Unprecedented Parallel Photosynthetic Losses in a Heterotrophic Orchid Genus

Heterotrophic plants are evolutionary experiments in genomic, morphological, and physiological change. Yet, genomic sampling gaps exist among independently derived heterotrophic lineages, leaving unanswered questions about the process of genome modification. Here, we have sequenced complete plastid genomes for all species of the leafless orchid genus Hexalectris, including multiple individuals for most, and leafy relatives Basiphyllaea and Bletia. Our objectives are to determine the number of independent losses of photosynthesis and to test hypotheses on the process of genome degradation as a result of relaxed selection. We demonstrate four to five independent losses of photosynthesis in Hexalectris based on degradation of the photosynthetic apparatus, with all but two species displaying evidence of losses, and variation in gene loss extending below the species level. Degradation in the atp complex is advanced in Hexalectris warnockii, whereas only minimal degradation (i.e., physical loss) has occurred among some "housekeeping" genes. We find genomic rearrangements, shifts in Inverted Repeat boundaries including complete loss in one accession of H. arizonica, and correlations among substitutional and genomic attributes. Our unprecedented finding of multiple, independent transitions to a fully mycoheterotrophic lifestyle in a single genus reveals that the number of such transitions among land plants is likely underestimated. This study underscores the importance of dense taxon sampling, which is highly informative for advancing models of genome evolution in heterotrophs. Mycoheterotrophs such as Hexalectris provide forward-genetic opportunities to study the consequences of radical genome evolution beyond what is possible with mutational studies in model organisms alone.