Fusarium crown rot (FCR), caused by various Fusarium species, is a chronic disease of cereals in many semi-arid regions worldwide. To clarify what effects drought-stress may have on FCR development, visual assessment, histological analysis and quantitative PCR were used to analyse the infection process of F. pseudograminearum in barley. This study observed for the first time that the severity of FCR symptom reflects the quantity of pathogens in infected tissues of barley under both drought-stressed and well-watered conditions. Drought-stress prolongs the initial infection phase but enhances the proliferation and spread of Fusarium pathogens after the initial infection phase. Under drought-stressed conditions, the invading hyphae were frequently observed to re-emerge from stomata and invade again the surrounding epidermis cells. Under the well-watered conditions, however, very few hyphae re-emerged from stomata and most infection was caused by hyphae intracellularly grown. It was also observed that drought-stress increased the length and density of trichomes dramatically especially in the susceptible genotypes, and that the length and density of trichomes were positively related to fungal biomass of F. pseudograminearum in plants.