Product Testing, Part 7: The role of confirmation testing – what to look for
Julia Stewart:
Hello, this is PMA PR Director Julia Stewart, and welcome back to PMA’s audio blog, “Ask Dr. Bob” with PMA’s Chief Science & Technology Officer Dr. Bob Whitaker. Bob and I have been talking about the challenges with product testing in produce. Bob, this time we promised to get into the role of confirmation testing. Why is it important for producers and buyers to understand the benefits and limitations of this type of testing, and what does it mean to supply chain logistics?
Bob:
In previous posts we discussed rapid tests such as PCR tests, and the benefits of those types of tests – mainly speed. Because of the pressures our industry is under to sample, test and get results back so that loads can be harvested or shipped, speed is king as they say and these DNA-based test methods have become the standard in the industry. But, we have also discussed some of the issues with these types of tests; mainly they are not always sensitive or accurate enough.
We have all heard of instances where initial positive samples fail to be confirmed on follow-up testing. It is extremely important to be sure a sample is in fact positive, because growers and processors are often making decisions not to ship or harvest thousands of dollars worth of products. And of course, adding another round of testing costs money and time. You all might recall that as little as a few years ago, some producers recalled product based on initial test results only to find out later that the original result might not really have been positive for a pathogen, but was simply a closely related non-pathogenic bacteria that shared many similar DNA fragments.
So, this is where “confirmation testing” enters the picture. Initial positive samples are often re-tested using another type of test. To be clear, this isn’t just running the same test again – this is using a completely different test to determine if the first was accurate or not. Some choose to use FDA recognized microbiological plating methods commonly referred to by many as “BAM testing” – “BAM” stands for FDA’s Bacteriological Analytical Manual. Others use another round of PCR-based testing focusing on a more diverse set of DNA primers characteristic of a specific pathogen. Sometimes, depending on the pathogen, there are other types of tests that can be used. For example, if an initial test indicates E. coli O157:H7, one might opt to use an immunological-based test to determine if shigatoxin, which is characteristic of E. coli O157:H7, is present.
Julia:
Wow, sounds like a very tricky issue. I see why you stress that anyone who is testing their products understand the benefits and limitations of each approach.
Bob:
That’s right Julia, there are pluses and minuses for any of the approaches I just described.
For example, using another round of PCR-based testing has the advantage of being fast. Once you have the “positive” results from the initial testing, you can have the lab immediately run a second round of testing using a different set of primers designed to the target the pathogen. You can have results back within another 12 to24 hours using this approach. The logic is that the more DNA primers you test for, the more likely it is that your original positive test was correct – and, correspondingly, the less likely that the positive test was just a close cousin of the pathogen that shared some of the pathogen’s same gene sequences. The drawback is that you have to be certain the primers you use really uniquely distinguish pathogens from non-pathogens. The science is not entirely conclusive on this matter yet, so you have to work with your labs to understand the specificity and selectivity of the probes used in your test and the relative significance of the number of unique primers used in building the confidence that your results are accurate.
On the other hand, BAM testing is kind of the gold standard for microbiologists. It’s based on the ability of bacteria to grow on different media “recipes”. Every microorganism’s DNA genetically codes for specific proteins or enzymes that allow that microorganism to use certain sugars and nitrogen sources to grow. Different bacterial species, of course, have different DNA and therefore differ in which sugars and nitrogen sources they can grow on. Over time, microbiologists have developed a complex set of testing media which can distinguish specific pathogens from related non-pathogenic strains. These media recipes are well recognized and accepted by scientists and can be a very powerful tool to identify specific pathogens.
So as a confirmation testing tool, BAM testing can be very specific and accurate. It can distinguish between closely related species of bacteria and at the end of the confirmation test, if the sample tested positive by PCR and BAM methods, you can be reasonably confident that you have a pathogen-positive sample and a live culture of the pathogen from the lab However, BAM testing isn’t without issue itself. These tests take time, anywhere from 48 to 96 hours depending on how you handle samples. It is also important to note, that there is also a degree of human discretion in reading BAM results, and bacterial colony shape and colors can be interpreted incorrectly on occasion.
Julia:
So, confirmation testing is an important tool to verify initial positive results and give the producer and the buyer confidence that products either are or are not truly contaminated with a pathogen. Thanks, Bob.
Next time, we’ll talk about a very complex area of concern: product sampling. So, listeners, please join us for what is sure to be an interesting discussion.
Remember you can email Bob at AskDrBob@pma.com. In addition to listening to these and other Ask Dr. Bob blog posts, we invite PMA members to visit our new online Food Safety Resource Center on PMA.com and check out the lab testing white paper in the Education section. Thanks!