Tidal fluctuation disturbances and amplified anthropogenic activities are defining characteristics of the intertidal zones of mountainous river estuaries. The accumulation and degradation of organic matter and nutrients in the sediments result in a complex element migration and transformation dynamics. Nonetheless, microbial transformation of dissolved organic sulfur (DOS) in the intertidal sediments upon tidal fluctuation remains poorly understood. Here, by taking a representative small mountainous river estuary in southeast China as an example, we synthesize evidence describing the composition of dissolved organic matter (DOM), microbial community structure and metabolic functions in sediments of variable depths (0-80 cm) at both high and low tide via FT-ICR-MS and metagenomic approach. Labile DOM, e.g., aliphatic and proteins were more inclined to be enriched in shallow sediments (0-30 cm). Upon tidal inundation, Thaumarchaeota was verified to facilitate the accumulation of recalcitrant organic matter through the mevalonate pathway, elevating the proportion of carboxyl-rich alicyclic molecules (CRAMs) and lignins in sediments. Whereas during ebb period, the microbial production of DOS through assimilated sulfate reduction (ASR) was signally intensified, contributing to the accumulation of sulfur-containing organic matter in deeper sediments. Based on the associations between Kyoto encyclopedia of Genes and Genomes modules and DOM formulas, cobalamin biosynthesis, ASR, and cysteine biosynthesis were observed positively correlated with the accumulation of sulfur-containing organic matter. Microbial community exhibited obvious taxonomic and functional variations between flood and ebb states. Nitrososphaerta in shallow sediments (0∼30 cm) was beneficial for the production of nitrogen-containing organic matter, while Bathyarchaeota and Chloroflexota in deep sediments (70-80 cm) predominantly governed the mineralization of organic matter. We firstly provided metagenomic evidence for the microbial transformation of sulfur-containing dissolved organic matter in the intertidal zone of a mountainous river estuary, which will be key to predicting coastal carbon storage and offer an important scientific basis for formulating intertidal ecosystem management and restoration strategies.