Transcriptomic comparison reveals modifications in gene expression, photosynthesis, and cell wall in woody plant as responses to external pH changes.

Soil acidification is one of the crucial global environmental problems, affecting sustainable land use, crop yield, and ecosystem stability. Previous research reported the tolerance of crops to acid soil stress. However, the molecular response of woody plant to acid conditions remains largely unclear. Rhododendron L. is a widely distributed woody plant genus and prefers to grow in acidic soils. Herein, weighted gene coexpression network analysis was performed on R. protistum var. giganteum seedlings subjected to five pH treatments (3.5, 4.5, 5.5, 6.0, 7.0), and their ecophysiological characteristics were determined for the identification of their molecular responses to acidic environments. Through pairwise comparison, 855 differentially expressed genes (DEGs) associated with photosynthesis, cell wall, and phenylpropanoid metabolism were identified. Most of the DEGs related to photosynthesis and cell wall were up-regulated after pH 4.5 treatment. Results implied that the species improves its photosynthetic abilities and changes its cell wall characteristics to adapt to acidic conditions. Weighted gene co-expression network analyses showed that most of the hub genes were annotated to the biosynthetic pathways of ribosomal proteins and photosynthesis. Expression pattern analysis showed that genes encoding subunit ribosomal proteins decreased at pH 7.0 treatment, suggesting that pH 7.0 treatment led to cell injury in the seedlings. The species regulates protein synthesis in response to high pH stress (pH 7.0). The present study revealed the molecular response mechanism of woody plant R. protistum var. giganteum to acid environments. These findings can be useful in enriching current knowledge of how woody species adapt to soil acidification under global environmental changes.

Alpine Rhododendron population contractions lead to spatial distribution mismatch with their pollinators under climate change.

The effect of global climate change on plant-pollinator interaction is not limited to changes in phenology and richness within communities but also includes the spatial mismatch caused by the inconsistency of geographical distribution changes. Subsequently, the pollinator interaction network may be remodeled or even disrupted. In this study, we simulated the suitable habitat niche of 15 Rhododendron species and their eight pollinator species as well as their overlapping versus geographical mismatch under the current and three future climate change scenarios in 2090s, using MaxEnt. Results showed that the suitable habitat of all Rhododendron species would decrease in 2090s. In particular, 10, 8, and 13 Rhododendron-pollinator assemblages would have a reduced spatial match region under the climate change scenarios, mainly due to the contraction of the suitable habitat of Rhododendron species. The results provide novel insights into the response of plant-pollinator interactions to global warming, useful to prioritize conservation actions of alpine plant ecosystems.

[Isolation and diversity analyses of endophytic fungi from Paris polyphylla var. yunnanensis].

The paper is aimed at studying the diversity of endophytic fungi community from Paris polyphylla var. yunnanensis, and to provide a scientific basis for the utilization value of the endophytic fungi as bioactive material resources. In the present study, endophytic fungi were isolated from roots, rhizomes and leaves of wild P. polyphylla var. yunnanensis collected from Baoshan, Heqing county and Songming city of Yunnan province, and identified and classified by morphological methods together with its ITS sequence analysis. Seven and forty-nine strains of endophytic fungi were isolated from P. polyphylla var. yunnanensis. They were identified belonging to 41 genus. In these 41 genus, 3 genus exist in root only, 12 genus only exist in rhizome and 8 genus only exist in leaf. There was difference in endophytic fungi isolated from different sample sites. Endophytic fungi diversity from rhizomes of Heqing site was the highest. Endophytic fungi similarity coefficient was low among different sites and tissues. Based on these results, it is reasonable to propose that endophytic fungi of P. polyphylla var. yannanensis from different tissue and different sample sites has a certain difference which is possibly relate to their different habitats, different structure and composition of each tissue.

Gene co-expression networks unravel the molecular responses of freshwater hydrophytes to combined stress of salinity and cadmium.

