Ndh Genes Research Articles

Transit From Autotrophism to Heterotrophism: Sequence Variation and Evolution of Chloroplast Genomes in Orobanchaceae Species.

The family Orobanchaceae including autotrophic, hemiparasitic, and holoparasitic species, is becoming a key taxa to study the evolution of chloroplast genomes in different lifestyles. But the early evolutionary trajectory in the transit from autotrophism to hemiparasitism still maintains unclear for the inadequate sampling. In this study, we compared 50 complete chloroplast genomes in Orobanchaceae, containing four newly sequenced plastomes from hemiparasitic Pedicularis, to elucidate the sequence variation patterns in the evolution of plastomes. Contrasted to the sequence and structural hypervariabilities in holoparasites, hemiparasitic plastomes exhibited high similarity to those of autotrophs in gene and GC contents. They are generally characterized with functional or physical loss of ndh/tRNA genes and the inverted small-single-copy region. Gene losses in Orobanchaceae were lineage-specific and convergent, possibly related to structural reconfiguration and expansion/contraction of the inverted region. Pseudogenization of ndh genes was unique in hemiparasites. At least in Pedicularis, the ndhF gene might be most sensitive to the environmental factors and easily pseudogenized when autotrophs transit to hemiparasites. And the changes in gene contents and structural variation potentially deeply rely on the feeding type. Selective pressure, together with mutational bias, was the dominant factor of shaping the codon usage patterns. The relaxed selective constraint, potentially with genome-based GC conversion (gBGC) and preferential codon usage, drive the fluctuation of GC contents among taxa with different lifestyles. Phylogenetic analysis in Orobanchaceae supported that parasitic species were single-originated while holoparasites were multiple-originated. Overall, the comparison of plastomes provided a good opportunity to understand the evolution process in Orobanchaceae with different lifestyles.

Hexavalent chromium leads to differential hormetic or damaging effects in alfalfa (Medicago sativa L.) plants in a concentration-dependent manner by regulating nitro-oxidative and proline metabolism

Chromium has been proven to be extremely phytotoxic. This study explored the impacts of increasing Cr(VI) exposure (up to 10 mg L−1 K2Cr2O7) on the growth and development of alfalfa plants and adaptation responses employed, in an environmentally relevant context. The threshold concentration of K2Cr2O7 in irrigation water beyond which stress responses are initiated is 1 mg L−1. Lower Cr(VI) exposure (0.5 mg L−1 K2Cr2O7) induced hormesis, evident through increased biomass and larger leaves, likely mediated by increased NO content (supported by elevated NR enzymatic activity and overexpression of NR and ndh genes). Elevated Cr(VI) exposure (5 and 10 mg L−1 K2Cr2O7) resulted in reduced biomass and smaller leaves, and lower levels of photosynthetic pigment (10 mg L−1 K2Cr2O7). Higher levels of lipid peroxidation, H2O2 and NO contents in these plants suggested nitro-oxidative stress. Stress responses included increased SOD and CAT enzymatic activities, further supported to some extent by MnSOD, FeSOD, Cu/ZnSOD and CAT transcripts levels. GST7 and GST17 gene expression patterns, as well as proline content, P5CS enzymatic activity and corresponding P5CS and P5CR gene expression levels emphasized the role of proline and GSTs in the adaptation responses. Results highlight the importance of managing Cr(VI) levels in irrigation water.

Phylogeny of Euclidieae (Brassicaceae) based on plastome and nuclear ribosomal DNA data

Euclidieae, a morphologically diverse tribe in the family Brassicaceae (Cruciferae), consists of 29 genera and more than 150 species distributed mainly in Asia. Prior phylogenetic analyses on Euclidieae are inadequate. In this study, sequence data from the plastid genome and nuclear ribosomal DNA of 72 species in 27 genera of Euclidieae were used to infer the inter- and intra-generic relationships within. The well-resolved and strongly supported plastome phylogenies revealed that Euclidieae could be divided into five clades. Both Cymatocarpus and Neotorularia are polyphyletic in nuclear and plastome phylogenies. Besides, the conflicts of systematic positions of three species of Braya and two species of Solms-laubachia s.l. indicated that hybridization and or introgression might have happened during the evolutionary history of the tribe. Results from divergence-time analyses suggested an early Miocene origin of Euclidieae, and it probably originated from the Central Asia, Pamir Plateau and West Himalaya. In addition, multiple ndh genes loss and pseudogenization were detected in eight species based on comparative genomic study.

