Advancing Rapid Microbial Testing
LABORATORY
So 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 ([email protected]), 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.
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 microbiological 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 ([email protected]), 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."
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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 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 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.
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Reality Check
Despite these developments, it’s critical to keep the big picture in perspective, says Michael Doyle ([email protected]), a food microbiologist and Director of the Center for Food Safety at the University of Georgia, Griffin. "Rapid methods to 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 103–105 target bacteria/mL sample to detect target organisms," he relates.
Scientists are getting the minimum down to 103/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 ([email protected]), 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 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.
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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 ([email protected]), 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."
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 interpreted 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 concentrations starting at 102/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.
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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.
by Linda L. Leake,
Contributing Editor,
Food Safety Consultant,
Wilmington, N.C.
[email protected]