Genome-wide identification and comparative analysis of DNA methyltransferase and demethylase gene families in two ploidy Cyclocarya paliurus and their potential function in heterodichogamy.

BACKGROUND: DNA methylation is one of the most abundant epigenetic modifications, which plays important roles in flower development, sex differentiation, and regulation of flowering time. Its pattern is affected by cytosine-5 DNA methyltransferase (C5-MTase) and DNA demethylase (dMTase). At present, there are no reports on C5-MTase and dMTase genes in heterodichogamous Cyclocarya paliurus. RESULTS: In this study, 6 CpC5-MTase and 3 CpdMTase genes were identified in diploid (2n = 2 × = 32) C. paliurus, while 20 CpC5-MTase and 13 CpdMTase genes were identified in autotetraploid (2n = 4 × = 64). 80% of identified genes maintained relatively fixed positions on chromosomes during polyploidization. In addition, we found that some DRM subfamily members didn't contain the UBA domain. The transcript abundance of CpC5-MTase and CpdMTase in male and female flowers of two morphs (protandry and protogyny) from diploidy was analyzed. Results showed that all genes were significantly up-regulated at the stage of floral bud break (S2), but significantly down-regulated at the stage of flower maturation (S4). At S2, some CpC5-MTase genes showed higher expression levels in PG-M than in PG-F, whereas some CpdMTase genes showed higher expression levels in PA-M than in PA-F. In addition, these genes were significantly associated with gibberellin synthesis-related genes (e.g. DELLA and GID1), suggesting that DNA methylation may play a role in the asynchronous floral development process through gibberellin signal. CONCLUSIONS: These results broaden our understanding of the CpC5-MTase and CpdMTase genes in diploid and autotetraploid C. paliurus, and provide a novel insight into regulatory mechanisms of DNA methylation in heterodichogamy.

Transcriptome Analysis Reveals the Role of GA3 in Regulating the Asynchronism of Floral Bud Differentiation and Development in Heterodichogamous Cyclocarya paliurus (Batal.) Iljinskaja.

Cyclocarya paliurus is an important medical plant owing to the diverse bioactive compounds in its leaves. However, the heterodichogamy with female and male functions segregation within protandry (PA) or protogyny (PG) may greatly affect seed quality and its plantations for medicinal use. To speculate on the factor playing the dominant role in regulating heterodichogamy in C. paliurus, based on phenotypic observations, our study performed a multi comparison transcriptome analysis on female and male buds (PG and PA types) using RNA-seq. For the female and male bud comparisons, a total of 6753 differentially expressed genes (DEGs) were detected. In addition, functional analysis revealed that these DEGs were significantly enriched in floral development, hormone, and GA-related pathways. As the dominant hormones responsible for floral differentiation and development, gibberellins (GAs) in floral buds from PG and PA types were quantified using HPLC-MS. Among the tested GAs, GA3 positively regulated the physiological differentiation (S0) and germination (S2) of floral buds. The dynamic changes of GA3 content and floral morphological features were consistent with the expression levels of GA-related genes. Divergences of GA3 contents at S0 triggered the asynchronism of physiological differentiation between male and female buds of intramorphs (PA-M vs. PA-F and PG-F vs. PG-M). A significant difference in GA3 content enlarged this asynchronism at S2. Thus, we speculate that GA3 plays the dominant role in the formation of heterodichogamy in C. paliurus. Meanwhile, the expression patterns of GA-related DEGs, including CPS, KO, GA20ox, GA2OX, GID1, and DELLA genes, which play central roles in regulating flower development, coincided with heterodichogamous characteristics. These results support our speculations well, which should be further confirmed.

Succession of the Resident Soil Microbial Community in Response to Periodic Inoculations.

To maintain the beneficial effects of microbial inoculants on plants and soil, repeated inoculation represents a promising option. Until now, the impacts of one-off inoculation on the native microbiome have been explored, but it remains unclear how long and to what extent the periodic inoculations would affect the succession of the resident microbiome in bulk soil. Here, we examined the dynamic responses of plant growth, soil functions, and the resident bacterial community in the bulk soil to periodic inoculations of phosphate-solubilizing and N2-fixing bacteria alone or in combination. Compared to single-strain inoculation, coinoculation better stimulated plant growth and soil nutrients. However, the benefits from inoculants did not increase with repeated inoculations and were not maintained after transplantation to a different site. In response to microbial inoculants, three patterns of shifts in the bacterial composition were observed: fold increase, fold decrease, and resilience. The periodic inoculations impacted the succession course of resident bacterial communities in bulk soil, mainly driven by changes in soil pH and nitrate, resulting in the development of three main cluster types throughout the investigation. The single and mixed inoculants transiently modulated the variation in the resident community in association with soil pH and the C/N ratio, but finally, the community established and showed resilience to subsequent inoculations. Consequently, the necessity of repeated inoculations should be reconsidered, and while the different microbial inoculants showed distinct impacts on resident microbiome succession, the communities ultimately exhibited resilience.IMPORTANCE Introducing beneficial microbes to the plant-soil system is an environmentally friendly approach to improve the crop yield and soil environment. Numerous studies have attempted to reveal the impacts of inoculation on the rhizosphere microbiome. However, little is known about the effectiveness of periodic inoculations on soil functioning. In addition, the long-term impact of repeated inoculations on the native community remains unclear. Here, we track the succession traits of the resident microbiome in the bulk soil across a growing season and identify the taxon clusters that respond differently to periodic inoculation. Crucially, we compare the development of the resident community composition with and without inoculation, thus providing new insight into the interactions between resident microbes and intruders. Finally, we conclude that initial inoculation plays a more important role in influencing the whole system, and the native microbial community exhibits traits of resilience, but no resistance, to the subsequent inoculations.

