The role of iron was evaluated with respect to the virulence of Trichomonas vaginalis in mice. Iron-supplemented and iron-depleted Diamond's trypticase–yeast extract–maltose (TYM) media were prepared by adding 360 µM of ferrous sulfate and 100 µM of 2,2′-dipyridyl. Trophozoites cultivated from normal TYM and iron-supplemented TYM media produced subcutaneous abscesses; however, trichomonads grown in an iron-deficient TYM medium failed to produce any pathology. In addition to the increased virulence of trophozoites in mice, iron affects the level of adherence and the cytotoxicity of trichomonads to HeLa cells, which are significantly reduced in trophozoites grown in iron-deficient medium. In conclusion, it is suggested that under iron-depleted conditions such as that induced by 2,2′-dipyridyl the virulence of T. vaginalis is reduced.