The Use of Risk Assessment Tools For Microbiological Assessment of Cleanroom Environments


Environmental monitoring describes the microbiological testing undertaken in order to detect changing trends of microbial count and microflora within cleanroom or controlled environments. The results obtained provide information about the physical construction of the room, the performance of the HVAC system, personnel cleanliness, gowning practices, the equipment and cleaning operations.

Over the past decade environmental monitoring has become more sophisticated in moving from random sampling, using an imaginary grid over the room and testing in each grid, to the current focus on risk assessment and the use of risk assessment tools to determine the most appropriate methods for environmental monitoring.

A risk assessment procedure is required to determine locations for environmental monitoring. Such risk based approaches are recommended in ISO 14698 and regulatory authorities are increasingly asking drug manufacturers about this subject.

Risk based approaches include FMEA (Failure Mode and Effects Analysis); FTA (Fault Tree Analysis) and HACCP (Hazard Analysis Critical Control Points), all of which employ a scoring approach 1 . At present, no definitive method exists and the various approaches differ in their process and the degree of complexity involved. However, the two most commonly used appear to be HACCP (which originated in the food industry) and FMEA (which was developed for the engineering industry) (Whyte and Eaton, 2004a).

These various analytical tools are similar, in that they involve:

  • Constructing diagrams of work flows
  • Pin-pointing areas of greatest risk
  • Examining potential sources of contamination
  • Deciding on the most appropriate sample methods
  • Helping to establish alert and action levels
  • Taking into account changes to the work process / seasonal activities

These risk assessment approaches are not only concerned with selecting environmental monitoring locations. They integrate the environmental monitoring system with a complete review of operations within the clean room to ensure those facilities, operations and practices are also satisfactory. The approaches recognise a risk, rate the level of the risk and then set out a plan to minimise, control and monitor the risk. The monitoring of the risk will help to determine the frequency, locations for and level of environmental monitoring (for example, refer to an article by Sandle [2003a], for a more detailed example).

This paper explores an example from three different techniques:

  • A simple conceptualisation of a risk using a table
  • FMEA

Example 1: The ‘Tabular Approach’ An example of analysing the risks which environmental monitoring are designed to detect, through a simple table, would be:

Area or Equipment: Sterility Testing Isolator
Risk: Contamination due to build up of microbial counts in the isolator environment
Failure or Situation: Failure to adequately clean after a use
EFFECT Minimising the risk (Mitigations to reduce risk) Monitoring

• If isolators are not cleaned regularly there is a possibility of micro-organisms remaining in the environment.


• Cleaning surfaces using water to remove dirt or spillages prior to the application of a suitable disinfectant.

• The disinfectant used must have a wide spectrum of efficacy but not be aggressive to the isolator material.

• The isolator should be designed so that it is easy to clean.



• An environmental monitoring programme (using settle plates, air samples, contact plates, swabs and finger plates) will show the areas of greatest risk. This data should be examined for trends.

• For out-of-limits environmental monitoring results appropriate corrective and preventative actions should be put in place.


Example 2: HACCP

The principles behind constructing an HACCP analysis are:

  1. Identify hazards (contamination risks) and assess their severity.
  2. Determine critical control points (CCP’s)
  3. Establish critical limits
  4. Establish system to monitor control of the CCP’s
  5. Establish the corrective action (when CCP is not under control)
  6. Establish procedures for verification to confirm that the HACCP system is working effectively
  7. Establish documentation and reporting systems for all procedures

Each of these ‘seven key points’ is a vital step in developing the risk assessment. The seven points include:

  1. To identify sources of contamination, construct a risk diagram / diagrams. This will show sources and routes of contamination. Examples include:

a) Areas adjacent to clean room / Isolator (e.g. airlocks, changing rooms)

b) Air supply

c) Room air

d) Surfaces

e) People

f) Machine

g) Equipment

2. Assess the importance of these sources and if they are / are not hazards that need to be controlled. Examples include:

a) The amount of contamination on, or in, the source that is available for transfer.

b) Ease by which the contamination is dispersed or transferred.

c) The proximity of the source to the critical point where the product is exposed.

d) How easily the contamination can pass through the control method.

