Unveiling Hidden Microbial Diversity in Mars 2020 Mission Assembly Cleanrooms with Molecular Insights into the Persistent and Perseverance of Novel Species Defying Metagenome Sequencing

Abstract:

Industrial cleanrooms, used for the assembly of space mission components, maintain rigorous cleaning regimens and nutrient-poor environments, resulting in low but persistent microbial loads. Detection of rare extremophiles is difficult in such environments because they are resistant, sparse, and DNA extraction and sequencing technologies are not advanced enough. Traditional metagenomics approaches are missing the rare species, as they can neither break up hard microbial cells nor separate small amounts of DNA. Additionally, there is no database reference for the new extremophiles. Within six months of monitoring Mars 2020 mission cleanrooms, 182 bacterial strains from 19 families have been identified: 14 novel Gram-positive species and eight spore-formers. ANI values show low similarity to known species and range from 77% to 94%. The bacteria were successfully cultured despite being present at an abundance of 0.001% in sequencing data. Functional studies showed capabilities such as nitrogen cycling, carbohydrate breakdown, and radiation resistance. Moreover, 12 biosynthetic gene clusters, including those associated with the production of ectoine and $\epsilon$-poly-L-lysine, hint at biotechnological applications. These results highlight the unappreciated microbial diversity in cleanrooms and point to the necessity of advanced detection methods to discover extremophiles that may have utility for biotechnology and guide future space exploration.

Detailed scripts and procedures are given on the Wiki page

Our analysis is distributed into three major parts

1. Discovery of novel species with extreme adaptations
2. Low abundance and persistence of hidden microbial communities in NASA cleanroom
3. Biotechnological prospects of cleanroom isolated strains

Here we have the overview of the workflow