Abstract EANA2025-127

The confidence that oceans of Icy Moons such as Europa and Enceladus meet conditions favourable for life to persist is reasonably high. These habitable conditions are necessarily informed by life on Earth and defined by the psychochemical limits within life as-we-know-it can persist. In particular, some terrestrial environments that share physicochemical characteristics with these exoenvironments are considered analogs, and the study of their communities, interactions, and functional capabilities serves to constrain the habitability potential of distant worlds. Also, extreme environments suit as natural reservoirs of biological models that can be tested under putative exoenvironments conditions.

As we move towards the post-genomic era, metagenomics is widely used to approach the study of these functional analogue environments. Similarly, whole-genome sequence of isolates is a common strategy to decipher the potential functions a microorganism can carry, which can later assist their lab-based cultivation under different conditions. However, the relevance of these functions in the context of the icy moons is scattered, and no accessible single platform allows for the annotation of astrobiology-relevant traits.

To ease the exploration of the astrobiological potential of these (meta)genomes, we developed Enceladupedia, a user-friendly web-based annotation tool that compiles functional features (genes) associated with the most favourable metabolisms and physiological traits relevant to the exooceans’ physicochemical conditions of Europa and Enceladus.

Enceladupedia builds upon a manually curated catalogue of 348 marker genes included in sixteen energy metabolic modules prioritized by their likelihood in each icy moon scenario, coupled with 256 genes linked to physiological adaptations, clustered into eleven categories, relevant to the most likely environmental stressors in the moons’ oceans such us starvation, osmotic stress, or high hydrostatic pressures. Key metabolic pathways identified include hydrogenotrophic methanogenesis and acetogenesis, which are particularly viable for Enceladus, while methane oxidation and sulfate reduction may be dominant in Europa's potentially oxidised ocean. Metal cycling might be favourable in both moons.

The Enceladupedia annotation workflow is fed by previously assembled sequences, and then, the pipeline performs Prodigal to detect open reading frames. These predicted protein sequences are then mapped against a custom target gene database via DIAMOND blastp, ensuring efficient identification of functionally relevant hits. The results are synthesized into both tabular outputs and interactive visualizations, including heatmaps showing module completeness and likelihood scores for Enceladus and Europa scenarios, as well as normalized gene abundance profiles. This approach not only streamlines functional annotation for astrobiological research but also enables intuitive comparative analysis across samples, guiding hypothesis generation about potential biosignature presence and community structure in metagenomes, and helps identify potential biological models for astrobiology lab-based experiments.

Enceladupedia’s web-based design emphasizes accessibility for non-bioinformaticians, allowing the community to leverage genomics tools, bridging theoretical astrobiology and practical bioinformatics applications. As return data from missions like Europa Clipper and JUICE become available, Enceladupedia’s flexible interface and updatable database structure aim to remain a useful resource for the evolving landscape of astrobiological studies.