The use of a scoring method can be very helpful.

3. Identify the methods that can be used to control these hazards. For example:

a) Air supply / HEPA filters

b) Dirty areas adjacent to cleanroom / Isolator (including differential pressures; airflow movement)

c) Room air (air change rates; use of barriers)

d) Surfaces (sterilisation; effectiveness of cleaning / disinfection procedures)

e) People (cleanroom clothing and gloves; room ventilation; training) f) Machines and equipment (sterilisation, effectiveness of cleaning, exhaust systems)

4. Determine valid sampling methods to monitor either the hazards or their control methods or both. For example:

a) HEPA filter integrity tests

b) Air supply velocity, air change rates

c) Room pressure differentials

d) Particle counts

e) Air samplers; settle plates, contact plates etc.

5. Establish a monitoring schedule with ‘alert’ and ‘action’ levels and corrective measures to be taken (for when these levels are exceeded). For example:

a) The greater the hazard, the greater the amount of monitoring required.

b) Trend analysis for alert and action levels: in or out of control.

6. Verify that the contamination control system is working effectively by reviewing key targets like product rejection rate, sampling results, control methods and so on. These may need to be modified over time. For example:

a) System for data review

b) Examine filling trials

c) Audit

d) Reassess hazards, effectiveness of control systems, frequency of monitoring, appropriateness of alert and action levels.

7. Establish and maintain documentation. For example:

a) Describe the steps being taken

b) Describe the monitoring p [procedures

c) Describe the reporting and review procedures Before implementing HACCP it is important to train all staff involved in the process and to use a multi-disciplinary team. For example, this may be made up of personnel from production, engineering, QC/QA, validation and so on.

Example 3: FMEA

FMEA schemes vary in their approach, scoring and categorisation. All approaches share in common a numerical approach. The example here, based on a sterility testing isolator, was to assign a score (from 1 to 5) to each of the following categories:

i) Severity

ii) Occurrence

iii) Detection


i) Severity is the consequence of a failure

ii) Occurrence is the likelihood of the failure happening (based on past experience)

iii) Detect is based on the monitoring systems in place and on how likely a failure can be detected.

By asking a series of questions each main part of the clean room or isolator system can be broken down into key parts. Such questions included:

i) What is the function of the equipment? How are its performance requirements?

ii) How can it fail to fulfil these functions?

iii) What can cause each failure?

iv) What happens when each failure occurs?

v) How much does each failure matter? What are its consequences?

vi) What can be done to predict or prevent each failure?

vii) What should be done if a suitable proactive task cannot be found?

The scoring is 1 (very good) to 5 (very bad). Therefore, a likelihood of high severity would be rated 5; high occurrence rated 5; but a good detection system would be rated 1.

Using these criteria a final FMEA score is produced:



From: severity score x occurrence score x detect score

Depending upon the score produced it can be decided whether further action is needed. There is no published guidance on what the score that dictates some form of action should be. A suggested score is 27 for the cut-off value where action was required. This was based on 27 being the score derived when the mid-score is applied to all three categories (i.e. the numerical value ‘3’ from severity (3) x occurrence (3) x detect (3)) and the supposition that if the mid-rating (or a higher number) was scored for all three categories then as a minimum the system should be examined in greater detail.

An example of one area of an isolator operation (the risks associated with the room in which the isolator is housed) is examined below.

Description of critical area: The isolator is situated in an unclassified room. There is not requirement to place a sterility testing isolator in a classified room.

