Viruses, bacteria, spores – our world is populated by organisms both large and small. This is not always beneficial to humans, especially when they are ill. It is therefore essential that medical devices have a certain degree of purity. However, if products are intended for sterile use, it is essential to assess and monitor their microbial and particle contamination – better known as bioburden and pyrogenicity.
What is “bioburden”? And why is it relevant?
Bioburden refers to all viable microorganisms on a specific surface or volume unit in the air. The composition of these germs can range from various bacteria to viruses and fungi. The decisive factor is that these microorganisms are viable, which also includes survival states such as encapsulated bacteria or fungal spores.
These microorganisms can be made visible by transferring them to a nutrient medium and stimulating growth under appropriate environmental conditions. Each individual cell then begins to metabolise and grow and divide. Typical patterns of colonies of the respective species then develop on the nutrient medium. The number of colonies present indicates the quantity of germs that were previously present – the bioburden is therefore expressed in CFU colony-forming units – per unit. This “unit” is usually related to a product or, in the case of flat textile fabrics, per m². However, it can also refer to CFU per m³ of air in the production area.
Germs are everywhere – even on medical devices. In the worst case, they can have fatal consequences. This is why it is necessary for many products to reduce or inactivate existing germs as far as possible. Three basic methods have been established to combat germs:
- Cleaning – mechanical/chemical removal of residues and particles from production, but also from any previous use of the medical device, including the removal of germs
- Disinfection – reduction, inactivation or killing of pathogenic microorganisms on an object or surface to prevent infection
- Sterilisation – killing or irreversibly inactivating microorganisms and spores on materials and objects, thereby eliminating germs capable of reproduction
It is important to understand that both disinfection and sterilisation are probability assumptions. The term “sterile” is defined as the absence of at least 10-6germs.
Depending on the environment in which the medical device is used and its intended purpose, it is necessary to assess how critical the existing bioburden is and what measures must therefore be taken to reduce and control it.
The necessity is most obvious for sterile products that are either used once or reprocessed before each use. However, even non-sterile products should have a certain level of cleanliness to meet general hygiene requirements. For reusable non-sterile products, such as a toilet chair in a hospital, particular attention must be paid to designing the product in such a way that surfaces can be cleaned and disinfected effectively.
What are the regulatory and normative requirements?
In principle, the requirement to control and minimise microbial contamination stems from the MDR – as a fundamental requirement that must first be considered applicable to every product. The general safety and performance requirements point 11 form the general framework for “infection and microbial contamination”.
- Devices and their manufacturing processes shall be designed as to eliminate or to reduce as far as possible the risk of infection to patients, users and, where applicable, other persons. […]
- Where necessary, devices shall be designed to facilitate their safe cleaning, disinfection, and/or re-sterilisation. […]
- Packaging systems for non-sterile devices shall maintain the integrity and cleanliness of the product and, where the devices are to be sterilised prior to use, to minimise the risk of microbial contamination; […]
This also makes it clear that knowledge and control of microbial contamination is necessary for both non-sterile and sterile products and that the packaging must be designed and manufactured in a suitable manner (=>; reference to sterile packaging).
The requirement for manufacturing under appropriate and controlled conditions is also explicitly stated in a separate subchapter; both points are reflected in the specific requirements of ISO 13485 (Ch. 6.4.2 and 7.5.5 and 7.5.6).
GSPR 11 is addressed in corresponding (harmonised) standards – these are a series of different standards for the validation of sterilisation, depending on the type of sterilisation:
- EN ISO 17665 “Sterilisation of health care products – Moist heat – Requirements for the development, validation and routine control of a sterilisation process for medical devices”
- ISO 11135 “Sterilisation of health care products – Ethylene oxide – Requirements for the development, validation and routine control of a sterilisation process for medical devices”
- ISO 11137-1 “Sterilisation of health care products – Radiation – Part 1: Requirements for the development, validation and routine control of a sterilisation process for medical devices”
All standards for the validation of sterilisation processes require the establishment and maintenance of a system for monitoring the microbial quality of the product in the relevant chapter 7 on product definition. The determination and definition of warning and action values for bacterial contamination, including specified recommendations for measures to be taken if the values are exceeded, has become established practice here.
To carry out the validation, it is necessary to apply further subordinate standards that specify, for example, requirements for sterilisation equipment or the bioindicators to be used, etc. One standard should be highlighted here in relation to microbial contamination, namely “ISO 11737-1 Sterilisation of health care products – Microbiological methods – Part 1: Determination of a population of microorganisms on products”. This is because all sterilisation processes aim to verify the “sterile” state, and deals with the inactivation of existing germs, which is more successful the lower the germ load – i.e. the bioburden – is before sterilisation.
Factors influencing low or high bioburden: How can bioburden be monitored?
In order to keep germs on products under control, it is good to know where the germs come from and why bioburden values can fluctuate. Microbes are so small that they cannot be seen, so in theory you have to consider where the germs occur: initially everywhere, on and in living organisms, including all production employees, on theoretically all surfaces and in the air, especially on the particle load contained in the air. Populations are particularly high where conditions are particularly favourable for germ growth: where organic material is available as a food source and sufficient heat and moisture are provided.
