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From: Food Quality & Safety magazine, August/September 2005

Microbiology

A Moving Target

by Mark A. DeSorbo

The ever-emerging microbiology macrocosm is a moving target that has an integrated blend of pathogens, hosts and bug bionetworks that clearly define roles among food safety management; government, industry and consumers. While a host of complexities can make the bull’s eye of food safety a tough hit, unity among the food safety powers that be could be the silver bullet.

Beneath the soil of a strawberry field lurks a silent killer. It’s not an insect or a voracious herbivore, but it packs a mighty bite and there’s no telling when it will strike. For a strawberry grower, black root rot is a moving target, as it is a very complex, wet-soil disease caused by various pathogens and environmental factors that keep even the savviest of horticulturists guessing.

That scenario is even more intense in the ever-emerging microbiology macrocosm, another moving target that has an integrated blend of pathogens, hosts and bug bionetworks that clearly define roles among food safety management; government, industry and consumers.

According to the recently released IFT Expert Report, “Emerging Microbiological Safety Issues: Implications for Control in the 21st Century,” a science-based approach is the best defense against food safety issues.

Nailing the bull’s eye of food safety involves understanding such complexities, the IFT reports indicates, as changes in demographics, geographic origin of food, production and processing, consumption patterns and, of course, the microorganisms themselves.

And keeping that target in the crosshairs can be difficult, for with those complexities comes a host of ideologies that herald a back-to-basics approach, automated DNA technologies and the need to combat emerging food safety threats and diseases freely with microbiological tools.

“Microbiological hazards are ever-changing, and the amount and complexity of data and the residual unknowns are growing at a rapid rate,” the IFT report concludes. “Each new scientific advance gives us the opportunity to add to our knowledge of foodborne illness using new techniques and researching new questions.”

At the same time, the report indicates, human susceptibilities are increasing, yet the ability to link food to adverse health outcomes continues to improve. “The human health and economic consequences of emerging microbiological food safety issues is immense,” the report says. “International coordination of food safety efforts should be encouraged.”

Technology Shoot Out

When it comes to popping a proverbial cap in pathogen’s posterior, food safety’s weapon of choice differs.

There are PCR (polymerase chain reaction) pistols, like the BAX detection and RiboPrinter microbial characterization systems from DuPont Qualicon (Wilmington, Del.) and the Genevision from Warnex (Laval, Quebec). New to the techno-scene are the re-badged BioSys automated systems, Envisio and Soleris, now manufactured by Centrus International Inc. (Kingsport, Tenn.), an Eastman subsidiary.

Of course, there are the good old standbys, like the ELISA—the classic six-shooter of speciation.

James Nokes, laboratory director for Microbac Laboratories Inc. (Maryville, Tenn.) believes PCR- and DNA-based technologies should be the way of the future.

“We’ll get away from the 8-hour test,” he says. “It will have to get faster.”

And in some cases, it already has. BioControl Systems, Inc. (Bellevue, Wash.) has unveiled several DNA-based products that yield quick results. Earlier this year, it unveiled its Assurance GDS for E. coli 0157: H7 test, which provides results in an eight-hour shift. Completing the system is the Assurance GDS Rotor-Gene, a multi-channel rotary cycler, which eliminates ambiguous melt curves and provides results in just 70 minutes.

BioControl also announced that its SimPlate Coliform / E. coli Color Indicator (CEc-CI) was also approved as AOAC Official Method 2005.03. SimPlate CEc-CI is a rapid method for quantifying both total coliforms and E. coli from food and environmental samples in only 24 hours.

PCR and ELISA methods, Nokes points out, are rather similar. It’s just the enrichment times that differ. “PCR has much fewer false positives than ELISA methods,” he adds. “With PCR, a positive is a positive. The FDA says show it, but our customers will take a positive PCR as a positive.” If you get a positive with an ELISA method, you have to validate it, he adds, and that means culturing in a Petri dish for several days.

While many food companies rely on PCR, in the eyes of federal regulators, a positive result using a DNA-based technology is “presumptive.”

The most popular tests among Microbac’s clientele, Nokes says, are E. coli and Salmonella and Listeria. “We can actually take that Listeria, find out the species and strain and track where it came from,” he adds.

The technology, he says, really shines when it is used to detect spoilage and monitor hygiene. The process applies PCR to the 16S gene reserve region that all bacteria have. A gel is then used to separate the amino acid to reveal the sequence.

“You don’t have to know what you’re looking for,” Nokes says. “You just know you have a problem and you learn what it is in two days as opposed to two or three weeks. Culture after culture after culture are just shots in the dark; but with this you can do fungal and yeast tests at the same time.”

But DNA testing isn’t always the best or only way, says Bruce Ritter, president and CEO of ELISA Technologies Inc., a Gainesville, Fla. maker of analytical test kits.

ELISA tests can detect specific protein targets like those that cause allergic responses. They are the best way to find and identify specific allergenic proteins in food, he says. For example, DNA tests can tell that it came from a cow, but cannot identify if it was from milk or meat.

“Every college teaches about genes and DNA, and that’s all well and good, but DNA tests cannot answer some of the questions that ELISA tests can,” Ritter says.

Immunoassays, he contends, are the tool of choice for detection of allergens in food because they can detect and identify the proteins that cause food allergies. As the list of allergens grows, he explains, there will be more tests developed for the allergenic markers.

“The ELISAs are faster, easier, more specific, more reliable and less expensive than DNA techniques,” he adds. “While everyone wants rocket science, sometimes all you need is a slingshot, and sometimes the simple answer is the best.”

