The root-knot nematode resistance (Mi) gene was screened in 25 tomato genotypes of Solanum lycopersicum, by amplification of REX-1 and Mi23 markers. Ten heterozygous tomato genotypes (Mimi), nine homozygous (MiMi) at the Mi locus and six lacking the Mi gene for resistance to root-knot nematode were identified using the marker REX-1. The results obtained with Mi23 marker confirmed the Mi gene status of the tomato genotypes, except for genotype Valouro RZ F1 that was homozygous (MiMi) and heterozygous (Mimi) at the Mi locus when using the REX-1 and Mi23 markers, respectively. The pathogenicity of Meloidogyne hispanica on the 25 tomato genotypes was assessed 60 days after inoculation with 5000 eggs on the basis of root gall index (GI) and reproduction factor (Rf). All the tomato genotypes were susceptible (excellent or good hosts), with GI > 4 and Rf > 2, except the genotype Rapit (Mimi), considered as resistant/hypersensitive (poor host). In this genotype, the nematode induced galls (GI = 4) on its roots and a small number of eggs were produced (Pf = 3085 ± 485). Significant differences in reproduction were detected between the Mi allelic conditions and genotypes within Mi allelic conditions. The increasing number of Mi alleles (0, 1 or 2) is associated with decreasing Rf, which suggests a possible dosage effect of the Mi gene. The variability observed in the Rf values for MiMi tomato genotypes may reflect an influence of the genetic background of the plants containing the Mi gene. Ten of the 25 tomato genotypes with Mi gene are commercially available. However, only Rapit can be used to control the three most common Meloidogyne spp. and inhibit the increasing of M. hispanica populations, and may have potential to be included in an integrated pest management programme. However, it is advisable to evaluate the pathogenicity of local populations of this nematode species associated with different environmental factors.