Salinization in freshwater lakes is becoming a serious global environmental problem, especially in lakes of plateaus such as south-western plateau of China. However, limited information is available about the molecular response of freshwater hydrophytes to salinity under multiple stress. In the present study, a weighted gene co-expression network (WGCNA) was used to identify the modules of co-expressed genes in the physiological and biochemical indicators of Pistia stratiotes to determine its molecular response to salinity (NaCl) alone and when combined with cadmium (Cd). The physiological and biochemical indicators showed that P. stratiotes improved its salt tolerance by enhancing photosynthetic abilities, reducing oxidative stress, and inducing osmoprotectant generation. Morever, addition of NaCl reduced the Cd accumulation in P. stratiotes. Transcriptome and WGCNA analysis revealed that the pathways of alpha-linolenic acid metabolism, ribosomal, flavonoid biosynthesis, and phenylpropanoid biosynthesis were significantly enriched in both treatments. Genes associated with photosynthesis-antenna proteins, nitrogen metabolism, and the acid cycle pathways were only expressed under salinity stress alone, while the proteasome pathway was only significantly enriched in the combined salinity and Cd treatment. Our findings provide novel insights into the effects of salinization on aquatic plants in freshwater ecosystems and the management of aquatic ecosystems under global change.

Seed morphology of Hypericum (Hypericaceae) in China and its taxonomic significance.

The seed morphology of 40 taxa within the genus Hypericum (Hypericaceae) from China, representing 9 sections of the genus, was examined using both Light and Scanning Electron Microscopy to evaluate the taxonomic relevance of macro- and micro-morphological features. Details articulating variation in seed size, color, shape, appendages, and seed coat ornamentation are described, illustrated, and compared, and their taxonomic importance is discussed. Seeds were generally brown in color and cylindric-ellipsoid to prolonged cylindric in shape. Seed size displayed wide variation, ranging from 0.37-1.91 mm in length and 0.12-0.75 mm in width. Seed appendages were observed as a characteristic morphological feature. Seed surface ornamentation has high phenotypic plasticity, and four types (reticulate, foveolate, papillose, and ribbed) can be recognized. In general, seed color and shape have limited taxonomic significance. However, some other features represent informative characters that can be used efficiently in distinguishing the studied taxa at the section and/or species levels. The findings illustrate that considerable taxonomic knowledge can be obtained by investigating the seed features of Hypericum, and the use of Scanning Electron Microscopy can reveal inconspicuous morphological affinities among species and play a role in taxonomic and systematic studies of the genus Hypericum. RESEARCH HIGHLIGHTS: Macro- and micro-morphological features of seeds of 40 Hypericum taxa from China were examined using Light and Scanning Electron Microscopy, providing the first broad study regarding seed morphology for Hypericum from China. Details and variations of seed size, shape, color, surface ornamentation, and appendages are fully presented. Seed features and their variation have important taxonomic significance at the section and/or species levels within Hypericum.

Transcriptomic and metabolomic analyses reveal the altitude adaptability and evolution of different-colored flowers in alpine Rhododendron species.

Understanding the molecular mechanisms and evolutionary process of plant adaptation to the heterogeneous environment caused by altitude gradients in plateau mountain ecosystems can provide novel insight into species' responses to global changes. Flower color is the most conspicuous and highly diverse trait in nature. Herein, the gene expression patterns, evolutionary adaptation and metabolites changes of different-colored flowers of alpine Rhododendron L. species along altitude gradients were investigated based on a combined analysis of transcriptomics and metabolomics. Differentially expressed genes were found to be related to the biosynthesis of carbohydrates, fatty acids, amino acids and flavonoids, suggesting their important roles in the altitude adaptability of Rhododendron species. The evolution rate of high-altitude species was faster than that of low-altitude species. Genes related to DNA repair, mitogen-activated protein kinase and ABA signal transduction, and lipoic acid and propanoate metabolism were positively selected in the flowers of high-altitude Rhododendron species and those associated with carotenoid biosynthesis pathway, ABA signal transduction and ethylene signal transduction were positively selected in low-altitude species. These results indicated that the genes with differentiated expressions or functions exhibit varying evolution during the adaptive divergence of heterogeneous environment caused by altitude gradients. Flower-color variation might be attributed to the significant differences in gene expression or metabolites related to sucrose, flavonoids and carotenoids at the transcription or metabolism levels of Rhododendron species. This work suggests that Rhododendron species have multiple molecular mechanisms in their adaptation to changing environments caused by altitude gradients.

Gene Co-expression Network and Regression Analysis Identify the Transcriptomic, Physiological, and Biochemical Indicators of the Response of Alpine Woody Plant Rhododendron rex to Drought Stress.

