Importance of Mycoplasma Detection in the Biopharmaceutical Industry

Besides promotion of global health, the biopharmaceutical industry plays a major role in strengthening the economic value of countries such as the United States. In the U.S. alone, the sector accounts for 4.5 million jobs, drawing professionals from diverse fields including engineering, technology, mathematics, and science (Uphoff & Drexler, 2013). For instance, Pfizer is a company based in the U.S., with a significant number of workers to manufacture its famous high-quality life-saving drugs. Manufacturing of the life-saving medicines of prescribed quality calls for high know-how of every stakeholder, more so, within the manufacturing sector. Some factors, however, may be stumbles towards achieving this. Often, low-quality medicines, which happen to be harmful to human health, have been produced whenever these aspects are underlooked. Mycoplasma contamination is one such factor that alters the quality of the biopharmaceutical products. It is, therefore, a frontline necessity to test and detect its presence in all stages of the cell culture, to ensure production of high-quality biopharmaceutical products.

Mycoplasma, a prokaryotic microorganism, is one of the standard contaminants found in the cell culture laboratories. Due to their microscopic nature, mycoplasmas can penetrate through filters used to prevent fungiform and bacterial contaminations. Young, Sung, Stacey, and Masters (2010) observe that a mycoplasma infection occurs when a previously infected culture is taken to the library. Even though mycoplasma contamination creates no turbidity, some outcomes, such as altered cell metabolism, chromosomal aberrations, morphological changes, and changed cell growth rates, can arise. Additionally, Armstrong, Mariano, and Lundin (2010) observe that mycoplasma infections can severely affect the reproducibility, consistency, and reliability of experimental results in the cell culture processes. Certainly, detecting the mycoplasma contaminations is critical since it exposes problems in these processes. However, a research conducted by Aldecoa-Otalora et al. (2009) shows that out of all the laboratory scientists, half fail to confirm the availability of mycoplasma in their cell compositions. This act is termed as unprofessionalism by Armstrong et al. (2010), who argue that although mycoplasmas are too microscopic to be viewed under a laboratory microscope; their contagion distress may be very significant.

Therefore, it is essential to examine the cell compositions, to confirm any manifestation of mycoplasma contamination, especially when biopharmaceuticals are to be made through this process. Ayling et al. (2011) assert that testing for the presence of mycoplasma contamination is a means of meeting regulatory guidelines set by the authorized pharmaceutical regulatory bodies. The leading organizations that lay precise technical documentation on how to detect mycoplasma include European Pharmacopeia, The U.S. Code of Federal Regulations, U.S. Pharmacopeia, and the International Conference on Harmonization (Uphoff, & Drexler, 2013). Both the U.S. and the European pharmacopoeia techniques employ indicator cell culture assay and broth culture test in mycoplasma detection. Whereas indicator cell culture assay performs fastidious mycoplasma detection, broth culture test, also known as agar, detects cultivable mycoplasma.

The prolonged timescale for the detection of mycoplasma using these two techniques is so far their main drawback. While as agar culture protocol takes four weeks, the indicator cell culture assay takes seven days (Young et al., 2014). The two techniques are applicable in the manufacturing of monoclonal antibodies among other biologics. However, their prolonged detection timeframe becomes a big problem for the manufacturers of products such as cytotoxic viral suspensions and cell therapies, which have short shelf-life. Following this challenge, more validated alternatives are accepted by the regulators. Conducting polymerase chain reaction (PCR) tests is one such step that can be taken to remedy the problem, as it mitigates the accuracy and speed shortcomings. Roche and Life Technologies are typical applicants of the PCR strategies in conducting mycoTOOL and MycoSEQ commercial tests respectively (Falagan-Lotsch et al., 2015). In both cases, the lowest amount of genomic DNA copies of Mycoplasma in a given sample sets the limit mark (Falagan-Lotsch et al., 2015). Contrastingly, the limit of detection of the slow cell cultures is the precise number of live cells capable of generating colony forming units (CFU) on the solid medium surface.

It is crucial to note that mycoplasma presence in cell lines can be detected using tests prescribed by pharmacopoeia; that is, culture and indicator techniques. Indicator method can be used to screen bulk vaccines, harvest, and virus seeds. The tests conducted using this method, however, depends primarily on the accessibility of standardized neutralizing anti-sera virus sufficiency within the system. Culture method is modified to determine the mycoplasma manifestation, which cannot be displayed by other tests due to their low-level nature (Young et al., 2010). Within the broth, the medium is the phenol red, which detects any change in pH if subcultured to agar. Mycoplasma contamination is denoted by a change in color in the indicator. Usually, the prime purpose for performing subcultures is to affirm the broth results. However, the nucleic acid burden contained in the bioproduct supplements and cell cultures can be detected using other testing methods, more so, if the mycoplasma contamination rate in the cell culture substrates is low. Under such cases, the perception of nonviable and viable matter must be addressed.

The efficacity and accuracy of microbiological processes to a particular test condition in the biopharmaceutical industry can only be determined, after the prevailing pace of mycoplasma contamination of the biological cell culture substrates and cell cultures are examined. A reduction in the rate of mycoplasma contamination, as envisioned in this study, has resulted from the implementation of control programmes with high strict qualities. Even though mycoplasma contamination detection is of great significance to the biopharmaceutical industry, its costly nature remains problematic. The advanced refinement and surveillance of raw materials such as media and sera have helped in mitigating possible entry of viable mycoplasma from these products. Alternative mycoplasma detection methods are fast emerging, and data recorded from previous tests aid their developments. According to Falagan-Lotsch et al. (2015), PCR, a rapid molecular base method of testing mycoplasma contamination, is the best method that gives enhanced speciation and reduced timeframe, which is of high significance while testing quality control of the biotherapeutics with rapid expansion. Nevertheless, a sound understanding of the outlined reference standards alongside the compendial techniques is the only way towards the formation of any suitable and qualified alternative method.

Conclusion

In conclusion, it is worth noting that mycoplasma detection has an ultimate vitality in the vaccine manufacturing and bio-therapeutic. The availability of NAT tests with GMP validation and streamlined assay validation methods gives manufacturers within the biopharmaceutical industry confidence in the results needed to produce and release the produced batches. The required time for safety testing, development, and selling of the biotechnology products is much reduced due to fast reproducibility resulting from the use of NAT-based techniques. Although mycoplasma contamination detection may be a bit expensive, its numerous benefits in the biopharmaceutical make the examination worthy.

References

Armstrong, S. E., Mariano, J. A., & Lundin, D. J. (2010). The scope of mycoplasma contamination within the biopharmaceutical industry. Biologicals, 38(2), 211-213.

Ayling, R. D., Hlusek, M., Churchward, C. P., McAuliffe, L., Nicholas, R. A., & Goncalves, R. (2011). Analysis of the mycoplasma species that affect birds by SDS PAGE and immune-blotting.

Falagan-Lotsch, P., Lopes, T. S., Ferreira, N., Balthazar, N., Monteiro, A. M., Borojevic, R., & Granjeiro, J. M. (2015). Performance of PCR-based and bioluminescent assays for mycoplasma detection. Journal of Microbiological Methods, 118, 31-36.

Uphoff, C. C., & Drexler, H. G. (2013). Detection of mycoplasma contaminations. In basic cell culture protocols. Humana Press.

Young, L., Sung, J., Stacey, G., & Masters, J. R. (2010). Detection of mycoplasma in cell cultures. Nature Protocols, 5(5), 929.