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Low organic matter supply and reduced spatiotemporal heterogeneity severely constrain biodiversity in groundwater. At the aquifer scale, spatial variation in the groundwater recharge rate is considered a key factor in generating groundwater patches with distinctly different food supplies and spatiotemporal heterogeneity. Our study addressed the role of groundwater recharge in sustaining biodiversity in a phreatic aquifer by testing differences in the density and species richness of invertebrate assemblages among 11 reference sites and 13 sites artificially recharged with storm water. The vertical distribution pattern of invertebrate assemblages was examined using well clusters at one shallow water-table recharge site and one nearby reference site. Groundwater recharge elevated dissolved organic C (DOC) concentrations in groundwater and increased spatiotemporal physicochemical heterogeneity. The thickness of the vadose zone (VZT) controlled DOC input to groundwater at recharge sites but did not reduce spatiotemporal heterogeneity. The higher density and richness of invertebrate assemblages at the well-cluster recharge site than at the well-cluster reference site also was controlled by VZT, suggesting that organic matter supply was a primary factor determining biodiversity patterns in groundwater. Invertebrate density increased and species composition shifted with increasing depth below the groundwater table at the shallow water-table well-cluster site. Some taxa, including several epigean species, preferentially occurred near the water table, whereas others, including several hypogean species, colonized deeper groundwater layers.