Open forests are among the world’s most species-rich ecosystems but these are now suffering from strong environmental pressure. With the increasing frequency of drought events accompanying climate change, open forests are now at high risk of canopy openness, an associated increase in nutrient load, and a consequent reordering of the plant community. Based on an 11-year field manipulative experiment, we examined whether the reintroduction of traditional forest uses, in particular the regular removal of litter which has been practiced for many centuries in Europe, may mitigate eutrophication and protect the ancient plant diversity of open forests. We examined vascular plant composition in 64, 3 × 3 m plots in 32 randomized blocks, each consisted of one litter-removal and one control plot in a subcontinental thermophilous oak forest dominated by Quercus petraea (Czech Republic, Central Europe). We analyzed how regular litter removal influenced the changes in topsoil nutrients and diversity and abundances of perennial, short-lived monocarpic and open oak forest plant species after eleven years of the annual application of this management. The regular litter removal significantly mitigated eutrophication by a reducing total nitrogen, and significantly decreased the amounts of base cations Ca2+ and Mg2+ in the topsoil. The studied plant species groups responded differently. Perennial and open oak forest species significantly increased their richness and abundances in the litter-removal plots and at the same time their diversity did not change significantly depending on inter-annual weather conditions. In contrast, litter removal negatively affected abundances of short-lived monocarpic species, whose richness was also considerably influenced by weather conditions (decreased the most in years with spring droughts). Moreover, we have found a protective effect of regular litter removal on drought-sensitive open oak forest species that survived in the litter-removal plots, whereas some completely disappeared from control sites during the period of repeated drought events. Our study thus demonstrates that litter removal is a tool with high potential in maintaining biodiversity in open forests affected by climate change.