The Bia Tástáil Chronicles:
Validation And Verification Of Microbiology Tests
Latest Insights From IAFP

July 18, 2019

(Photograph is provided courtesy of Arek Socha of Pixabay.)

The annual convention of the International Association for Food Protection is being held next week – from July 21 through July 24 – in the Midwest of the United States of America.  Leaders of the food industry, the manufacturers of rapid test kits, independent labs, and others will gather in Louisville, Kentucky.  A number of symposia and meetings will be held at the Kentucky International Convention Center.

One of the symposia will be one focused on “Microbiological Method in Food Manufacturing:  Are Your Methods ‘Fit Enough’ For Purpose?”

In this initial edition of The Bia Tástáil Chronicles, two of three presenters provide a preview of this symposium.

J. David Legan of Eurofins Microbiology Laboratories is one of those presenters.  His speech is entitled “The Testing Lab Dilemma”.

Dr. J. David Legan.

“My presentation will focus on the dilemma faced by independent testing labs to make certain that the testing methods in use are actually suitable,” stated Dr. Legan.  “While there are a number of validation schemes, that is just part of the story.  Life can get very complicated.  What is critical is to be able to make good decisions based on independent testing.  Part of the dilemma is to make the best selection from among different methods so as to get reliable results.”

Dr. Legan has a degree in Applied Biology from the University of Bath in the United Kingdom;  he earned his PhD in Food Technology at the University of Reading, also in the United Kingdom.  He initially worked in research in the areas of microbiology and pathogen modeling in Great Britain.  He then worked for a number of years focused on food testing and food preservation research in corporate labs at Nabisco and Kraft Foods.  Today, he works at an independent testing lab in Madison, Wisconsin, that was acquired last year by Eurofins Microbiology Laboratories.

Two of the elements to be considered in testing food products, according to Dr. Legan, are the method selection chosen by the testing lab or the end user as well as the method validation selected by the testing kit manufacturer.

“Overall, there are several key steps to ensure food safety through rapid testing,” explained Dr. Legan.  “First, determine the risk you are managing.  Second, choose a test kit that meets that risk.  And third, make sure that the rapid test kit has been through an appropriate validation process.”

“For example, you may choose a specific rapid test to ascertain the risk of Salmonella, E. coli, and/or Listeria,” stated Dr. Legan.

“The method developer decides what validation scheme to follow as well as deciding which review body to send for analysis – AOAC International, Health Canada, AFNOR (French Association for Normalization) Group, and other review bodies.  AOAC International and AFNOR Group are two of the key review bodies worldwide,” stated Dr. Legan.

He indicated that the method developer also “selects food products from separate categories for testing and works with independent labs to obtain data on method performance with each food category.  They then provide the independent test results to the validation body, e.g. AOAC International.  That organization then reviews the data to determine if the method is sufficiently robust, precise and reliable for use.  In particular, a validation body strives to make certain that the testing method is sensitive enough and excludes organisms that might be present from normal handling.”

“To make sure test results are accurate,” Dr. Legan explained, “The rapid tests that are developed need to work well even if you don’t know the heritage of the sample or the contents of the sample to be tested.”

Rapid microbiology test kits need to take into account the shelf life of the specific food product being tested.
“You don’t want to wait for a long time [for results] for a food product with a short shelf life,” stated
Dr. David Legan.  “A hamburger, for example, has a shelf life measured in days as compared to a cookie
that can have a longer shelf life.” (Photograph is provided courtesy of Moerschy from Pixabay.)

“You also need to make sure that the rapid test is suitable for the product being analyzed,” Dr. Legan continued.  “With pizza, for example, you need to take into account each of the individual components.  A rapid test needs to be able to detect the pathogen of concern in the presence of the bakery, cheese, vegetables, and meat components of the pizza.”

“In most cases, the rapid tests for food-related pathogens are not developed directly by food companies,” stated Dr. Legan.  “Instead, the food companies are consulted as the rapid tests are developed by diagnostic firms.  The test method selection affects cost, speed, and sensitivity.”

“Typically, the faster a test can produce results the more it uses technology that may cost more than previous testing kits,” explained Dr. Legan.  “The ultimate goal of a rapid test is to identify a risk before a food product is consumed by the public.”

Dr. Legan stated that “Food safety protections have been enhanced as better testing methods have been developed.”

As an example, he pointed to one of the pathogens that can impact food safety.

“When Listeria first appeared, testing methods were very slow – from several weeks to several days.  As tests became more advanced, timing decreased from several days to testing today that provides results typically within one to two days.”