Comparative Plastid Genomics of Neotropical Bulbophyllum (Orchidaceae; Epidendroideae).

Pantropical Bulbophyllum, with ∼2,200 species, is one of the largest genera in Orchidaceae. Although phylogenetics and taxonomy of the ∼60 American species in the genus are generally well understood, some species complexes need more study to clearly delimit their component species and provide information about their evolutionary history. Previous research has suggested that the plastid genome includes phylogenetic markers capable of providing resolution at low taxonomic levels, and thus it could be an effective tool if these divergent regions can be identified. In this study, we sequenced the complete plastid genome of eight Bulbophyllum species, representing five of six Neotropical taxonomic sections. All plastomes conserve the typical quadripartite structure, and, although the general structure of plastid genomes is conserved, differences in ndh-gene composition and total length were detected. Total length was determined by contraction and expansion of the small single-copy region, a result of an independent loss of the seven ndh genes. Selection analyses indicated that protein-coding genes were generally well conserved, but in four genes, we identified 95 putative sites under positive selection. Furthermore, a total of 54 polymorphic simple sequence repeats were identified, for which we developed amplification primers. In addition, we propose 10 regions with potential to improve phylogenetic analyses of Neotropical Bulbophyllum species.

The complete chloroplast genome sequence of Taxillus yadoriki (Loranthaceae): a hemi-parasitic evergreen shrub in East Asia

Taxillus yadoriki (Loranthaceae) is a hemiparasitic evergreen shrub distributed in Korea and Japan. We report the complete chloroplast genome of T. yadoriki to provide insight into the phylogenetic relationship of Loranthaceae. This genome is 122,192 bp long, with two IR regions (22,756 bp each) that separate a large single-copy (LSC) region (70,628 bp) and a small single-copy (SSC) region (6052 bp). It contains 109 genes that encode 68 proteins, 8 rRNAs, and 33 tRNAs. All of ndh genes have been lost and the SSC region consists of only four genes similar to other Taxillus species. In ML phylogenetic, monophyly of Taxillus was strongly supported with high bootstrap value and formed a sister group with Scurrula.

Plastome structure and phylogeny of Gnetum luofuense C.Y. Cheng (Gnetaceae, Gnetales)

This article describes the complete chloroplast genome of Gnetum luofuense. The G. luofense plastome was 114,795 bp in length, containing a large single copy region (66,103 bp) and a small single copy region (9438 bp), separated by two inverted repeat regions (19,627 bp). The genome lost all ndh genes and contained 116 genes, including 68 protein-coding genes, 40 tRNA genes, and eight rRNA genes. The GC content was 33.3%, 12 genes all contained an intron, ycf3 gene contained two introns while rps12 was a transpliced gene. Phylogenetic analysis using 61 concatenated protein-coding genes suggests that G. luofuense with the rest of other gnetophytes were sister to or nested within all conifers.

Insights Into Chloroplast Genome Evolution Across Opuntioideae (Cactaceae) Reveals Robust Yet Sometimes Conflicting Phylogenetic Topologies.

