A QUESTION OF CONFIDENCE – how food microbiology labs can ensure their results are reliable

| November 18, 2014

http://www.dreamstime.com/royalty-free-stock-photography-young-male-researcher-chemistry-lab-image19001167Supply chains have become ever longer and more complex as food production has developed into a truly global industry. The need to monitor the safety of food at every stage means that microbiological testing remains a key weapon in the fight against food contamination and the volume of testing undertaken continues to grow. Today’s laboratories are equipped with state of the art facilities operated by highly trained staff, but just how reliable are their results? A recent study of lab accuracy suggests that there is no room for complacency, but there is a lot that testing labs can do to make sure that their customers have confidence in the results they produce.

In the early 1980s many food manufacturers still relied on microbiological testing as an important tool for ensuring the absence of foodborne pathogens in their products and for monitoring microbiological quality. But this all changed as HACCP-based food safety management systems replaced the much less effective quality control through testing approach. Microbiological testing retained an important role, but as a means of verifying the effectiveness of the HACCP system rather than a frontline defence against pathogens and spoilage organisms. That being so one might expect that the number of microbiological tests carried out by the food industry would have fallen, but the opposite seems to be the case. According to a market report from Strategic Consulting, Microbiology Testing in the Global Food Industry- 8th Edition, the food industry worldwide is likely to have carried out almost a billion microbiology tests in 2013 and the volume of testing has increased by 128% over the last 15 years. The number of pathogen tests has increased even more, and now accounts for almost a quarter of the total. The value of the global food microbiology testing market is now estimated to be about $3 billion and has risen by 40% in five years.

There are a number of reasons for this rapid growth. One is simply that the food industry itself has grown substantially. But the extended supply chains that are necessary in modern food production systems and repeated safety problems, such as the 2011 E. coli outbreak in Germany, have also increased the demand for testing significantly. Microbiological tests are now applied at many stages in the supply chain, from primary production to retail, and are used more and more to monitor the safety and quality of ingredients as well as to verify that HACCP systems are working as they should. New rapid methods have also made testing a viable option in applications where traditional culture-based methods take too long to produce a result.

Laboratories progress but old problems remain

The laboratory infrastructure serving the food industry has changed dramatically since the 1980s. At that time many food manufacturers and retailers maintained in-house lab facilities for microbiology, but this is no longer the case. Much testing is now outsourced, many small independent laboratories have closed and the sector is now dominated by a small number of much larger centralised facilities, which are often part of a multinational chain. Much improved sample collection and logistics allow testing to be done in labs located further away from their customers, sometimes in a different country. The economies of scale created by these changes have allowed more investment in sophisticated instrumentation and automated preparation systems, reducing the number of technicians required and allowing labs to focus on training their staff to a higher standard.

All this means that the food microbiology labs serving the food industry are almost unrecognisable in comparison to those operating in 1980 and a comprehensive modernisation process has taken place. But many of the routine methods in use by labs today have changed little over the last thirty years. Despite the introduction of DNA-based techniques, many are still based on culturing the target microorganisms and then applying further biochemical or immunological tests to confirm their identity. These traditional methods have their roots in the medical microbiology of the nineteenth century and not only take time, but are also subject to a wide range of influences that can alter the result. The composition of the culture medium, sterilisation cycles, fluctuations in temperature within incubators, and of course the skill of the staff performing the test can all have an effect. The microorganisms themselves may also decline to cooperate. There is convincing evidence that some pathogens may enter a ‘viable but non-culturable’ (VNC) state in certain foods, making their detection by conventional methods almost impossible. These factors can cause considerable variation in the results obtained by different labs analysing the same samples, or even variable results from within the same laboratory.

Systems designed to address these problems have necessarily become a vital part of the successful operation of a food microbiology lab. Nevertheless, results of an American Proficiency Institute study presented at a scientific meeting in May show that potentially dangerous false negative results returned by US labs from proficiency testing samples (see below) were surprisingly common, especially for Campylobacter (9.1%) and Salmonella (4.9%). A lower, but still significant rate of false positives was also noted. If similar figures apply in Europe and other regions, as is quite probable, it would be a cause for some concern.

