In this third instalment of our Recommended Research Papers series, I have included a collection of six articles. All of them have piqued my interest as they tackle some extremely pertinent issues in our industry.
We all know from experience the sheer number of things that could go wrong within a cleanroom. Managers must be on their toes when it comes to the upkeep of their facilities and the operations that ensure standards are met. It pays to be a couple of steps ahead with a comprehensive strategy in place should you be faced with a potential catastrophe.
Preventative measures often built around making educated decisions is paramount to mitigating the risks affiliated with cleanroom facilities. These articles contain information that serves as excellent guidelines.
Start-up cleanrooms, initially and after a worst-case event
This paper gives sound advice on entry and cleaning procedures in the event of an unscheduled shutdown or failure to the air handling unit in a pharmaceutical cleanroom. The level of cleaning and monitoring required before the cleanroom is re-opened for use will depend on the length of the shutdown and the type or cause of failure. There is a great deal of information in this article and it is a must-read for both cleanroom operators and managers alike.
Abstract
Environmental control of classified areas within a biopharmaceutical facility is maintained by systems controlling humidity, air temperature, air exchanges, filtration and pressure differentials and by practices such as room cleaning, limited access and facility flow. The failure of any of these systems has the potential to impact the clean state of a Classified Area.
Many facilities have procedures in place to conduct the day-to-day operations but fail to have procedures in place to handle catastrophic or non-routine events. These disturbances should be defined as part of the Facility Cleaning Procedures in order to provide guidance to operators in the event they occur during off-shift or to provide a routine response to a non-routine event. The information in this article will provide an understanding of how these events may affect the cleanroom so measures may be taken to prevent issues leading to shutdown, costly discrepancy investigations, and potential lost product.
Source references and availability
Authors: Polarine J and Kroeger B
Published: July 2017
Available from ResearchGate.
Biodecontamination of Cleanrooms and Laboratories Using Gassing Systems
This paper offers an excellent comparison of the two primary methods of laboratory and cleanroom fumigation; formaldehyde and hydrogen peroxide vapour. Detailing the methods used for both systems, it goes on to list the pros and cons of each technique, after which the author looks at the regulatory requirements for the validation of gassing/fumigation systems. He finishes by summarising the other technologies that are available.
Abstract
Cleanrooms, laboratory areas, isolators and biosafety workspaces (microbiological safety cabinets) require a level of cleanliness and microbial control (achieved through disinfection) according to the intended use of the area. With cleanrooms this relates to microbial levels according to the class or grade of the room; with containment laboratories and biosafety cabinets this is with the intention of eliminating specific pathogens.
Source references and availability
Author: Tim Sandle
Published: January 2017
Available from Institute of Validation Technology.
Evaluation of the recovery rate of different swabs for microbial EM
This article was of immediate interest to me, as I spent many years using swabs for the environmental monitoring of cleanroom equipment. Contact or Rodac plates are suited for flat surfaces, but not for uneven or hard to reach surfaces. Many different swab tip materials are used from various manufacturers, on different surfaces, tested both in vitro and in situ. In the abstract you will see below, the most important advice I can take from this study is the importance of sticking to swab usage as per manufacturer instructions for optimal recovery.
Abstract
Environmental microbiological monitoring provides information on the hygiene condition of pharmaceutical clean rooms and equipment for manufacturing of drug products. Different methods can be used to recover microorganisms. For surfaces, normally contact plates (e.g., RODAC or dipslides) are used; however, when surfaces are uneven, swabs should be used. In the present study three different swabs were evaluated for their ability to recover microorganisms from different surfaces. Thereby two methods and two approaches were evaluated. Swab samples were either directly stroked on agar or the swab was eluted, membrane-filtrated, and the filter placed on an agar plate.
Experimentally, artificial inoculated surfaces typically found in clean rooms (in vitro study) and naturally inoculated floors (in situ study) were sampled. Thus with this approach the most convenient swab and the most suitable recovery methods under laboratory as well as real clean room conditions were evaluated. With this set-up, we found the most suitable swab for our environmental monitoring not only by using artificial inoculated surfaces but also under more realistic clean room conditions, which is most important for microbiological environmental monitoring sampling.
Source references and availability
Authors: M Goverde, J Willrodt, A Staerk
Published: August 2016
Available from NCBI .
Growth promotion efficiency of dehydrated Agar settle plates used for monitoring viable particles in grade A environments
This is the first of two short articles regarding the recovery of microorganisms on environmental monitoring media. This one looks at the effects of dehydrated agar on the recovery of microorganisms on settle plates, following exposure for various lengths of time in a laminar air flow unit.
Abstract
Background: Settling method of air sampling was introduced by Koch in 1881 by measuring the number of bacteria that settled onto solid medium, and nowadays is one of the controls established by GMP standards to monitor processes performed in grade A environments. Settle plates do not measure the number of microorganisms in the air but measure the number of microorganisms settling from air onto a known surface area in a known time. Even today the wrong idea that sedimentation plates are susceptible to dehydration and lose the effectiveness to recover microorganisms still persists. Whyte and Niven demonstrated that microorganisms commonly found in cleanrooms are little affected by the drying caused by exposure of settle plates up to 6 hours of exposure to unidirectional flow. However, the current GMP standards do not allow more than 4 hours of exposure and the effect of dehydration with respect to local sampling conditions should be checked. In this study, the effect of dehydration of a grade A laminar flow and humidity onto 90 mm settle plates was evaluated at 4, 6 and 8 hours.
