Background and aims <p>Litter in the riparian zone can fall vertically into the stream or slide laterally into the stream after transient terrestrial retention. However, the difference in the decomposition process between these two types of litter and the influence of terrestrial<b>–</b>aquatic microbial community coalescence on litter decomposition remain unclear.</p> Methods <p>To address this gap, we used litterbags to simulate lateral litter (with 14-day terrestrial retention) and vertical litter, evaluating their decomposition differences and regulatory mechanisms.</p> Results <p>After 117&#xa0;days of stream decomposition, the cumulative mass loss of the lateral input litter was 22.76%, whereas that of the vertical input litter reached 29.73%. After entering the stream, the lateral input litter had a greater nitrogen content, dehydrogenase activity and fungal biomass than did the vertical input litter, indicating the greater involvement of microorganisms in its decomposition. However, the abundances of microbial decomposition functional pathways in the vertical input litter were greater than those in the lateral input litter, suggesting that the microorganisms in the vertical input litter presented greater decomposition abilities. Different initial microorganisms resulted in the colonization of distinct aquatic microorganisms on litter. The decomposition abilities of the initial and subsequently colonized microorganisms are significantly correlated.</p> Conclusion <p>Initial soil microorganisms may antagonize the colonization of efficiently decomposing aquatic microorganisms on litter, resulting in slower decomposition of lateral litter. The coalescence of terrestrial and aquatic microbial communities has a nonnegligible negative impact on litter by altering the microbial community composition, thereby reducing leaf litter nutrient subsidies to streams.&#xa0;</p> Graphical Abstract <p></p>