Abstract: Reference crop evapotranspiration (ET0) is a basic parameter for formulating irrigation systems and a necessary variable for drought assessment. In order to comprehensively explore the temporal and spatial variation and the main causing factors of ET0 in the humid subtropical regions of Central and Southern China, we calculated ET0 using Penman-Monteith method. The Mann-Kendall test, Pettitt, sliding t and cumulative anomaly mutation method were used to explore the variation of meteorological elements and ET0. The Pearson correlation analysis was applied for the analysis of driving factors of ET0 based on the daily meteorological data of 246 stations in the region, together with atmospheric circulation indices from 1961 to 2020. Results indicated that Tmean, Tmin, and Tmax showed an upward trend (0.028, 0.029, 0.019 ℃/a), while n, us, and RHmean showed a fluctuating downward trend (-0.018 h/year, -0.005 m/(s·year), and -0.052 %/year). The mutation points of meteorological elements in sequence were in 1996, 1997, 1996, 1980 and 1988, 1981 and 1988, 2002.The overall trend of ET0 decreased with a tendency rate of -0.45 mm/year. A significant decrease in ET0 in summer would lead to a decrease in annual ET0. The spatial distribution of ET0 at the annual and seasonal scales was relatively complex. The correlation coefficients between ET0 and NAHAI, NANRP, APVA, NAPVA, PPVI, AEPVI and AO, AAO, NAO, EM, TSN, SOI were -0.86~0.85 and -0.07~0.09, respectively. The decline trend of ET0 was reflected in time and space, with a close relation to the changes of meteorological elements. The Pacific polar vortex intensity index (PPVI) was regarded as the most relevant factor affecting the change of ET0. The above conclusions are of great significance to provide scientific support of formulating irrigation systems.