Environmental monitoring (EM) is a tool utilized by the pharmaceutical industry to measure and monitor microbial bioburden levels in a manufacturing facility. Periodic sampling of the air and surfaces within controlled environments allows the microbiologist to determine the approximate number and types of microorganisms present in a given area over a defined period of time. This data is analyzed through a process known as trending. Trending is performed in order to detect fluctuations in microbial levels, and allow firms to ensure that these levels remain within acceptable parameters. “Traditional” methods of trending EM data often include the use of excursion rates. “Excursion rate”, for the purpose of this article, is defined as the percentage of the total number of collected EM samples with counts exceeding either the alert or action level.
Alert levels are defined as an established level of microorganisms that gives early warning of a potential drift from normal operating conditions.1 Action levels are defined as an established level of microorganisms that, when exceeded, indicate that a process is outside of its normal operating range.1 Exceeding an alert level does not necessarily warrant immediate investigation and corrective action, although some sort of follow-up action is generally required in order to determine the cause of the exceeded level, its impact, and to ensure that the affected area has returned to its normal operating conditions. Exceeding an action level usually requires an immediate investigation and subsequent corrective or preventative action (CAPA).
Trending by excursion rate alone is limited in terms of the amount and type of information it provides. Excursions result from a single sampling event. They represent only the samples that exceeded a pre-defined microbial count at a specific location at a specific point in time. Similarly, excursion rates do not take into account any results obtained from other samples collected in the same area during that particular sampling event. In short, trending by excursion rate alone only demonstrates how many individual samples collected over a selected period of time exceeded a pre-determined microbial count.
While this information is helpful in terms of identifying areas where elevated microbial levels may exist or a potential drift from normal operating levels, it does not provide much information about the overall state of microbial control across the facility environment. Microbial populations are dynamic. Levels change over time due to factors such as seasonal variation, changes in staffing or the presence of contractors, new equipment, etc. In any given EM sample set, there are usually microbial recoveries present on individual samples that did not exceed the established alert or action limits. The number of these will vary with each sample set. This information is important for maintaining effective microbial control, but it is not visible when trending solely by excursion rates.
In contrast to excursion rates, the contamination rate takes into consideration the total number of recoveries across all samples collected in a defined area as opposed to the number of colony forming units (CFU) on a single sample. It is qualitative as opposed to quantitative; the number of CFU recovered on individual samples is irrelevant. Utilizing contamination rates in trending allows the microbiologist to see fluctuations in the overall microbial levels across the entire sampled area. This provides a more accurate reflection of the environment as it actually exists.
USP General Chapter <1116> currently recommends the use of contamination rates for EM trending in aseptic processing environments.2 However, this technique can easily and effectively be applied to any controlled environment used for any operation, from Grade A to Controlled Not Classified (CNC). The utilization of contamination rates in conjunction with excursion rates increases the amount and quality of the information generated. The impacts and effects of sub-alert trends, seasonal variations and aberrant events which were largely unseen when trending using excursion rates alone, are suddenly highly visible. The power and granularity of the existing trending program is increased, allowing it to become the truly useful and informative tool it should be.
Why Is the Use of Contamination Rates Outside of the Clean Room Environment So Useful?
In a Grade A aseptic processing environment where the action limit for microbial counts is usually 1 CFU,2,3 excursion rates and contamination rates will be virtually identical. However, as classification levels decrease (e.g., Grade B through CNC), the value of adding contamination rates to the trending program quickly becomes evident. As the area classification level decreases, the allowable level of microbial bioburden increases. The established alert and action levels also increase. This means that, while it is expected that there will be higher levels of microbial contamination present, an individual plate counts must be higher in order to trigger an excursion. The fluctuations occurring in microbial counts that do not exceed any established excursion level, also known as subalert trends, are equally or more important in terms of maintaining consistent and effective overall contamination control. The utilization of contamination rates raises the visibility of these sub-alert trends and other aberrations that are generally unseen when performing “traditional” trend analysis alone.
