The plastid is a semiautonomous organelle with its own genome. Plastid genomes have been widely used as models for studying phylogeny, speciation and adaptive evolution. However, most studies focus on comparisons of plastid genome evolution at high taxonomic levels, and comparative studies of the process of plastome evolution at the infrageneric or intraspecific level remain elusive. Holcoglossum is a small genus of Orchidaceae, consisting of approximately 20 species of recent radiation. This made it an ideal group to explore the plastome mutation mode at the infrageneric or intraspecific level.

It is estimated that floral deception has evolved in at least 7500 species of angiosperms, of which two thirds are orchids. Epipactis veratrifolia (Orchidaceae) is a model system of aphid mimicry as aphidophagous hoverflies lay eggs on false brood sites on their flowers. To understand the evolutionary ecology of floral deception, we investigated the pollination biology of E. veratrifolia across 10 populations in the Eastern Himalayas. We reconstructed the phylogeny of Epipactis and mapped the known pollination systems of previously studied species onto the tree.

DNA barcoding is expected to be one of the most promising tools in biological taxonomy. However, there have been no agreements on which core barcode should be used in plants, especially in species-rich genera with wide geographical distributions. To evaluate their discriminatory power in large genera, four of the most widely used DNA barcodes, including three plastid regions (matK, rbcL, trnH-psbA) and nuclear internal transcribed spacer (nrITS), were tested in seven species-rich genera (Ficus, Pedicularis, Rhodiola, Rhododendron,Viburnum, Dendrobium and Lysimachia) and a moderate size genus, Codonopsis. All of the sequences from the aforementioned seven large genera were downloaded from NCBI. The related barcodes for Codonopsis were newly generated in this study. Genetics distances, DNA barcoding gaps and phylogenetic trees of the four single barcodes and their combinations were calculated and compared in the seven genera. As for single barcode, nrITS has the most variable sites, the clearest intra- and inter-specific divergences and the highest discrimination rates in the seven genera. Among the combinations of barcodes, ITS+matK performed better than all the single barcodes in most cases and even the three- and four-loci combinations in the seven genera. Therefore, we recommend ITS+matK as the core barcodes for large plant genera.

Acanthochlamys bracteata (Velloziaceae) is a resurrection plant with cold tolerance. Herein, a chromosome-level reference genome of A. bracteata based on Nanopore, Illumina, and Hi-C data is reported. The high-quality assembled genome was 197.97 Mb, with a scaffold N50 value of 8.64 Mb and a contig N50 value of 6.96 Mb. We annotated 23,509 protein-coding genes. Eight contracted gene families and three expanded gene families were detected. Repeat sequences accounted for approximately 28.63% of the genome. The LEA1 and Dehydrin gene families, which are involved in desiccation resistance, expanded in A. bracteata. We identified genes involved in chilling tolerance, COLD1.

Four new species of Orchidaceae from China, Heminium lijiangense, Peristylus fasciculatus, Platanthera milinensis, and Ponerorchis gongshanensis, together with a new country record, Peristylus tenuicallus, are described and illustrated based on morphological and/or phylogenetic analyses. Heminium lijiangense is closely related to H. elisabethae but differs from it by having the dorsal sepal ovate-orbicular and lip mid-lobe distinctly shorter than lateral lobes. P. fasciculatus is close to Peristylus tradescantifolius but is distinguished from it by having several fascicled and straight, root-like tubers (vs. one or two oblongoid tubers), old stems usually persistent, middle lobe of lip narrowly ligulate-lanceolate and half as long as the lateral lobes (vs. middle lobe deltoid, about a third as long as the lateral lobes or less), a raised callus at the base of each lateral lobe (vs. callus absent), spur gradually attenuate toward the apex (vs. spur clavate). Platanthera milinensis is similar to P. stenochila by sharing small green flowers and lip without a spur, but differs in having a creeping rhizome, a corymbose inflorescence, and a broadly ovate and slightly 3-lobed lip. Ponerochis gongshanensis is similar to P. faberi in its small flowers, but differs in having a linear leaf c. 3 mm wide (vs. leaf 5-13 mm wide), in the lip having collar-like raised margins on the sides of the spur entrance, and a mid-lobe which is notched at the apex but not divided into two divergent lobules that are nearly as large as the lateral lobes, as in P. faberi. All the proposed species obtained high support in phylogenetic analysis as new species. The recently described genus Apetalanthe is reduced to synonymy of Ponerorchis and a new combination is made.

