[LABORATORY]
by Linda L. Leake
Purdue University microbiology professor Arun Bhunia (standing) and
postdoctoral researcher Padmapriya Banada are using a
system that analyzes scattered laser light to quickly identify
bacteria in food samples.
Photo by Tom Campbell, courtesy of Purdue University
Advancing Rapid
Microbial Testing
S
o many rapid microbial tests, so little time. That’s
at once the dream come true and dream to come
of scientists everywhere. Faster methods and
continually emerging technologies have revolutionized microbial testing in recent years, slashing first
hours, then days, off the time to get results.
Rapid testing is gaining momentum nationally and
internationally as an area of research and as a reliable
tool to monitor the kinds, numbers, and metabolites
of microorganisms related to food spoilage, food
preservation, food fermentation, and food safety, says
Daniel Y. C. Fung (danfung@k-state.edu), Professor
of Food Science in the Dept. of Animal Sciences and
Industry at Kansas State University, Manhattan,
and a leading authority on rapid microbial testing.
Faster methods and continually emerging technologies
have revolutionized microbial testing in recent years.…
Routinely traversing the globe to give lectures, conduct workshops, and discuss the role of rapid testing of
food for safety and security, as well as of water, environmental, air, industrial, and medical specimens, Fung lives
and breathes 24/7 with his finger on the pulse of what’s
happening all over the planet relative to this hot topic.
“No one knows exactly how many microbiopg
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logical tests are done annually in the world, but the
number is increasing,” Fung says. “Based on data
available at this moment, about 33% of all testing
is done in North America, 33% in Europe, and 33%
in the rest of the world. But in the next 10–15 years,
the dynamics will change to 25% in North America,
25% in Europe, and 50% in the rest of the world.”
He adds that his native China, with a population
of 1.3 billion, will set a bold new pace. “When
China opens up to rapid testing, the market will
be huge. Southeast Asia, South America, Africa,
the Middle East, and the Pacific Rim will also
contribute to market growth in the years ahead.”
Multi-Billion-Dollar Industry
According to the market report Food-Micro 2005
published by Strategic Consulting, Inc., Woodstock,
Vt. (www.strategic-consult.com), the global
market currently includes more than 625 million
food industry microbiological tests completed in
2005, with a market value exceeding $1.65 billion.
“The food sector represents the largest market
segment within the industrial microbiology market
and represents almost 50% of the total market,” says
Thomas Weschler (weschler@strategic-consult.com),
SCI President. “The food sector is more than double
the size of any other industrial segments, including the
pharmaceutical, beverage, environmental, industrial
processing, and personal-care products sectors.”
Since 1998, the market value for food microbiology
has grown significantly and has had an annual growth
rate of 9.2%, Weschler points out. “Based on SCI
research, the food microbiology testing market is
expected to grow to 822.8 million tests in 2010 with a market value of $2.4 billion,” he relates. “This represents a
projected annual growth rate of 5.6% in testing volume.”
Driving this growth worldwide is an increase in food
consumption, consumer demand, industry’s food safety
priorities, and regulation, Weschler says. The acceleration of the conversion of traditional microbiological
testing methods to rapid methods is a function of those
phenomena. It’s no surprise that, despite the higher
[LABORATORY]
Rapid Microbial Testing continued...
Samples are added to
the Saber Detection
System for automated
sample processing.
cost per test, these newer methods
are being used more frequently
because they provide faster results
and/or ease-of-use benefits
compared to traditional testing.
Nonetheless, traditional methods
still account for approximately
65% of the microbiological tests
performed worldwide in the food
market. Rapid counterparts, including convenience-based (Petrifilm,
for example), immunoassay-based,
and molecular-based methods,
account for 35%, amounting to
220 million tests/year. Some
50% or more of all tests are run
to determine non-pathogens or
indicator organisms, according
to Food-Micro 2005.
“By 2010, much will have
changed,” says Weschler.
“Traditional methods will still be the
predominant ones used, accounting
for 428.2 million tests/year. However,
in 2010 traditional methods will
represent only 52% of all tests
conducted, which is a reduction
of 12.4% from 2005 based on
percentage of tests performed.”