Material and Methods: Agar Trypcase Soja 3P™ (TSA3) plates (BioMérieux, Marcy l'Etoile, France) were used. Plates were subjected to an exposure period of 4, 6 and 8 hours into a Grade A Laminar Flow Cabinet (flow rate 0.42 m/s) and a working environmental humidity of 38–40% in order to dehydrate the media in a real environment. The plates were then placed for a 2-hours exposure time in an unclassified environment (administrative area and quality control laboratory, 10 locations) along with control plates that had not undergone the prior dehydration process. Plates were then incubated at 30–35°C for 72 hours.
Results: No significant differences were observed between the recoveries of the dehydrated plates at 4, 6 and 8 hours and the control plates.
Source references and availability
Authors: A Rodriguez, I De Toro, M Mediavilla, M. Fiñana, R Maldonado, D Rodriguez-Rodriguez, L Leyva
Published: May 2017
Available from Cytotherapy.
Validation of Collection Efficiency of Contact Plates Used on Surface Sampling in Cell Therapy Cleanrooms
The second article by the same team looks at the recovery of microorganisms on contact plates used on surfaces containing disinfectant residues. It shows the importance of having the appropriate neutralisers in your contact plates to neutralise the disinfectant residues. Without the neutralisers, there will be a risk of false negative results giving an inaccurate view on both the cleanliness of the surfaces and the effectiveness of your cleaning regime.
Abstract
Background: Microbiological sampling methods used in pharmaceutical cleanrooms should efficiently collect and count microorganisms and must be adequately validated to ascertain their collection efficiency. Disinfectants used in cleaning and disinfection procedures in controlled environment areas may leave residues on surfaces that could interfere with the ability to promote the growth of microorganism in the media used in surface sampling methods. Therefore, the culture media must be able to neutralize these residues. Most manufacturers add neutralizing agents such as lecithin, polysorbate 80 (Tween 80), L-histidine and sodium thiosulfate, among others. Even so, the capacity of these neutralizing agents depends on the type and concentration of the disinfectant used and this capacity must be checked under real conditions by the user. In this study, the neutralizing capacity of Count-Tact™ 3P agar plates (BioMérieux) was checked against three disinfectant agents.
Material and Methods: The following ready-to-use biocides were assessed: InSpec™ HA premium (200 ppm hypochlorous acid), InSpec™ OX premium (6% hydrogen peroxide and 4% PAA. Redditch Medical, UK) and Klercide™ 70/30 D.Ethanol (Ecolab Contamination, UK). Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Candida albicans and Aspergillus niger (Bioball™ 30 ufc SingleShot, BTF BioMérieux) were used for growth promotion tests. Additionally, two microorganisms that usually grow in cleanrooms areas were selected: Kocuria rhizophila (Bioball™ 30 ufc SingleShot) and Micrococcus luteus (prepared in our laboratory; strain isolated from environmental monitoring control). A stainless steel surface was sprayed with each of the biocides, allowing them to dry until completely evaporated. The contact plates were then faced to the surface for a contact time of 10 seconds. Using a laminar flow cabinet, the mentioned contact plates were inoculated with the indicated microorganisms. The plates were finally incubated at 30–35°C for 24 hours. Control plates without exposure to biocide agents were also seeded.
Results: All recoveries met the criteria established in Eur. Ph. 2.6.12: “not differ by a factor greater than 2 from the calculated values for a standardised inoculum”.
Source references and availability
Authors: A Rodriguez, I De Toro, M Mediavilla, M. Fiñana, R Maldonado, D Rodriguez-Rodriguez, L Leyva
Published: May 2017
Available from Cytotherapy.
Revision of viable EM in a development pilot plant based on quality risk assessment: a case study
I do love a good case study when it is directly relevant to my line of work. This is a classic example of one. When designing and implementing an environmental monitoring regime it is a regulatory expectation the sampling locations are based on a risk assessment method – in this case Hazard Analysis Critical Control Point (HACCP) was used. A total of eleven hazards were identified, which included factors such as temperature, presence of water, frequency of cleaning and number of personnel operating in that area.
A score depending on severity of risk was assigned to each section. For example for temperature, a lower score was assigned for lower temperatures as there was less chance for the proliferation of microbes. A higher temperature had a higher score as microbes were more likely to proliferate. These hazards were then applied to the existing regime. Do read this paper yourself, as the detail will give you food for thought!
Abstract
In this case study, the principles of Quality Risk Management were applied to review sampling points and monitoring frequencies in the Hormonal Tableting unit of a Formulation Development Pilot Plant. In the cleanroom area premises, different functions are located, and therefore a general method was established for risk evaluation based on the Hazard 2 analysis and critical control points (HACCP) method. The point of this is to evaluate these premises (i.e. production area itself and ancillary clean areas) from a microbial load and state to observe if the existing monitoring program met the emerged advanced monitoring practice.
Source references and availability
Author: Ildikó Ziegler (formerly Mohammed-Ziegler)
Published: January 2017
Available from ResearchGate.
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