Collabieae (Orchidaceae) is a long neglected tribe with confusing tribal and generic delimitation and little-understood phylogenetic relationships. Using plastid matK, psaB, rbcL, and trnH-psbA DNA sequences and morphological evidence, the phylogenetic relationships within the tribe Collabieae were assessed as a basis for revising their tribal and generic delimitation. Collabieae (including the previously misplaced mycoheterotrophic Risleya) is supported as monophyletic and nested within a superclade that also includes Epidendreae, Podochileae, Cymbidieae and Vandeae. Risleya is nested in Collabiinae and sister to Chrysoglossum, a relationship which, despite their great vegetative differences, is supported by floral characters. Ania is a distinct genus supported by both morphological and molecular evidence, while redefined Tainia includes Nephelaphyllum and Mischobulbum. Calanthe is paraphyletic and consists four clades; the genera Gastrorchis, Phaius and Cephalantheropsis should be subsumed within Calanthe. Calanthe sect. Ghiesbreghtia is nested within sect. Calanthe, to which the disputed Calanthe delavayi belongs as well. Our results indicate that, in Collabieae, habit evolved from being epiphytic to terrestrial.

The plastid genome (plastome) of heterotrophic plants like mycoheterotrophs and parasites shows massive gene losses in consequence to the relaxation of functional constraints on photosynthesis. To understand the patterns of this convergent plastome reduction syndrome in heterotrophic plants, we studied 12 closely related orchids of three different lifeforms from the tribe Neottieae (Orchidaceae). We employ a comparative genomics approach to examine structural and selectional changes in plastomes within Neottieae. Both leafy and leafless heterotrophic species have functionally reduced plastid genome. Our analyses show that genes for the NAD(P)H dehydrogenase complex, the photosystems, and the RNA polymerase have been lost functionally multiple times independently. The physical reduction proceeds in a highly lineage-specific manner, accompanied by structural reconfigurations such as inversions or modifications of the large inverted repeats. Despite significant but minor selectional changes, all retained genes continue to evolve under purifying selection. All leafless Neottia species, including both visibly green and nongreen members, are fully mycoheterotrophic, likely evolved from leafy and partially mycoheterotrophic species. The plastomes of Neottieae span many stages of plastome degradation, including the longest plastome of a mycoheterotroph, providing invaluable insights into the mechanisms of plastome evolution along the transition from autotrophy to full mycoheterotrophy.

Neottieae comprise about 150-200 species and are distributed mainly in temperate and subtropical zones of the northern hemisphere. Mycoheterotrophy is common in Neottieae. Based on three DNA markers and a broad sampling of Neottieae, these results indicate that Neottieae is strongly supported as monophyletic and Palmorchis is sister to the remaining genera of Neottieae. Holopogon and Neottia s.s. are deeply nested within Listera. The habit of leafless mycotrophy has independently evolved at least three times in Neottieae, one in Cephalanthera, another in Neottia s.l. and the third in the clade formed by Limodorum and Aphyllorchis.

The implantation process is complex, requiring reciprocal interactions between implantation-competent blastocysts and the receptive uterus. There were reports to show that some microRNAs (miRNAs) may play a key role during embryo implantation in mouse. However, the miR-320 expression profiles in the rat uterus during peri-implantation are unknown. In the present study, we found that the expression level of miR-320 was lower on day 5 of gestation (g.d. 5) in rats than g.d.3 and g.d.4 and restored gradually from g.d.6. MiR-320 was specifically localized in glandular and luminal epithelia and decidua. The expression of miR-320 was not significantly different in the pseudopregnant uterus and decreased in the uteri of rats subjected to activation of delayed implantation. Artificial decidualization and treatment with progesterone increased the miR-320 expression. Thus, miR-320 was differentially expressed in the rat uterus during implantation. The expression level was affected by active blastocysts and decidualization during the window of implantation. Steroid hormones, progesterone stimulated miR-320 expression.

Calypsoinae is a small subtribe in Orchidaceae (Epidendroideae) characterized by diverse trophic strategies and morphological characters. Calypsoinae includes 13 genera, four of which are leafless and mycoheterotrophic. Mycoheterotrophic species in the leafless genus Corallorhiza are well suited to studies of plastome evolution. However, the lack of plastome sequences for other genera in Calypsoinae limits the scope of comparative and phylogenetic analyses, in particular our understanding of plastome evolution. To understand plastid genome evolution in Calypsoinae, we newly sequenced the plastomes of 12 species in the subtribe, including representatives of three mycoheterotrophic genera as well as five autotrophic genera. We detected two parallel photosynthetic losses in Corallorhiza. Evolutionary analyses indicated that the transition to obligate mycoheterotrophy leads to the relaxation of selection in a highly gene-specific pattern.