Increasing severity of drought stress due to global change and extreme weather has been affecting the biodiversity, function, and stability of forest ecosystems. However, despite being an important component in the alpine and subalpine vegetation in forest ecosystems, Rhododendron species have been paid rare attention in the study of molecular mechanism of tolerance or response to drought. Herein, we investigated the correlation of transcriptomic changes with the physiological and biochemical indicators of Rhododendron rex under drought stress by using the co-expression network approach and regression analysis. Compared with the control treatment, the number of significantly differentially expressed unigenes (DEGs) increased with the degree of drought stress. The DEGs were mainly enriched in the cell wall metabolic process, signaling pathways, sugar metabolism, and nitrogen metabolism. Coupled analysis of the transcriptome, physiological, and biochemical parameters indicated that the metabolic pathways were highly correlated with the physiological and biochemical indicators under drought stress, especially the chlorophyll fluorescence parameters, such as the actual photosynthetic efficiency of photosystem II, electron transport rate, photochemical quenching coefficient, and the maximum quantum efficiency of photosystem II photochemistry. The majority of the response genes related to the metabolic pathways, including photosynthesis, sugar metabolism, and phytohormone signal pathway, were highly expressed under drought stress. In addition, genes associated with cell wall, pectin, and galacturonan metabolism also played crucial roles in the response of R. rex to drought stress. The results provided novel insight into the molecular response of the alpine woody species under drought stress and may improve the understanding of the response of forest ecosystems to the global climate change.

The complete chloroplast genome sequence of endangered plant Trachycarpus nanus (Arecaceae).

Trachycarpus nanus is an endangered plant that is endemic to southwest of China. In the present study, the complete chloroplast genome of this species was assembled and characterized using whole genome next-generation sequencing. The complete chloroplast genome showed a circular genome of 158,713 bp size with 36.6% GC content. The genome is of typical structure and contain a pair of inverted repeat (IR) regions with 27,240 bp, separated by one large single-copy (LSC) with 86,395 bp, and one small single-copy (SSC) regions with 17,838 bp. The genome contained 132 genes, including 86 protein-coding genes, 8 rRNA genes and 38 tRNA genes. A phylogenetic tree reconstructed based on 21 chloroplast genomes reveals that Trachycarpus nanus is most related with Chamaerops humilis. The information provides important genetic basis for the species' future studies on phylogenetic and utilization.

Genetic Diversity and Population Structure of Rhododendron rex Subsp. rex Inferred from Microsatellite Markers and Chloroplast DNA Sequences.

Genetic diversity is vital to the sustainable utilization and conservation of plant species. Rhododendron rex subsp. rex Lévl. is an endangered species endemic to the southwest of China. Although the natural populations of this species are facing continuous decline due to the high frequency of anthropogenic disturbance, the genetic information of R. rex subsp. rex is not yet elucidated. In the present study, 10 pairs of microsatellite markers (nSSRs) and three pairs of chloroplast DNA (cpDNAs) were used in the elucidation of the genetic diversity, population structure, and demographic history of 11 R. rex subsp. rex populations. A total of 236 alleles and 12 haplotypes were found. A moderate genetic diversity within populations (HE = 0.540 for nSSRs, Hd = 0.788 for cpDNA markers), high historical and low contemporary gene flows, and moderate genetic differentiation (nSSR: FST = 0.165***; cpDNA: FST = 0.841***) were detected among the R. rex subsp. rex populations. Genetic and geographic distances showed significant correlation (p < 0.05) determined by the Mantel test. The species exhibited a conspicuous phylogeographical structure among the populations. Using the Bayesian skyline plot and species distribution models, we found that R. rex subsp. rex underwent a population demography contraction approximately 50,000-100,000 years ago. However, the species did not experience a recent population expansion event. Thus, habitat loss and destruction, which result in a population decline and species inbreeding depression, should be considered in the management and conservation of R. rex subsp. rex.

The complete chloroplast genome of a species Cansjera rheedei (Opiliaceae).

In this study, Cansjera rheedei J. F. Gmelin is an important role in the phylogeny and evolution of Opiliaceae plant. The chloroplast genome of C. rheedei is 144,306 bp in size, with an average GC content of 37.5%. The complete chloroplast genome has a typical quadripartite structure, including a large single copy (LSC) region (82,773 bp) and a small single copy (SSC) region (9745 bp), which were separated a pair of inverted repeats (IRs, 25,894 bp). This plastome contained 101 different genes, including 67 protein-coding genes (PCGs), 30 tRNA genes and four rRNA genes. The chloroplast genome of C. rheedei has completed that will be based on the phylogeny and genomic studies in the family Opiliaceae.