Chloroplast genomes (plastomes) are frequently treated as highly conserved among land plants. However, many lineages of vascular plants have experienced extensive structural rearrangements, including inversions and modifications to the size and content of genes. Cacti are one of these lineages, containing the smallest plastome known for an obligately photosynthetic angiosperm, including the loss of one copy of the inverted repeat (∼25 kb) and the ndh gene suite, but only a few cacti from the subfamily Cactoideae have been sufficiently characterized. Here, we investigated the variation of plastome sequences across the second-major lineage of the Cactaceae, the subfamily Opuntioideae, to address (1) how variable is the content and arrangement of chloroplast genome sequences across the subfamily, and (2) how phylogenetically informative are the plastome sequences for resolving major relationships among the clades of Opuntioideae. Our de novo assembly of the Opuntia quimilo plastome recovered an organelle of 150,347 bp in length with both copies of the inverted repeat and the presence of all the ndh gene suite. An expansion of the large single copy unit and a reduction of the small single copy unit was observed, including translocations and inversion of genes, as well as the putative pseudogenization of some loci. Comparative analyses among all clades within Opuntioideae suggested that plastome structure and content vary across taxa of this subfamily, with putative independent losses of the ndh gene suite and pseudogenization of genes across disparate lineages, further demonstrating the dynamic nature of plastomes in Cactaceae. Our plastome dataset was robust in resolving three tribes with high support within Opuntioideae: Cylindropuntieae, Tephrocacteae and Opuntieae. However, conflicting topologies were recovered among major clades when exploring different assemblies of markers. A plastome-wide survey for highly informative phylogenetic markers revealed previously unused regions for future use in Sanger-based studies, presenting a valuable dataset with primers designed for continued evolutionary studies across Cactaceae. These results bring new insights into the evolution of plastomes in cacti, suggesting that further analyses should be carried out to address how ecological drivers, physiological constraints and morphological traits of cacti may be related with the common rearrangements in plastomes that have been reported across the family.

The Complete Plastomes of Five Hemiparasitic Plants (Osyris wightiana, Pyrularia edulis, Santalum album, Viscum liquidambaricolum, and V. ovalifolium): Comparative and Evolutionary Analyses Within Santalales.

Most species of Santalales (the sandalwood order) are hemiparasites, including both facultative and obligate hemiparasites. Despite its rich diversity, only a small fraction of the species in the sandalwood order have sequenced plastomes. The evolution of parasitism–associated plastome reduction in Santalales remains under-studied. Here, we report the complete plastomes of three facultative hemiparasites (Pyrularia edulis, Cervantesiaceae; Osyris wightiana, and Santalum album, Santalaceae), and two obligate hemiparasites (Viscum liquidambaricolum and Viscum ovalifolium, Viscaceae). Coupled with publicly available data, we investigated the dynamics of plastome degradation in Santalales hemiparasites. Our results indicate that these hemiparasites can be characterized by various degrees of plastome downsizing, structural rearrangement, and gene loss. The loss or pseudogenization of ndh genes was commonly observed in Santalales hemiparasites, which may be correlated to the lifestyle shift from photoautotroph to hemiparasitism. However, the obligate hemiparasites did not exhibit a consistently higher level of gene loss or pseudogenization compared to facultative hemiparasites, which suggests that the degree of plastome reduction is not correlated with the trophic level facultative or obligate hemiparasitism. Instead, closely related taxa tend to possess highly similar plastome size, structure, and gene content. This implies the parasitism-associated plastome degradation in Santalales may evolve in a lineage-specific manner.

Variation in RNA‐editing sites of chloroplast protein‐coding genes in early‐maturity mutant induced by carbon‐ion beam in Sweet Sorghum

AbstractUsing prediction and experimental determination, we compared the RNA‐editing sites in chloroplast protein‐coding genes in an early‐maturity mutant (KFJT‐1) induced by carbon‐ion beam irradiation in sweet sorghum and its wild type (KFJT‐CK). Twenty‐two and 20 editing sites were identified in the KFJT‐CK and KFJT‐1 chloroplast, respectively, and the differential editing sites between KFJT‐CK and KFJT‐1 were mainly in mat, ndh and rpo genes. Furthermore, mutations in RNA‐editing sites in KFJT‐1 affected the structure of the encoded protein, which implying that RNA‐editing events might influence the function of essential genes. Finally, a comparative analysis of the RNA‐editing events between KFJT‐1 and other 10 Poaceae species revealed that the RNA‐editing sites in matK‐1002, rpoC1‐1163 and rpoC2‐242 were unique to KFJT‐1. This suggested that RNA‐editing events due to carbon‐ion beam irradiation affected the structure of the encode proteins, thereby influenced their biological functions. This study is the first to report on the variation in chloroplast RNA‐editing patterns induced by carbon‐ion beams irradiation, and its effect on the chloroplast in KFJT‐1 at the post‐transcriptional level.

