The aim of the present study was to estimate the molecular prevalence of T. gondii along various segments of the genital tract of confirmed chronically infected ewes. Genital tracts were collected from 42 ewes; meat samples from the same ewes were previously confirmed positive for T. gondii DNA. The whole DNA was extracted from 4 parts of the genital tract (ovary, horns, body of the uterus and vagina). PCR was used to amplify a 114 base-pairs of T. gondii B1 gene. For all studied samples, 95.2 % had at least one infected genital part. Toxoplasma gondii infection was confirmed by sequencing 20 amplicons randomly chosen. The majority of infected animals has 4 T. gondii-infected genital anatomical parts. Principal component analysis (PCA) revealed a cluster of adult animals (>24 months) with positive PCR in the ovaries and the vagina and another cluster of Barbarine animals having positive PCR in the horns and body of the uterus. General linear model confirmed PCA results and showed a significant higher prevalence of T. gondii in the ovaries and vagina of older animals (p = 0.001 and p = 0.02, respectively) and a higher prevalence of T. gondii in the horns of the uterus of Barbarine animals (p = 0.03). Toxoplasma gondii seems to highly persist along the various segments of the ewe's genital tract but further investigations are necessary to link such prevalence with the pathological implications.