Developments in real-time measurement technologies for bioburden applications are providing tangible benefits in the biopharmaceutical industry. Benefits include increased agility and efficiency to support decision-making, aid in the identification of elusive root causes, and enabling development of risk mitigation strategies among others. This article shares a case study in which a rapid micro- biological methodology (RMM) was leveraged as a tool to support a forensic evaluation for recovery of environmental conditions in a manufacturing facility after an unplanned business disruptive event. After failed attempts of traditional environmental air microbial verification tests, a rapid microbiological methodology system was used to enable spatially-segmented and finite-element focused scanning approaches for real-time measurement of biological particles in air media. These approaches supported identification of probable contamination focus areas which guided formulation and execution of an assertive cleaning strategy that recovered environmental conditions and solved an investigation, ultimately allowing the facility to resume commercial manufacturing operations.
Case Study Background
The island of Puerto Rico was hit by category 4 hurricane Maria in September 2017. The passing of the hurricane was so destructive that the power utilities blackout period is considered – at the moment of writing – the second largest globally (and counting) and the biggest by far in US history.1 During post-hurricane recovery efforts there were different initiatives which involved use of RMM technologies in a prioritized approach to reduce risks and expedite remediation of key manufacturing areas. Recovery of other areas, that were not necessarily initial priority timewise, followed to an extent recovery approaches typically used for planned disruptive events such as shutdowns for maintenance, construction, and/or repairs. Probably due to the severity of the atmospheric event and length of associated power disruption, a few of these latter areas failed traditional environmental microbial verification tests which triggered further investigation.
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In order to help identify root cause and tailor remediation activities, RMM was leveraged for real-time measurement of environmental air biological particles incorporating spatially-segmented and finite- element approaches in support of a forensic evaluation. These efforts intended to identify microbial contamination focus areas, and thus, guide cleaning and recovery efforts.
Apparatus
A rapid microbiological method (RMM) system was leveraged for the forensic activities (refer to Figure 1). This technology – capable of instantaneous microbial detection in air media – was designed for pharmaceutical applications and is able to measure both airborne microbes and non-viable particles continuously and in real-time using an optically-based system2 which allows for simultaneous particle sizing and fluorescence sensing of an induced stream of air. In specific terms of principle of measurement, this technology incorporates Mie scattering3 theoretical framework to interrogate particle size (spherical shape estimation) using a laser beam and a lens and photodiode optical assembly to generate a Mie scattered light path. Concurrently, a fluorescent light path coupled with long pass filter and photomultiplier tube module assembly determines fluorescence in particles which is attributed to viable or biological source. Reported results include particle count, size, and viable/non-viable classification.
Spatially-Segmented and Finite- Element Forensic Evaluation Using RMM Technology
The RMM system described was leveraged for in-situ, real-time biological particulate counting in air media. The spatially-segmented and finite-element approaches used intended to identify probable contamination focus areas – relying on relative measurements for biological particles – in aims to guide prospective cleaning efforts. In specific, the RMM technology was used for:
- Environmental air static scanning using a grid approach to map bioburden at room floor level
- Environmental air ‘dynamic doors’ approach to map bioburden exchange through access and cabinet doors
- Focused scanning of specific areas using flex tubing connected to system to map contribution of elements
- Scanning of ceiling fixtures (lamps, conditioned air entries, etc.) to map bioburden at room ceiling level
- Environmental air static re-scanning after remediation and cleaning activities but before sampling for traditional microbiological quality verification tests
Next we expand on each of the applications listed and share actual sample data results for illustrative purposes.
Environmental Air Static Scanning Using Grid Approach
In order to assess potential contamination focus areas, it was decided to divide the room using a grid approach (refer to Figure 2). This would allow focusing environmental interrogation by segmenting room space, which also aligned to the objective of identifying areas within the room that required additional attention during remediation activities. Since there was no previous baseline or characterization performed with the RMM system for acceptable conditions of environmental air static behavior (i.e., previous biological particle levels that typically resulted in passing of compendial environmental microbial quality verification test), hints for probable contamination focus areas were primarily expected from any significant disparity in values observed when comparing grid results for biological count. Figure 2 shows a simplified schematic of the room grid division and shares initial measured values for biological average count at each measured grid.
