DataSheet_1_Phylogenetic position and plastid genome structure of Vietorchis, a mycoheterotrophic genus of Orchidaceae (subtribe Orchidinae) endemic to Vietnam.pdf

The orchid genus Vietorchis comprises three species, all discovered in the 21 century. Each of these species is achlorophyllous, mycoheterotrophic and is known to be endemic to Vietnam. The type species of the genus, V. aurea, occurs in a single location in northern Vietnam within a lowland limestone karstic area. Vietorchis furcata and V. proboscidea, in contrast, are confined to mountains of southern Vietnam, far away from any limestone formations. Taxonomic placement of Vietorchis remained uncertain for the reason of inconclusive morphological affinities. At the same time, the genus has never been included into molecular phylogenetic studies. We investigate the phylogenetic relationships of two species of Vietorchis (V. aurea and V. furcata) based on three DNA datasets: (1) a dataset comprising two nuclear regions, (2) a dataset comprising two plastid regions, and (3) a dataset employing data on the entire plastid genomes. Our phylogenetic reconstructions support the placement of Vietorchis into the subtribe Orchidinae (tribe Orchideae, subfamily Orchidoideae). This leads to a conclusion that the previously highlighted similarities in the rhizome morphology between Vietorchis and certain mycoheterotrophic genera of the subfamilies Epidendroideae and Vanilloideae are examples of a convergence. Vietorchis is deeply nested within Orchidinae, and therefore the subtribe Vietorchidinae is to be treated as a synonym of Orchidinae. In the obtained phylogenetic reconstructions, Vietorchis is sister to the photosynthetic genus Sirindhornia. Sirindhornia is restricted to limestone mountains, which allows to speculate that association with limestone karst is plesiomorphic for Vietorchis. Flower morphology is concordant with the molecular data in placing Vietorchis into Orchidinae and strongly supports the assignment of the genus to one of the two major clades within this subtribe. Within this clade, however, Vietorchis shows no close structural similarity with any of its genera; in particular, the proximity between Vietorchis and Sirindhornia has never been proposed. Finally, we assembled the plastid genome of V. furcata, which is 65969 bp long and contains 45 unique genes, being one of the most reduced plastomes in the subfamily Orchidoideae. The plastome of Vietorchis lacks any rearrangements in comparison with the closest studied autotrophic species, and possesses substantially contracted inverted repeats. No signs of positive selection acting on the protein-coding plastid sequences were detected.

兰属（Vietorchis）包含3个物种，所有物种均发现于21世纪。该属所有物种均无叶绿素、为菌异养型，且仅特有分布于越南。本属的模式种为金黄兰（V. aurea），仅分布于越南北部一处低地石灰岩喀斯特地貌区域。与之相对，V. furcata与V. proboscidea仅见于越南南部山地，与任何石灰岩地貌均相距甚远。由于形态亲缘关系尚不明确，兰属（Vietorchis）的分类地位长期处于未定状态；同时，该属从未被纳入分子系统发育研究（molecular phylogenetic studies）范畴。本研究基于三类DNA数据集对兰属（Vietorchis）的两个物种（V. aurea与V. furcata）的系统发育关系展开探究：（1）包含两个核基因区域（nuclear regions）的数据集，（2）包含两个质体基因区域（plastid regions）的数据集，（3）基于完整质体基因组（entire plastid genomes）数据的数据集。系统发育重建（phylogenetic reconstructions）结果支持将兰属（Vietorchis）归入兰亚族（Orchidinae，兰族Orchideae，兰亚科Orchidoideae）。这一结论表明，此前被指出的兰属与树兰亚科（Epidendroideae）、香荚兰亚科（Vanilloideae）部分菌异养属之间的根状茎形态相似性，实为趋同演化（convergence）的案例。兰属（Vietorchis）在兰亚族中深度嵌入，因此原成立的Vietorchidinae亚族应被视为兰亚族（Orchidinae）的同物异名（synonym）。在系统发育重建结果中，兰属（Vietorchis）与光合属Sirindhornia构成姊妹群。Sirindhornia仅分布于石灰岩山地，据此可推测，与石灰岩喀斯特地貌的关联是兰属（Vietorchis）的祖征（plesiomorphic）。花形态特征与分子数据一致支持将兰属（Vietorchis）归入兰亚族（Orchidinae），并强烈支持该属隶属于该亚族内的两个主要演化支之一。然而，在该演化支内，兰属（Vietorchis）与其他任何属均无紧密的结构相似性；尤为特别的是，此前从未有研究提出兰属（Vietorchis）与Sirindhornia存在近缘关系。本研究还组装了V. furcata的质体基因组（plastid genome），其全长为65969 bp，包含45个独特基因，是兰亚科（Orchidoideae）中质体基因组缩减程度最高的类群之一。与近缘已研究的自养物种（autotrophic species）相比，该质体基因组未发生重排，且反向重复序列（inverted repeats）大幅收缩。未检测到编码蛋白质的质体序列（protein-coding plastid sequences）受到正选择（positive selection）的迹象。