The air in subterranean karst cavities is often depleted in methane (CH₄) relative to the atmosphere. Karst is considered a potential sink for the atmospheric greenhouse gas CH₄ because its subsurface drainage networks and solution-enlarged fractures facilitate atmospheric exchange. Karst landscapes cover about 14% of earth’s continental surface, but observations of CH₄ concentrations in cave air are limited to localized studies in Gibraltar, Spain, Indiana (USA), Vietnam, Australia, and by incomplete isotopic data. To test if karst is acting as a global CH₄ sink, we measured the CH₄ concentrations, δ¹³C_CH4, and δ²H_CH4 values of cave air from 33 caves in the USA and three caves in New Zealand. We also measured CO₂ concentrations, δ¹³C_CO2, and radon (Rn) concentrations to support CH₄ data interpretation by assessing cave air residence times and mixing processes. Among these caves, 35 exhibited subatmospheric CH₄ concentrations in at least one location compared to their local atmospheric backgrounds. CH₄ concentrations, δ¹³C_CH4, and δ²H_CH4 values suggest that microbial methanotrophy within caves is the primary CH₄ consumption mechanism. Only 5 locations from 3 caves showed elevated CH₄ concentrations compared to the atmospheric background and could be ascribed to local CH₄ sources from sewage and outgassing swamp water. Several associated δ¹³C_CH4 and δ²H_CH4 values point to carbonate reduction and acetate fermentation as biochemical pathways of limited methanogenesis in karst environments and suggest that these pathways occur in the environment over large spatial scales. Our data show that karst environments function as a global CH₄ sink.