“Without question, the enhanced testing for Listeria has saved lives,” stated Dr. Legan.

“Testing is most effective when it forms part of a well-designed food safety plan,” concluded Dr. Legan.  “Our goal as a testing lab is to protect our customers and their consumers from harm.”

Joining Dr. Legan at this symposium on “Microbiological Method in Food Manufacturing:Are Your Methods ‘Fit Enough’ For Purpose?” will be Morgan Wallace of Rheonix.His speech is entitled “Comparison of ‘Fitness for Purpose’ in Established Validations Schemes:Is There a Best Approach?”

Dr. Morgan Wallace.

“My talk will focus on food safety testing from the method developer perspective,” explained Dr. Wallace.  “This includes a historical view of methods validations.”

Dr. Wallace has degrees in Microbiology and Cell Science and Political Science from the University of Florida in the United States.  He earned his PhD in Food Science and Human Nutrition with a specialization in Food Microbiology, also at the University of Florida. 

He initially worked in antimicrobial resistance research at the Antimicrobial Resistance Research Unit, USDA Agricultural Research Service (USDA-ARS) in Athens, Georgia.  This research unit studies antimicrobial resistance in bacteria from veterinary and food sources using the same methods that the Centers for Disease Control and Prevention (CDC) use to study bacteria of human clinical origin.

The focus of Dr. Wallace at the USDA-ARS was the study of pathogens originating from animals, food sources, and food production facilities.  One of the goals of this research was to determine how pathogens became resistant to antimicrobial agents.

He continued his research efforts in the Microbial Food Safety Research Unit at the USDA-ARS facility in Wyndmoor, Pennsylvania, located in suburban Philadelphia.  At this locale, Dr. Wallace focused on pathogens contaminating ready-to-eat meat products.  In particular, he researched Listeria in hot dogs.

From the USDA-ARS, he moved on to working at DuPont Qualicon, now Hygiena (Qualicon Diagnostics LLC).  In this corporate environment, Dr. Wallace ran the validations group.  This included field trials for molecular diagnostic test kits for pathogens such as Salmonella, pathogenic E. coli, and Listeria, among others.

“Much of my work focused on real-world applications to track pathogens with more rapid and sensitive techniques,” explained Dr. Wallace.  “We strived to reduce false positives and false negatives in food safety testing processes.”

Today, Dr. Wallace works at Rheonix, a business headquartered in New York State.  This company has created a rapid test, the Beer SpoilerAlert™ Assay, that “allows breweries to test for organisms that can spoil beer at any point in the manufacturing process,” according to the firm’s website.

Rheonix also has a created a second rapid test, the Listeria PatternAlert™ Assay, that “enables 7-hour pattern identification direct from an enriched sample,” according to the business.

“Listeria is a cold-loving, damp area-loving organism,” stated Dr. Wallace.  “It can grow at much lower temperatures than many other pathogens.  Listeria is well-adapted to thrive in food manufacturing and storage facilities.  Many businesses have re-designed food storage equipment to remove recessed hiding places, cracks, and crevices.  Lots of environmental testing is conducted by manufacturers to enhance the safety of food during manufacturing and in storage.”

“When I first started working in the field of food diagnostics, you could get approval for one test for a specific pathogen for all foods,” explained Dr. Wallace.  “That view has changed through the years.”

Dr. Wallace provided an example involving lettuce.

“Years ago, the idea that one test could identify a specific pathogen in all produce, for example, may have seemed rational, but did not work well in practice.  A scheme may have been developed to test for a specific pathogen in all types of lettuce, yet a method that worked well in identifying a specific pathogen in romaine lettuce and iceberg lettuce may not work well with red-leaf lettuce.”

“Both AOAC International and AFNOR Group decided that the all-category approach did not work,” explained Dr. Wallace.

“One of the key factors in determining whether a test meets the ‘Fitness for Purpose’ goal is to determine where food products are similar enough that the same test for a specific pathogen can be used for several specific products rather than just one food product,” stated Dr. Wallace.

Using hot dogs to illustrate his point, Dr. Morgan Wallace stated: “From a molecular standpoint, both
a cooked turkey hot dog and a cooked pork hot dog are similar enough that the same test could
be used to identify a specific pathogen in either variety of cooked hot dog.  The same test may not,
though, work well for a raw sausage.” (Photograph is provided courtesy of Jim Walker from Pixabay.)

