Succession of the bacterial community from a spacecraft assembly clean room when enriched in brines relevant to Mars

Interplanetary spacecraft are built in a spacecraft assembly facility (SAF), a clean room designed to reduce microbial contamination that could confound life detection missions or influence native ecosystems. The frigid hyperarid near-surface environment of Mars has ample hygroscopic Mg and Na salts of chloride, (per)chlorate and sulphate that may deliquesce to form dense brines, liquids with low water activity, and freezing points <0°C. The current study sought to define the climax microbial community after 6 mo of enrichment of SAF floor wipe samples in salt plains medium supplemented with 50% (w/v; ~2 M; a_w = 0.94) MgSO₄ or 20% (w/v; ~1.9 M; a_w = 0.91) NaClO₃. After 1 wk, 4 wk and 6 mo of incubation, metagenomic DNA extracts of the enriched SAF microbial community were used for high-throughput sequencing of 16S rRNA genes and subsequent phylogenetic analyses. Additionally, dozens of bacterial strains were isolated by repetitive streak-plating from the climax community after 6 mo of enrichment. Early in the enrichment, staphylococci greatly dominated and then remained abundant members of the community. However, actinobacteria succeeded the staphylococci as the dominant taxa as the cultures matured, including Arthrobacter, Brachybacterium and Brevibacterium. A diverse assemblage of bacilli was present, with Oceanobacillus being especially abundant. The SAF culture collection included representatives of Brachybacterium conglomeratum, Brevibacterium sediminis, Oceanobacillus picturae and Staphylococcus sciuri. These were characterized with biochemical and physiological tests, revealing their high salinotolerance. Shannon diversity indices were generally near 2, reflecting modest diversity at several levels of identity and the community structures were uneven throughout. However, minor members of the community seem capable of the ecosystem functions required for biogeochemical cycling. For instance, organisms capable of all the functions of the N cycle were detected. The microbial assemblage in SAFs is the most likely to be transported by spacecraft to another world. While individual microbial populations may exhibit the qualities needed for survival at the near-surface of Mars, certainly entire communities with the capacity for complete biogeochemical cycling, would have a greater chance of survival and proliferation.