Eucalyptus globulus Labill. is widely cultivated and used by industry but its productivity is currently restricted by drought events, so research focussing on supporting programs to breed adapted germplasm is needed. In the present work we monitored severity of acute drought stress (7 and 11 days after water withholding) and relief (2 h and 3 days after rewatering) by quantifying several biochemical markers of oxidative stress and DNA methylation patterns in leaves. Water withholding imposed a mild oxidative stress as estimated by redox shifts in the major antioxidant pools and increased lipid peroxidation. At the DNA level, global 5-methylcytosine distribution increased over the dehydration period especially in vascular tissue as estimated by immunolocalisation. Using methylation-sensitive RAPD analysis, which discriminates methylation changes occurring in specific DNA sequences, we found a high number of specific demethylation events also taking place. Immunolocalisation indicated a rapid reduction in global DNA methylation 2 h after rehydration; however, a large number of de novo methylation events were still detected by methylation-sensitive RAPD. These events were associated with decreased lipid peroxidation and high cellular GSH pools relative to unstressed plants. Our results indicate the parallel induction of redox and complex DNA methylation changes occurring during stress imposition and relief.