Environmental Air ‘Dynamic Doors’ Scanning Approach
In order to provide a detailed interrogation of the different elements with potential contribution to room environmental bioburden, it was decided to perform ‘dynamic doors’ measurements. For this approach, each access door was individually opened and closed while the RMM system was measuring nearby in order to assess the degree of bioburden variability due to the air exchange disturbance. This activity was also performed with the doors of each cabinet within the room. Once again, the objective was to identify probable contamination focus areas in order to guide additional cleaning efforts. Figure 3 shares sample pictures and results for these activities.
Focused Scanning of Floor-Level Elements Using Flex Tubing Extension
In order to complete the picture for the different barrier elements and the level of bioburden exchange with the room before remediation activities, it was decided to further focus scanning by using a flex tubing extension connected to the RMM technology and interrogate the elements’ perimeters. Figure 4 illustrates some of the focused perimeter scanning activities and shares results for some of the floor- level elements interrogated.
Focused Scanning of Ceiling-Level Fixtures Using Flex Tubing Extension
A similar approach (as in previous section) was used for ceiling-level elements. The perimeter of ceiling-level fixtures (lamps, air ducts, explosion vent panels) was scanned using the flex tubing extension connected to the RMM system. Figure 5 shares pictures and sample results from this part of the exercise.
Basis of Analysis and Pre/Post Remediation Comparison of Biological Particle Measurements
The analysis of the results for the scanning activities – summarized in previous sections of this article – helped identify probable contamination focus areas. From Figure 2, for example, areas with relatively higher biological average count where identified (individual grids highlighted in yellow). Also, when considering results from a holistic perspective, it was possible to further narrow probable contamination sources. For example, the relatively higher bioburden levels measured for grids 1 and 6 (Figure 2) during environmental static scanning are most probably connected to the relatively higher values obtained when scanning the perimeter of lamps labeled L15 and L16 in Figure 5. The relatively higher bioburden values measured in grid 5 (Figure 2) possibly relate to deficient perimeter sealing for some of the explosion vent panels. Similarly, relatively higher bioburden values for grids 23 thru 25 (Figure 2) are connected to the relatively higher bioburden values measured during ‘dynamic doors’ scanning for door labeled‘Utility Room Door’in Figure 4 that connects to a room with dust collection equipment, and also the contribution from air exchange through the double doors room entrance (Figure 4).
The identification and narrowing of probable contamination focus areas within the room enabled the formulation of an assertive cleaning strategy resulting in the implementation of tailored remediation activities that increased effectiveness of the efforts. In order to assess the effectiveness of the remediation activities, static environmental scanning was repeated using the previous grid approach. This activity allowed for comparison of bioburden levels before and after remediation and provided information to help decision-making– either clean again or proceed to verification test sampling – for reduced risk of failing the traditional environmental microbial quality verification test. Figure 6 share sample results as well as pre/post comparison of results after static re-scanning activities. The overall biological average particle count measured with RMM decreased from 36 before remediation activities to 23 after implementation of an assertive cleaning strategy, representing a reduction of over 36% for biological particle counts. Based on results, the room was sampled and passed the traditional environmental microbial quality verification test receiving approval to resume production.
Concluding Remarks
This article shared an industrial case study in which an alternative rapid microbiological methodology was used during a forensic evaluation. Spatial segmentation of room and finite elements focused scanning approaches were leveraged to help identify probable contamination focal points and support the selection and implementation of remediation activities for a manufacturing facility after the passing of hurricane María. The real-time biological particles measurement activities helped identify probable contamination focus areas by delimiting grids for room static environmental air scanning complemented with ‘dynamic doors’ scanning approach and with focused scanning of elements’perimeters at the floor and ceiling levels using a flex tubing extension connected to the RMM apparatus. The identification of probable biologically-contaminated sources allowed for an improved remediation strategy and the implementation of tailored cleaning activities which increased effectiveness of the overall recovery efforts, ultimately enabling passing of traditional environmental microbial tests and allowing resume of commercial manufacturing operations and continued supply of pharmaceutical products to patients.
