We investigated the influence of different levels of osmotic stress on growth and development in selected wild almond species(eight Prunus spp.) grown in vitro. The study, while endorsing the efficacy of in vitro screening of auxiliary buds of wild almond for osmotic stress tolerance, showed species variability in its response to osmotic stress. Osmotic stress reduced growth and development of all the species. However, the putative tolerant Prunus spp. showed better performance than the putative susceptible genotypes. On average there was an 80% decrease in shoot dry weight at-1.2 MPa. Reduction in shoot weight was more common in osmotic stress-susceptible species in the section labeled ‘Euamygdalus’. The tolerant Prunus species produced smaller changes in biochemical responses than the sensitive cultivars for malondialdehyde content, catalase activity, relative permeability of protoplast membranes, and net photosynthetic rate. The tolerant species maintained cell integrity better than drought sensitive species. Wild almond species in the section labeled ‘Spartioides’(Prunus arabica(Olivier) Neikle, Prunus glauca (Browicz) A.E. Murray, Prunus scoparia Spach) and ‘Lycioides’(Prunus lycioides Spach, Prunus reuteri Bossi. et Bushe) were best adapted to osmotic stress. Increase in chlorophyll concentration and leaf thickness under high osmotic stress can be considered as preliminary selection parameters for osmotic stress tolerance in Prunus spp. The study confirmed the efficacy of the in vitro method for screening of large number of genotypes for osmotic stress tolerance in wild almond species. We investigated the influence of different levels of osmotic stress on growth and development in selected wild almond species (eight Prunus spp.) Grown in vitro. The study, while endorsing the efficacy of in vitro screening of auxiliary buds of wild almond for osmotic stress tolerance , showed the species variability in its response to osmotic stress. However, the putative tolerant Prunus spp. showed better performance than the putative susceptible genotypes. On average there was an 80% decrease in shoot dry weight at-1.2 MPa. Reduction in shoot weight was more common in osmotic stress-susceptible species in the section labeled ’Euamygdalus’. The tolerant Prunus species produced smaller changes in biochemical responses than the sensitive cultivars for malondialdehyde content, catalase activity, relative permeability of protoplast membranes, and net photosynthetic rate. The tolerant species maintained cell integrity better th an drought sensitive species. Wild almond species in the section labeled ’Spartioides’ (Prunus arabica (Olivier) Neikle, Prunus glauca (Browicz) AE Murray, Prunus scoparia Spach) and ’Lycioides’ (Prunus lycioides Spach, Prunus reuteri Bossi. et Bushe ) in best adapted to osmotic stress. Increase in chlorophyll concentration and leaf thickness under high osmotic stress can be considered as preliminary selection parameters for osmotic stress tolerance in Prunus spp. The study confirmed the efficacy of the in vitro method for screening of large number of genotypes for osmotic stress tolerance in wild almond species.