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Florestina is shown to consist of six annual species occurring mostly in arid and semiarid regions of Mexico. Florestina species are morphologically similar and consequently phylogenetic relationships within the genus are poorly understood. We present a phylogenetic study based on morphological characters, DNA sequences of nuclear non-coding spacers (ETS and ITS) and chloroplast non-coding spacers (rpl32-trnL and trnC-petN). The ETS and ITS spacer-based phylogenies allowed several well-supported conclusions: (1) the genus Florestina is monophyletic and Palafoxia is its closest relative; (2) Florestina latifolia and F. platyphylla form a strongly supported clade; (3) four taxa that are morphologically very similar, F. liebmannii, F. pedata, F. simplicifolia, and F. tripteris, are phylogenetically closely related and based on the sequence data we suggest that these should be recognized as only two species, one comprising F. pedata and F. simplicifolia, which shows wide morphological variation throughout its distributional range; and the other comprising F. liebmannii and F. tripteris; (4) F. lobata and F. purpurea are species very distinct from the remainder of the species in Florestina. Our phylogenetic analyses suggest that hybridization and introgression may be involved in the evolutionary history of Florestina.