Liparis aureolabella and L. mengziensis, two new species from the karst region of southwestern China, and L. bingzhongluoensis, a new species from montane region in Yunnan, are described and illustrated. L. aureolabella is easily distinguished from its relatives by having abaxially purple leave with purple reticulate veins prominent adaxially, a lip auriculate at base, and falcate-lanceolate pollinia. Liparis mengziensis is closely related to L. petiolata and L. auriculata, but differs from them by having an ovate to broadly ovate leaf, purple lip and apex connate along the margins. Liparis bingzhongluoensis is similar to Liparis nanlingensis, but the new species is characterized by having a lip with two transparent ridges on its disc, longitudinally concave basal callus and triangular column wings. Phylogenetic analyses based on nuclear ribosomal ITS and plastid matK sequences showed that L. aureolabella and L. mengziensis are nested with L. petiolata or L. auriculata in a monophyletic clade. L. bingzhongluoensis is sister to a clade formed by L. nanlingensis, L. tsii, L. sasakii and L. krameri. Moreover, morphological comparisons strongly support that the three species as separated species newly to science.

Gastrodia longistyla, a new species of Orchidaceae from Yunnan Province, China, is described and illustrated. It is morphologically similar to Gastrodia peichatieniana, but can be easily distinguished from the latter by having a rhombic epichile, long column (6.0-7.5 mm long), and a needle-shaped appendage (1.8-3.2 mm in length) at the base of the stigma. Identification key and colour photographs are provided. A preliminary risk-of-extinction assessment, according to the IUCN Red List Categories and Criteria, is given for the new species. The plastome of G. longistyla is 30464 bp in length with GC content approximately 24.8%, and the plastome does not contain some housekeeping genes, such as matK, rpl16, or all photosynthesis genes. In addition, the G. longistyla plastome lacks an IR region. This indicates that the plastome is in the last stage of degradation.

A new species, Neottia wuyishanensis (Neottieae, Orchidaceae), is described and illustrated from Wuyishan, Fujian, eastern China, based on morphological characters and molecular data. Molecular phylogenetics indicate that N. wuyishanensis is nested within the clade formed by Neottia fugongensis and Neottia nujiangensis. Morphologically, N. wuyishanensis is similar to N. fugongensis and N. pseudonipponica, but differs from them by having indistinct auricles at the base, and in the shape and size of lobelets.

In a previous study, via microRNA microarray analysis we found that miR-98 is differentially expressed in rat uteri during the peri-implantation period (unpublished data). However, the role of miR-98 in rat embryo implantation remains elusive. Here, we found that the level of miR-98 is lower on day 5 and 6 of gestation (g.d. 5-6) than that on g.d. 3-4 and g.d. 7-8 in rat. MiR-98 expression is significantly decreased by delayed implantation. Down-regulation of miR-98 promotes ESC proliferation and inhibits apoptosis. Up-regulation of miR-98 displays opposite effects. Further investigation revealed that miR-98 can bind to the 3'-untranslated region (3'-UTR) of B-cell lymphoma-extra large (Bcl-xl) to inhibit Bcl-xl translation. Collectively, down-regulation of miR-98 in rat uterus during the receptive phase is linked to the increase of cell proliferation via targeting Bcl-xl.

Subtribe Orchidinae (Orchidaceae, Orchidoideae) are a nearly cosmopolitan taxon of terrestrial orchids, comprising about 1800 species in 47 to 60 genera. Although much progress has been made in recent years of phylogenetics of Orchidinae, considerable problems remain to be addressed. Based on molecular phylogenetics, we attempt to illustrate the phylogenetic relationships and discuss generic delimitation within Orchidinae. Seven DNA markers (five plastid and two nuclear), a broad sampling of Orchidinae (400 species in 52 genera) and three methods of phylogenetic analysis (maximum likelihood, maximum parsimony and Bayesian inference) were used.

Five new species (Gastrochilus yei, Gastrochilus minimus, Luisia simaoensis, Taeniophyllum xizangense, Tuberolabium subulatum) and two newly recorded species (Cleisostoma tricornutum, Luisia inconspicua) of Vandeae (Orchidaceae) from China are described and illustrated. Gastrochilus yei is similar to G. affinis and G. nepalensis, but differs from them by having an epichile not lobed, the apex of the hypochile not bilobed, and a tine on the apex of the leaf. Gastrochilus minimus is similar to G. acinacifolius, but can be distinguished from the latter by having a flabellate epichile that is densely hirsute on the adaxial surface and an inconspicuous central cushion; in addition, the hypochile of G. minimus has a keel that extends to the apex of the epichile. Taeniophyllum xizangense is similar to T. stella and T. radiatum, but it is distinguished from them by having much bigger flowers, inflorescences densely covered with short-bristly hairs, papillae on the external surface of sepals, and bigger triangular-ovate viscidium. Luisia simaoensis is similar to L. magniflora and L. ramosii, but can be easily distinguished from them by having lateral sepals longer than dorsal sepals and petals, lip with irregular and waved margins, and lip with bilobed apex. Luisia inconspicua is moved from Gastrochilus to Luisia based on phylogenetic analyses of plastid matK sequence data. Tuberolabium subulatum is similar to T. carnosum, but it can be easily distinguished from the latter by having an inflorescence much shorter than the leaves, yellow sepals and petals, and many small papillae outside the lip lobes.