Lentil is an important winter-sown legume for semi-arid and temperate areas, food consumption of seed is considerable in several countries of Mediterranean Basin. In Central and Southern Italy different lentil landraces are cultivated within specific marginal areas and commercialized with a recognizable geographical indication of origin. Considering the extensive germplasm and the economic importance of lentil in several rural areas, detailed knowledge of existing genetic variation from different regions is the first important step both for conservation and exploitation of genetic resources, allowing to develop breeding programs. In field experiments over three cropping seasons (2002-2005), 25 lentil accessions from Plant Genetic Institute of National Research Council (Bari), representing part of a large germplasm collection from different areas, were carried out at the University of Reggio Calabria in order to characterize the agronomic performances in a semi-arid environment and to study genetic variability. For this purpose, 10 AFLP primer combinations and 6 SSR markers were used. The agronomic results highlighted the influence of different climatic conditions on phenological, biometrical and yielding traits. A considerable production level of lentil (2,55 t ha-1) and a low yield variability in the three years was observed, showing the high adaptability of the germplasm tested to semi-arid environment. The earliness and the plant height appeared as the most important traits negatively correlated to grain yield; in particular the earliness was confirmed as suitable mechanism of escape from abiotic stress. Genetic characterization showed that a few number of microsatellites and primer combinations are able to provide significant insights on genetic diversity combining the 25 accessions in 3 large clusters that mainly mirror their geographic origin. Principal Component Analysis that consider genetic as well as morphological and agronomic data allowed the discrimination of groups with higher similarity and the Laird variety was far from all the other accessions. The macrosperma accession MG 106401 from Algeria results the closest to Laird because of their late-flowering. The fingerprinting of these elite genotypes and the evaluated suitability of agronomic traits could usefully represent strategic tools for new targeted lentil breeding programs in Mediterranean area.