface water (Chalmers et al. 2000; Lee et al. 2002).
A likely source of contamination of aquatic systems is cattle manure
and agricultural runoff. The intestinal cells of cattle, swine and
deer lack the highly specific surface receptors that the toxin
requires in order to attach and enter the cell. Therefore these
animals do not exhibit disease when infected (Pruimboom-Brees et al.
2000) and can be carriers, shedding the organism in their feces.
O157:H7 persists in cattle manure (Wang et al. 1996; Bolton et al.
1999; Fukushima et al. 1999; Osek 2002) and manure-amended soil (Jiang
et al. 2002) and experiments with models have suggested that it may
leach through soil (Gagliardi and Karns 2000). Meat can become
contaminated during slaughter, and the organisms can be thoroughly
mixed into beef when it is ground. Bacteria present on cows’ udders or
on equipment may get into raw milk. Although the number of organisms
required to cause disease is not known, it is suspected to be very
A major source of
human infection is undercooked ground beef; other sources include
consumption of unpasteurized milk and juice, raw sprouts, lettuce and
salami, as well as contact with infected live animals. Waterborne
transmission occurs through swimming in contaminated lakes or pools,
or drinking inadequately treated water. The organism is easily
transmitted from person to person and has been difficult to control in
child daycare centers. Infection often causes severe, acute bloody
diarrhea (although nonbloody diarrhea is also possible) and abdominal
cramps. Usually little or no fever is present, and the illness
resolves in 5 to 10 days. It can also be asymptomatic.
Rapid methods to
detect E. coli O157:H7
are important to identify the source of outbreaks. Both molecular and
culture-based methods have been used for the detection of
E. coli O157:H7.
Culture-based methods developed for clinical samples have been applied
to environmental samples. These methods rely on enrichment cultures
followed by confirmation based on metabolic and antigenic properties.
A disadvantage of this approach is the lack of complete correlation of
these antigenic and metabolic properties with Shigella toxin (stx)
production (Karch and Bielaszewska 2001). Also, culture-based methods
for the detection of O157:H7 are slow and labor intensive, so they are
not ideal for analysis of the large numbers of samples that must be
tested when possible environmental sources of an outbreak are being
As the infectious
dose is very small and the number of cells contaminating environmental
samples or food may be low, immunomagnetic capture with anti-O157
antibody has been suggested as a means to concentrate and detect the
target cells (Pyle et al. 1999). However, this approach is limited to
cells displaying a specific antigen, making it unsuitable for other
STEC. Molecular approaches for bacterial detection avoid the need for
culture and can be designed to be more specific. Primers targeting
stx1, stx2 and other genes specific to
E. coli O157:H7 have been
used in PCR and real time PCR (Olsvik and Strockbine 1993; Fratamico
et al. 2000; Fortin et al. 2001; Li and Drake 2001; Ibekwe et al.
2002; Ibekwe and Grieve 2003). Specific, rapid diagnosis of
E. coli O157:H7 is
possible using PCR techniques (Al-Ajmi et al., 2006; Holicka et al.,
Help confirm the disease causing agent
Shorten the time required to confirm a clinical
diagnosis of E. coli
Help ensure that horse populations are free of
E. coli O157:H7
Early prevention of spread of this bacterial strain
Minimize human exposure to this bacterial strain
Al-Ajmi, D., Padmanabha, J., Denman, S.E., Gilbert, R.A., Al Jassim,
R.A. and McSweeney, C.S.(2006) Evaluation of a PCR detection method
for Escherichia coli O157:H7/H- bovine faecal samples. Lett Appl
Holicka, J., Guy,
R.A., Kapoor, A., Shepherd, D. and Horgen, P.A. (2006) A rapid (one
day), sensitive real-time polymerase chain reaction assay for
detecting Escherichia coli O157:H7 in ground beef. Can J Microbiol.
Rectal swab, or 0.2 ml feces, or 0.2 ml bacterial culture,
Contact Zoologix if advice is needed to determine an appropriate specimen type for a specific diagnostic application. For specimen types not listed here, please contact Zoologix to confirm specimen acceptability and shipping instructions.
For all specimen
types, if there will be a delay in shipping, or during very warm
weather, refrigerate specimens until shipped and ship with a cold pack
unless more stringent shipping requirements are specified. Frozen
specimens should be shipped so as to remain frozen in transit. See
shipping instructions for more
2 business days
Qualitative real time