Weed control is essential for a successful establishment and growth of tree seedlings in former agricultural land. Weed control methods are effective but can be costly in terms of time, damage to nontarget vegetation, or increased soil erosion. Alternatively, some living mulches can exclude undesirable vegetation, protect the soil, compete minimally with associated trees, and supplement soil nitrogen, but there is a lack of knowledge on living mulch systems in Mediterranean afforestation. Thus, the objective of the present study was to evaluate the effects on Holm oak (Quercus ilex L.), mastic tree (Pistacia lentiscus L.), wild olive (Olea europaea L. var. sylvestris Brot.) and terebinth (Pistacia terebinthus L.) seedlings of wrinkled medick (Medicago rugosa Desr.) mulch. Survival, growth, photosynthesis, foliar nutrient and soil parameters were measured during the first year. 36 months after planting, seedlings in the living mulch had survival rates of between 60% for mastic tree and 8.3% for Holm oak, compared with survival rates of 70% in the mechanical treatment for mastic tree and 2% for Holm oak. Photosynthesis and foliar nutrient concentrations were improved by the living mulch treatment. The soil under the living mulch had higher CEC, soil organic matter levels and nitrogen content in comparison to the cultivated soil. The response of living mulch differs between species and environmental conditions but our study suggests a positive effect due to soil protection. Living mulch may be a promise alternative for use in Mediterranean afforestation programs.