Exploration of the Fungal Cause of Facial Eczema (Pithomyces chartarum) Using Modern Molecular Techniques

Abstract

Facial eczema, a disease caused by the ingestion of toxigenic spores produced by Pithomyces chartarum, is extremely costly for New Zealand’s agriculture sector. Much effort has been made to prevent disease outbreaks; however current preventative measures are often ineffective, costly, and labour intensive. This is both due to limitations with the methods themselves, and the use of spore numbers to estimate facial eczema risk. The issue with the latter is that it detects P. chartarum presence too late for the use of preventative measures to be effective. Despite the need for changes in facial eczema management, very little research has been done into P. chartarum and how it produces the facial eczema causing toxin, sporidesmin, using modern molecular tools. To address this problem, we developed a polymerase chain reaction (PCR) based detection system, explored the relationship between gene expression and toxin production in P. chartarum, and developed a ddPCR-based assay to aid further transcriptomic study. The generation of genomic and transcriptomic data allowed for exploration of a common assumption, that the biosynthetic gene clusters which produce ETP toxins, like sporidesmin, underwent horizontal gene transfer. The PCR detection system we devised is specific to toxigenic P. chartarum, as it targets genes (Spd 1, Spd 5, Spd 13, Spd 17, and Spd 20) in the sporidesmin biosynthetic cluster. The transcriptomic data suggests that the relationship between toxin production and the expression of the P. chartarum biosynthetic cluster is different to that of other characterised ETP biosynthetic clusters, a trend which can be further explored using the ddPCR assay. The structure of the sporidesmin biosynthetic cluster and the phylogenetic distribution of ETP producing species both support the theory of horizontal ETP gene cluster transfer. The development of an effective, fast, and scalable detection system for toxigenic P. chartarum can be used to optimise the timing of preventive measures and revive previously unfeasible measures. The genomic and transcriptomic information presented in this study is useful in both understanding P. chartarum as the causal agent of facial eczema, and in addressing evolutionary questions which broaden our understanding of fungal genomics.