All types of rapid methods
will make significant gains in
usage during the coming five-year
period, Weschler continues.
“When combined, the annual test
Recently Released Rapid Tests
• Oxoid Ltd., Hampshire, UK
(www.oxoid.com), has designed
the Oxoid Listeria Rapid Test
for the detection of Listeria
species in many different foods,
including dairy, raw vegetables,
meats, poultry, and fish. The
procedure includes the use
of two enrichment broths for
the maximum recovery and
growth of Listeria followed by an
immunoassay using the testing
device. After adding the sample
to the testing device, results
are available in about 20 min. A
blue line in the result window on
the testing device is a positive
result and indicates the presence of Listeria flagella antigen.
• 3M, St. Paul, Minn.
(www.3m.com), introduced
the 3M Petrifilm EL Plate, a
sample-ready culture medium
that allows for the detection
of Listeria species, including
Listeria monocytogenes.
The results can be interpg
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preted quantitatively, which
could help identify hot spots
and contamination sources. It
provides results in about 30 hr
from the time of sample collection. An additional attribute of
this testing kit is that is requires
no enrichment, which means
less risks of cross-contamination due to culture transfers.
• Whatman plc, Florham
Park, N.J. (www.whatman.
com), offers the FTA® Concentrator-PS, a portable two-stage
filter co-developed with the
Food and Drug Administration
that can detect Cryptosporidium,
Giardia, Cyclosporia, and other
dangerous parasites in fresh
produce and water supplies.
The field test, which will be
used primarily by regulatory
agencies and municipalities,
takes as little as 15 min, and
definitive results can be
obtained the same day.
The device uses FTA, a
proprietary chemically treated
matrix that rapidly isolates
and protects nucleic acids at
room temperature. The sample
is passed through the matrix,
concentrating and destroying parasites. The device
immobilizes and preserves
the organisms’ DNA, which
can be tested with a simple
polymerase chain reaction
to confirm the presence of
any number of parasites.
• Sword Diagnostics, Inc.,
a new company based at the
National Center for Food Safety
and Technology, Chicago, Ill.
(www.sworddiagnostics.com),
announces its groundbreaking benchtop rapid test for
environmental Listeria that
provides results in just 16 hr.
The Saber Detection System
combines immunoassay capture
with Raman detection, an
advanced optical technique
that measures wavelength
shifts. This system has the
ability to detect the presence
of microorganisms in concentra-
volume of rapid methods will almost
double from current levels and reach
394.6 million tests in 2010,” he elaborates. “The gain in market value for
rapid methods will be even more
pronounced than the testing-volume
increases, since the rapid methods
have much higher average prices
per test than traditional methods.
“With such solid growth
prospects, the food sector
is a market segment that all
diagnostic manufacturers need to
understand and, where they are
not currently present, possibly
enter,” Weschler advises.
Reality Check
Despite these developments, it’s
critical to keep the big picture in
perspective, says Michael Doyle
(mdoyle@uga.edu), a food microbiologist and Director of the Center
for Food Safety at the University of
Georgia, Griffin. “Rapid methods to
tions starting at 10 2 /mL.
• Purdue University
researchers have developed
a new system Bacteria Rapid
Detection Using Optical Scattering Technology that analyzes
scattered laser light to quickly
identify bacteria for applications
in medicine, food processing,
and homeland security,
reportedly at one-tenth the cost
of conventional technologies
(Bayraktar et al., 2006).
With this system, photons
bounce off of the colony, and
the pattern of scattered light
is projected onto a screen
behind the Petri dish. This
“light-scatter pattern” is
recorded with a digital camera
and analyzed with sophisticated
software to identify the types
of bacteria. Using this method,
it takes less than 5 min to
identify harmful organisms.
To date, the system has been
proven to accurately identify six
species of Listeria, plus select
species of Salmonella, Vibrio,
Escherichia coli, and Bacillus.
cheaper than employing and training a person
to do less-expensive testing methods.”
