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Volume 33, Issue 7
Safeguarding against microbial contamination requires rapid detection and innovative technology.
Guarding against microbial contamination remains a challenge for biomanufacturers, particularly in the purification steps of protein therapeutics. From the integrity of filtering processes to the aseptic fill/finish of the end product, each step must be optimized to prevent contamination as well as remove microbial contaminants.
Despite there being established validated cleaning processes, there still remains demand for higher purity and increased sterility assurance in biopharmaceutical production. This means biomanufacturers may be required to reassess their procedures and technologies as effective microbial contamination control in biomanufacturing is a critical requirement.
Safeguarding against microbial contamination faces inherent challenges in all stages of biomanufacturing because the manufacturing environment is conducive to microbial growth, consisting of three critical conditions-moisture, nutrients, and other favorable factors, such as temperature, pH, and osmolarity, points out Paul Lopolito, technical service senior manager at STERIS.
“All three exist within upstream and downstream biopharmaceutical manufacturing and present difficulties to safeguarding against microbes. Controls are required to limit the introduction of microbes from cell lines, raw materials, packaging, environment, equipment, utilities, workflow, and personnel,” Lopolito says.
However, a reduction in personal interaction with product through isolators and restricted access barrier systems, as well as more efficient cleaning and disinfection programs, significantly limits the introduction of extrinsic microbes Lopolito adds. “The key is to understand the microbial hazards throughout each process step and to continuously strive to reduce the highest risk items. The risks may be compounded as the equipment and facilities age,” he notes.
Analytical assays are a critical tool for detecting microbial contaminants as well as identifying them. Traditional methods, such as bioburden testing and viable testing using contact (RODAC) plates, settle plates, and swabbing, adequately demonstrate microbial control, says Beth Kroeger, technical service senior manager, STERIS. Newer microbial detection methodologies with rapid results, however, are becoming more reliable. In some instances, organoleptic means may detect contamination even before the quantitative results are available, Kroeger asserts.
An example of the benefits of innovative microbial detection technology is maintaining a visual examination of the cell culture during a passage, or in bioreactor monitoring, and not relying solely on automation for cell counts with instrumentation. “Operators should still be able to identify contamination well before a sample is identified as contaminated based on cell characteristics, culture conditions, and growth attributes, such as oxygen consumption and dissolved oxygen levels,” Kroeger explains.
Meanwhile, in addition to isolating the process or use of restricted access barrier systems (RABS), the process itself should be safeguarded. This can be accomplished by having robust systems and procedures in place for control of raw materials used in the process, materials entering the cleanroom, equipment design, and personnel training and management, notes Kroeger.
“A material review board should approve raw materials used in the process and establish suitable microbial specifications. Raw materials should be assessed during process validation using multiple lots and/or bioburden results if the specification is anything other than 0 colony-forming unit (cfu)/mL. If the process capability is not able to remediate a microbial challenge, it is better to find that out in cleaning or process validation and not during routine manufacturing,” Kroeger explains.
“An example of this is chromatography resin. If the specification on the certificate of analysis is < 100 cfu/mL and lots having 0 cfu/mL are tested, ensure a lot that challenges this specification is included in the cleaning validation or process validation to prove the process is robust enough to clear any microbial issues,” Kroeger adds.
Kroeger also recommends that items entering the cleanroom and, ultimately, the Grade A/B area should be double wrapped in a protective barrier and sterilized by autoclave, irradiation, or other similar means. In addition, as the items move from a controlled non-classified area to an aseptic processing area or Grade A/B area, a layer of protective wrap may be removed, or the wrapped item may be disinfected. Disinfection should include the use of a sporicidal agent that is sprayed on the items, or by wiping the outer wrap as an item moves from one area to a stricter classified area via material airlocks (MALs).
“Typically, it is a combination of removal of barrier wrapping and wiping to transfer the materials into the aseptic processing area. Large data sets such as pressure, humidity, temperature, and environmental monitoring should be trended with the use of more automated environmental monitoring techniques to aid in timely risk assessments regarding the process and the product,” Kroeger states.
“Equipment maintenance is also a crucial practice to ensure reliability. Routine maintenance activities, such as passivation and derouging, may be necessary to keep equipment in optimal condition and not contribute to microbial contamination. Advances in formulated alkaline detergents and acid detergents can also be effective in microbial and biofilm removal. In addition, integration of equipment from vendors within the aseptic area has advanced over the past 10 years, providing more options and effectual connections versus the sterile welds of the past,” Kroeger emphasizes.
“Finally, operators should be trained periodically in aseptic technique and on basic microbial control with an understanding of how contamination can enter the environment and the product,” Kroeger adds.
Innovations in analytical assays and bioprocessing technologies offer better capabilities for maintaining a sterile bioprocessing environment. “There has been progress in single-use disposable technology and development of continuous manufacturing systems, which reduce or eliminate operator interaction and cross-contamination between different products,” Lopolito says.
Additionally, there have also been developments in viable/non-viable particulate monitoring systems. “These alternative rapid microbial tests have been difficult to integrate within the environmental monitoring programs because the industry standards have been based on older methodologies, and direct correlation between new and traditional methods is difficult,” Lopolito elucidates.
Further, regulatory agencies and corporate initiatives encourage the use of rapid methods for faster decision making and to prevent loss of product. “Perhaps this rapid technology may be used to detect contamination and divert potentially contaminated media to waste rather than feed in a bioreactor, or to alert of a change in environmental conditions that would be unfavorable to processing,” Kroeger says.
In terms of future innovation, there is always a need for fast and reliable prescreening test methods for various microbial and chemical hazards. The control strategy often involves screening or evaluating all materials introduced, testing for the absence or quantity of microbes, and designing critical process, such as equipment cleaning or facility disinfection, to mitigate the risk. Thus, the need for fast and reliable prescreening is crucial.
“Effective use of prescreening tools may expedite results and reduce overall testing. Positive prescreening test results may then be confirmed using traditional test methods,” Lopolito concludes.
Vol. 33, No. 7
When referring to this article, please cite it as F. Mirasol, “Detecting Microbial Contamination in Bioprocessing,” BioPharm International 33 (7) 2020.