To improve our understanding of the origin and evolution of mycoheterotrophic plants, we here present the chromosome-scale genome assemblies of two sibling orchid species: partially mycoheterotrophic Platanthera zijinensis and holomycoheterotrophic Platanthera guangdongensis. Comparative analysis shows that mycoheterotrophy is associated with increased substitution rates and gene loss, and the deletion of most photoreceptor genes and auxin transporter genes might be linked to the unique phenotypes of fully mycoheterotrophic orchids. Conversely, trehalase genes that catalyse the conversion of trehalose into glucose have expanded in most sequenced orchids, in line with the fact that the germination of orchid non-endosperm seeds needs carbohydrates from fungi during the protocorm stage. We further show that the mature plant of P. guangdongensis, different from photosynthetic orchids, keeps expressing trehalase genes to hijack trehalose from fungi. Therefore, we propose that mycoheterotrophy in mature orchids is a continuation of the protocorm stage by sustaining the expression of trehalase genes. Our results shed light on the molecular mechanism underlying initial, partial and full mycoheterotrophy.

Two new Platanthera (Orchidinae, Orchideae, Orchidoideae) species, P. guangdongensis and P. zijinensis, from the Guangdong Province in China are described and illustrated based on morphological and molecular analyses. Platanthera guangdongensis is leafless and resembles P. fujianensis, but it differs in terms of the shorter and/or smaller inflorescence, floral bract, dorsal sepal, lateral sepal, petal, and ligulate lip. Platanthera zijinensis is morphologically similar to P. minor, but it differs by the ovate-oblong to ovate floral bract, colour of flower, ovate dorsal sepal, oblong lateral sepal, ovate-oblong lip, and curved forward spur. The phylogenetic analyses based on nuclear ribosomal ITS and chloroplast (rbcL and matK) sequence data support that P. zijinensis and P. guangdongensis being sisters to each other and genetically sister to P. minor. The chloroplast rbcL of P. guangdongensis is a pseudogene, supporting its characterization as a holomycotrophic orchid.

Four new Viola species from southern China, i.e. Viola qingruii, V. heyuanensis, V. chaozhouensis, and V. longissima, are described and illustrated based on morphology and molecular phylogeny. Viola qingruii is widely distributed in the coastal mountains of southeastern China, while V. heyuanensis, V. chaozhouensis, and V. longissima, are all narrow endemics in Guangdong Province, China. These species are described and compared with the morphologically related species V. lucens, V. tenuis, V. guangzhouensis and V. huizhouensis. Phylogenetic analysis based on ITS confirmed that all of the four new species belong to Viola sect. Plagiostigma subsect. Diffusae. An identifcation key is provided to distinguish the four new species from other species of V. subsect. Diffusae in southern China. In addition, the origin and evolution of characters such as aerial stems, stolons, elaiosomes and tuberculate seeds in Viola are briefly discussed.