Potato late blight caused by the Oomycete Phytophthora infestans is the most important disease of this crop world-wide. Late blight in Colombia presents high levels of incidence and severity in the potato-growing areas, taking epidemic proportions and causing a complete lost of production when fungicides are not applied at a frequent rate. Knowledge of genotypic characteristics of plant pathogens is a main condition to design and apply control measures for crop diseases. In this research, mitochondrial haplotypes, mating type and variation level of ITS and 28S ribosomal DNA sequences were evaluated in P. infestans populations of 40 isolates obtained in potato crops from the Colombian provinces of Antioquia, Boyaca, Cundinamarca, and Norte de Santander. Results indicated that all P. infestans isolates in the four populations showed the same mitochondrial haplotype (IIa) and had mating type A1, which suggests they possibly belong to the clonal lineage EC-1. Regarding ITS and 28S rDNA analysis, all Colombian isolates presented the same sequences for both regions and had high similarity with isolates from other hosts and regions of world. These results suggest that P. infestans populations attacking potato in four provinces of Colombia are mainly clonal and there is not evidence of their sexual reproduction.