There’s additional good news for most of the
food industry, Donnelly says—there’s not much
difference in results between expensive tests
and inexpensive tests, so users can decide what
is most appropriate for their specific budget. »»
The blue line in this rapid test indicates the presence of Listeria.
Photo courtesy of Kansas State University
detect pathogens are still not rapid,” he purports.
“Most methods require 8–24 hr to get results.
We need to get it down to less than 15 min.”
Progress is being made toward that goal,
he says. “The Food and Drug Administration, U.S. Dept. of Agriculture, and the food
industry have initiatives to promote food safety
programs based on Hazard Analysis and Critical
Control Points (HACCP) principles,” he points out.
“That work can benefit greatly from real-time
microbial tests that take minutes, not all day.”
Thus far, Doyle says, no test is sensitive
enough to match the regulatory gold standard of
one detectable target organism in a 25-g sample.
“We currently need 10 3 –10 5 target bacteria/mL
sample to detect target organisms,” he relates.
Scientists are getting the minimum down
to 10 3 /mL, but we still need better ways to
concentrate bacteria to increase sensitivity,
Doyle emphasizes. “Breakthroughs in this regard
should be forthcoming in the next few years.”
Catherine Donnelly (catherine.donnelly@
uvm.edu), Professor of Microbiology in the Dept.
of Nutrition and Food Sciences at the University
of Vermont, Burlington, is energized by current
rapid test happenings and the prospects of
new developments. “It’s a really exciting time
to be a food microbiologist,” she says. “The
advantages testing technology provide are
outstanding. Rapid tests are user friendly,
accurate, and approved by AOAC International.
There are a variety of options for users.”
Embracing reality in her renowned work
with Listeria, Donnelly is quick to point out
that rapid microbial testing is characterized by good news and bad news.
“The good news is the technology,” she
explains. “The bad news is that some equipment and instruments come with a hefty price
tag. Some rapid tests are more cost effective
than others. But it’s important to keep in
mind that some expensive instruments are
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Top-Ten Attributes of an Ideal Rapid Test
Daniel Y. C. Fung, Professor of Food Science, Kansas State University,
Manhattan, says that many factors contribute to making a rapid test ideal
to use.
1. Accurate for the intended purposes. It considers sensitivity, minimal
detectable limits, specificity of test system, versatility, potential applications, and comparison to reference methods.
2. Speedy and productive. It includes time in obtaining results and the
number of samples processed per run, per hour, and per day.
3. Cost effective. The initial, per-test, reagent, and labor costs are
reasonable.
4. Acceptable. It is acceptable by the scientific community and regulatory agencies.
5. Simple to operate. The test takes into consideration sample preparation, operation of test equipment, and computer versatility.
6. Amenable to training. Location (on or off site), length of time, and
qualifications of operator are all factors.
7. User-friendly reagents. Preparation, stability, availability, and
consistency of reagents need to be considered.
8. Great company reputation.
9. Exceptional technical service. Speed, availability, cost, and scope
all count.
10. Optimum utility and space requirements.
Rapid Microbial Testing continued...
Enrichment Essentials
The all-important enrichment step—the time it takes to
grow pathogens to detectable levels—is the greatest
hindrance to rapid microbial testing, says Dong-Hyun Kang
(foodsafety_2001@yahoo.com), Associate Professor of
Food Science in the Dept. of Food Science and Human
Nutrition at Washington State University, Pullman.
“Most enrichment steps take so much time that
we don’t get test results for 10–24 hr,” Kang says.
“If we can find a really good stimulant or a bacterial
target for a pathogen we can decrease the enrichment time to 4–6 hr. So it’s critical to focus more
research on reducing the enrichment time.” FT
REFERENCE
Bayraktar, B., Banada, P.P., Hirleman, E. D., Bhunia, A.K., Robinson,
J.P. and Rajwa, B. 2006. Feature extraction from light-scatter patterns of
Listeria colonies for identification and classification. J. Biomedical Optics.
11(3): 034006.
Linda L. Leake, Contributing Editor,
Food Safety Consultant,
Wilmington, N.C.
• llleake@aol.com
Denver Instruments
1/2 Horiz No Bleed
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