BACKGROUND: ONE OF THE MAJOR TECHNICAL CHALLENGES IN PLANETARY PROTECTION FOR THE EUROPA LANDER AND OTHER NASA DEEP SPACE MISSIONS IS IN REDUCING THE BIOBURDEN OF TERRESTRIAL MICROORGANISMS ASSOCIATED WITH SOLID ROCKET MOTOR (SRM) INSULATION AND OTHER MATERIALS THAT MAY HAVE MICROBIAL CONTAMINANTS. AN ADDITIONAL CHALLENGE IS IN THE IDENTIFICATION OF APPROPRIATE SRM INSULATION MATERIALS THAT MEET THE THERMAL MECHANICAL AND RHEOLOGICAL SPECIFICATIONS FOR EUROPA LANDER AND OTHER MISSIONS. IN PREVIOUS EXPERIMENTS WE HAVE IDENTIFIED ADHESIVES THAT REDUCE BACILLUS SPORE VIABILITY THAT ARE COMMONLY USED IN SRM INSULATION ASSEMBLY. IN THIS PROPOSAL WE WILL COMBINE OUR COLLECTIVE EXPERTISE IN MICROBIOLOGY AND MATERIAL ENGINEERING TO IDENTIFY THE IDEAL COMBINATIONS OF ADDITIVES ADHESIVES POLYMERS AND INSULATION MATERIALS FOR USE IN THE EUROPA LANDER MISSION THAT COULD ALSO BE APPLICABLE TO OTHER DEEP-SPACE MISSIONS. SPECIFIC OBJECTIVES: 1) DETERMINE THE REDUCTION IN BACILLUS SPECIES SPORE VIABILITY IN RESPONSE TO ADHESIVE FORMULATIONS APPLIED TO SRM CANDIDATE INSULATION MATERIALS. THESE STUDIES WILL INCLUDE MULTIPLE ADHESIVE FORMULATIONS ALREADY IN USE FOR SPACECRAFT ASSEMBLY AND EVALUATE THE EFFECTS OF SPORE INOCULUM TIME OF EXPOSURE AND INSULATION COMPOSITION ON THE DEGREE OF BIOBURDEN REDUCTION THAT CAN BE ACHIEVED. WE WILL CONDUCT THESE ASSAYS USING STANDARD NASA PROTOCOLS WITH FOUR DIFFERENT BACILLUS SPECIES ISOLATED FROM SPACECRAFT ASSEMBLY ENVIRONMENTS AND WILL USE CRYOGRINDING TO PROCESS INSULATION MATERIALS PRIOR TO QUANTITATIVE DETERMINATION OF SPORE VIABILITY ON LABORATORY MEDIA. 2) EVALUATE BIOBURDEN ASSOCIATED WITH EUROPA LANDER MATERIALS. IN ADDITION TO SRM INSULATION THERE ARE OTHER MATERIALS THAT MUST BE EVALUATED FOR THEIR INHERENT BIOBURDEN. IN CONSULTATION WITH SCIENTISTS AT MARSHALL SPACE FLIGHT CENTER AND THE JET PROPULSION LABORATORIES OTHER MATERIALS ARE EXPECTED TO BE IN NEED OF EVALUATION FOR THEIR ASSOCIATED BIOBURDEN IN PARTICULAR LINERS O RINGS AND LUBRICANTS. USING THE SAME METHODOLOGIES APPLIED TO SRM INSULATION THE BIOBURDEN ASSOCIATED WITH EACH OF THESE MATERIALS WILL BE QUANTITATIVELY DETERMINED IN REPLICATE SAMPLES. 3) CONDUCT CULTURE-BASED AND METAGENOMIC ANALYSES OF THE MICROBIOTA ASSOCIATED WITH MATERIALS OF PARTICULAR CONCERN FOR EUROPA LANDER PLANETARY PROTECTION. MONITORING BACILLUS SPECIES SPORE VIABILITY IS A CRITICAL INDICATOR OF BIOBURDEN REDUCTION BUT OTHER ORGANISMS MAY BE ASSOCIATED WITH EUROPA LANDER MATERIALS. WE WILL THEREFORE USE A COMBINATION OF METHODS THAT INCLUDES ANALYSIS OF MICROBIAL CULTURES OBTAINED FROM PREVIOUS OBJECTIVES AND CULTURE-INDEPENDENT INVESTIGATION OF 16S RRNA GENE PCR AMPLICONS THAT ARE TYPICAL OF MICROBIOME STUDIES AS WELL AS A SHOTGUN METAGENOMIC APPROACH THAT WILL COVER A GREATER BREADTH OF THE PHYLOGENETIC DIVERSITY PRESENT IN INSULATION MATERIALS. APPROPRIATENESS OF THIS RESEARCH FOR THIS PROGRAM ELEMENT: THIS RESEARCH FOCUSES ON ONE OF THE MAJOR CHALLENGES IN PLANETARY PROTECTION FOR THE EUROPA LANDER MISSION IN PREVENTING FORWARD CONTAMINATION DUE TO MICROORGANISMS ASSOCIATED WITH SRM INSULATION AND OTHER EUROPA LANDER MATERIALS.