Reclaimed water (RW), treated residential and municipal wastewater repurposed for beneficial use, has emerged as an alternative water source amid growing global pressures on freshwater resources. However, RW commonly contains elevated nutrients, heavy metals, pharmaceuticals, and microplastics that threaten aquatic ecosystems when discharged. Constructed wetlands (CWs), engineered ecosystems designed to mimic natural wetlands (NWs), are often incorporated into wastewater treatment plants as a tertiary water treatment step. However, the extent to which CWs replicate the chemical characteristics and biogeochemical processes of NWs remains poorly understood. We compared chemical signatures and microbial nutrient acquisition between CWs and NWs and hypothesized that (1) chemical signatures and microbial extracellular enzyme activity (EEA) differ between CWs and NWs, and (2) EEA varies in response to water chemistry. We sampled 9 CWs and 7 NWs during the summer rainy season across north-central Florida, USA. We measured physicochemical parameters in situ and collected water samples to quantify carbon, nitrogen, and phosphorus forms, dissolved organic matter (DOM) quality, and microbial EEA. Multivariate analysis revealed distinct chemical signatures between wetland types (R² = 0.50, p < 0.001). CWs showed high phosphorus and carbon levels and fresh, microbially derived DOM, whereas NWs were characterized by more humified DOM. EEA differed among wetland types, reflecting differences in water chemistry and DOM characteristics. CWs showed higher organic matter-degrading EEA (17,016 ± 11,617 nmol h⁻¹ L⁻¹), indicating greater microbial investment in organic matter decomposition. NWs showed higher carbon-, nitrogen-, and phosphorus-acquiring EEA (70.7 ± 48.7, 201 ± 78.1, and 527 ± 102 nmol h⁻¹ L⁻¹, respectively), indicating greater microbial investment in nutrient acquisition. Our findings suggest that CWs may not fully replicate chemical characteristics or biogeochemical processes of NWs, highlighting the need to understand the mechanisms underlying nutrient treatment performance as CWs and RW are increasingly used in global water conservation efforts.