Managing quality

The most obvious way of minimising errors and variation in laboratory test results is the development of an effective quality management system. The same quality principles can be applied to lab testing as to any other industrial process where there are inputs and outputs and procedures that can be documented. A quality management system for a food microbiology laboratory should cover every activity involved in the testing process from receipt of samples to reporting results. But there are some aspects of operating food microbiology testing that are of particular importance for building confidence in the integrity of those results.

Most basic of these is the accommodation and facilities available for the laboratory. In the past some microbiology labs were set up in unsuitable locations, sharing facilities with chemical analysis, or occupying areas within factories lacking adequate space, or even proper separation from production areas. This is largely a thing of the past, not least for health and safety reasons, and the growth of third party testing has helped to improve the design and fitting out of food microbiology laboratories. No matter how well equipped and staffed a lab is, reliable performance cannot be guaranteed if the basic accommodation is inadequate. Equally important is the choice and operation of laboratory equipment. All instruments and other equipment must be suitable for the purpose for which they will be used and must be maintained, calibrated and operated according to set procedures. Maintenance, regular calibration and operation of equipment also need to be recorded and documented to demonstrate that each item is operating within the desired limits. For example, a small rise in autoclave temperature during a pre-set sterilising cycle can have an adverse effect on culture medium and could result in false negative results. As modern food microbiology evolves into a less hands-on operation and more instrumentation finds its way onto the laboratory benches, this is becoming increasingly important.

Staff working in the laboratory are clearly a vital part of the operation. Responsibilities must be clearly defined and each technician must have received adequate, documented training to perform their roles. While on-the-job training is extremely useful, certain minimum qualifications and skills have to be demonstrable. It is also important to ensure that training is provided when new methods or equipment are introduced into the laboratory. It is vital that the test results generated by the lab are completely traceable from sample reception and identification right through the analytical procedure to the reporting stage. When a non-compliance or an unexpected result is indicated, it must be possible to follow the testing process fully to determine whether that result is genuine, or the result of an error or malfunction. While this can be done using a paper trail, many laboratories now use an integrated computer-based record system such as LIMS (Laboratory Information Management System), often directly linked to automated instruments and equipment to minimise the need for manual record keeping.

Choosing the right test methods

Outbreaks picWhile all of these factors are important, the real key to reliable results is often the choice of test method. Chosen test methods must be appropriate for the purpose and for the food material being tested. They must also have been properly validated, which simply means that sufficient objective evidence has been collected and examined to confirm that the method is fit for its intended use. The simplest way to achieve this is to adopt recognised standard methods, such as those published by the International Standards Organisation (ISO). These have the considerable advantage of having been exhaustively validated during their development and are tried and trusted. Current ISO methods exist for most of the tests routinely carried out in food microbiology laboratories, both for pathogen detection and enumeration of spoilage organisms and hygiene indicators. There are also other sources of validated methods, such as the Bacteriological Analytical Manual (BAM) published by the FDA in the US, and methods developed by recognised scientific organisations, professional bodies and trade associations.

It is still feasible for a laboratory to use non-standard methods, or to develop in-house methods for specific purposes, but the laboratory must properly validate these. Method validation can be a lengthy and costly business and usually involves testing using standard reference cultures, comparisons with other validated methods, determining reproducibility, inter-laboratory comparisons and assessment of the factors that might affect test results and of measurement uncertainty. It is not surprising that many labs prefer to adopt standard methods whenever they are available. There is an argument that this reliance on published standards may be slowing progress in method development, especially in terms of rapid methods based on molecular biology and other innovative technologies. This may be true to some extent, but most food manufacturers and consumers would probably agree that the integrity of laboratory test results is paramount.

Widespread use of ISO and other standard methods has certainly not halted a trend towards the adoption of new rapid methods. While most ISO methods are culture-based, many allow the use of immunological and other rapid tests for confirmatory testing in place of traditional biochemical tests. Many rapid microbiology tests and commercial instruments have also been thoroughly assessed and have received certification from bodies such as the AOAC in the USA, NordVal in Scandinavia and French-based AFNOR, which grants its NF VALIDATION mark to properly validated alternative food microbiology methods in Europe. More than a hundred alternative methods have already been granted the NF VALIDATION mark and they include a number of molecular detection methods based on PCR and other techniques. This means that laboratories can adopt methods based on new technologies without having to undertake prohibitively expensive and time consuming validation, although some work to demonstrate that the method works properly in the environment of a specific lab will still be necessary, as will an assessment of measurement uncertainty.

