What Are Microbiology Tests And USP 1117 Guidelines?
What are regulatory microbiology tests? And why are microbiology tests necessary for your medical device or product?
Microbiology testing identifies the presence and the type of microbes in a manufacturing environment, medical product, or medical device. Microbiological testing is both a regulatory requirement and a risk mitigation strategy for aseptic manufacturing environments. Specific regulatory testing for microbiology may include microbial enumeration testing, particulate analysis, yeast analysis, antifungal activity assessment, growth promotion testing, microbial limits testing, zone of inhibition testing, water analysis, bacteriostasis/fungistasis testing, bacteria identification, yeast identification, fungi identification, gram-negative staining, product inoculations, biological indicator tests, and BI incubation time reduction studies. Many microbiology tests, particularly microbial enumeration testing, are similar to bioburden tests. You can find distinctions between microbiology and bioburden testing HERE.
As medical devices and products must be sterile to be used within a human body, microbiological testing is imperative for preventing product-induced infections at the time of use.
There are two methods used to create a sterile medical product. First, a final chemical or heat sterilization process is used on packaged medical products or devices to kill any live microorganisms picked up during the manufacturing process for terminal sterilization. The second method utilizes aseptic processing through the entire medical device or product manufacturing process. Aseptic processing requires the exclusion of microorganisms from the manufacturing methods. Thus, microorganisms must be prevented from entering open containers or product materials during aseptic processing. As a result, microbial tests are used to monitor clean rooms and facilities that manufacture products using aseptic processing.
How is equipment for microbiological testing controlled?
Most microbiology equipment (incubators, water baths, and autoclaves) is subject to standard validation practices of incoming qualification, operational qualification, and performance qualification. Additionally, periodic calibration (generally annually) is commonly required. New equipment is tested according to a protocol approved by the quality assurance unit (QAU). In addition, cleaning and sanitization of equipment such as incubators, refrigerators, and water baths should be performed regularly to minimize the potential for contamination in the laboratory. Door seals of incubators and refrigerators should be cleaned and checked for the state of repair. Instruments (such as pH meters and spectrophotometers) are calibrated regularly and tested to verify performance on a routine basis. Calibration frequency and performance verification vary based on the type of instrument and the importance of that equipment to the data generated in the laboratory. Equipment that is difficult to sanitize (such as refrigerators and incubators) should be dedicated to aseptic operations (such as storage of media for testing and incubating sterility test samples) and live culture operations to minimize the potential for accidental contamination of the tests. Autoclaves are central to microbial testing and must have proper validation in place to demonstrate adequate sterilization for a variety of operations. Autoclave resources must be available (and validated) to sterilize waste media (if performed in that laboratory) as well as the media prepared in that laboratory. The choice of one or several autoclaves is not driven by a need to separate aseptic and live operations (everything in the properly maintained autoclave is sterile after the cycle) but rather driven by resource considerations (see below).
Is Laboratory layout and operation Important for microbiology testing?
The layout of the laboratory or contract testing facility has an impact on microbiology testing results. Laboratory layout and design must ensure that cross-contamination of microbial cultures is minimized. Additionally, microbial samples must be handled in an environment that makes contamination unlikely (biological safety cabinets). The laboratory is divided into “clean” aseptic areas and “dirty” microbial culture areas for microbiological testing. This reduces the risk of false-positive test results. Areas, where sterile product samples are handled and incubated should be completely free of live cultures. If complete separation of “dirty” and “clean” culture zones cannot be accomplished, aseptic practices get to be employed to reduce accidental contamination risk. Aseptic practices include protective clothing barriers, sanitization and disinfection procedures, and biological safety cabinets designated for only aseptic operations. Some sterile samples will demonstrate microbial growth and require further analysis of the contaminants. If microbial growth is detected, the sample is swiftly taken to the “dirty” live culture section of the laboratory or designated biological safety cabinet. All subculturing, staining, microbial identification, or other microbial investigational operations are completed in the live culture section of the laboratory or “dirty” biological safety cabinets. Note that technicians should not open any sample with clear microbial colony growth in a “clean” zone. Keeping staff working in “clean” zones separate from staff working in “dirty” zones minimizes the risk of cross-contamination between laboratory zones.
