Trichothecenes are terpene-derived secondary metabolites produced by multiple genera of filamentous fungi, including many plant pathogenic species of Fusarium. These metabolites are of interest because they are toxic to animals and plants and can contribute to pathogenesis of Fusarium on some crop species. Fusarium graminearum and F. sporotrichioides have trichothecene biosynthetic genes (TRI) at three loci: a 12-gene TRI cluster and two smaller TRI loci that consist of one or two genes. Here, comparisons of additional Fusarium species have provided evidence that TRI loci have a complex evolutionary history that has included loss, non-functionalization and rearrangement of genes as well as trans-species polymorphism. The results also indicate that the TRI cluster has expanded in some species by relocation of two genes into it from the smaller loci. Thus, evolutionary forces have driven consolidation of TRI genes into fewer loci in some fusaria but have maintained three distinct TRI loci in others.