Temporal and spatial distribution characteristics of agricultural water requirement are of great significance to the management of regional drought prevention, mitigation and optimal allocation of irrigation water. Guizhou is a typical region suffering from droughts all around the year and seasonal water shortage of agriculture limits the sustainable agricultural development. In order to investigate the temporal and spatial distribution of agricultural irrigation water requirement and irrigation requirement index in Guizhou Province, in this study, the effective precipitation (Pe), the crop evapo-transpiration (ETc) of rice, maize, winter wheat, oilseed rape and tobacco in different growth stages were estimated by using effective precipitation coefficient, FAO Penman-Monteith equation and crop coefficient method based on the meteorological and statistical data of planting crops area between 1953 and 2012 in Guizhou Province. The annual net agricultural irrigation water requirement (IR) and irrigation requirement index (IRI) were statistically analyzed. Results showed that IR and IRI in the 9 cities of Guizhou province were in the range of 314.48-742.57 mm and 0.34-0.68, respectively. During the last 6 decades, IR and IRI had shown fluctuation characteristics with multi peaks distribution. The peak value demonstrated a decrease trend with the prolonged fluctuation period. The temporal turning stage of IR and IRI was between 1993 and 2000. Before that stage, IR and IRI declined. IR and IRI showed seasonal characteristics of the highest IR in summer and highest IRI in winter. IR showed high value in the East and low value in the West district, while the high value of IRI was in the East and Northwest district, but the low value was in the Southwest district of Guizhou Province. 9 cities could be divided into three risk regions, and the high risk in water shortage lied in the East and Northwest district, which was the focus of regional drought management. The construction of water pond project and the drought resistance plans should be made and carried out to alleviate the problem of engineering water shortage in these regions. The increments of IR and IRI were mainly attributed to the climate change, especially to the change of precipitation. The results of crop evapotranspiration (ETc) showed that the mean ETc of rice, maize, oilseed, winter wheat rape, and tobacco were 738.1, 482.4, 367.4, 347.9, and 450.4 mm, respectively. And the IR showed that the mean IR of rice, maize, oilseed, winter wheat rape, and tobacco were 362.2, 142.4, 208.4, 213.1, and 102.6 mm, respectively. Regional water resources transferred from the southern area to the northern side could be carried out to improve the basic conditions of regional agriculture production. This study can provide support for the identification of seasonal water shortage in Guizhou Province, the layout of agricultural water conservancy project, and the adjustment of planting structure and cropping pattern.