Current technologies for bioethanol production rely on the use of freshwater for preparing the fermentation media and use yeasts of a terrestrial origin. Life cycle assessment has suggested that between 1,388 to 9,812 litres of freshwater are consumed for every litre of bioethanol produced. Hence, bioethanol is considered a product with a high-water footprint. This paper investigated the use of seawater-based media and a novel marine yeast strain ‘Saccharomyces cerevisiae AZ65’ to reduce the water footprint of bioethanol. Results revealed that S. cerevisiae AZ65 had a significantly higher osmotic tolerance when compared with the terrestrial reference strain. Using 15-L bioreactors, S. cerevisiae AZ65 produced 93.50 g/L ethanol with a yield of 83.33% (of the theoretical yield) and a maximum productivity of 2.49 g/L/h when using seawater-YPD media. This approach was successfully applied using an industrial fermentation substrate (sugarcane molasses). S. cerevisiae AZ65 produced 52.23 g/L ethanol using molasses media prepared in seawater with a yield of 73.80% (of the theoretical yield) and a maximum productivity of 1.43 g/L/h. These results demonstrated that seawater can substitute freshwater for bioethanol production without compromising production efficiency. Results also revealed that marine yeast is a potential candidate for use in the bioethanol industry especially when using seawater or high salt based fermentation media.