Environmental monitoring of pharmaceutical cleanrooms provides assurance that our production environment is in control, and in compliance. A variety of test methods are performed to monitor the number of total particles in the air, the number of viable (alive) particles in the air and the number of viable organisms on surfaces, including personnel working in the controlled rooms.
The number of particles in the air can be determined very quickly. Particle counters are used to collect a known quantity of air and then print out the breakdown of the particle sizes and the number of each size particle in that sample. Results print out within minutes. This allows for quick reactions, and allows areas with high particle counts to be shut down, investigated and their condition corrected during the production of a batch. In addition, drug compounds produced while areas are out of specification can be segregated.
Unfortunately, these results do not tell us when we have viable organisms in the environment. Methods for testing for viable organisms require that the organisms be collected, either with an air sampler or with swabs and/or contact plates. The sampling media is then incubated, typically for three to seven days, and then colonies are counted. This delay to results delays reaction to contamination issues and can make investigations very difficult. By that time, the rooms in question have typically been cleaned numerous times, so re-sampling results are almost always meaningless and determining the root cause of the contamination is difficult. Since real-time response is not possible, batches are jeopardized.
However, rapid microbiological methods may provide a solution. For some time, the industry has been interested in using RMM for sterility testing, to expedite product release. While a desirable goal, it comes with a great deal of additional regulatory scrutiny. Some companies have been successful in implementing RMM for release testing in the U.S., but acceptance of rapid methods for finished product testing is still not widespread and regulatory opinions on the topic vary. At the very least, a regulatory filing is needed to change from the traditional sterility test to an alternative method.
As rapid microbiological methods evolve, their use to perform environmental monitoring could provide a path of less resistance from a regulatory standpoint. Many of the technologies that have been developed for product testing may have significant merit for performing environmental monitoring (Table 1). It remains to be seen whether there will be a significant market for this kind of service (Box, bottom).
|bioMerieux||BacT/ALERT||Detects CO2 from growing organisms; used for qualitative product testing and analysis of swab samples|
|BioVigilant||IMD-A 200-1, IMD-A 220-4||Optical technology; instantaneous detection of particles and microorganisms in air samples|
|Celsis||RapiScreen, AMPiScreen||ATP bioluminescence; qualitative product testing and analysis of swab samples|
|Chemunex||Chem Scan RDI||Solid-phase cytometry; quantitative testing of water, filterable products|
|Lonza||microCompass||Real-time reverse transcriptase PCR assay; quantitative testing of water, filterable products, swab samples|
|Millipore||Milliflex Quantum||Fluorescent staining based system; water, filterable products, swab samples|
|Pall||Pallchek||ATP bioluminescence; quantitative of water, filterable product testing|
|Rapid Micro Biosystems||Growth Direct System||Automated non-destructive growth-based system; quantitative testing of water, filterable products, swab samples|
Case Study: Use of a Rapid Method to Analyze Swab Samples
A study was performed at the MicroWorks laboratory in Crown Point, Indiana, to determine if it might be feasible to utilize an alternative technology to analyze swab samples. Two different test methodologies were performed.
The purpose of surface sampling as part of an overall environmental monitoring program is to track the level of surface contamination in the facility to ensure that cleaning and sanitization is effective. Swabs are often used for sampling irregular or hard-to-reach surfaces and critical surfaces where contact plates are not practical. In addition, cleaning hold time studies are often performed using swabs. Sanitizers collected from surfaces can be neutralized and dilutions can be done when highly contaminated areas are sampled.
Surface samples are most frequently incubated at 30-35˚C for at least three days or 20-25˚C for five to seven days, or at some combination of the two temperatures.
For this study, we used the MicroWorks Swab Sampling System (MSSS 001). The swab system consists of a sterile, double-bagged, gamma-irradiated product containing a calcium alginate swab and a sodium citrate diluent. These sterile packages are designed to be taken into cleanrooms and/or isolator systems.
The instrument utilized for this study was the BacT/ALERT system. This automated microbial detection system is built on bioMerieux’s colorimetric technology that detects microorganism growth by tracking CO2 production. To utilize this system, sterile liquid media bottles are inoculated with the test sample and the bottle is loaded into the instrument. A variety of different media are available depending on the product being tested and the target organisms. If microorganisms are present in the test sample, carbon dioxide is produced as the microorganisms metabolize the substrates in the culture medium. With CO2 production, the color of the sensor in the bottom of each culture bottle changes from dark to light.