Dissolved organic matter (DOM) is integral to fluvial biogeochemical functions, and wetlands are broadly recognized as substantial sources of aromatic DOM to fluvial networks.Yet how land use change alters biogeochemical connectivity of upland wetlands to streams remains unclear. We studied depressional geographically isolated wetlands on the Delmarva Peninsula (USA) that are seasonally connected to downstream perennial waters via temporary channels. Composition and quantity of DOM from 4 forested, 4 agricultural, and 4 restored wetlands were assessed. Twenty perennial streams with watersheds containing wetlands were also sampled for DOM during times when surface connections were present versus absent. Perennial watersheds had varying amounts of forested wetland (0.4-82%) and agricultural (1-89%) cover. DOM was analysed with ultraviolet-visible spectroscopy, fluorescence spectroscopy, dissolved organic carbon (DOC) concentration, and bioassays. Forested wetlands exported more DOM that was more aromatic-rich compared with agricultural and restored wetlands. DOM from the latter two could not be distinguished suggesting limited recovery of restored wetlands;DOM from both was more protein-like than forested wetland DOM. Perennial streams with the highest wetland watershed cover had the highest DOC levels during all seasons; however, in fall and winter when temporary streams connect forested wetlands to perennial channels, perennial DOC concentrations peaked, and composition was linked to forested wetlands. In summer, when temporary stream connections were dry, perennial DOC concentrations were the lowest and protein-like DOM levels the highest. Overall, DOC levels in perennial streams were linked to total wetland land cover, but the timing of peak fluxes of DOM was driven by wetland connectivity to perennial streams.Bioassays showed that DOM linked to wetlands was less available for microbial use than protein-like DOM linked to agricultural land use. Together, this evidence indicates that geographically isolated wetlands have a significant impact on downstream water quality and ecosystem function mediated by temporary stream surface connections.This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.