Abstract There is increasing evidence that mobile genetic elements can drive the emergence of pathogenic fungal species by moving virulence genes horizontally. The 14 kbp ToxhAT transposon has been shown to be moving the necrotrophic effector, ToxA, horizontally between fungal species that infect Triticum aestivum (wheat), namely Parastagonospora nodorum , Pyrenophora tritici-repentis , and Bipolaris sorokiniana . All three species utilise the ToxA protein to infect wheat. Previous genomic evidence found ToxhAT in distinct chromosomal positions in two isolates of B. sorokiniana , indicating that the transposon is still active in this species. Here we confirm the movement of ToxhAT using long-read Nanopore MinION sequencing of eight novel and one previously published B. sorokiniana isolates. One event of independent transposition of ToxhAT was observed, and target site duplications of “TA” were identified, confirming this was an autonomous movement facilitated by a yet unidentified transposase. Whole genome analysis revealed that ToxhAT is a passenger embedded in a much larger, conserved 170–196 kbp mobile genetic element. This element, termed Sanctuary , belongs to the newly described Starship transposon superfamily. This classification is based on the presence of short direct repeats, empty insertion sites, a putative tyrosine recombinase gene and other features of Starship transposons. We also show that ToxhAT has been independently captured by two different Starships , Sanctuary and Horizon which share little to no sequenced identity, outside of ToxhAT. This classification makes Horizon and Sanctuary part of a growing number of Starships involved in the horizontal gene transfer of adaptive genetic material between fungal species. Importance The work presented here expands our understanding of a novel group of mobile genetic elements called Starships that facilitate the horizontal exchange of virulence genes in fungal pathogens. Our analysis shows that Sanctuary and ToxhAT are likely active and autonomous transposons in the B. sorokiniana genome. We also show that the smaller ToxhAT transposon has been independently captured by two different Starships , viz. Sanctuary in B. sorokiniana and Horizon in P. tritici-repentis and P. nodorum. Outside of ToxhAT these two Starships share no sequence identity. The capture of ToxhAT by two different mobile elements in three different fungal wheat pathogens demonstrates how horizontal transposon transfer is driving the evolution of virulence in these important wheat pathogens.

Sanctuary: AStarshiptransposon facilitating the movement of the virulence factor ToxA in fungal wheat pathogens