Tag Archives: microorganisms

Robin Stombler, Auburn Health Strategies
In the Food Lab

Five Questions Food Facilities Should Ask About Testing

By Robin Stombler
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Robin Stombler, Auburn Health Strategies

The FDA issued the first of several final regulations aimed at modernizing the food safety system through the use of hazard analysis and risk-based preventive controls. Inherent in this system are a number of requirements that eligible food facilities must follow, such as developing a written food safety plan, monitoring, corrective actions and verification. Laboratory testing is an essential component as well.

Robin Stombler presented “Laboratory Oversight and FSMA: Why and When” at the Food Labs Conference in Atlanta, GA | March 7–8, 2016So, what should food facilities know about laboratory testing within the context of the preventive controls for human food final rule?  First and foremost, the final rule states, “facilities have a responsibility to choose testing laboratories that will produce reliable and accurate test results.”  While a future regulation is expected to address the need for accredited laboratories and model laboratory standards, the preventive controls rule adopts other requirements pertaining to testing. Here are five questions that food facilities should ask about testing and the preventive controls rule.

1. What is the difference between pathogens and microorganisms?

The final rule defines “pathogen” to mean a microorganism that is of public health significance. A microorganism is defined as “yeasts, molds, bacteria, viruses, protozoa and microscopic parasites, and includes species that are pathogens.” Microorganisms that are of public health significance and subject food to decomposition or indicate that the food is adulterated or is contaminated with filth are considered “undesirable.”

2. How must food facilities account for pathogens?

Food facilities must prepare and implement a written food safety plan. One component of the food safety plan must include a written hazard analysis. This analysis must identify known or reasonably foreseeable hazards. These hazards may be biological, which includes parasites, environmental pathogens and other pathogens.

In another example, the food safety plan must include written verification procedures. This is to demonstrate that the facility is verifying that its preventive controls are implemented consistently and are significantly minimizing or preventing the hazards. These verification procedures are intended to be appropriate to the particular food facility, the food in question, and the nature of the preventive control and its role within the facility’s food safety system. With this in mind, facilities must conduct activities such as product testing for a pathogen or an appropriate indicator organism or other hazard, and environmental monitoring.

3. Are there written procedures specific to product testing?

Yes. Procedures for product testing must be scientifically valid and must identify the test microorganisms or other analytes. The procedures for identifying samples, including their relationship to specific lots of products, must be written and implemented. The procedures for sampling, including the number of samples and the sampling frequency, must be outlined. The facility must recognize the laboratory conducting the testing as well as describe the tests that are performed and the analytical methods used. Corrective action steps must also be included.

4. What are the procedures for environmental monitoring?

Similar to product testing, these procedures must be scientifically valid, identify the test microorganisms, and be put in writing. For routine environmental monitoring, the location from which the samples are collected and the number of sites that are tested must be stated. The final rule indicates that the “number and location of sampling sites must be adequate to determine whether preventive controls are effective.”  Written procedures must also identify the timing and frequency for collecting and testing samples. Again, similar to product testing, the laboratory conducting the testing and the tests and analytical methods used must be divulged. Corrective action procedures must also be included.

5. How does the supply-chain program incorporate testing?

A receiving facility is required to document a written supply chain program in its records. A component of that program includes documentation of sampling and testing performed as a supplier verification activity. The documentation must include identification of the raw material or other ingredient (including, if appropriate, lot number) and the number of samples tested. It also means that the tests conducted and the analytical methods used must be identified. The date the test is conducted as well as the date of the test report must be provided, and the identity of the laboratory performing the testing must be revealed. Any corrective actions that were taken in response to a hazard detection must also be reported.

This Q&A provides a glimpse into how the preventive controls final rule for human food incorporates laboratory testing. For more details, access the final rule.

Using ATP-based Methods for Cleaning and Sanitation Verification

By Camila Gadotti, M.S., Michael Hughes
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There are several factors that must be considered when selecting a reliable and accurate system for detecting adenosine triphosphate.

A common way to assess the effectiveness of cleaning and sanitation programs in food manufacturing facilities is through the use of methods that detect adenosine triphosphate (ATP). Methods based on ATP detection are inexpensive and rapid, and provide the ability to perform onsite in real-time. There are several manufacturers of ATP-based methods, but choosing the most reliable one can be a daunting task. This article will discuss how these methods work and which factors should be considered to make an informed purchasing decision.

