Carotenoid complement of rust spores: Variation among species and pathotype.

Rust fungi, which are responsible for some of the world's most destructive plant diseases, are named for the distinctive rusty colour of one or more of their spore forms. Rust spore pigments are thought to provide protection against UV radiation and oxidative stress, and may act as virulence factors. However, with the exception of daisy rust spores, the identity and relative abundance of the carotenoids in the rust spore cytoplasm have not been investigated using modern analytical methods, and little is known about the dependence of the carotenoid complement on species, pathotype, spore-colour mutations and season. We developed and validated a method to separate, identify and quantify rust carotenoids by reversed-phase high-performance liquid chromatography (RP-HPLC) combined with mass spectrometry. The carotenoids identified were lycopene, γ-carotene, β-carotene and phytoene. Rates of carotenoid degradation depended greatly on spore storage conditions, with freezing at -80 °C providing optimal stability. Carotenoid profiles of 103 isolates from 14 rust species were compared, showing that the ratio γ-carotene:β-carotene varied substantially among species. Total carotenoid content was generally lower in spring than in autumn (Sydney, Australia)-possibly due to differences in solar exposure-but the percentage of individual carotenoids was relatively stable. Among the colour mutants tested, chocolate mutants of Puccinia graminis f. sp. tritici (wheat stem rust) contained no carotenoid pigments, while albino mutants of P. striiformis f. sp. tritici (wheat stripe rust) contained only phytoene, a colourless carotenoid. We discuss our results in terms of the biogenesis and biological functions of carotenoids in rust fungi.