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Microbes need energy to grow, reproduce, repair damage, maintain their metabolisms, and interact with their environment. Phototrophic microbes can harness the power of sunlight while chemotrophs derive energy from chemical compounds. Thermodynamic calculations can tell us whether a chemotrophic metabolic reaction will yield energy in an aqueous environment depending on fluid composition, temperature, and pressure. If the calculation reveals that energy is not available for a reaction, the reaction can be ruled out as a viable metabolic strategy in that system. Similarly, energy supplies can be quantified for energy-yielding reactions to generate hypotheses about how chemotrophic microbes harness energy in a system. Because of its usefulness for interpreting chemotroph metabolic strategies, several recent studies have quantified microbial energy supplies in natural systems and growth experiments using free and open-source software tools developed for the Water-Organic-Rock-Microbe (WORM) Portal online computing environment [1, 2, 3, 4]. The WORM Portal is an NSF-funded geochemical modeling platform for researchers, students, and the public that can be accessed for free through an internet browser. The WORM Portal comes pre-packaged with computational Jupyter notebook tools and educational demos covering a variety of topics in geobiology and geochemistry. In this presentation, we will demonstrate how the WORM Portal can be used to quantify microbial energy supplies, chemical affinities, and power (energy over time) in water samples and growth media under ambient conditions and elevated temperatures and pressures, and how you can apply the WORM Portal to quantify energy supplies in your own systems of interest. [1] Alain et al. (2022). Sulfur disproportionation is exergonic in the vicinity of marine hydrothermal vents. Environmental Microbiology, 24(5), 2210-2219. [2] Howells et al. (2025). Energetic and genomic potential for hydrogenotrophic, formatotrophic, and acetoclastic methanogenesis in surface-expressed serpentinized fluids of the Samail Ophiolite. Frontiers in Microbiology, 15, 1523912. [3] Parsons et al. "Hydrothermal Seepage of Altered Crustal Formation Water Seaward of the Middle America Trench, Offshore Costa Rica." Geochemistry, Geophysics, Geosystems 25.1 (2024): e2023GC011246. [4] Rhim et al. (2024). Mode of carbon and energy metabolism shifts lipid composition in the thermoacidophile Acidianus. Applied and Environmental Microbiology, 90(2), e01369-23. 

The availability of chemical energy supplies is fundamental to environmental and planetary habitability. However, the presence of a chemical energy supply does not guarantee the presence of microorganisms capable of consuming it. In this study, chemical energy supplies available in Yellowstone National Park (YNP) hot springs were calculated, and the results indicate that ammonia oxidation, calculated using total dissolved ammonia, is one of the major energy supplies. Nevertheless, known ammonia-oxidizers (AO) are only present in a small fraction of the hot springs tested. Where AO are present, they do not dominate the microbial communities (relative abundances <5%), even in cases where total dissolved ammonia oxidation is the richest energy supply. The AO in YNP hot springs are predominantly ammonia-oxidizing archaea (AOA), which tend to favor environments with low total ammonia (sum of NH3 and NH4+) concentrations, despite the requirement of ammonia (NH3) as a substrate. Hot spring pH and temperature determine the ratio of NH3 to NH4+ and, consequently, NH3 availability to resident AOA. In this study, total ammonia measurements were collected from YNP hot spring samples using ion chromatography in coordination with biological sampling. DNA was extracted from simultaneously collected samples for 16S rRNA gene sequencing and analysis, and for the identification of known AOA. The WORM-portal (https://worm-portal.asu.edu/) was used to speciate the total ammonia measurements into ammonia and ammonium activities. By performing speciation calculations, we identified a potential lower limit for substrate (NH3) availability and a potential upper limit for NH4+ concentrations for the YNP hot spring AOA. Thus, the niche for AOA across YNP hot springs is dictated by the form of the total dissolved ammonia present, not by the energy supply available for total dissolved ammonia oxidation.