FMEA schematic:

Process step Failure Mode Significance of failure Severity of consequence (score)

Loading isolators pre-sanitisation / performing sterility testing

That contamination from the room could enter transfer or main isolators

Reduced efficency of transfer isolator sanitisation / contamination inside main isolator


Measures to detect failure Occurrence (score) Detection systems Detection (score)

Would be shown from reduced evaporation rate for isolator sanitisation / poor environmental monitoring results in main isolator / potential sterility test failures / sanitisation cycle has been validated using biological indictors of 106 spores


Isolator room is monitored monthly for viables and particles / staff wear overshoes on entry / Dycem mat in place / entry to room has controlled access / environmental monitoring performed inside main isolator / isolators are at positive pressure to the room and air is HEPA filtered


FMEA score: 3 x 1 x 1 = 3

Analysis: There is no problem considered from the room environment. Entry to the room is controlled; the sanitisation cycle has been challenged with a level of microorganisms far greater than would ever be found in the environment (spores of Geobacillus stearothermophilus); all items entering the isolator are sanitised (using a chlorine dioxide based sporicidal disinfectant) and the isolator itself is an effective positive pressure barrier to the outside (at >15 pascals).

As detailed earlier, environmental monitoring is performed inside the isolator during testing. This monitoring, which has an action level of 1 cfu, is designed to detect any potential contamination inside the isolator environment.

Numerical approaches

A third component of the risk assessment approach is to evaluate the risk once an activity has taken place. By using a largely numerically driven set of tools then repeatability and reproducibility can be ensured. Examples of individual out of limits results and data sets relating to an operation are examined below, using examples from an aseptic filling process. Following this an example of an overall assessment of different processes over time are explored. Numerical approaches are useful in applying a level of consistency between one decision and another.


The use of risk assessment approaches is an important current GMP topics in microbiological environmental monitoring. This paper has outlined some possible tools for such a risk assessment approach. However, each suite of clean rooms or isolator will be subtly different. The microbiologist needs to consider each aspect of the environment and decide what level of monitoring best suits their system, and then to justify the techniques used and the locations selected. The approach adopted should be detailed in a written rationale and approved by senior management. After this a rigorous and defensible system will be in place to satisfy regulatory expectations, and to aid the user in risk assessing problematic environmental monitoring situations and results.

References and further reading

BS EN ISO 14698 – 1:2003: ‘Cleanrooms and associated controlled environments – Biocontamination control – Part 1: General principles and methods’

ISO 14644-1 Cleanrooms and Associated Controlled Environments – Classification of Air Cleanliness

ISO 14644-2 Cleanrooms and Associated Controlled Environments – Specifications for Testing and Monitoring to prove continued compliance with ISO 14644-1

PDA Technical Report No. 13 (revised): ‘Fundamentals of an Environmental Monitoring Programme’, September / October 2001

Reich, et al. (2003): ‘Developing a Viable Microbiological Environmental Monitoring Program for Nonsterile Pharmaceutical Operations’, Pharm. Technol., March, pp92- 100

Sandle, T. (2003a): ‘The use of a risk assessment in the pharmaceutical industry – the application of FMEA to a sterility testing isolator: a case study’, European Journal of Parenteral and Pharmaceutical Sciences, 2003; 8(2): 43-49

Sandle, T (2003b): ‘Selection and use of cleaning and disinfection agents in pharmaceutical manufacturing’ in Hodges, N and Hanlon, G. (2003): ‘Industrial Pharmaceutical Microbiology Standards and Controls’, Euromed Communications, England

Whyte, W. (2001): ‘Cleanroom Technology: Fundamentals of Design, Testing and Operation’

Whyte, W. and Eaton, T. (2004a): ‘Microbiological contamination models for use in risk assessment during pharmaceutical production’, European Journal of Parenteral and Pharmaceutical Sciences, Vol. 9, No.1, pp11-15

Whyte, W. and Eaton, T. (2004b): ‘Microbiological risk assessment in pharmaceutical cleanrooms’, European Journal of Parenteral and Pharmaceutical Sciences, Vol. 9, No.1, pp16-23

Whyte, W. and Eaton, T. (2004c): ‘Assessing microbial risk to patients from aseptically manufactured pharmaceuticals’, European Journal of Parenteral and Pharmaceutical Sciences, Vol. 9, No.3, pp71-79