Relationship of Mutation of Kodon S 315 T katG Mycobaterium Tuberculosis Gene with Multidrug Resistance Tuberculosis in South Sumatera

MDR TB is a condition of M. tuberculosis that is resistant to isoniazid and rifampicin. This occurs because of certain genomic mutations in certain Mtb specific genes. Some of the genes that play a role in the INH mutation are the katG gene, the inhA gene, the ahpC gene, the ndh gene and the kasA gene. The most frequent mutation gene is the katG gene, around 50-80%, and the most mutations (64%) are found in the S 315T codon. The aim was to determine the relationship of S315T genotype in the M. tuberculosis katG gene with the incidence of Mtb resistance in MDR TB patients in South Sumatra. This type of research is an observational cross sectional study approach, conducted in Palembang RSMH in collaboration with the Clinical Microbiology Laboratory of FK UNSRI Palembang from January - July 2019. The sample of the case group (MDR TB) was 50 people while the control group (TB SO) was 21 people. Each group was tested for S315T genotype katG M. tuberculosis gene using the PCR-RFLP method with the Mspl enzyme. Visualization of the katG gene in codon 315 in this study was only found in 40 samples, 26 samples in the case group (MDR TB) and 14 samples in the control group (TB SO). Results Genetic frequency distribution in the case group (MDR TB) was 65.38% wildtype and 34.62% mutant, whereas in the control group (TBSO) it was 92.86% wildtype and 7.14% mutant. The 315 coded KatG gene mutation in this study was not related to the incidence of MDR TB, with p = 0.07, an odds ratio of 6.8 (0.77 - 61.40, 95% CI). The conclusions of this study found 315G katG M. tuberculosis codon mutation mutations of 34.62% but not significantly related to the incidence of MDR TB in South Sumatra.

Genome Sequencing of the Endangered Kingdonia uniflora (Circaeasteraceae, Ranunculales) Reveals Potential Mechanisms of Evolutionary Specialization.

SummaryKingdonia uniflora, an alpine herb, has an extremely narrow distribution and represents a model for studying evolutionary mechanisms of species that have adapted to undisturbed environments for evolutionarily long periods of time. We assembled a 1,004.7-Mb draft genome (encoding 43,301 genes) of K. uniflora and found significant overrepresentation in gene families associated with DNA repair, underrepresentation in gene families associated with stress response, and loss of most plastid ndh genes. During the evolutionary process, the overrepresentation of gene families involved in DNA repair could help asexual K. uniflora reduce the accumulation of deleterious mutations, while reducing genetic diversity, which is important in responding to environment fluctuations. The underrepresentation of gene families related to stress response and functional loss of ndh genes could be due to lack or loss of ability to respond to environmental changes caused by long-term adaptation to a relatively stable ecological environment.

Complete chloroplast genome sequence of Pedicularis verticillata and chloroplast genome reconstruction of P. cheilanthifolia (Orobanchaceae)

We here report the complete chloroplast genome of Pedicularis verticillata and the reconstructed chloroplast genome of Pedicularis cheilanthifolia, the first such genome reported for this genus. The P. verticillata genome is 142,733 bp, including a 24,984-bp inverted repeat (IR) region, 89,723-bp large single-copy (LSC) region, and 12,042-bp small single copy (SSC) region, and the P. cheilanthifolia genome is 147,807 bp (IRs: 24,916 bp; LSC: 82,718 bp; SSC: 15,257 bp). The chloroplast genome of P. verticillata contains 116 genes, including 71 coding genes, 8 rRNAs, and 37 tRNAs. In the genomes of both species, 10 ndh genes are lost and ndhB was identified in the IR region. Additionally, rps16 is also lost in P. verticillata but not in P. cheilanthifolia. The maximum-likelihood phylogenetic tree showed that Pedicularis is a monophyletic and sister group to other hemi-parasitic species in Orobanchaceae.

