Op Ex & Lean Six Sigma

Framework for Fast Microbiological Assessment

Encyclopedia of Rapid Microbiological Methods, Volume 4: The Next Generation of Rapid Method Applications, Validation Strategies and Regulatory Acceptance

By Michael J. Miller, Ph.D.

Editor’s note: In January 2013, Volume 4 of the successful PDA-DHI technical book series, Encyclopedia of Rapid Microbiological Methods, was released. In this review, long-time proponent of rapid methods and editor of the encyclopedia, Dr. Michael J. Miller, provides an expert synopsis of the book’s relevant and timely content.

Volume 4 of the Encyclopedia of Rapid Microbiological Methods engages readers with an excellent introduction by Dr. Bryan Riley, New Drug Microbiology Staff at the Food and Drug Administration’s (FDA’s) Center for Drug Evaluation and Research, and the Agency’s expert on rapid method technologies. Dr. Riley explains that modern approaches to process control — including Process Analytical Technology (PAT) — require the availability of results in real time (or at least close to real time) to enable the operator to immediately use the test results and intervene if necessary to make process-optimizing decisions and adjustments. Although real-time results are only currently available for a limited category of microbiological tests, there are many microbiological methods that are significantly more rapid than the traditional test methods. 

Dr. Riley continues, offering that the rapid methods available today vary a great deal in their mechanisms of operation. Some of these methods, he notes, still rely on a period of microbial growth using traditional media, but reduce their time to result by using an alternate method of microbial detection. Other rapid methods do away with growth entirely and utilize a stain or inherent microbial auto fluorescence to detect microorganisms; even down to the level of a single microbial cell. Furthermore, some of the available methods are quantitative, some are qualitative, and vary in their time to result (from real-time to several days) but all of these methods seem to have found a niche in the pharmaceutical microbiologist’s arsenal.
Current rapid microbiological test methods are now able to start providing some of the advantages long enjoyed by industry colleagues in clinical and food microbiology labs. Pharmaceutical microbiologists would be well served by considering which of their samples would provide a benefit with a more rapid result, then assessing the current alternate microbiological methods to see if any of them fit their needs.

Featuring up-to-the-minute advances and details regarding quality control and choosing appropriate methods, Volume 4 covers future-use scenarios, current and emerging technologies, mass spectrometry, genotypic methods for identification, new case studies and application of USP and other guidelines. The book offers valuable assessments and reviews of environmental monitoring, validation, sterility testing, Mycoplasma testing, and the application of rapid microbiological methods as they relate to both bio-processing, product-specific method advances and regulatory considerations.
For instance, guidance on the application of modern microbial methods is covered in Chapter 1, which discusses the Quality Control testing of probiotics, including master and working cell banks, release and stability testing, and viable cell counts. It also details identification and strain typing, absence of bacterial pathogens, antibiotic resistance, adherence to the intestinal wall and acid and bile resistance.

Aligning RMM to suit

Considerations when aligning a rapid microbiological method to suit an end-user’s particular needs is reviewed in Chapter 2. Topics range from the drivers for rapid methods, time savings, same-day results, sample compatibility and automation, to using a qualitative method as a screening tool, validation, identification, integration with LIMS and other data management platforms, as well as false positives, false negatives and detection limits.
In Chapter 3, a close look is taken at the future of rapid and automated microbial identification systems. Here the Encyclopedia provides an overview of technologies based on the growth of microorganisms, the detection of cellular components, optical spectroscopy, nucleic acid amplification and MEMS. Included are examples outlining the utilization of biochemical and carbohydrate substrates, fatty acid analysis, MALDI-TOF and SELDI-TOF mass spectrometry, FT-IR, elastic and inelastic light scattering, ribotyping, PCR, microfluidics and microarrays.

A more detailed case study in Chapter 4 covers the use of MALDI-TOF mass spectrometry for the identification of microorganisms. Discussions on sample preparation, OQ, PQ, accuracy, precision, robustness and computer validation are also covered. In Chapter 5, another case study on microbial identification focuses on genotypic methods, amplification of DNA, automation and validation (accuracy, precision, robustness and specificity). A discussion of the new method’s ability to comply with GMP principles is covered as well.

A review of workflow/applications for a rapid growth-based microbial identification system are reviewed by its supplier in Chapter 6. The chapter’s contributor also provides validation considerations, as well as an overview of enhancements to the supplier’s existing technology.

New growth-based method

A case study of a new growth-based rapid microbiological method that detects the presence of specific organisms and provides an estimation of viable cell count is provided in Chapter 7. The chapter thoroughly covers sample preparation, assay workflow, and the method’s applicability to a wide-range of microorganisms. Additionally, data from a validation case study, inclusivity and exclusivity testing, and a comparison to USP <61> for aerobic counts, yeast and mold, and Gram-negative bile tolerant microorganisms also provide readers valuable insight into the viability and efficacy of this new method.

An evaluation of a relatively new growth-based rapid method that utilizes a membrane filtration workflow coupled with a viability staining technique is featured in a revealing end-user case study in Chapter 8. The chapter’s author provides a review of the technology and evaluates the method’s results. This is followed by a discussion of the system’s applicability when it comes to monitoring mammalian cell cultures, as well as additional benefits the method can offer to biopharmaceutical processors.

In Chapter 9, an optical spectroscopy rapid method supplier details the technical and applicational aspects of its environmental monitoring system and how their technology can help users comply with validation recommendations of USP <1223>. Accuracy, precision, limit of detection/quantification, linearity, range and robustness are some of the parameters that were examined.