Microbial contamination of samples is always possible unless careful aseptic precautions are taken for staff, samples, and raw materials. Indeed, any samples not taken aseptically are unreliable and compromise the microbiological results. Thus, any environmental sampling methods require aseptic handling in the loading and unloading of sampling instruments. As an alternative to cleanrooms, isolators may be used for sterile microbiological testing. Isolators have lower levels of environmental contamination than operated clean rooms. However, isolators can have false-negative results due to the chemical disinfection of materials brought into or using within the isolator.
How are laboratory technicians trained and contamination controlled for microbiology testing?
All staff, supervisors, and managers must have a core education background in microbiology or a closely related biological science. These individuals will be assigned responsibilities within their microbiological skill and experience level. Additionally, microbiologists will complete experiments using standard operating procedures (SOPs). SOPs describe the methodology that the microbiologist will follow to ensure accurate and reproducible results. Microbiologists will be given SOPs that fit within their competency level. Specific course work, relevant experience, continuing education, and accredited certifications all support establishing laboratory competency. Regular training is encouraged for all laboratory jobs.
Periodic performance assessments ensure technician consistency with SOPs. Performance tests are designed to evaluate key microbiology laboratory competencies. Plating, aseptic technique, and data documentation may be competencies a performance test evaluates. Managerial microbiologists will have training in additional areas such as leadership, laboratory safety, technical report writing, and interpretation of regulatory guidelines. Note that laboratory supervisors and managers should have a level of competence in microbiology at least as high as those they supervise.
Aseptic Sample Handling
Live microorganisms in microbiology samples are sensitive to the handling and storage conditions they receive. Handling and storage conditions that should be monitored include sample source, container composition, storage time, and storage temperature. Best results come from minimizing the time between sample collection and sample assessment. If the samples must be transported a distance for testing, ensure that transport and storage conditions are suitable for that test and sample. All microbiological specimens should be taken using aseptic techniques to avoid inaccurate results. Microbiology samples should have information on the specimen source, the date the sample was taken, the date of specimen submission, the person or department submitting the sample, and any potentially hazardous materials associated with the specimen. When samples are received, the testing facility should acknowledge the samples’ receipt, number, and identity. If microbiological testing and sampling occur in the same facility, staff assigned to sampling activities, particularly those in support of aseptic processing, should avoid working in areas with live microbial cultures. This prevents cross-contamination between aseptic “clean” and microbial “dirty” areas of the laboratory.
Summary
Overall, laboratory equipment and technician control for microbial testing are vital to accurate results. The testing laboratory’s layout, equipment qualification, and regular laboratory technician training all ensure that microbiological testing is completed time and time accurately again. Whether you choose to do microbiology testing in-house or to outsource your regulatory testing, make sure you establish procedures that provide for the control of equipment and laboratory technicians performing microbial assays. Should you decide to outsource your microbiological or aseptic environmental monitoring testing for your medical device or medical product, choose a contract testing organization that can support your needs.
Ethide Labs is a contract testing organization that specializes in Microbiology Testing and Environmental Monitoring. Ethide Labs provides in-vitro cytotoxicity tests in-house and outsources in-vivo cytotoxicity work for toxicity testing of medical devices, products, and drugs. Ethide Labs also offers Sterilization Validations, Bioburden Testing, Bacterial Endotoxin Testing, EO Residual Testing, Package Integrity Testing & Cytotoxicity Testing services for medical device companies and allied industries. Ethide is an ISO 13485 certified facility.
References
United States Pharmacopeial Convention. <1117> Microbiological Best Laboratory Practices. Rockville, MD, USA. 2021. (USPC <1117>).