Complete plastid genome of an alpine plant Gentiana filistyla (Gentianaceae) and phylogenetic analysis

The complete plastid genome of Gentiana filistyla, occurring in Hengduan Mountains with high altitudes, was determined and analyzed in this work. It had a circular-mapping molecular with the length of 139,476 bp, has similar gene composition with G. section Kudoa but contains eight less ndh genes than the available species from sections Cruciata, Microsperma and Frigidae. Phylogenetic analysis showed that G. filistyla clustered together with section Kudoa rather than sections Microsperma and Frigidae. The plastome provided in this work would be useful for elucidation of phylogenetics and evolution in Gentiana.

The complete chloroplast genome of a hemiparasitic plant Santalum boninense (Santalaceae), endemic to the Bonin (Ogasawara) Islands

The complete chloroplast genome sequence of a hemiparasitic plant Santalum boninense (Santalaceae) was determined and described in this study. The chloroplast genome was 144,263 bp in length, consisting of a large single-copy region (83,912 bp), a small single-copy region (11,349 bp), and inverted repeats (24,501 bp). The chloroplast genome contained 124 genes including 72 protein-coding genes, 35 tRNA genes, eight rRNA genes, and nine pseudogenes. All the ndh genes and infA gene were found to lose function by gene loss or pseudogenization. GC content of the whole chloroplast genome was 38.0%.

Plastome Evolution and Phylogeny of Orchidaceae, With 24 New Sequences.

In order to understand the evolution of the orchid plastome, we annotated and compared 124 complete plastomes of Orchidaceae representing all the major lineages in their structures, gene contents, gene rearrangements, and IR contractions/expansions. Forty-two of these plastomes were generated from the corresponding author's laboratory, and 24 plastomes—including nine genera (Amitostigma, Bulbophyllum, Dactylorhiza, Dipodium, Galearis, Gymnadenia, Hetaeria, Oreorchis, and Sedirea)—are new in this study. All orchid plastomes, except Aphyllorchis montana, Epipogium aphyllum, and Gastrodia elata, have a quadripartite structure consisting of a large single copy (LSC), two inverted repeats (IRs), and a small single copy (SSC) region. The IR region was completely lost in the A. montana and G. elata plastomes. The SSC is lost in the E. aphyllum plastome. The smallest plastome size was 19,047 bp, in E. roseum, and the largest plastome size was 178,131 bp, in Cypripedium formosanum. The small plastome sizes are primarily the result of gene losses associated with mycoheterotrophic habitats, while the large plastome sizes are due to the expansion of noncoding regions. The minimal number of common genes among orchid plastomes to maintain minimal plastome activity was 15, including the three subunits of rpl (14, 16, and 36), seven subunits of rps (2, 3, 4, 7, 8, 11, and 14), three subunits of rrn (5, 16, and 23), trnC-GCA, and clpP genes. Three stages of gene loss were observed among the orchid plastomes. The first was ndh gene loss, which is widespread in Apostasioideae, Vanilloideae, Cypripedioideae, and Epidendroideae, but rare in the Orchidoideae. The second stage was the loss of photosynthetic genes (atp, pet, psa, and psb) and rpo gene subunits, which are restricted to Aphyllorchis, Hetaeria, Hexalectris, and some species of Corallorhiza and Neottia. The third stage was gene loss related to prokaryotic gene expression (rpl, rps, trn, and others), which was observed in Epipogium, Gastrodia, Lecanorchis, and Rhizanthella. In addition, an intermediate stage between the second and third stage was observed in Cyrtosia (Vanilloideae). The majority of intron losses are associated with the loss of their corresponding genes. In some orchid taxa, however, introns have been lost in rpl16, rps16, and clpP(2) without their corresponding gene being lost. A total of 104 gene rearrangements were counted when comparing 116 orchid plastomes. Among them, many were concentrated near the IRa/b-SSC junction area. The plastome phylogeny of 124 orchid species confirmed the relationship of {Apostasioideae [Vanilloideae (Cypripedioideae (Orchidoideae, Epidendroideae))]} at the subfamily level and the phylogenetic relationships of 17 tribes were also established. Molecular clock analysis based on the whole plastome sequences suggested that Orchidaceae diverged from its sister family 99.2 mya, and the estimated divergence times of five subfamilies are as follows: Apostasioideae (79.91 mya), Vanilloideae (69.84 mya), Cypripedioideae (64.97 mya), Orchidoideae (59.16 mya), and Epidendroideae (59.16 mya). We also released the first nuclear ribosomal (nr) DNA unit (18S-ITS1-5.8S-ITS2-28S-NTS-ETS) sequences for the 42 species of Orchidaceae. Finally, the phylogenetic tree based on the nrDNA unit sequences is compared to the tree based on the 42 identical plastome sequences, and the differences between the two datasets are discussed in this paper.

