Rapid Salmonella Detection
Using the R.A.P.I.D. LT
Food Security System
idahotechnology.com
About 40,000 cases of Salmonella poisoning are reported in the
United States each year. Of these cases, approximately 400 people
die.1 Food producers need to protect themselves by ensuring that
processed and prepared foods are not contaminated with Salmonella. They
need to be able to detect Salmonella earlier and, in many cases, increase
the quantity of testing
without sacrificing sensitivity or specificity. Meeting both technical and
budget requirements can
be very challenging.
To enable faster, less
expensive testing without sacrificing quality,
Idaho Technology developed the Salmonella
Food Security System
(FSS), a PCR-based detection method that rapidly and specifically
identifies Salmonella
species in food. The system consists of the
R.A.P.I.D.® LT thermocy-cler and the AOAC-ap-proved Salmonella LT
test kit. Thermocycling
takes only 35 minutes,
and the entire procedure takes 17 hours. The method involves a 16-hour
sample enrichment, mechanical lysis to release DNA, DNA amplification
(PCR), internal amplification controls, and automatic result interpretation by
software. Samples can be tested individually or pooled for an increased
throughput. The system may also be used and is AOAC approved with the
Matrix MicroScience Pathatrix, a magnetic bead-based technology used to
selectively bind Salmonella in the presence of complex matrices.
The FSS is approved for both single and pooled samples. Post-enrich-ment sample pooling is an excellent way to increase throughput and decrease costs for mostly negative samples. It is cost efficient when more
than 90% of samples are negative. In this pooling scheme, samples are individually enriched and then pooled into a composite sample. The composite sample is then tested in the same way as an individual sample. If the
composite sample is called positive, the individually enriched samples may
be tested to determine which of those samples is positive. This process
takes less than an hour due to the short 35-minute PCR run.
To qualify for AOAC approval, the system was evaluated for sensitivity,
specificity, ruggedness, and stability of reagents. Three food types were
used––ham, chicken, and chocolate––and test results were compared to
reference methods. Each food type was divided into two portions: one portion of the food type was not inoculated, and the second portion was inoc-
ulated in a large batch so as to provide
samples for testing by both the FSS and
reference methods. Each food matrix was
inoculated with a different Salmonella en-terica serova, each chosen because they
have been responsible for foodborne illness or associated with recent outbreaks:
Salmonella Enteritidis with cooked ham;
Salmonella Typhimurium with raw chicken;
and Salmonella Senftenberg with chocolate. Testing was performed with both
pooled samples ( 50 samples per food
type combined in one container) and individual samples ( 25 samples per food
type).
In both individual and pooled samples
with and without the Pathatrix, the FSS
demonstrated 100% sensitivity and 100%
specificity for the three different food
types. It also proved to be robust and reproducible. The ruggedness study
demonstrated that the system produced
consistent results even with variability in
reagent preparation time and pipetted
sample volumes. Lot-to-lot and shelf-life
studies demonstrated consistent results
with several lots of reagents produced at
different times.
This system represents a significant
improvement over standard methods in a
number of ways:
• It is significantly faster, providing results in about 17 hours as opposed to
72 hours for the USDA and FDA BAM
methods,
2,
3 and can perform real-time
PCR and provide automated results in
35 minutes after enrichment and sample processing.
• It is equivalent to current USDA and
FDA BAM official methods used to detect low levels of Salmonella in food
(the system can detect a single bacterium in a 25-gram food sample).
• Pooling samples yielded the same results as individual samples and
demonstrated no false positives or
false negatives. Pooling enables more
testing at less cost.
• The system is easy to use, with fewer
steps and minimal sample handling.
This method promises to help food
producers improve the quality and quantity of testing to protect their companies
and their consumers from Salmonella
contamination.
References
1. http://www.cdc.gov/nczved/dfbmd/
disease_listing/ salmonellosis_gi.html#8
2. http://www.cfsan.fda.gov/~ebam/bam- 5.html
3. http://www.fsis.usda.gov/PDF/MLG_ 4_03.pdf