ATP is the universal energy currency in all living cells. It is present in all viable microorganisms (with the exception of viruses) and in foodstuffs. High amounts of ATP can be found in some fresh foods like vegetables, while other foods, especially highly processed foods such as fats, oils or sugar, contain very low amounts of this molecule. It is also important to know that ATP can be found in the environment in its free form hours after a cell has died.1 An ATP bioluminescence assay operates on the principle that ATP in food/food residues and microorganisms, in the presence of a luciferin/luciferase complex, leads to light emission. This light can be measured quantitatively by a luminometer (light-detecting instrument), with results available in 10–40 seconds. The amount of light emitted is proportional to the amount of ATP on a surface and hence its cleanliness. The light emitted is typically measured in relative light units (RLUs), calibrated for each make of instrument and set of reagents. Therefore, the readings obtained from assessing the cleaning of food manufacturing facilities need to be compared with baseline data representing acceptable clean values.

Varying Optical Components

Luminometers have evolved over the years from very large and cumbersome in size to small handheld models that can be used anywhere within a manufacturing facility. Although several components are housed inside these instruments, the optical component is the most important part of a luminometer. Used to detect light coming from the ATP/luciferin/luciferase reaction, the optical component is the defining factor related to luminometer reliability, sensitivity and repeatability. Good luminometers use a photomultiplier tube (PMT) in the light detection system; however, as part of the drive toward cheaper and smaller instruments, some manufacturers have replaced PMTs with less-sensitive photodiode-based systems. When using photodiodes, the swab chemistry must be adapted to produce more intense light. This results in a shorter duration of light, decreasing the time window allotted to place the swab in the luminometer and obtain an accurate read. A PMT, however, multiplies the electrical current produced when light strikes it by millions of times, allowing this optical device to detect a single photon. This approach emits light over a longer period of time. Although the weight of the system is also dependent on factors such as the battery, case and the display screen, a luminometer constructed with a photodiode will generally weigh less than a luminometer constructed with a PMT, since the former is smaller than the latter.

Sensitivity Testing

When an ATP hygiene monitoring system has poor sensitivity or repeatability, there is substantial risk that the test result does not truly represent the hygienic status of the location tested. Therefore, it may provide false positives leading to unnecessary chemical and labor costs and production delays, or false negatives leading to the use of contaminated pieces of equipment. A system that is sensitive to low-level contamination of a surface by microorganisms and/or food residues allows sanitarians to more accurately understand the status of a test point. The ability of a system to repeat results gives one peace of mind that the result is reliable and the actions taken are appropriate. To test different ATP systems for sensitivity, one can run the following simple test using at least eight swabs per system:

•    Make at least four serial dilutions of a microbial culture and a food product in a sterile phosphate buffer solution.
•    Using an accurate pipette, dispense 20 μl of these dilutions carefully onto the tip of the swabs of each ATP system and read the swabs in the respective luminometer, following the manufacturer’s instructions.
•    Use caution when dispensing the inoculum onto the swab head to prevent any sample loss or spillage. In addition, it is very important the swabs are inoculated immediately prior to reading, which means that each swab should be inoculated one at a time and read in the respective luminometer. Repeat this process for all the swabs.


To test different ATP systems for sensitivity, one can run a simple test using at least eight swabs per system. Photo courtesy of 3M

The most sensitive system will be the one that results in the most “fail results” (using the manufacturers’ recommended pass/caution/fail limits).

One can also test different ATP systems for repeatability by the following test:

•    Prepare a dilution of a standard ATP positive control or a food product such as fluid milk in a sterile phosphate buffer. If using a standard ATP positive control, follow the manufacturer’s direction to prepare dilution. If using fluid milk, add 1 ml of milk into 99 ml of phosphate buffer.
•    Using an accurate pipette, dispense 20 μl of this standard onto the tip of the swabs of each ATP system and read these swabs in their respective luminometer, following the manufacturer’s instructions.
•    Prepare and read at least 10 swabs for each system you are evaluating, and capture the results on a digital spreadsheet.
•    Once all 10 swab results (for each system) are in the spreadsheet, calculate the mean (=average) and standard deviation (=stdev) for each system’s data set. Divide the standard deviation by the mean and transform the result in percentage; this value is called the coefficient of variation percentage (CV%).
The test with the lowest CV% is the most repeatable and will provide the most reliable information to help make the correct decisions for a food manufacturing facility.

Choosing the Right ATP System

There are many ATP systems available on the market to support cleaning and sanitation verification in manufacturing plants. Some systems are more reliable than others and will provide results that are meaningful, accurate and repeatable. Be sure, therefore, not to choose a system solely based on its price. Check for the quality of the instrument, ask the sales representative what kind of optical device is used in the construction of the instrument and, moreover, perform an evaluation running tests for both sensitivity and repeatability. It is also important to consider the functionality and usability of the software provided with the system to ensure that the software can be used to customize sample plans, store test results and produce reports and graphs.


  1. Jay, J. M., ‎Loessner, M. J., & Golden, D. A. (2008). Modern Food Microbiology.


About the Author:

Camila Gadotti, M.S., is a field technical service professional and Michael Hughes is a technical service professional with 3M Food Safety.