EXAMPLE 1: Which Operator Presents the Higher Risk?
Consider the following scenario. Two aseptic processing operators perform the same job function in a Grade C environment. Operator A works on the first shift, and Operator B the second. Both don full aseptic gowning before performing their tasks, and both operators are plated at the end of their shift prior to de-gowning. A total of six (6) contact plate samples are collected from each operator at the end of each shift (e.g.; both gloves, both sleeves, top of the suit zipper, and face mask). An action level of >5 CFU/plate is established for the individual sampling sites.
Over a period of 6 months, assuming a 5-day work week, both operators would have generated a total of 780 gown samples each (26 weeks x 5 days x 6 samples). Operator A has exceeded the action limit a total of eight (8) times. Using the following formula to calculate the excursion rate,
(Number of Excursions/Total Number of Samples Collected) x 100
Operator A demonstrates an excursion rate of 1.03%. Operator B has recorded no excursions at all, yielding an excursion rate of 0.00%. Which operator has the greater likelihood to transmit microbial contamination from their gowning to the product?
Based on the excursion rate data alone, Operator A would be the most likely as Operator B recorded no excursions. However, an analysis of the contamination rate shows that Operator A had 52 plates out of the total 780 with a microbial count of >0 CFU, including the 5 plates that exceeded the action limit of >5 CFU/plate. Using the following formula to calculate the contamination rate,
(Number of Samples with Counts 0 CFU)/Total Number of Samples Collected) x 100
Operator A demonstrates a contamination rate of 6.67%. Operator B, who had an excursion rate of 0.00%, had 122 plates out of the total 780 with a microbial count of >0, but none of those 122 plates had greater than 5 CFU/plate. This results in a contamination rate of 15.64% for Operator B. The contamination rates reveal that it is Operator B that poses the greater transmission risk because microbial contamination is present more often on Operator B’s gowning materials even though Operator B exceeded no established alert or action limits.
The information provided by this analysis could potentially lead to a decision to retrain Operator B in gowning and aseptic technique, or to re-evaluate the duties this operator performs. It would also provide critical data to be utilized as part of a root cause analysis, for example, in the event of a sterility failure of the aseptically processed product. This is an example of how contamination rates can be used to analyze EM data from a different perspective and with more granularity.
Ability to Apply Mitigations and Corrective Actions More Effectively and Efficiently
The following example demonstrates how the use of contamination rates can impact a root cause analysis investigation, resulting in more efficient and effective CAPA.
EXAMPLE 2: A Single Exceeded Action Limit in a Processing Room
An action-level excursion has occurred on a viable air sample in a processing room. Adjacent to this processing room are an equipment airlock and a second processing room. All three rooms open into the same common controlled corridor. No excursions have occurred in either adjacent room or in the corridor. An investigation is initiated, and as an immediate corrective action, the decision is made to perform an additional sanitization in the processing room where the excursion occurred, but what about the two adjacent rooms?
Because the traditional system of trending by excursion rate alone only considers counts obtained on a single sample, comparing the level of microbial bioburden in the two adjacent rooms and the connecting corridor is not a simple endeavor. The following information is not immediately available:
- In the affected room, was the sample that exceeded the action limit the only sample from that room that demonstrated microbial recoveries? If not, did the other collected samples indicate that microbial contamination is present throughout the room as opposed to being restricted to the area of the affected sample?
- What about the levels in the other two rooms and the corridor? Although no excursions occurred, were the number of recoveries higher in these rooms also? It is quite possible that the overall microbial bioburden in those three areas is elevated, but the number of CFUs on the collected samples have simply not exceeded the alert or action limit. If so, the overall microbial bioburden present is higher, and the risk that another excursion will occur increases.
If contamination rates are utilized as part of the trending program, this information will be readily available to the investigator. These details are critical to accurately determining the root cause of the excursion, as well as the appropriate CAPA.