Demonstrating competence

An excellent way to check the reliability of a lab’s test results is to take part in an external quality assurance scheme, usually referred to as proficiency testing. This involves receiving samples prepared by a third party, which contain known pathogens or levels of microbiological contamination, and testing them according to standard laboratory procedures. The results are then returned to the body that prepared the samples for assessment. Proficiency testing is valuable as a means of alerting management to potential quality issues in the laboratory and as an objective assessment of the competence of the laboratory. An example of such a scheme is the Food Examination Performance Assessment Scheme (FEPAS) run by the Food and Environment Research Agency in the UK. The FEPAS scheme sends out samples to its members periodically that are carefully prepared to mimic real food samples – even to the extent of containing background microflora – and have been inoculated with known pathogens, indicators, or spoilage organisms. The laboratory then tests the samples according to its usual procedures and submits the results. These are analysed statistically and an assessment report is sent back to the lab. Most food microbiology laboratories now undertake external proficiency testing, often as a requirement of the quality management system.

Accreditation

Over the last twenty years it has become almost essential for food microbiology laboratories to demonstrate their competence and the reliability of their results by gaining some form of third-party accreditation. To gain accreditation the laboratory will need to ensure that all aspects of its operation meet the requirements of a recognised standard. Once that has been achieved, the operation is inspected and assessed by an appropriate certification organisation to confirm that the laboratory complies with the standard. Once the certificate is issued, the laboratory is accredited to the standard and will be periodically re-assessed to ensure that it continues to comply. Laboratories are typically accredited for specific tests rather than for the full range of services they offer. This means that potential customers need to check that their chosen lab is accredited for all the tests that they need, but accreditation is a very strong indicator that the laboratory is properly equipped, staffed and managed and will provide a reliable service. Many national food safety authorities, large food manufacturers and retail chains insist that only accredited labs conduct any testing done on their behalf.

The accreditation standard chosen by most food microbiology laboratories is ISO/IEC 17025:2005 General requirements for the competence of testing and calibration laboratories, which covers the general requirements of running a laboratory competently and responsibly. It is wide ranging and includes requirements applying to all aspects of the laboratory operation, but it is a general standard and not one designed specifically for food laboratories. This can make interpretation of its requirements problematic. Fortunately, guidance on interpretation for food microbiology labs has been published, notably by European co-operation for Accreditation (EA), which issued guidance document EA – 4/10 Accreditation for Microbiological laboratories, designed principally for food and environmental testing labs, in 2002. There are also a number of other accreditation schemes designed specifically for food microbiology laboratories. An example is the UK-based Camden Laboratory Accreditation Scheme, or CLAS. CLAS accreditation also meets the requirements of broader food industry standards, such as the BRC Global Standard – Food. More than 260 labs in the UK and elsewhere are currently accredited under the CLAS scheme

It is clear that much has been done to build confidence in food microbiology test results over the last thirty years. Today’s lab is likely to be well equipped, have some form of accreditation and a comprehensive quality management system designed to protect the integrity of the test procedures and the overall laboratory operation. The methods in use will be proven, the technicians well trained and their competence demonstrated by taking part in proficiency testing. But it is unreasonable to expect 100% accuracy from even the best run laboratory. There are so many variables that might affect the results, not least the diversity inherent in microbial populations, that it is virtually impossible to take all of them into account. New DNA-based detection methods may help to improve the reliability of routine test methods by focusing on fixed genetic markers rather than variable phenotypic characteristics, but molecular biology methods are also prone to uncertainty. For example, components in some foods can interfere with the PCR process, or with extraction of microbial DNA. As these methods develop, quality assurance measures to cope with sources of uncertainty will need to develop too. It is unlikely that food microbiology tests will ever be fool proof and a focus on managing quality will remain a key aspect of running a successful lab.

Sources of more information

FEPAS web pages

EA Accreditation guidance for microbiology laboratories

 

Based on an article first published in Food Engineering and Ingredients (September 2013)

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