Premise of research. Interspecific hybridization was hypothesized to explain the remarkable floral phenotype variation found in overlapping areas of distribution for several Anacyclus species. We aimed to investigate genome size in natural populations of three diploid Anacyclus species with special emphasis in their contact areas to explore patterns of variation as additional evidence supporting current hybridization in these areas.

Methodology. Flow cytometry was used to estimate the genome size of 564 individuals of the species complex of A. clavatus, A. homogamos, and A. valentinus from 30 sites. Additionally, genome size variation of 173 first-generation synthetic hybrids between these three species was also studied and compared with the estimates obtained in sympatric sites.

Pivotal results. Differences in genome size between A. clavatus and A. valentinus were significant in nonoverlapping areas of species distribution, whereas in overlapping areas, the variation increased, preventing a clear differentiation between species. In sympatric sites of A. clavatus and A. valentinus, individuals with intermediate genome sizes between them were also observed and were significantly similar to those obtained from the first-generation experimental hybrids between these species. Genome sizes of A. clavatus and A. homogamos did not differ enough to allow discrimination between these species.

Conclusions. The patterns of genome size variation observed in sympatric populations of A. clavatus and A. valentinus support the occurrence of current gene flow between these species and the existence of contact areas in overlapping distribution areas where phenotypic and genomic variation increases.