<p><strong>Plastome data provide insights into intra and interspecific diversity and <em>ndh</em> gene loss in <em>Capparis</em> (Capparaceae)</strong></p>

The genus Capparis is well known for its medicinal properties and many species of the genus are widely used in crude drug preparation. The present study is an effort to understand the inter and intraspecific plastome variations within genus Capparis using two newly sequenced plastomes. Plastome representatives from four taxa of Capparis were compared to understand the gene composition and arrangements within the genus. The LSC and SSC regions exhibited high nucleotide variation in Capparis species. The loss of two ndh genes from C. spinosa var. herbacea is reported for the first time. Moreover, we also report pseudogenization of seven genes from the genome of Capparis spinosa var. herbacea. In addition, rpl32-ccsA, atpF-atpH, petA-psbJ, trnL-trnF, trnH-psbA spacer region and one gene ycf1 were identified as potential barcodes for the genus Capparis based on inter and intraspecific variations.

Ectopic Expression of Rv0023 Mediates Isoniazid/Ethionamide Tolerance via Altering NADH/NAD+ Levels in Mycobacterium smegmatis.

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) accounts for nearly 1.2 million deaths per annum worldwide. Due to the emergence of multidrug-resistant (MDR) Mtb strains, TB, a curable and avertable disease, remains one of the leading causes of morbidity and mortality. Isoniazid (INH) is a first-line anti-TB drug while ethionamide (ETH) is used as a second-line anti-TB drug. INH and ETH resistance develop through a network of genes involved in various biosynthetic pathways. In this study, we identified Rv0023, an Mtb protein belonging to the xenobiotic response element (XRE) family of transcription regulators, which has a role in generating higher tolerance toward INH and ETH in Mycobacterium smegmatis (Msmeg). Overexpression of Rv0023 in Msmeg leads to the development of INH- and ETH-tolerant strains. The strains expressing Rv0023 have a higher ratio of NADH/NAD+, and this physiological event is known to play a crucial role in the development of INH/ETH co-resistance in Msmeg. Gene expression analysis of some target genes revealed reduction in the expression of the ndh gene, but no direct interaction was observed between Rv0023 and the ndh promoter region. Rv0023 is divergently expressed to Rv0022c (whiB5) and we observed a direct interaction between the recombinant Rv0023 protein with the upstream region of Rv0022c, confirmed using reporter constructs of Msmeg. However, we found no indication that this interaction might play a role in the development of INH/ETH drug tolerance.