Chapters 10 and 11 deliver a comprehensive evaluation of an optical spectroscopy technology for the real-time and continuous monitoring of airborne microorganisms in cleanroom and isolator environments. The narrative, supported with extensive photography, exposes readers to an in-depth study of real-time monitoring during an aseptic fill. Among other things, these chapters detail the transfer of sterilized components, interventions and the reactions to and outcomes of an isolator glove breach.

A rapid method for the release testing of both sterile (sterility testing) and non-sterile products (bioburden assessments) is found in Chapter 12. Here, an ATP bioluminescence technology is validated, and the authors discuss their qualification workflow and results. An end-user’s validation approach for a rapid, growth-based detection system and its application as an alternate sterility test for cellular immunotherapy products is the focus of Chapter 13. Challenges associated with the conventional method are outlined, a discussion followed by user’s approach to feasibility testing, method validation, and navigating the regulatory path for commercial approval.

Chapter 14 contains an informative case study by an end-user of a rapid, solid-phase cytometry technology. The author describes his company’s validation strategy, including accuracy, precision, limit of detection, robustness, ruggedness and the use of statistical models when comparing the results to the acceptance criteria. Additional considerations include the use of stressed cells and matrix effects on the overall validation plan.
An ATP bioluminescent system supplier provides an alternative to the conventional sterility test in Chapter 15. Highlights include validation strategies, the use of challenge microorganisms, system suitability, understanding background values, how to conduct product specific feasibility testing and much more. Chapter 16 focuses on the statistics of validating an alternative sterility test. Subjects include probabilities and multiplicity, limit of detection and a comparison of what is statistically different versus what is statistically equivalent. This is a must read for anyone wanting to validate rapid sterility tests and how to design the studies and use statistics to justify the results.

An overview of a validation approach for a next-generation ATP monitoring technology is covered by Chapter 17. This chapter’s contributor describes the principles associated with the new method and how to evaluate limit of detection and validation strategies for a wide-range of fluid samples. Chapter 18 provides readers with a novel qPCR-based system for the detection of specific microorganisms. An overview of the technology is provided by its supplier and includes a preliminary study on validation parameters, specificity, limit of detection and data analysis.

Detecting Mycoplasma is an important element of downstream biopharmaceutical processing and Chapter 19 addresses the use of rapid methods within this context. These contributors provide an overview of Mycoplasma and the importance of its timely and accurate detection. Additionally, the authors describe traditional and alternative detection methods, and include a case study outlining the use of a nucleic acid amplification platform. A review of regulatory author requirements for nucleic acid amplification systems is also discussed.

A new nucleic acid amplification and microarray-based rapid method for the detection of Mycoplasma is highlighted in Chapter 20. Here, the contributors detail technology workflow and make the case why this method has advantages over traditional methods. Mycoplasma detection regulatory guidelines are also examined which will help those less familiar with the compliance issues associated with this important process element.

Chapter 21 provides an overview of rapid viral detection methods. This chapter’s authors debate classic versus molecular biology approaches and discuss the prospects for viral safety testing. Their experiences with Vesivirus, MVM and other public viral incidents are covered, as well as other topics associated with the future direction of molecular-based detection methods.

Speaking of future directions, Chapter 22 investigates a promising new microbiology technology wave: alternative/ rapid methods for the QC laboratory. Regulations, required skill sets for pharma microbiologists and the microbiological curricula will provide readers with what they need to know about this innovative technological advance.

Finally, Chapter 23 goes into great depth discussing the real-world application of rapid microbiological methods for bioprocessing. Similarly biopharmaceutical manufacturing, regulations are reviewed; as are testing requirements and contamination events which set the stage for evaluating a wide range of rapid method applications.

These are very exciting times for the rapid detection, quantification and characterization of microorganisms. The information presented in Volume 4 of the Encyclopedia of Rapid Microbiological Methods provides the industry with a comprehensive collection of technology reviews and validation strategies designed to encourage today’s microbiologists to move away from centuries-old techniques and to embrace the next generation of novel, more-sensitive and rapid microbial detection platforms. The Encyclopedia will provide a framework for all industry, clinical and government sectors required to evaluate the environment, products, processes and test samples for the presence of microorganisms to embrace the rapid methods that are available today, and what will be coming in the years ahead.

To order Volume 4, contact PDA at https://store.pda.org/ProductCatalog/Product.aspx?ID=1899. For volumes 1-3 go to https://store.pda.org/ProductCatalog/Product.aspx?ID=513.”

About the Author

Dr. Michael J. Miller is an internationally recognized microbiologist and subject matter expert in pharmaceutical microbiology and the design, validation and implementation of rapid microbiological methods. Author of more than 100 technical publications and presentations in the areas of rapid microbiological methods, PAT, ophthalmics, disinfection and sterilization, Dr. Miller is the editor of PDA’s Encyclopedia of Rapid Microbiological Methods; is president of Microbiology Consultants, LLC (http://microbiologyconsultants.com); and maintains http://rapidmicromethods.com, a website dedicated to the advancement of rapid methods. In his current role, Dr. Miller consults with multinational companies in providing technical, quality and regulatory solutions in support of RMMs, sterile and non-sterile pharmaceutical manufacturing, contamination control, isolator technology, validation and microbiological PAT.

Published in the March 2013 issue of Pharmaceutical Manufacturing magazine

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