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Reference evapotranspiration (ET0) plays an important role in studies on hydrological cycles and environmental change. In present research, the temporal changing characteristics of climatic variables, as well as growing season ET0 and attribution analysis of ET0 trend from 1961 to 2016 were investigated at 99 meteorological stations across the Western-Sichuan Plateau, Sichuan Basin, Yunnan-Guizhou Plateau, and Guangxi Basin in Southwest China (SC). The results showed that the change point for ET0 series was detected in 1996 by the Cramer's test method. Growing season ET0 declined significantly (P < 0.01) by 10.25 mm/decade during 1961-1996, while it increased significantly (P < 0.05) during 1997-2016 by 8.12 mm/decade. Maximum temperature (T-max) and minimum temperature (T-min) exhibited a significant increasing trend during 1961-2016, while relative humidity (RH), wind speed (WS) and sunshine duration hours (SD) showed a significant downward trend in SC. ET0 was highly sensitive to RH, followed by SD, T-max T-min and WS, whilst the sensitivity of ET0 to climatic variables fluctuated during the growing season. The results of attribution analysis showed that the decline in SD as well as WS allocated the significant decrease in ET0, offsetting the impact of increased temperature during 1961-1996. However, decreased RH and increased air temperature commonly reversed the trend in ET0 during 1997-2016 in SC. The contribution rate of each climatic variable exhibited great variations both temporally and spatially due to the differences in geographical location and climatic conditions. The increase in ET0 will result in higher water consumption for the growth of crops, especially for Westem-Sichuan Plateau and Yunnan-Guizhou Plateau, appeared to have experienced a more significant increase (P < 0.05) from 1997 to 2016. The results can provide a guideline for the regional agricultural production administration and allocation of water resources in the context of ongoing climate change.