Plastid phylogenomic data yield new and robust insights into the phylogeny of Cleisostoma–Gastrochilus clades (Orchidaceae, Aeridinae)

The Cleisostoma–Gastrochilus clades are among the most speciose and diverse groups of Asian orchids and are a taxonomically problematic group. Phylogenetic relationships among the genera of these clades have remained unresolved with traditional sequences from one or a few markers. We present large-scale phylogenomic data sets, incorporating complete chloroplast genome sequences from 53 species (including 41 species sequenced in this study), to compare plastome structure and to resolve the phylogenetic relationships of these clades. The plastomes of Cleisostoma–Gastrochilus clades possessed the quadripartite structure and plastome genes of typical angiosperms with sizes ranging from 142 to 149 kb and encoding a set of 118–120 genes. Unusual structural features were detected in the plastome of Uncifera acuminata, including the presence of a large 17-kb inversion (19 genes) in the Large Single-Copy region and the loss of the rpl32 gene in Cleisostoma fuerstenbergianum. The pseudogenization of ndh genes was widespread in these clades. Phylogenomic analyses, including 68 plastid protein-coding genes, showed that these clades can be subdivided into three major groupings and six subgroupings: Vandopsis undulata, the Gastrochilus clade (including the Trichoglottis and Gastrochilus subclades) and the Cleisostoma clade (including the Vandopsis, Diploprora, Cleisostoma and Schoenorchis subclades). Two genera, Vandopsis and Cleisostoma, were not monophyletic. A new genus, Cymbilabia, was proposed to avoid non-monophyly of Vandopsis. Our results demonstrate the power of plastid phylogenomics to improve the phylogenetic relationships of intricate groups and provide new insight into plastome evolution in Orchidaceae.

Complete Chloroplast Genome of Paphiopedilum delenatii and Phylogenetic Relationships among Orchidaceae.

Paphiopedilum delenatii is a native orchid of Vietnam with highly attractive floral traits. Unfortunately, it is now listed as a critically endangered species with a few hundred individuals remaining in nature. In this study, we performed next-generation sequencing of P. delenatii and assembled its complete chloroplast genome. The whole chloroplast genome of P. delenatii was 160,955 bp in size, 35.6% of which was GC content, and exhibited typical quadripartite structure of plastid genomes with four distinct regions, including the large and small single-copy regions and a pair of inverted repeat regions. There were, in total, 130 genes annotated in the genome: 77 coding genes, 39 tRNA genes, 8 rRNA genes, and 6 pseudogenes. The loss of ndh genes and variation in inverted repeat (IR) boundaries as well as data of simple sequence repeats (SSRs) and divergent hotspots provided useful information for identification applications and phylogenetic studies of Paphiopedilum species. Whole chloroplast genomes could be used as an effective super barcode for species identification or for developing other identification markers, which subsequently serves the conservation of Paphiopedilum species.

Plastome evolution and phylogeny of subtribe Aeridinae (Vandeae, Orchidaceae)

Subtribe Aeridinae (Vandeae, Epidendroideae, Orchidaceae) consists of 83 genera and 2,345 species. The present study completely decoded the plastomes and nuclear ribosomal (nr) RNA gene clusters of seven species of Aeridinae belonging to Gastrochilus, Neofinetia, Pelatantheria, and Thrixspermum and compared them with existing data to investigate their genome evolution and phylogeny. Although no large structural variations were observed among the Aeridinae plastomes, 14 small inversions (SI) were found in Orchidaceae for the first time. Therefore, the evolutionary trends and usefulness of SI as molecular identification markers were evaluated. Since all 11 ndh genes in the Aeridinae plastome were lost or pseudogenized, the evolutionary trends of ndh genes are discussed at the tribe and family levels. In the maximum likelihood tree reconstructed from 83 plastome genes, the five Orchidaceae subfamilies were shown to have diverged in the following order: Apostasioideae, Vanilloideae, Cypripedioideae, Orchioideae, Epidendroideaeae. Divergence times for major lineages were found to be more recent, 5–10 Mya, than previous studies, which only used two or three genes. Vandeae, which includes Aeridinae, formed a sister group with Cymbidieae and Epidendreae. The Vandeae, Cymbidieae, and Epidendreae lineages were inferred to have diverged at 25.31 Mya; thus, numerous speciation events within Aeridineae occurred since then. Furthermore, the present study reconstructed a phylogenetic tree from 422 nrITS sequences belonging to Aerdinae and allied taxa and uses it to discuss the phylogenetic positions and species identities of five endangered species.