The first and most important measure for keeping the germ load within a defined range is the appropriate design of the production area. Without controlled conditions, the bioburden cannot be monitored. A clean room in accordance with ISO 14644-1 Class 7 or 8 is not always necessary, but the minimum requirements for a controlled area are:
- A closed area with overpressure control, air conditioning and air filters. This can also be a laminar flow unit.
- A hygiene concept for staff that includes work clothing (including caps, beard protection, special shoes, etc.), hygiene rules such as hand washing, hand disinfection, rules for handling cosmetics, smoking, short fingernails, procedures for respiratory diseases, etc., and regular training.
- A concept for cleaning and disinfecting rooms and surfaces.
- Suitable structural/spatial facilities for the entry of personnel and materials (e.g. contaminated transport packaging – and the bioburden on it – should not be brought into the controlled area)
The next step is to define the appropriate specifications for the relevant bioburden limits for the materials, auxiliary materials and operating supplies used in production (at the very least, these should not be ignored). Work equipment such as rulers used in the controlled production area should also be cleaned regularly and disinfected if necessary. The specifications also include suitable requirements for raw material packaging to ensure that the goods can be properly transferred to the controlled production area.
Don’t forget: keep an eye on your suppliers! As a manufacturer, it is your duty to assess, redefine and monitor the necessary hygiene conditions for materials or product precursors at your suppliers.
The actual production of the medical device is also very important. If work steps are required that cause high contamination with organic residues, particles or lubricants that have no place on your finished medical device, you should introduce and validate suitable cleaning and/or disinfection steps.
Mechanical production processes involving high temperatures (e.g. plastic injection moulding) result in products with significantly lower bacterial contamination than manual manufacturing processes (e.g. the manual production of cotton swabs) requiring a high level of personnel. The germ load is never constant; it is also subject to seasonal fluctuations, which can lead to different environmental conditions depending on the building’s facilities, which in turn affect contamination in production: temperature, humidity, employees’ susceptibility to colds, etc.
How can the bioburden be monitored?
Once the potential sources of bioburden contamination in the product have been identified, it is important to assess the effectiveness of the defined procedures. It is essential to carry out regular bioburden measurements at all points in the production chain. The most important value is, of course, the bioburden on the product, but this will certainly be subject to fluctuations and cannot be controlled without measurement results for the relevant influencing factors. Other useful sampling points should be selected depending on the product and the defined threshold values, starting with door handles, work surfaces and equipment, and extending to the palms of employees’ hands.
EN ISO 11737-1:2018 + A1:2021 describes in detail how the tests are to be carried out, how the bioburden should be determined and characterised, and how the methods are to be validated. Chapter 8.6 contains the requirement to define bioburden levels, with a supplement in Annex A, which explicitly explains the warning and action limits.
Controlling the bioburden is most important when sterilisation is carried out using radiation, as this method is very directly linked to the actual microbial load for achieving the SAL. In the latest edition of ISO 11137-1 from April 2025 (not yet harmonised, but certainly required by the notified bodies as state of the art), the specification of warning and action values has even been included in the normative part, together with the requirement to provide a characterisation of the bioburden. To this end, as explained in Annex A, the manufacturer should ensure that it employs personnel with sufficient expertise in microbiology, sterilisation safety and the statistical evaluation of data (the alarm and action ranges are usually determined using standard deviations). The warning and action values must be defined by the respective manufacturer on a risk basis and should not be defined lightly and without sufficient expertise.
What are pyrogens and what do they have to do with bioburden?
In medicine, pyrogens (from the ancient Greek pyr = fire and gennáein = to generate) are substances that, when administered parenterally (i.e. via the gastrointestinal tract), are capable of causing fever. In the manufacture of medical devices, there are two types of pyrogens that may be of importance to manufacturers:
Firstly, tiny particles of the material used in medical devices or other chemical substances resulting from production can cause fever reactions. This material-related pyrogenicity should be addressed in the biological evaluation (see also ISO 10993-11, Annex G) and can only be detected by means of rabbit tests (according to Ph. EUR or USP). For monitoring in production, this means that it is essential to keep an eye on the number of other contaminants and particles in addition to the microbial load (bioburden).
On the other hand, a reaction occurs in the body due to endotoxins produced by Gram-negative bacteria. These substances produced by the bacteria remain on the products even after the germs have been killed during sterilisation. It is therefore necessary to know the pyrogenic load present on the products, to control it, to keep it as low as possible and also to regularly provide evidence that no undesirable endotoxins remain. This can be done with the Limulus Amebocyte Lysate LAL test, which is described in the European Pharmacopoeia. Since 2021, there has also been an international standard – ISO/TR 21582:2021 Pyrogenicity – Principles and methods for pyrogen testing of medical devices– which, however, has not yet gained widespread recognition and has not yet been harmonised in the EU or recognised by the FDA.
For some products, pyrogen-free status is desirable or even required instead of or in addition to sterility, especially for products that come into contact with the body. In this case, it is necessary to examine production processes and possible contamination during manufacturing very carefully, incorporate suitable cleaning steps if necessary, and verify a certain level of purity as part of a validation process. A single negative LAL test or rabbit test is not sufficient for permanent proof of pyrogen-free status.
Bioburden and pyrogenicity – both topics are becoming increasingly important in the context of MDR certification, whether in the context of reprocessing validation or a new perspective on production processes. Even if counting may be tedious, don’t turn a blind eye to it! We are happy to assist you with detection and monitoring. Users and patients will thank you for it.
Â
