4

Sterilization and Cleaning Methods

The procedures used to ensure spacecraft cleanliness and, ultimately, to achieve the desired sterilization standards begin during the design and manufacturing of spacecraft components. Afterward, when the components are being assembled, further cleaning and sterilization protocols are implemented. Unfortunately, it is not currently practical to sterilize an entire spacecraft at one time, post-assembly, while at the same time protecting all of its diverse components and sensors from damage or failure. The different sensitivities of internal components to sterilizing procedures require that many of the parts be sterilized individually, using a procedure compatible with their function. For complex scientific missions, therefore, whole-spacecraft sterilization is not an option—a single sterilization procedure would be limited by the spacecraft’s most sensitive component. As a result of this constraint, many spacecraft components are sterilized individually and then assembled in clean rooms using rigorous procedures that minimize recontamination.

CLEANING AND STERILIZATION STANDARDS

NASA’s current planetary protection requirements for Mars missions are derived from the procedures applied to the Viking landers. Missions not carrying life-detection experiments must be cleaned to ensure that the spacecraft’s total bioload does not exceed 300,000 spores and that the density of spores on the spacecraft’s surfaces does not exceed 300 m-2.Missions with life-detection experiments must undergo additional procedures to ensure that the total bioload does not exceed 30 spores. The effectiveness of the various procedures currently used by NASA and its contractors to meet these bioload standards is determined by the use of reference organisms, including Bacillus subtilis (var. niger), Bacillus pumilis, and Bacillus stearothermophilus. Bacillus spp. (endospore formers) were originally selected as a microbiological indicator of sterilization success on the basis of their enhanced resistance to heat, desiccation, and radiation.

ACHIEVING THE STANDARDS

The twofold approach to the control of forward contamination used by the Viking mission—careful cleaning of the spacecraft, followed by active bioload reduction through heat sterilization (see Box 1.1 in Chapter 1)—forms the basis for the procedures currently in use. All missions are carefully cleaned and then those with life-detection experiments undergo sterilization.

The Viking landers were assembled in clean rooms (see Box 4.1 for a description of current clean-room procedures). During assembly, microbial assays (see Chapter 5) were conducted to establish that the average and total burden of spores on the lander’s accessible surfaces were 300 m-2and 300,000, respectively.1 Current practice requires that those parts of the spacecraft not meeting the requisite bioload standards be washed with isopropyl alcohol and/or a sporicide (ethanol, 65 percent; isopropanol, 30 percent; and formaldehyde, 5 percent) to reduce their bioburden. Decontaminated surfaces are then retested for their contaminating microbiological burden.

Once the landers had been assembled and sealed inside their bioshields, the bioload was further reduced by dry heating the whole spacecraft to at least 111.7 °C for some 30 hours. This procedure was credited with reducing the lander’s bioburden by a factor of 104. Future spacecraft can be designed to maximize accessibility of their components for pre- and post-assembly bioload reduction. However, some components are hermetically sealed before assembly, so cleaning/sterilizing procedures must be conducted before sealing to prevent recontamination. The sterilization procedures commonly applied in a variety of applications to sealed and unsealed components are listed in Table 4.1. It is worth noting that many of these procedures can have negative impacts on spacecraft performance and may increase mission cost.



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Preventing the Forward Contamination of Europa 4 Sterilization and Cleaning Methods The procedures used to ensure spacecraft cleanliness and, ultimately, to achieve the desired sterilization standards begin during the design and manufacturing of spacecraft components. Afterward, when the components are being assembled, further cleaning and sterilization protocols are implemented. Unfortunately, it is not currently practical to sterilize an entire spacecraft at one time, post-assembly, while at the same time protecting all of its diverse components and sensors from damage or failure. The different sensitivities of internal components to sterilizing procedures require that many of the parts be sterilized individually, using a procedure compatible with their function. For complex scientific missions, therefore, whole-spacecraft sterilization is not an option—a single sterilization procedure would be limited by the spacecraft’s most sensitive component. As a result of this constraint, many spacecraft components are sterilized individually and then assembled in clean rooms using rigorous procedures that minimize recontamination. CLEANING AND STERILIZATION STANDARDS NASA’s current planetary protection requirements for Mars missions are derived from the procedures applied to the Viking landers. Missions not carrying life-detection experiments must be cleaned to ensure that the spacecraft’s total bioload does not exceed 300,000 spores and that the density of spores on the spacecraft’s surfaces does not exceed 300 m-2.Missions with life-detection experiments must undergo additional procedures to ensure that the total bioload does not exceed 30 spores. The effectiveness of the various procedures currently used by NASA and its contractors to meet these bioload standards is determined by the use of reference organisms, including Bacillus subtilis (var. niger), Bacillus pumilis, and Bacillus stearothermophilus. Bacillus spp. (endospore formers) were originally selected as a microbiological indicator of sterilization success on the basis of their enhanced resistance to heat, desiccation, and radiation. ACHIEVING THE STANDARDS The twofold approach to the control of forward contamination used by the Viking mission—careful cleaning of the spacecraft, followed by active bioload reduction through heat sterilization (see Box 1.1 in Chapter 1)—forms the basis for the procedures currently in use. All missions are carefully cleaned and then those with life-detection experiments undergo sterilization. The Viking landers were assembled in clean rooms (see Box 4.1 for a description of current clean-room procedures). During assembly, microbial assays (see Chapter 5) were conducted to establish that the average and total burden of spores on the lander’s accessible surfaces were 300 m-2and 300,000, respectively.1 Current practice requires that those parts of the spacecraft not meeting the requisite bioload standards be washed with isopropyl alcohol and/or a sporicide (ethanol, 65 percent; isopropanol, 30 percent; and formaldehyde, 5 percent) to reduce their bioburden. Decontaminated surfaces are then retested for their contaminating microbiological burden. Once the landers had been assembled and sealed inside their bioshields, the bioload was further reduced by dry heating the whole spacecraft to at least 111.7 °C for some 30 hours. This procedure was credited with reducing the lander’s bioburden by a factor of 104. Future spacecraft can be designed to maximize accessibility of their components for pre- and post-assembly bioload reduction. However, some components are hermetically sealed before assembly, so cleaning/sterilizing procedures must be conducted before sealing to prevent recontamination. The sterilization procedures commonly applied in a variety of applications to sealed and unsealed components are listed in Table 4.1. It is worth noting that many of these procedures can have negative impacts on spacecraft performance and may increase mission cost.

