The effects of local climate parameters, seasonality and sources of air masses on the stable water isotopes [hydrogen (delta H-2) and oxygen (delta O-18)] in precipitation were investigated along an altitudinal gradient (381-1118 m a.s.l.) in southern Bohemia, Central Europe, from December 2021 to November 2023. The relationship between the observed delta H-2 and delta O-18 values was consistent with the Global Meteoric Water Line. The isotopic composition of precipitation changed with increasing altitude by -6.5 and -1.2 parts per thousand km(-1) for delta H-2 and delta O-18, respectively. The delta values differed between seasons, with the values being most enriched in summer and most depleted in winter. An analysis of the air mass trajectories using the Hybrid Single-Particle Lagrangian Integrated Trajectory Model showed that the main sources of precipitation were the North Atlantic (from 44% in spring to 70% in fall and winter), the Arctic Ocean (from 15% in summer to 38% in spring) and the Mediterranean Sea (from 12% in winter to 34% in summer). Throughout the study period, average delta values differed significantly between air masses (p < 0.05) and along the altitudinal gradient, with the most enriched values (from -56 to -37 parts per thousand for delta H-2 and from -8.2 to -5.5 parts per thousand for delta O-18) and the most depleted values (from -91 to -84 parts per thousand for delta H-2 and from -12.7 to -12.0 parts per thousand for delta O-18) occurring in the Mediterranean and Arctic air masses, respectively. However, for individual daily samples, strong correlations occurred between the delta values and air temperature (the strongest), humidity, precipitation, time of sunshine and solar radiation, while the influence of air mass directions was not significant. A stepwise regression analysis showed that most of the variation in daily delta values (43-48% and 47-52% of delta H-2 and delta O-18, respectively) was explained by the combination of air temperature and humidity, precipitation amount, and season.