The main intrinsic benefit of the RMM technology presented comes from its capability to provide instant sampling and measurements for environmental air biological particles. If traditional microbial tests would have been used to guide the remediation process, each test would have required anywhere from 3 to 7 days of incubation and reading of incubation plates. Test(s) failures typically trigger investigation activities and associated incremental costs, while the manufacturing area would have been kept at idle during the extended issue resolution period adding costs due to loss of opportunity. This case study demonstrates how leveraging the right technology and expertise can prove fruitful. As we all know, technology keeps evolving at a fast pace and there are considerable opportunities with potential to increase the efficiency and effectiveness of business processes. Keeping abreast of contemporary technologies and matching them to business unmet needs can unleash measurable benefit and – when strategically adopted – increase competitiveness both in the short- and long-term horizons.
Acknowledgements
Many colleagues were directly and/or indirectly involved supporting the activities described in this article. This script only summarizes and shares a few samples of the actual work that collectively spanned for weeks. Although not possible to mention everyone, we would like to acknowledge the support and/or sponsorship of the following colleagues and leaders, without which the work and/or article publication would not have been possible: Gratitude goes to Josué Armaiz, Elizabeth Ortiz, Iván Vega, Ricardo Colón, Víctor Cartagena, Alexander García, Ebed Rodríguez and Benjamín Vélez. We would also like to acknowledge Jeffrey Weber and Joanny Salvas for providing awareness of RMM technologies throughout the Pfizer Network. Thank you all.
References
- Irfan, U. (2018). Puerto Rico’s blackout is now the second largest on record worldwide. Electronically retrieved April 15, 2018 from Vox website at https://www.vox. com/2018/4/13/17229172/puerto-rico-blackout-hurricane-maria.
- BioVigilant Technical Paper (N.S.). Applications Concept: Use of an IMD-A® System for Personnel Training in Cleanroom and Aseptic Manufacturing Environments (page 1). Electronically retrieved on February 9, 2018 from BioVigilant website at http://biovigilant. com/wp-content/uploads/2014/07/Training.pdf.
- Hahn, D.W. (2009). Light Scattering Theory. Electronically retrieved on March 2nd, 2018 from University of Florida website at http://plaza.ufl.edu/dwhahn/Rayleigh%20and%20 Mie%20Light%20Scattering.pdf.
Author Biography
José-Miguel Montenegro-Alvarado is Manager of Process Analytical Technology (PAT) projects as part of Pfizer’s Global Technology Services (GTS) / Global Engineering (GE) / Manufacturing Process Analytics and Control (MPAC) Team. Based in Vega Baja, Puerto Rico in his current role he is responsible for technical support and facilitates deployment of PAT at Pfizer’s Global Solids Manufacturing (GSM), Active Pharmaceutical Ingredient (API), and Consumer Healthcare (CH) sites.
Montenegro’s academic background includes Bachelor’s and Master’s degrees in Chemical Engineering at the University of Puerto Rico – Mayagüez with a minor equivalency in Economics. His professional career in the pharmaceutical industry started in 2001 at the Searle & Co. Caguas site after industrial internships in medical devices with Baxter and Techno-Plastics Industries. In 2007 Montenegro was recruited by Pfizer Center Functions as part of the Process Analytical Sciences Group (PASG). Throughout time Montenegro has interfaced with over 20 Pfizer sites in 4 different continents including United States, Puerto Rico, Australia, Argentina, Brazil, Italy, Mexico, Spain and Venezuela. In 2010 Montenegro transitioned into his current role as Manager – PAT projects.