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Preventing the Forward Contamination of Europa TABLE 4.1 Common Sterilization Procedures Procedure—Target Techinique—Problems Dry heat—exterior/interior 105-180 °C for 1 to 300 hours—Problems caused by thermomechanical incompatibility between materials can lead to the faliure of electronic components. Wet heat—exterior/interior 120-134 °C for 3 to 20 minutes—Problems can be caused by steam (e.g., corrosion and water absorption). Alcohol wipes—exterior surfaces Isopropyl or ethyl alcohol swabbing—Problems arise because interior and encased surfaces (e.g., electronic components) are inaccessible. Ethylene dioxide—exterior/internal exposed surfaces Toxic gas, 40 to 70 °C—Problems arise because the gas can only reach exposed surfaces and because it is absorbed by some types of polymers (e.g., rubbers and polyvinyl chloride). Gamma radiation—exterior/subsurface Typically, 2.5 Mrad—Problems encountered include optical changes in glasses and damage to electronics and solar cells. Beta radiation—exterior/near-surface 1 to 10 MeV—Problems arise because of limited penetration. Hydrogen peroxide plasma—exterior/internal exposed surfaces 6 mg/l H202 concentrated at 58%—Problems can be encountered because the unexposed surfaces remain untreated. Ultraviolet—exterior surfaces 5,000 to 20,000 J/m2—Problems arise because unexposed surfaces remain untreated. Methyl bromide—exterior/internal exposed surfaces Toxic gas—Problems can be encountered because unexposed surfaces remain untreated and because the gas catalyzes chemical reactions between metal and other components. These treatments can be highly effective, but they have their limitations in certain circumstances. Important factors influencing germicidal activity include the following: the types of microorganisms; the number of organisms; the intrinsic resistance of the organisms; the amount of organic soil on the item to be sterilized; the type and concentration of germicide; the time and temperature of exposure; and the compatibility between of the device being sterilized and the technique being used.

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Preventing the Forward Contamination of Europa BOX 4.1 Current Clean-Room Procedures Clean rooms are highly controlled environments accessible only to trained personnel following strict and unambiguous cleanliness protocols. Representative standard NASA clean-room protocols include the following: During assembly, workers are required to wear full face shield suits; No human contact directly with spacecraft is permitted. Latex gloves are worn in the clean room, and spacecraft are not seeded with tracer organisms to facilitate monitoring; Cameras are used to observe and monitor assembly; Clean-room air passes through high efficiency particulate air (HEPA) filters and dehumidifiers to minimize airborne microbial contamination and corrosion, respectively; Surface particles are removed by vacuuming; Witness plates are regularly collected and stored; Contact between hardware and biologically relevant materials is minimized; and Surface areas of the spacecraft are monitored periodically for their microbiological burden, during and after assembly. Sterile cotton swabs are used to collect contaminating surface microorganisms, which are subsequently cultured and counted. Unfortunately, clean rooms do not guarantee contamination-free assemblies. Mistakes happen, and clean hardware may not remain clean. Thus, good in-process cleaning procedures are necessary. REFERENCE 1 Viking '75 Project, Pre-launch Analysis of Probability of Planetary Contamination, Volumes II-A and II-B, M75-155-01 and M75-155-02, Jet Propulsion Laboratory, Pasadena, Calif., 1975.