More Accurate and Definitive Root Cause Analysis and CAPA
Where did the contamination originate in the scenario presented in Example #2? If the overall levels are normal in the affected room, and the excursion occurred on a single sample, this could indicate an isolated event. However, if other levels are elevated throughout the room, is it possible, for example, that a sanitization was missed, or a HEPA filter is malfunctioning in the room?
If the levels are also elevated in the adjacent corridor, it becomes increasingly likely that direct or indirect transfer into the room occurred either on an operator or on a piece of equipment, such as a cart. If the levels are elevated in the corridor and both adjacent rooms, it is still possible that the contamination was transferred into the room, but the likelihood also increases that there may be a larger systemic problem (e.g., an issue with the HVAC system, sanitization or gowning program, or procedures not being properly followed).
In this manner, the analysis of contamination rates can often determine the focus of the investigation, and provide empirical data to support the conclusions. The accuracy of the root cause identification impacts the efficacy of the applied CAPA. For each of the possible root causes just described, the required CAPA would be very different.
More Efficient and Cost Effective Application of CAPA
Because microbial contamination can be transferred from one area to another by both direct and indirect means, it is important to consider these factors when determining corrective actions. When microbial contamination levels increase, the likelihood of transmission to other areas also increases.
Knowing where microbial contamination levels are elevated aids in determining the type and scope of the corrective action to be applied. If elevated levels are restricted only to the room where an excursion occurred, it may be necessary to perform the additional sanitization in that room only. If levels are elevated overall, it may be prudent to sanitize the adjacent areas as well (either as a corrective or preventative action) even though no excursions may have occurred in these areas. The contamination rates can then be monitored to determine if additional sanitizations are required, or if the microbial levels have returned to an acceptable state of control. The end result is more effective overall contamination control. The quality and integrity of the product is protected while material and labor costs associated with performing sanitizations in areas where none is required are eliminated.
Greater Ability to Proactively Mitigate
When trending solely by excursion rates, action is triggered by a specific event, namely an exceeded limit. Therefore, any resulting action is decidedly reactive as opposed to proactive. Excursions are frequently not detected until several days after sample collection. As previously mentioned, trending solely by excursion rates does not provide specific information regarding the possibility that overall microbial levels are increasing in other areas of the facility. A sudden increase in excursion rate is frequently a direct result of an increase in the overall microbial contamination rate that has been occurring for some time.
If contamination rates are continuously assessed over time, changes to the state of microbial control are more easily detected. Increases in populations can be detected earlier as patterns and sub-alert trends become visible. As a result, proactive mitigations can be applied before an established alert or action level is exceeded. This is demonstrated in the following example.
EXAMPLE 3: A Summer Heatwave
A biological drug substance facility is operating during a summer heatwave. Operators have been sweating heavily before they enter the building. Once inside, they go to the locker rooms, don their gowning materials and begin their work. The firm has been asked by the local power company to raise the facility room temperatures to the extent possible in order to relieve stress on the power grid. As a result, operators are reporting that they are uncomfortable and sweating in their gowning materials. This is found to be a particular issue in the downstream processing rooms, where aseptic connections are made. Sanitization procedures and schedules have not been altered.
The facility trending program utilizes both excursion and contamination rates. A review of the recent environmental monitoring data shows that while only one alert level excursion has occurred in the area since the heatwave began, the overall contamination rates have begun to rise, and much more rapidly than usual. Bodily shedding generally increases when sweating occurs. The moisture on the operators’ skin causes the gowning materials to adhere, creating more friction as the operators move and thus compounding the problem. Therefore, it is suspected that the increase in bodily shedding is responsible for the increase in contamination rate. This is quickly verified by the microbiology laboratory as an examination of the microbial recoveries shows that most are Gram-positive cocci, which represent many of the most common human skin floras.
Although no excursions other than the single alert-level excursion has occurred, based on the information provided by the contamination rate, the firm decides proactively mitigate the situation. Possible actions might include increasing the sanitization frequency, or arranging a meeting to advise all personnel with access to the production areas that the risk of a batch contamination has increased as there is a higher level of microbial contamination present in the area as a result of the heatwave. The operators can then increase their diligence in their gowning, general hygiene and other contamination control procedures.
