The vegetation of the southern fields in the Atlantic Forest underwent several changes during the ice ages of the Lower and Middle Holocene period . The decrease in temperature and humidity during these periods favored the expansion of grassy vegetation areas, which formed large continuous areas in the southern Brazilian states . This condition favored the wide distribution of typical plant species of grassy vegetation areas and, according to Mayle et al. , among these species, the most abundant in these regions were those from the Poaceae and Asteraceae families; the latter being the family of A. flaccida. Behling  proposed that in the glacial periods during the Lower and Middle Holocene, the Araucaria Forest was isolated in refuges located in the river valleys. It is not difficult to conclude that these valleys may have also acted as a barrier to gene flow between populations of the species from grassy vegetation fields. Considering this context, it is possible that the genetic proximity observed between the A. flaccida from Guarapuava, Xanxerê, and Campos Novos populations is partly due to them having been somewhat continuous in the past. When analyzing the topography of the southern states of Brazil (Fig 6), it is possible to observe that such a situation was probable, because among these populations there are no river valleys that could have maintained forests acting as a barrier for the populations during cold and dry periods in the past. Additionally, the Uruguay River valley (Fig 6) may have acted as an important barrier for gene flow among the populations of Paraná/Santa Catarina and Rio Grande do Sul.