The EU target to replace 10 percent of transport fuels by renewables by 2020 requires additional water. This study calculates water footprints (WFs) of transport modes using first generation bio-ethanol, biodiesel or bio-electricity and of European transport if 10 percent of transport fuels is bio-ethanol. Results are compared with similar goals for other regions. It is more efficient to use bio-electricity and bio-ethanol than biodiesel. Transport by train or car using bio-electricity (8–19 and 11–13 litres per passenger km) is more water efficient than transport by car (36–212) or airplane (65–136) using bio-ethanol. For cars, there is a factor of ten between water-efficient cars using bio-ethanol and water-inefficient cars using biodiesel. Biofuel-based freight transport is most water-efficient by ship or train; airplanes are least efficient. Based on first generation biofuels, the EU goal for renewable transport energy results in a WF of 62 Gm3 per year, 10 percent of the current WF. Differences in transport energy use and in production systems result in a broad range of annual transport-related WFs: from 60 m3 per capita in Bulgaria to 500 m3 in Finland. If similar targets are applied in other regions, the additional WF of North America and Australia will be 52 percent of the present regions WFs. The global WF for biofuel-based transport in this scenario will be 9 percent of the current global WF. Trends towards increased biofuel application enhance the competition for freshwater resources.