This ability to proactively mitigate alone provides a number of important benefits.
Better protection of the process, product and patient from microbial contamination
Proactive mitigation provides better overall protection of the process, product, and ultimately the patient. The ability to discover and proactively mitigate rising microbial bioburden levels before process tolerances or environmental specifications are exceeded ensures a more steady state of microbial control in the processing environment, and that that microbial bioburden remains at acceptably low levels.
Fewer EM excursions result in fewer EM investigations and less required CAPA
EM excursion investigations are notoriously difficult and laborious to perform. This is compounded by the fact that most exceeded alert or action levels are not detected until at least two to three days (at minimum) after the actual sampling event has occurred. In a highvolume manufacturing facility, this usually means that more than one batch may be adversely impacted by a single excursion, substantially expanding the scope of the investigation. If an EM excursion was successfully averted due to the proactive measures, resources dedicated to performing the ensuing investigation can be utilized elsewhere. No additional CAPA is required, eliminating any costs associated with that remedial action and verification of efficacy.
Increased assurance of product quality and patient safety
All EM investigations included in a product batch record must be closed before the batch can be released to the market. Depending on the criticality of the EM excursion and its potential impact, failure to adequately identify the root cause and batch impact can result in batch rejection. Batch rejection is extremely costly in terms of materials and labor, and negatively impacts the production schedule. There is also the possibility that batches already released to the market could be affected, resulting in complaints, adverse events and/or product recalls. If the ability to proactively mitigate ensures that all processing activities take place within the established EM limits, the level of product quality assurance and patient safety is increased by lowering risks due to microbial contamination.
Increased assurance of uninterrupted supply to the market
Most importantly, proactive mitigations facilitate successful production and release of product. Batch rejection or delayed release can adversely impact supply to the market, which then negatively impacts patients who rely on that product. In some cases, this can quickly become a matter of life and death.
Other Important and Tangible Benefits
The ability to analyze the EM data through a more granular and robust trending program improves the ability to evaluate the efficacy of other microbiological control programs. For example, opportunities for improvement in the facility gowning and sanitization programs become evident. Using contamination rates in trending can reveal the individual rooms or monitoring sites that demonstrate a greater range of fluctuation, or consistently higher rates of contamination overall. This provides empirical data that may be used to justify increased or decreased monitoring and sanitization frequencies, or monitoring locations. Similarly, there is also increased ability to identify and address specific operator performance issues, and use this information to enhance the training program.
Summary
Combining the use of contamination rates with the existing “traditional” method of trending by excursion rate can substantially increase the power and usefulness of any facility EM trending program. Greater and earlier visibility of changes to the existing state of control allows for proactive mitigation. Proactive mitigation, in turn, leads to better and more consistent overall microbial control.
The ability to proactively mitigate can also result in fewer EM excursions, which leads to a reduction in the number of EM investigations. This results in time and resource savings that can be applied to other areas of the business. When investigations are required, the increased visibility of parameters such as sub-alert trends and the relative distribution of microbial populations helps to improve the accuracy of root cause analysis. More accurate root cause analysis allows for more effective and efficient CAPA to be implemented. This results in additional time, resource and material cost savings.
The increase in the amount and quality of the information generated by the enhanced EM trending program results in better decision making regarding contamination control measures. Better overall contamination control leads to higher levels of product quality assurance and patient safety. It also helps to ensure consistent supply to the market.
References
- Parenteral Drug Association (PDA), PDA Technical Report#13 (Revised), Fundamentals of an Environmental Monitoring Program, 2001
- United States Pharmacopoeia, USP General Chapter 1116, “Microbiological Control and Monitoring of Aseptic Processing Environments” 2017
- EudraLex—EU GMP Volume 4 “Good Manufacturing Practice (GMP) Guidelines-Annex 1: Manufacture of Sterile Medicinal Products”, 2009