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International audience; Ascochyta blight, caused by Ascochyta fabae (teleomorph: Didymella fabae) has decreased faba bean production in Tunisia and worldwide. The teleomorph has recently been observed in Tunisia, raising new questions of how to control this major disease. Isolates (317) of the pathogen were obtained between 2011 and 2013, from four geographical regions in Tunisia (Beja, Bizerte, Jendouba and Tunis). The 240 isolates obtained in 2012 were compared for mating type frequency and genetic variation by using ten polymorphic SSR markers. Of these isolates, MAT1-2 was more common (2:1) in Tunisia than MAT1-1, but this ratio can change according to population. Low to high genetic variation was detected between locations and among pathogen populations. Beja showed the greatest genotypic richness (R = 0.42), followed by Tunis (R = 0.25), Bizerte (R = 0.13), and Jendouba (R = 0.11). Indices of association (I-A) and Rd were significantly different from 0 in all the populations, suggesting high multilocus linkage disequilibrium and confirming clonal populations. Population structure of the isolates was inferred using Bayesian analyses, Principal Component Analysis (PCA), and Minimum Spanning Networks, which all revealed that the populations from each location were not distinct. Evaluating changes in seasonal genetic diversity showed low to high variances of F-ST values between the two cropping seasons in all regions. However, the PCA analysis failed to separate the A. fabae isolates sampled during the two successive seasons into two groups, indicating that these populations did not constitute distinct genetic groups. These results suggest that gene flow was limited among populations, even those separated by short geographic distances. Future studies should enlarge the number of samples of representative populations, to overcome the limitations of a small sample size and to provide a more accurate assessment of A. fabae population structure.