The plot thickens: Haploid and triploid-like thalli, hybridization, and biased mating type ratios in Letharia

<p>The study of the reproductive biology of lichen fungal symbionts has been traditionally challenging due to their complex and symbiotic lifestyles. Against the common belief of haploidy, a recent genomic study found a triploid-like signal in <em>Letharia</em>. Here, we used genomic data from a pure culture and from thalli, together with a PCR survey of the MAT locus, to infer the genome organization and reproduction in <em>Letharia</em>. We found that the read count variation in the four <em>Letharia</em> specimens, including the pure culture derived from a single sexual spore of <em>L. lupina</em>, is consistent with haploidy. By contrast, the <em>L. lupina </em>read counts from a thallus' metagenome are triploid-like. Characterization of the mating-type locus revealed a conserved heterothallic configuration across the genus, along with auxiliary genes that we identified. We found that the mating-type distributions are balanced in North America for <em>L. vulpina</em> and <em>L. lupina</em>, suggesting widespread sexual reproduction, but highly skewed in Europe for <em>L. vulpina</em>, consistent with predominant asexuality. Taken together, we propose that <em>Letharia</em> fungi are heterothallic and typically haploid, and provide evidence that triploid-like individuals are rare hybrids between <em>L. lupina</em> and an unknown <em>Letharia</em> lineage, reconciling classic systematic and genetic studies with recent genomic observations.</p>