A molecular and morphological phylogenetic analysis of South American Podocominae is herein performed. Taxa of both South American and Australasian Podocominae as well as members of other subtribes of Astereae were included in the analyses. Sequence data from the nuclear internal transcribed spacer (ITS), and the plastid trnL–trnF intergenic spacer regions were used in the molecular analysis to assess relationships among the South American genera of Podocominae. The trnL–trnF intergenic spacer provided only six informative characters. In contrast, the ITS region was more variable. Our results showed that: 1) neither the South American genera of Podocominae nor the whole subtribe Podocominae are monophyletic; 2) South American and Australasian genera, initially placed in the subtribe Podocominae on the basis of morphological characters, show no evidence of close relationship in our phylogenetic analyses based on molecular data; 3) Our results do not confirm Podocoma in its current concept as monophyletic; 4) Asteropsis, Inulopsis, Microgyne, and Sommerfeltia form a close assemblage; 5) Microgyne is monophyletic when molecular and morphological phylogenetic analyses are performed, thus supporting previous morphology based studies; and 6) the morphological characters used to define the subtribe Podocominae are homoplastic in the context of the present morphological and DNA sequence based phylogenetic analyses.