Is Your Pathogen Test the Best it Can Be?

In the past, general discussion of testing would likely bring to mind a lab setting in which technicians were sampling product to identify the presence of pathogens, allergens, or even pesticides. Today, the first thought, and discussion among both industry and consumers, is more likely to be that of individuals being tested for COVID-19. While the pandemic has changed our world, there are some new understandings that have come out of it that can be applied to other areas.

Testing is one of those. Take, for instance, the increased discussion of test “sensitivity” and “specificity.” With the continuing introduction of new viral and antibody tests for COVID, these words have become increasingly important to distinguish the efficacy of the various tests. And the concepts are just as critical in microbiological testing.

In simple terms, “The sensitivity of a test tells us how well the test identifies a pathogen, and the specificity of a test tells us how well it determines the absence of a pathogen,” said Auburn Health Strategies President Robin Stombler. “Both are important factors to consider when utilizing a test for detection of specific food pathogens and spoilage microorganisms.”

Providing further definition, Safe Food Alliance Senior Director of Analytical Services Thomas Jones explained: Analytical sensitivity refers to the lowest number of organisms that a method can detect; sensitivity can also refer to the inclusivity of the method, or its ability to detect all of the microorganisms in the category being tested (ex: all of the strains of Salmonella). Specificity is the ability of the method to distinguish the target from similar but genetically distinct non-target organisms (ex: Salmonella from other enteric microbes).

“Ideally, we want a test method that does not waste time or resources generating false-positive results (good specificity) while being able to detect even a single viable pathogen in the sample (high sensitivity),” he said.

Building on the concept of false results, when false positives occur, pathogen-free foods may be rejected erroneously, but when false negatives occur, foods that contain harmful pathogens may be shipped to consumers, Stombler said. “In real terms, a false-positive result may mean that a food product is withheld from market, leading to economic losses for the food producer or manufacturer. A false-negative result may risk public health with the distribution of a potentially contaminated food,” she said.

Because microbial detection, whether viral or bacterial, requires sensitive and selective methods, “it is crucial to pick the correct test platform and validated methods for your product,” added Cherney Technical Liaison Lauren Ellie. So, it is important that your in-house or outsourced lab ensures sample integrity, follows best practices by utilizing the proper media for enrichment, and follows specified incubation times and temperatures to be able to detect pathogens. “The testing platform utilized needs to be sensitive and specific to the pathogen you are testing for,” she said.

DETECTION DIFFICULTIES. The greatest challenges of COVID-19 testing tend to be related to all the unknowns of the newly emerged virus. But in food, the difficulties often revolve around the challenges of detection within the food. As noted in the USDA Food Safety and Inspection Service (FSIS) presentation, “Understanding and Evaluating Microbiological Sampling and Testing,” pathogens are hard to detect because:

• They are typically not evenly distributed.
• They occur at low levels.
• They are often injured when found in the product.
• Detection may be inhibited by material in the food product; its food matrix.

The fact that pathogens are typically low in number and sporadic in distribution makes sampling a major challenge, Jones said. And because the specific food types or matrices can interfere, the analytical methods must be validated for each food type. Additionally, he said, “Pathogens can enter into a dormant state where they may still be viable and capable of causing disease, but not capable of growing and being detected by the test method.”

Similarly, during processing of the product, bacterial cells are typically injured, which can add another layer of difficulty in testing, Ellie said. Other obstacles can be related to product type that includes the presence of antimicrobial compounds, such as natural or added preservatives, oily product types, and products with a lot of particulate matter, she said.

Even monitoring of the environment of food facilities to assure recognized testing standards are followed can cause challenges, Stombler said.

Sampling. According to FSIS, all sampling plans have limitations. Thus, the relative rigor of the sampling program must be evaluated, with the best sampling plans providing the opportunity for, but no guarantee of, detection.

Ellie recommends that sampling plans take into account the product type and production volume, adding that it is important to know and understand pathogen risk based on water activity, pH, salt content, etc. “Be conscious of the potential sources of pathogen contamination that can result via cross contamination from raw materials, as well as the potential for environmental contamination,” she said. “Good manufacturing practices, Hazard Analysis Critical Control Point (HACCP), and a strong environmental monitoring program can help reduce the possibility of contamination.”

Because bacteria are not necessarily evenly dispersed throughout a product, “pathogen detection can feel like you’re searching for a needle in a haystack,” Ellie said. “And obviously you can’t test 100% of your product, so ultimately it comes down to sample size taken versus production size.”

As such, when pathogen testing your product, it is crucial to obtain a representative sample, Ellie said. Pathogens are more likely to be evenly dispersed in a liquid that is easily homogenized, while in solid foods, pathogens are not necessarily present throughout the entire product. “It’s important to understand when conducting sampling that you need get the correct volume of sample relative to the amount of product produced, as well as a good representative sample for your plan to be comprehensive and thorough.”

Jones also discussed the use of a two-class plan for pathogens that varies by the amount of sample enriched/tested. In general, he said, the more non-uniform the microbial distribution in the food, the larger the sample that is needed to enrich/test for pathogens. Then, he said, “Method performance can be evaluated by validation studies involving sample spiking with live microorganisms and by comparing the test results obtained with any known published data for that food type, such as prevalence studies.”

If sampling is not done correctly, the subsequent testing will not be meaningful, Jones added, noting that:

The sample has to be taken with proper aseptic technique and be representative of the lot to be tested.

• The methods used to analyze the sample should be official methods that are properly validated and conducted by an accredited laboratory.
• End product and environmental testing alone are not enough. “The preventive controls mandated by FSMA and at the heart of food safety programs like HACCP must be in place to ensure food safety,” he said.

As such, Stombler said, “Accurate and reliable test results should be the foundation for every food laboratory. While laboratory standards exist, there is no mandate or assurance that food laboratories follow them.” As such, she recommends that food manufacturers ask any laboratory which is testing its food for human consumption, “Are you accredited to ISO/IEC 17025:2017?”

This is becuase laboratories that are accredited to ISO/IEC 17025:2017 demonstrate an adoption of recognized standards, including the use of quality controls, proficiency testing, and competence assessment, she said. “Accreditation to this standard also indicates that a laboratory has proven its ability to perform tests consistently.”

Whether your lab is in-house or outsourced (and whether testing for pathogens or COVID-19), it is critical that you always strive for accurate and reliable test results — no matter how challenging the course. Even in the fast-paced world in which we live where time is money, there is no short cut when dealing with food safety.

The author is Senior Advisor and Former Editor of QA magazine. She can be reached at llupo@gie.net.