A conserved tetrapolar mating system permits sexual hybridization and recombinant host phenotypes in wheat- and barley-adapted Puccinia striiformis

Host specialization can restrict gene flow in fungal pathogens, yet sexual compatibility may persist when mating occurs on a shared alternate host. We tested this possibility by crossing wheat-adapted Puccinia striiformis f. sp. tritici and barley-adapted P. striiformis f. sp. hordei on Berberis vulgaris. Despite contrasting host specialization and genome-wide sequence differentiation, reciprocal crosses produced 18 viable aeciospore-derived progeny. Chromosome scale parental assemblies revealed physically unlinked homeodomain (HD) and pheromone receptor (PR) mating-type loci, consistent with a conserved tetrapolar mating system. All progeny inherited one HD haplotype from each parent, representing all four parental HD haplotype combinations. Phenotyped progeny displayed host-interaction profiles distinct from either parent, including compatibility with selected genotypes of both wheat and barley. These findings demonstrate that host-associated differentiation in P. striiformis can coexist with incomplete intrinsic reproductive isolation between the tested parental isolates, allowing sexual hybridization to generate novel host phenotypes.