A comprehensive metagenomics framework to characterize organisms relevant for planetary protection

Abstract

Background Clean rooms of the Space Assembly Facility (SAF) at the Jet Propulsion Laboratory (JPL) at NASA are the final step of spacecraft cleaning and assembly before launching into space. Clean rooms have stringent methods of air-filtration and cleaning to minimize microbial contamination for exoplanetary research and minimize the risk of human pathogens, but they are not sterile. Clean rooms make a selective environment for microorganisms that tolerate such cleaning methods. Previous studies have attempted to characterize the microbial cargo through sequencing and culture-dependent protocols. However, there is not a standardized metagenomic workflow nor analysis pipeline for spaceflight hardware cleanroom samples to identify microbial contamination. Additionally, current identification methods fail to characterize and profile the risk of low-abundance microorganisms. Results A comprehensive metagenomic framework to characterize microorganisms relevant for planetary protection in multiple cleanroom classifications (from ISO-5 to ISO-8.5) and sample types (surface, filters, and debris collected via vacuum devices) was developed. Fifty-one metagenomic samples from SAF clean rooms were sequenced and analyzed to identify microbes that could potentially survive spaceflight based on their microbial features and whether the microbes expressed any metabolic activity or growth. Additionally, an auxiliary testing was performed to determine the repeatability of our techniques and validate our analyses. We find evidence that JPL clean rooms carry microbes with attributes that may be problematic in space missions for their documented ability to withstand extreme conditions, such as psychrophilia and ability to form biofilms, spore-forming capacity, radiation resistance, and desiccation resistance. Samples from ISO-5 standard had lower microbial diversity than those conforming to ISO-6 or higher filters but still carried a measurable microbial load. Conclusions Although the extensive cleaning processes limit the number of microbes capable of withstanding clean room condition, it is important to quantify thresholds and detect organisms that can inform ongoing Planetary Protection goals, provide a biological baseline for assembly facilities, and guide future mission planning.

Contamination analysis of Arctic ice samples as planetary field analogs and implications for future life-detection missions to Europa and Enceladus

“…In the controls, Acinetobacter was more abundant in the DNA-extraction control. Members from the Acinetobacter genus have been identified before as part of the kitome (i.e., contaminants from DNA extraction reagents) 41 , and as contaminants of the NASA Space Assembly Facility 42 , 43 , where the last cleaning steps of space vehicles take place before launch. Finally, the Acinetobacter case shows how our method allows not only the reduction, identification, and monitoring of likely contaminants but also a close trace to their possible origin.…”

Supervised Machine Learning Enables Geospatial Microbial Provenance

“…Species classified by Cassandra can be further used to monitor pathogenic outbreaks as well as the spread of AMR using additional resources. Databases like Global Biodiversity Information Facility (GBIF) [ 49 ], TerrestrialMetagenomeDB [ 50 ], and Microbe Directory (MD2) [ 51 ] can be used as ancillary information for defining phenotypes, and other attributes of the bioindicators species [ 52 ]. Thus, the use of this tool can help improve source tracing and phylogenetic reconstruction to determine disease transmission in geographical locations in the situations of biocrime and biothreat.…”

A history of the MetaSUB consortium: Tracking urban microbes around the globe

“…We have already collected samples from 21 monuments and landmarks from 12 different cities, including surface samples from the ancient cities of Tel Megiddo in northern Israel, which follows some of the protocols we developed when doing metagenomics on the Dead Sea Scrolls ( Anava et al., 2020 ). Some of our methods for k-mer mapping are now also used in projects with NASA for determining levels of possible biological contamination for spacecraft ( Danko et al., 2021b ).…”