“Method development is critical to determine the limits of which food products can be tested with the same scheme,” explained Dr. Wallace.  “Both validation and verification are important aspects for rapid testing of food products.”

“Validation of a testing method – confirming that the testing method works to detect the targets in the manner specified – is done through entities like AOAC International and AFNOR Group.”

“Verification of that testing method is achieved when an individual end user can get the same correct results on a consistent basis.”

“Another aspect on whether a test meets the ‘Fitness for Purpose’ standard is determining the breaking point of the assay level of a test for a specific pathogen,” stated Dr. Wallace.  “This is important as the developer of a test verifies that the negatives found through the test are truly negatives and to ensure that the test works well in the real world.  Where alternative methods differ in performance from reference methods need to be identified and where the alternative method does not perform as well as the reference method, variations on the alternative method can be developed to improve performance.”

“There can be a tremendous difference between testing in a laboratory setting using simplified materials versus testing using samples from a manufacturing plant,” explained Dr. Wallace.  “A test must be able to work well in the real world – not just in a lab using pure cultures.”

Dr. Wallace noted the development of rapid microbiology test kits can be costly.

“Developing these tests can be expensive – it can be both labor-intensive and time-intensive,” stated Dr. Wallace.  “You have a group of highly-trained individuals working together to show that a test has been able to be validated to show that it works well.  Determining what protocol is best is one of the most important aspects of the validation method development process.”

“What diagnostic method providers are able to claim as far as method applicability is different from validating organization to validating organization and has changed over time,” noted Dr. Wallace.  “Not too long ago, method providers could make broader claims than were warranted, but now much narrower claims are allowed.  The problem with narrow claims, though, is that since it is not possible to fully validate all of the tens of thousands of food products, end users will need to test finished products with less than full validations.  Careful evaluations of what products are similar enough to lump with other products must be conducted so that appropriate methods with sufficient validation can be used.”

“Typically, in the United States,” explained Dr. Wallace, “It will be a combination of decisions made by food businesses, validating organizations, independent labs, and governmental regulators to determine how much validation is needed for a rapid test to meet standards of ‘Fitness for Purpose’.”

“The condition of the sample as well as the sample’s size may also impact ‘Fitness for Purpose’,” continued Dr. Wallace.  “Whether the sample is fresh, frozen, partially-frozen, raw, or cooked can impact test results.  A method that works well with a sample that is 25 grams in weight may not work well for a sample that weighs 375 grams.”

“In many cases, industry labs and independent labs operate very differently from government-run labs,” stated Dr. Wallace.  “Many of the labs operated by businesses are staffed 24 hours a day, while the labs run by governmental regulators may only operate during regular business hours.”

Part of that difference is the focus of each entity as they strive to confirm the safety of food.

“The mandate of governmental regulators is not to get the product out-the-door,” explained Dr. Wallace.  “The time needed to get results from a test, though, are far more critical for food producers.  A truck filled with product often cannot move from the loading dock until the test results come back.  Businesses cannot allow food products to go into commerce until and unless the test results show that the food products are safe for consumption.”

“Rapid tests approved for use in Europe may be different from rapid tests approved for use in the United States,” continued Dr. Wallace.  “An example is how testing is done for Listeria.  There are dozens of different species of Listeria, with six core species.  But only Listeria monocytogenes makes humans sick.  It is the species of Listeria that causes the infection listeriosis.”

“In the United States, there is zero tolerance for any Listeria monocytogenes at all to be found in ready-to-eat food products available for human consumption,” stated Dr. Wallace.  “Rapid tests can either target all species of Listeria as an indicator that L. monocytogenes may be present, or can target only the pathogen L. monocytogenes.”

“In Europe, though, a rapid test may need to determine that the level of the L. monocytogenes is below what is considered a safe level in ready-to-eat foods where the L. monocytogenes is not able to grow and make someone sick,” explained Dr. Wallace.

In addition to Drs. Legan and Wallace, Mr. Bradley Ziebell of Conagra Brands will also be making a presentation at this symposium of the International Association for Food Safety.  The title of his presentation will be “Performance Verification and Application of Rapid Pathogen Test Methods:  A Food Company Perspective”.

The three individuals are scheduled to make their presentations from 1:30 PM to 3:00 PM on Monday, July 22nd.  The symposium will be held in the M112 Conference Theater at the Kentucky International Convention Center.

Do you have questions about the rapid microbiology testing industry?

Governmental regulations?  Company operations?

Your questions may be answered in future news columns.

Contact Richard McDonough at biatastailchronicles@gmail.com.

© 2019 Richard McDonough