Characterization of the complete chloroplast genome sequence of submerged macrophyte Stuckenia pectinata (Potamogetonaceae) and its phylogenetic position.

Stuckenia pectinata is widely distributed submerged macrophyte in the world. Herein, the complete chloroplast genome of this species was assembled and characterized using whole genome next-generation sequencing. The complete chloroplast genome showed a circular genome of 156,669 bp size with 36.5% GC content. The genome is of typical structure and contain a pair of inverted repeat (IR) regions with 26,074 bp, separated by one large single-copy (LSC) with 86,285 bp, and one small single-copy (SSC) regions with 18,236 bp. De novo assembly and annotation showed the presence of 131 unique genes with 85 protein-coding genes, 38 tRNA genes, and eight rRNA genes. A maximum-likelihood phylogenomic tree reconstructed based on 15 chloroplast genomes reveals that S. pectinata is most closely related to Zostera marina.

The complete chloroplast genome of Cinnamomum pittosporoides reveals its phylogenetic relationship in Lauraceae.

Cinnamomum pittosporoides is an important timber plant endemic to southwest of China. In the present study, we sequenced the complete chloroplast genome of C. pittosporoides and used the data to reveal the species phylogenetic in Lauraceae. The complete chloroplast genome showed a circular genome of 152,730 bp size with 39.2% GC content. The genome is of typical structure and contains a pair of inverted repeat (IR) regions with 20,074 bp, separated by one large single-copy (LSC) with 93,722 bp and one small single-copy (SSC) regions with 18,860 bp. The genome contained 116 genes, including 82 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. A phylogenetic tree reconstructed based on 26 chloroplast genomes reveals that C. pittosporoides is most related with C. chago in Lauraceae.

Characterization of the complete chloroplast genome sequence of wetland macrophyte Typha orientalis (Typhaceae).

Typha orientalis is an important wetland macrophyte native to the eastern parts of Asia and Oceania. Herein, the complete chloroplast genome of this species was assembled and characterized using whole-genome next-generation sequencing. The complete chloroplast genome showed a circular genome of 160,969 bp size with 36.6% GC content. The genome is of typical structure and contains a pair of inverted repeat (IR) regions with 26,691 bp, separated by one large single-copy (LSC) with 89,118 bp, and one small single-copy (SSC) regions with 18,469 bp. The genome contained 132 genes, including 86 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. A phylogenetic tree reconstructed based on 15 chloroplast genomes reveals that T. orientalis is most related to Typha latifolia.

Transcriptome Analysis of Cinnamomum chago: A Revelation of Candidate Genes for Abiotic Stress Response and Terpenoid and Fatty Acid Biosyntheses.

Cinnamomum chago, an endangered species endemic to Yunnan province, possesses large economic and phylogenetic values in Lauraceae. However, the genomic information of this species remains relatively unexplored. In this study, we used RNAseq technology to characterize and annotate the C. chago transcriptome and identify candidate genes involved in special metabolic pathways and gene-associated simple sequence repeats (SSRs) and single-nucleotide polymorphism (SNP). A total of 129,097 unigenes, with a mean length of 667 bp and an N50 length of 1,062 bp, were assembled. Among these genes, 56,887 (44.07%) unigenes were successfully annotated using at least one database. Furthermore, 47 and 46 candidate genes were identified in terpenoid biosynthesis and fatty acid biosynthesis, respectively. A total of 22 candidate genes participated in at least one abiotic stress response of C. chago. Additionally, a total of 25,654 SSRs and 640 SNPs were also identified. Based on these potential loci, 55 novel expressed sequence tag (EST)-SSR primers were successfully developed. This work provides comprehensive transcriptomic data that can be used to establish a valuable information platform for gene prediction, signaling pathway investigation, and molecular marker development for C. chago and other related species. Such a platform can facilitate further studies on germplasm conservation and utilization of Lauraceae species.

Physiological epicotyl dormancy and its alleviation in seeds of Yunnanopilia longistaminea: the first report of physiological epicotyl dormancy in China.

