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Oil and gas development can result in natural gas migration into shallow groundwater. Methane (CH 4 ), the primary component of natural gas, can subsequently react with solutes and minerals in the aquifer to create byproducts that affect groundwater chemistry. Hydro-biogeochemical processes induced by fugitive gas from leaky oil and gas wells are currently not well understood. We monitored the hydro-biogeochemical responses of a controlled natural gas release into a well-studied Pleistocene beach sand aquifer (Canadian Forces Base Borden, Ontario, Canada). Groundwater samples were collected before, during, and up to 700 days after gas injection and analyzed for pH, major and minor ions, alkalinity, dissolved gases, stable carbon isotope ratios of CO 2 and CH 4 , and microbial community composition. Gas injection resulted in a dispersed plume of free and dissolved phase natural gas, affecting groundwater chemistry in two distinct temporal phases. Initially (i.e. during and immediately after gas injection), pH declined and major ions and trace elements fluctuated; at times increasing above baseline concentrations. Changes in the short-term were due to invasion of deep groundwater with elevated total dissolved solids entrained with the upward migration of free phase gas and, reactions that were instigated through the introduction of constituents other than CH 4 present in the injected gas (e.g. CO 2 ). At later times, more pronounced aerobic and anaerobic CH 4 oxidation led to subtle increases in major ions (e.g. Ca 2+ , H 4 SiO 4 ) and trace elements (e.g. As, Cr). Microbial community profiling indicated a persistent perturbation to community composition with a conspicuous ingrowth of taxa implicated in aerobic CH 4 oxidation as well anaerobic S, N and Fe species metabolism., (Copyright © 2019 Elsevier B.V. All rights reserved.)