While having knowledge of the most frequently identified bacteria is informative, it is important to note that these Top 15 bacteria identified from manufacturing environments, as cited in the original post, only represent approximately 50% of the microbes that are
identified during routine monitoring and investigations . These microorganisms
represent the diversity of bacteria from across the Pharmaceutical, Medical
Device, Personal Care and Cosmetics, Neutraceutical and Biotechnology
production facilities; the analysis did not include the fungi and yeast that
are also isolated from these environments.  As determined by Accugenix, the remaining 50% of the bacteria comprise 1,964 unique species which were identified in Environmental
Monitoring programs — a number that implies that following the environmental
ecology by methods such as colony morphology and Gram staining, as suggested,
would be daunting.

Pharmaceutical environmental monitoring programs have resulted in the
identification of 1,979 unique bacterial species by Accugenix over the last
five years. Indicating how critical it is that the identification libraries or
databases, against which you compare your data whether generated by phenotypic,
prototypic or genotypic methods, contain all relevant species or the
interpretation of your data is not reliable.  It is also critical to realize that this list
of the Top 15 frequently encountered bacteria is a reflection across all
industries and manufacturing sites.  Specific
manufacturing facilities may have different distribution of frequently
occurring microorganisms, depending on their environment.

According to the FDA Guidance for Industry, “Sterile Drugs Products
Produced by Aseptic Processing — Current Good Manufacturing Practices”,
September 2004, “The goal of microbiological monitoring is to reproducibly
detect microorganisms for purposes of monitoring the state of environmental control.
Consistent methods will yield a database that allows for sound data comparisons and interpretations.”   Thus, it is recognized that not only is the
library database of great importance, but also the methods used for
identification are essential to accurate and reproducible identification.

It is known that DNA Sequencing rapidly provides data for identification that is
substantially more accurate, robust and reproducible than relying solely on
visual phenotypic characteristics since the sequence-based result is not
dependent on growth conditions or ancillary testing.  This is so well understood and accepted that the FDA recommended the use of genotypic methods in their Aseptic Processing 2004 update to the guidance document. The FDA states, “Genotypic methods have been shown to be more accurate and precise than traditional biochemical and
phenotypic techniques. These methods are especially valuable for investigations
into failures… Sterility test isolates should be identified to the species
level.  Microbiological monitoring data should be reviewed to determine if the organism is also found in the laboratory and production environments, personnel, or product bio-burden.  Advanced identification methods (e.g., nucleic-acid based) are valuable for investigational purposes.  When comparing results from environmental
monitoring and sterility positives, both identifications should be performed
using the same methodology.”

In his response posted May 30, Dr. Miller re-iterated these points, “Not one
identification technology will give you 100% accurate identifications each
time…(i.e., you may not get a match if the organism is not in the system’s
database/library). That being said, genotypic methods, such as gene sequencing,
have been shown to provide more accurate results than phenotypic or biochemical
methods… Newer technologies, such as MALDI-TOF mass spectrometry also provide
very accurate results, as long as there is a match in the database…” These
statements are quite accurate.  By combining accurate, refined sequencing or mass spectrometry technologies with extensive, curated databases, the frequency of accurate identification is greatly enhanced.

About Barry A. Friedman, PhD LLC

Regulatory Compliance, Expert Witness, Pharmaceuticals, Biotechnology, Aseptic Processing, GMP, GLP, Quality Control, Positron Emission Tomography (PET), Microbiology, Process Validation, Validation of Analytical and Microbiological Methods
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