Integrated biosynthetic, resistance, and sensing systems in non-spore-forming bacteria from low-biomass environments
To ensure crew and mission safety and to prevent forward contamination of extraterrestrial environments, spacecraft components are assembled, processed, and cleaned within rigorously controlled facilities under strict adherence to established good manufacturing practices and protocols. Some microorganisms can tolerate cleanroom-associated stressors, such as desiccation and minimal nutrient availability, and may therefore remain detectable despite routine cleaning. At the NASA MSFC, three previously unidentified non-spore-forming bacterial isolates were recovered from cleanroom surfaces where components were processed. Genome-resolved analyses showed that all three isolates encode biosynthetic gene clusters, including carotenoid pathways, NRPS-like regions, epsilon-poly-L-lysine biosynthesis, genes associated with predicted stress tolerance such as heavy metal detoxification, and antimicrobial resistance determinants. Isolate PPS120 carries an acquired class A beta-lactamase (blaFONA-2) consistent with an extended beta-lactam-resistance phenotype, while PPS72 harbors mercury reduction genes that may contribute to tolerance of mercury-associated chemical stresses. Integrated genomic, phenotypic, and biochemical analyses supported classification of the isolates as three novel species, designated Paenarthrobacter huntsvillensis sp. nov. PPS72 (=DSM 120864, =ATCC TSD-592), Mycetocola huntsvillensis sp. nov. PPS117 (=DSM 120865, =ATCC TSD-593), and Erwinia huntsvillensis sp. nov. PPS120 (=DSM 120870, =ATCC TSD-594). Species delineation was supported by gyrB divergence (<90% identity) and genome-wide thresholds (Average Nucleotide Identity <89%; digital DNA-DNA hybridization <36%), with core-genome and marker-gene phylogenies placing each isolate on a distinct, well-supported branch relative to the closest described taxa. The identification of these three novel non-spore-forming bacteria underscores the value of genomic and functional screening for improving our understanding of cleanroom microbiota. These findings have broader implications for planetary protection by expanding genomic reference collections that may support future microbial surveillance and risk assessment. Such efforts may contribute to planetary protection and space habitat biosafety initiatives, while also identifying candidate biosynthetic pathways for future functional characterization and exploration of their biotechnological potential.