Yunnanopilia longistaminea is an endangered monotypic species belonging to Opiliaceae. This edible plant is an important germplasm source with a high economic value in China if propagation were less difficult. Seed dormancy and germination of this species were investigated to improve propagation. Considering seeds have a fully developed embryo and mature and are dispersed in summer, and radicles and epicotyls emerge the following autumn and next spring, respectively, we hypothesized that Y. longistaminea seeds may undergo physiological epicotyl dormancy. Seed moisture content and viability decreased as dehydration occurred. Thus, the seeds may be recalcitrant. The seed germination of this species involves two stages: radicle emergence and epicotyl (shoot) emergence. The optimum temperature was 28 °C and 28 °C/20 °C to radicle emergence. The optimum GA3 solution for the seeds undergoing shoot emergence was 100 mg L-1. The percentages of shoot emergence in seven and 14 days stratification at 5 °C were slightly higher than those in other groups. This study is the first to describe physiological epicotyl dormancy in Y. longistaminea seeds. Under natural conditions, seeds are subjected to Y. longistaminea a autumn → winter → spring temperature. Warm moist conditions and cold stratification can improve radicle emergence and alleviate epicotyl dormancy, respectively. The duration of cold stratification also significantly affects the epicotyl dormancy release of Y. longistaminea. Optimal dormancy breakage methods are warm (28 °C/20 °C) → cold (5 °C) → GA3(100 mg L-1) → warm (28 °C/20 °C).

De Novo Assembly of Transcriptome and Development of Novel EST-SSR Markers in Rhododendron rex Lévl. through Illumina Sequencing.

Transcriptome sequences generated by next-generation sequencing (NGS) technologies can be utilized to rapidly detect and characterize a large number of gene-based microsatellites from different plants. Rhododendron rex Lévl. is a perennial woody species from the family Ericaceae and an endangered plant with high ornamental value endemic to Southwestern China. Nevertheless, the genetic and genomic information of R. rex remain unknown. In this study, we performed transcriptome sequencing for R. rex leaf samples, and generated large transcript sequences for functional characterization and development gene-associated SSR markers. A total of 164,242 unigenes were assembled and 115,089 (70.07%) unigenes were successfully annotated in public databases. In addition, a total of 15,314 potential EST-SSRs were identified, and the frequency of SSRs in the R. rex unigenes was 9.32%, with an average of one EST-SSR per 5.65 kb. The most abundant type was repeated di-nucleotide (54.63%), followed by mono- (26.03%) and tri-nucleotide (18.51%) repeats. Based on the SSR-containing sequence, 100 primer pairs were randomly selected and synthesized and used for assessment of the polymorphism. Thirty-six primer pairs were polymorphic and revealed polymorphism among 20 individuals from four R. rex populations. A total of 197 alleles were identified, with an average of 5.472 alleles per locus. The Polymorphism Information Content ranged from 0.154 to 0.870, with a mean of 0.482. The newly developed EST-SSR markers exhibited high transferability (58.33-83.33%) among the six subgenera. Thus, these novel EST-SSR markers developed would provide valuable sequence resources for population structure, genetic diversity analysis, and genetic resource assessments of R. rex and its related species.

Genetic diversity and population structure of an extremely endangered species: the world's largest Rhododendron.

Comprehensive studies on the genetic diversity and structure of endangered species are urgently needed to promote effective conservation and management activities. The big tree rhododendron, Rhododendron protistum var. giganteum, is a highly endangered species with only two known endemic populations in a small area in the southern part of Yunnan Province in China. Unfortunately, limited information is available regarding the population genetics of this species. Therefore, we conducted amplified fragment length polymorphism (AFLP) analysis to characterize the genetic diversity and variation of this species within and between remaining populations. Twelve primer combinations of AFLP produced 447 unambiguous and repetitious bands. Among these bands, 298 (66.67 %) were polymorphic. We found high genetic diversity at the species level (percentage of polymorphic loci = 66.67 %, h = 0.240, I = 0.358) and low genetic differentiation (Gst = 0.110) between the two populations. Gene flow between populations (Nm) was relatively high at 4.065. Analysis of molecular variance results revealed that 22 % of the genetic variation was partitioned between populations and 78 % of the genetic variation was within populations. The presence of moderate to high genetic diversity and low genetic differentiation in the two populations can be explained by life history traits, pollen dispersal and high gene flow (Nm = 4.065). Bayesian structure and principal coordinate analysis revealed that 56 sampled trees were clustered into two groups. Our results suggest that some rare and endangered species are able to maintain high levels of genetic diversity even at small population sizes. These results will assist with the design of conservation and management programmes, such as in situ and ex situ conservation, seed collection for germplasm conservation and reintroduction.