Lock and Load Your Lab

The proving grounds, ultimately, are the laboratories, and a lab is only as good as its people and quality system, says Dr. Jeffrey L. Kornacki, president and senior technical director of Kornacki Food Safety Associates (Madison, Wis.).

Kornacki, who worked for several years as a microbiologist, now works as a consultant, and assists food companies with everything from sanitary facility and equipment design to lab quality systems to the good old fashioned microbiology basics.

“Product residue gets embedded and it’s almost impossible to clean,” says Jeffrey Kornacki. “The principles of sanitary design have been around for a long time. There needs to be a new cleaning and sanitizing paradigm that goes beyond throwing aqueous solutions on surfaces.”

Moreover, several types of food prep equipment apparently never had any design input from microbiologists.

“I was in a sandwich-making plant and the slicer part of a sandwich maker had 60 moving parts. How do you clean something like that? That’s a serious problem and it was designed by non-microbiologist,” Kornacki points out.

Without cleanability, there is greater bacteria growth, and when it comes to emerging foodborne pathogens, Kornacki says it is still unclear what, in fact, causes illness and more methods are needed to detect those sources.

“There is no effective tissue method to identify and recover these organisms,” he says. “We need to get to a point with actionable results without a culture, and there’s a lot of pressure to do that. There are people who say they have those types of assays, but I’ve been burned before. It’s got to be culturally confirmed. Some of these PCR-based tests are great. The rate of false-positives is minimal, but the government wants to see a culture, and that has tremendous consequences. The last thing you want to be is wrong. You have to make absolutely sure that it’s a positive.”

There also needs to be unity in the field of applied microbiology, and he says that means establishing a lab quality system accredited by ISO 17025, a standard that outlines general requirements for the “competence of testing and calibration laboratories.”

According to ISO, about half of the 50,000 laboratories worldwide are accredited under the standard, which is a must for labs in the European Union. The standard was updated from the 1999 version to make it compatible with ISO 9001:2000, which sets requirements for management systems in businesses and other organizations. The changes were a must because more businesses are opting for ISO 9001:2000 accreditation, including those that testing and calibration laboratories serve.

The modifications place more requirements on top management in their oversight of such laboratories. The new ISO standard also requires accredited laboratories to demonstrate a commitment to continually improve the effectiveness of management systems. A laboratory must also demonstrate they have a program to continually improve customer satisfaction and communications.

“You gain confidence in testing by having a comprehensive lab quality system that allows you to stand behind your results,” Kornacki says.

Key elements to having a solid lab quality system are knowledge of the method and having it written out when a positive result is reported. “Labs are very unique,” he says, adding that it isn’t enough to just say that a method or lab practice is FDA or BAM approved.

Training is also a key element. Technicians must be competent, and that proficiency hangs on clearly-written testing requirements, Kornacki says. “The technicians must read (testing requirements), observe it and then perform it under supervision to prove they can do it well,” he adds.

While equipment calibration and maintenance schedules are givens, and include everything from thermometers to large pieces of equipment, contamination control must also be a priority.

A lot can go wrong quickly if contamination control protocols are not in place. Kornacki recalls one particular lab study for a company that believed it had created Salmonella-resistant eggs, and it involved inoculating yokes with a particular strain.

During that experiment, the tray of plates was tipped over. The technician quickly picked the plates up, sanitized her hands and then set up a separate set of plates, contaminating that experiment with her hands.

The arrangement of test tubes can also usher in cross-contamination. “There needs to be a separation between critical areas to avoid cross-contamination,” he adds.

Proper sampling and comprehensive tracking of samples is also important, for data from botched sample is of no value.

“ABC Company samples may be in the same rack as samples for XYZ Company, and you test both and both tests come back positive,” Kornacki says. “Then you have to ask yourself, which one was positive first. You have to document what you’re doing so you can save yourself the embarrassment and your client the frustration.”

Many hassles, he says, can be avoided by simply validating the media. On another occasion, some plates turned green after samples were inoculated, and it was determined that the method used to inoculate the samples on the plates was not acceptable, and therefore, the samples had to be redone.

“That’s what good labs do. It’s difficult to admit mistakes, but be honest, because you’ll build a bond with your client,” Kornacki says. “A good lab will also know the limits of what it can handle. Know your limits; don’t say ‘yes’ to everything.”

And, of course, you can’t forget the basics, he says, adding that although such a review can be mundane, it is very important to embrace the basics of food microbiology.

“For example, Listeria grows in cold, wet environments. We say that Salmonella can’t grow in a dry product, but what we really should say is that bacteria cannot grow below a certain level of moisture, but it can survive,” Kornacki says. “The basic concept of growth and survival should not be ignored.”

The writers of the IFT report agree, saying that food safety needs to focus on further controlling processing conditions through the application of a science-based HACCP plan.

Furthermore, while it may be difficult to imagine a food safety system that reacts quickly and successfully to emerging microbiological food safety issues, the IFT report indicates that the flexibility in responding to new information and hazards and the sharing of data is essential.

A food safety system, the IFT authors contend, must emphasize validation and verification of methods to assure food safety.

“The complex interrelationship of the pathogen, host and microbial ecology ensures a role for everyone in food safety management—industry, regulatory agencies, public health officials and consumers,” the report concludes. “A flexible, science-based approach that relies on all parties to fulfill their role is our best weapon against emerging microbiological food safety issues.” –FQ

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