The scope of mycoplasma contamination within the biopharmaceutical industry

doi:10.1016/j.biologicals.2010.03.002

Biologicals

Volume 38, Issue 2, March 2010, Pages 211-213

https://doi.org/10.1016/j.biologicals.2010.03.002 Get rights and content

Abstract

Mycoplasma is well recognized as one of the most prevalent and serious microbial contaminants encountered within the manufacturing of biopharmaceuticals from the research phase to clinical development and production. The potential for mycoplasma contamination within cell culture systems was first identified by Robinson et al. in 1956 [1]. Presently, contamination rates in established cell cultures have been reported between 15 and 35% with considerably higher occurrence cited in certain selected populations [2]. In the last few years, there has been an expansion of diagnostic approaches for mycoplasma detection with the development and validation of rapid microbiological methods. The objective of this study was to determine current levels of mycoplasma infection of cell cultures, cell substrates and biologicals within a client based population. Retrospective comparison of 40,000 sample results was done to determine total contaminations rates amongst four (4) individual analytical assays. The establishment of reference data, such as existing contamination rates, becomes important in the critical appraisal of rapid microbiological methods for the detection of mycoplasma.

Introduction

The emergence of rapid methods for detecting mycoplasma provides the opportunity to reduce testing times of both cell culture systems and their derived biopharmaceutical products to a matter of hours compared to the minimum standard time of 28 days with the current regulatory approved direct culture based methods [3], [4], [5], [6]. Ideally, these emerging testing methods will provide the necessary real time results for those human cell-based therapies with short half lives. In addition, the incorporation of these assays for intermediate product testing allows earlier identification of mycoplasma contamination thereby expediting forward process decision making or corrective action to preserve downstream product integrity. The current challenges to the industry and regulatory agencies are validating and determining the applicability of these methods to the various subtypes of test articles used and generated within the manufacturing process. Integral to this process is an understanding of the current scope of mycoplasma contamination related to these various test articles. This study was undertaken to provide the mycoplasma contamination rates within four (4) individual mycoplasma detection assays offered by Bionique Testing Laboratories (BTL) in which the test articles are representative of cell cultures, cell substrates and end products seen in the production of biopharmaceuticals.

Section snippets

Methods

We retrospectively analyzed data from mycoplasma testing results at our facility to determine levels of contamination within cell cultures, substrates and biologicals. 10,000 consecutive samples were selected from each of four (4) individual detection assays to compare the incidence of mycoplasma contamination within the constituent subpopulations of raw materials, cell lines and cell culture derivatives used within biopharmaceutical manufacturing. The four assays were as follows:

•
Direct DNA

Statistical analysis

The GraphPad Prism 5 software (2007 GraphPad Software Inc. La Jolla, CA USA) was used for analysis of the data. We compared changes over time in the rates of contamination using a chi-square test for trend. We compared the rates of mycoplasma contamination among the four assay groups by 1-way ANOVA, followed by Bonferroni post tests for pairwise comparisons. For all tests, a P value of <0.05 was considered statistically significant.

Incidence

Analysis of data from the US Food and Drug Administration and three independent contract testing laboratories showed a declining incidence of mycoplasma contamination within cell cultures (P < 0.0001, chi-square test for trend) (Table 1). Based on Bionique Testing Laboratories’ observations of contamination rates within the M-250 assay cited in Table 1 for the years 1994 and 2009, there was a 1.6 fold greater occurrence in 1994 compared with the 2009 rate (95% confidence interval: 1.5–1.8).

Analysis of mycoplasma contamination in individual assays

Of the

Conclusions

Recognition of the current rates of mycoplasma contamination of cell cultures, cell culture substrates and biologicals within the biopharmaceutical industry is essential in determining the accuracy and efficacy of rapid microbiological methods to specific test articles and conditions. Implementation of more stringent quality control programs has resulted in a decline in the mycoplasma contamination rates as depicted in this study. That said, mycoplasma contamination remains a significant and

Acknowledgements

We thank the staff of Bionique Testing Laboratories for their contributions in the preparation of this manuscript. In addition, we appreciate Dr. John Ryan for his critical review of the material and Dr. Larry Johnson of Trudeau Institute for providing the statistical analysis.

References (7)

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Mycoplasma contamination of cell cultures: incidence, sources, effect, detection, elimination, prevention
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CBER et al.
Points to consider in the characterization of cell lines used to produce biologicals

There are more references available in the full text version of this article.

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The scope of mycoplasma contamination within the biopharmaceutical industry

Abstract

Introduction

Section snippets

Methods

Statistical analysis

Incidence

Analysis of mycoplasma contamination in individual assays

Conclusions

Acknowledgements

Contamination of human cell cultures by pleuropneumonialike organisms

Science

Mycoplasma contamination of cell cultures: incidence, sources, effect, detection, elimination, prevention

Cytotechnology

Points to consider in the characterization of cell lines used to produce biologicals