Earth’s largest aquifer ecosystem resides in igneous oceanic crust, where chemosynthesis and water-rock reactions provide the carbon and energy that support an active deep biosphere. The Calvin Cycle is the predominant carbon fixation pathway in cool, oxic, crust; however, the energy and carbon metabolisms in the deep thermal basaltic aquifer are poorly understood. Anaerobic carbon fixation pathways such as the Wood-Ljungdahl pathway, which uses hydrogen (H2) and CO2, may be common in thermal aquifers since water-rock reactions can produce H2in hydrothermal environments and bicarbonate is abundant in seawater. To test this, we reconstructed the metabolisms of eleven bacterial and archaeal metagenome-assembled genomes from an olivine biofilm obtained from a Juan de Fuca Ridge basaltic aquifer. We found that the dominant carbon fixation pathway was the Wood-Ljungdahl pathway, which was present in seven of the eight bacterial genomes. Anaerobic respiration appears to be driven by sulfate reduction, and one bacterial genome contained a complete nitrogen fixation pathway. This study reveals the potential pathways for carbon and energy flux in the deep anoxic thermal aquifer ecosystem, and suggests that ancient H2-based chemolithoautotrophy, which once dominated Earth’s early biosphere, may thus remain one of the dominant metabolisms in the suboceanic aquifer today.