Legacy per- and polyfluoroalkyl substances (PFASs), mainly long-chain ones, are being substituted by short chain homologues and novel fluorinated alternatives, whereas their occurrence, spatial distribution, sources and substitution characteristics are not well understood. For the first time, the occurrence and replacing trend of the legacy and novel fluorinated alternatives were examined in the surface water from Fen and Wei rivers, which are the two major rivers located in the underdeveloped and ecology vulnerable areas of central and western China. Results showed that the contamination of legacy and emerging PFASs in both river basins was widespread, and mainly caused by industrial activities. In both rivers, perfluorohexane sulfonic acid (PFHxS), as a substitute for perfluorooctane sulfonic acid (PFOS), was predominant in the urban areas. In the Fen River, more substitutes of PFOS, such as 6:2 fluorotelomer sulfonate (6:2 FTS) and 6:2 chlorinated polyfluorinated ether sulfonate (6:2 Cl-PFESA), were distinct, while significant replacing for PFOA with short-chain perfluoroalkyl carboxylic acids (C4-C7) and Ammonium salt of 4,8-dioxa-3H-per-fluorononanoate (ADONA) was observed in Wei River. Besides, advanced oxidation experiment indicated that there were unknown PFASs which could be the precursors of perfluorocarboxylic acids in Wei River. Isomeric analyses indicated that there was contribution of telomerization related sources for PFOA in both rivers, whereas PFOS was mainly from ECF. The estimated total mass discharge of PFASs derived from Wei and Fen River to the Yellow river were 239 and 62.6 kg/year, respectively.