Real-Time PCR Test Kit
Provides Faster Time to
Results
Salmonella is a bacterium that causes one of the most common enteric (intestinal) infections in the United States—salmonellosis. In
some states, salmonellosis is the most commonly reported cause
of enteric disease, and overall it is the second most common bacterial
foodborne illness reported (usually slightly less frequent than
Campylobacter infection). The reported incidence of Salmonella illnesses
is about 14 cases per each 100,000 persons, amounting to approxi-
mately 30,000 confirmed cases of salmonellosis yearly in the U.S. according to
the CDC in 2005. In this year alone, just
over 36,000 cases were reported from
public health laboratories across the nation, representing a 12% decrease compared with the previous decade, but a 1.5
percent increase over 2004 according to
the CDC last year. As only about 3 percent of Salmonella cases are officially reported nationwide, and many milder
cases are never diagnosed, the true incidence is undoubtedly much higher. The CDC estimates that 1.4 million
cases occur annually.
Cargill Chocolate Products, a producer of chocolate for the chocolate
industry, sought to implement an ISO method for the detection of
Salmonella in their finished products as well as raw materials, including cocoa
liquor, cocoa butter, milk powders (whole and skim milk, lactose and
whey powder) and emulsifiers. The company’s products range from plain,
milk and white chocolate for liquid delivery to solid products in block-form, buttons, sprinkles, drops, organic chocolate, vegetable fat-based
fillings, yoghurt compounds and fillings. Cargill is one of the world’s
major originators and processors of cocoa beans, manufacturing cocoa
products as well as industrial chocolate, which are used by food and
confectionery companies to produce cakes, biscuits, confectionery and
other products for consumers around the world.
With 400 employees at six European production facilities, it is imperative for Cargill Chocolate Products to maintain a strict control safety regimen for detection of microbial pathogens. As Salmonella is such a major
problem for food processors and manufacturers, they needed to have the
most robust testing method available for their raw materials and finished
products.
foodscience.bio-rad.com
A Search for a Better Method
Previously, the company was using a Salmonella-derivative method
that allowed for the generation of results in 48 hours, including a pre-en-richment in milk for 20 hours, doubling of cell numbers for 8 hours in
Rappaport-Vassiliadis media and incubation on XLD and Hektoen plates
for 20 hours. An additional 24 hours was then needed for the confirmation of suspicious colonies with a rapid biochemical identification kit.
Although their microbial laboratories were accredited according to
ISO17025 and the Salmonella ring tests were extremely reliable, there
was a general discomfort among some of their customers as the test
used was not an approved ISO method.
PCR technology has allowed a tremendous increase in detection sensi-
tivity and, therefore, a PCR-based test
was investigated. As luck would have it,
cocoa molecules can inhibit PCR and,
once again, the company had to embark
on a search for possible solutions to the
problem of Salmonella testing in choco-late-based products.
A collaboration between Cargill and
Bio-Rad Laboratories solved the problem.
The use of the iQ-Check™ Salmonella I kit
from Bio-Rad succeeded for most of the
matrixes. Although the use of cocoa powder inhibited the PCR, the new generation
Bio-Rad kit, together with the expertise
and experience of Cargill Chocolate Products, was able to overcome this additional
challenge. Today, no interference from either cocoa liquor or cocoa powder matrix
is noticed in the testing for Salmonella.
This real-time PCR kit allows rapid
control and reaction times in HACCP procedures. Using gene-specific probes and
primers enhances reproducibility while increasing sensitivity and specificity and
PCR internal controls ensure accurate results. As a high-throughput methodology,
94 samples can be processed simultaneously, and the test can be performed in
parallel with other iQ-Check tests on the
96-well real-time instrument (tests are
available for E. coli O157:H7, Listeria spp.
and Listeria monocytogenes). Bio-Rad
also offers a smaller 48-well real-time instrument, for laboratories with lower
throughput. Ready-made reagents allow
for simple extraction, amplification and
detection. As a result of these advantages,
tested products can be released more
quickly, saving storage costs for the company. The test ensures reliable results in
less than 24 hours, one-third the time of
Cargill’s former method.
Redouan Sadiky, in charge of all
Cargill Chocolate Products European laboratories, states, “The Cargill method is
very easy for a technician to learn and
provides us quicker results compared
with our old method. Raw material is released one day quicker, resulting in a
lower raw material stock need and cost
savings. Our customers are more confident with the PCR-based method compared with our internal method.”
After the success of the iQ-Check™
Salmonella II kit in the French factory,
Cargill Chocolate decided to implement
the Bio-Rad test and corresponding
equipment in each of the other chocolate
factories (in Belgium, the UK, and Germany) to perform Salmonella testing in
both raw materials and finished products.