Genome-Wide Identification MIKC-Type MADS-Box Gene Family and Their Roles during Development of Floral Buds in Wheel Wingnut (Cyclocarya paliurus).

MADS-box transcription factors (TFs) have fundamental roles in regulating floral organ formation and flowering time in flowering plants. In order to understand the function of MIKC-type MADS-box family genes in Cyclocarya paliurus (Batal.) Iljinskaja, we first implemented a genome-wide analysis of MIKC-type MADS-box genes in C. paliurus. Here, the phylogenetic relationships, chromosome location, conserved motif, gene structure, promoter region, and gene expression profile were analyzed. The results showed that 45 MIKC-type MADS-box were divided into 14 subfamilies: BS (3), AGL12 (1), AP3-PI (3), MIKC* (3), AGL15 (3), SVP (5), AGL17 (2), AG (3), TM8 (1), AGL6 (2), SEP (5), AP1-FUL (6), SOC1 (7), and FLC (1). The 43 MIKC-type MADS-box genes were distributed unevenly in 14 chromosomes, but two members were mapped on unanchored scaffolds. Gene structures were varied in the same gene family or subfamily, but conserved motifs shared similar distributions and sequences. The element analysis in promoters' regions revealed that MIKC-type MADS-box family genes were associated with light, phytohormone, and temperature responsiveness, which may play important roles in floral development and differentiation. The expression profile showed that most MIKC-type MADS-box genes were differentially expressed in six tissues (specifically expressed in floral buds), and the expression patterns were also visibly varied in the same subfamily. CpaF1st24796 and CpaF1st23405, belonging to AP3-PI and SEP subfamilies, exhibited the high expression levels in PA-M and PG-F, respectively, indicating their functions in presenting heterodichogamy. We further verified the MIKC-type MADS-box gene expression levels on the basis of transcriptome and qRT-PCR analysis. This study would provide a theoretical basis for classification, cloning, and regulation of flowering mechanism of MIKC-type MADS-box genes in C. paliurus.

Geographic Variation in the Chemical Composition and Antioxidant Properties of Phenolic Compounds from Cyclocarya paliurus (Batal) Iljinskaja Leaves.

Cyclocarya paliurus has been widely used as an ingredient in functional foods in China. However, the antioxidant properties of phenolic compounds and the effect of the plant origin remain unclear. The present study evaluated the geographical variation of this plant in term of its phenolic composition and antioxidant activities based on leaf materials collected from five regions. high-performance liquid chromatography (HPLC) analysis showed that there are three major components, quercetin-3-O-glucuronide, kaempferol-3-O-glucuronide, and kaempferol-3-O-rhamnoside, and their contents varied significantly among sampling locations. The investigated phenolic compounds showed substantial antioxidant activities, both in vitro and in vivo, with the highest capacity observed from Wufeng and Jinzhongshan. Correlation analysis revealed that quercetin and kaempferol glycosides might be responsible for the antioxidant activities. Our results indicate the importance of geographic origin, with sunny hours and temperature as the main drivers affecting the accumulation of C. paliurus phenolics and their antioxidant properties.

Antidiabetic Effect of Cyclocarya paliurus Leaves Depends on the Contents of Antihyperglycemic Flavonoids and Antihyperlipidemic Triterpenoids.

Cyclocarya paliurus has been used commonly to treat diabetes in China. However, the effective components and the effect of plant origin remain unclear. In this study, C. paliurus leaves with different chemical compositions were selected from five geographical locations, and their effects on streptozotocin (STZ)-induced diabetic mice were evaluated with both ethanol and aqueous extracts. Glucose levels, lipid levels, and biomarkers of liver and kidney function were measured. The principal components of both C. paliurus ethanol and aqueous extracts from different geographical locations differed quantitatively and qualitatively. Results showed that C. paliurus extracts with better antihyperglycemic effects were characterized by higher contents of total flavonoids, especially quercetin-3-O-glucuronide and kaempferol-3-O-glucuronide. Furthermore, significantly negative correlations were found between triterpenoids contents and lipid levels. These results revealed the potential antihyperglycemic capacity of C. paliurus flavonoids and the antihyperlipidemic effect of C. paliurus triterpenoids. Thus, we suggest that the composition of C. paliurus compounds might help to design therapeutic alternatives for the treatment of diabetes mellitus. However, geographic origins and the extraction solvents can also affect the effectiveness of the treatment as these factors influence the chemical compositions and thereby the biological activities.

Chemical Fingerprint and Multicomponent Quantitative Analysis for the Quality Evaluation of Cyclocarya paliurus Leaves by HPLC-Q-TOF-MS.

Cyclocarya paliurus is an edible and medicinal plant containing various bioactive components with significant health benefits. A combinative method using high-performance liquid chromatography (HPLC) fingerprint and quantitative analysis was developed and successfully applied for characterization and quality evaluation of C. paliurus leaves collected from 18 geographical locations of China. For the fingerprint analysis, 21 common peaks were observed among the 18 samples, and these peaks were identified by high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS), while a simultaneous quantification of 16 markers was conducted to interpret the variations of contents of these bioactive compounds among the C. paliurus leaves from different geographical locations. Quantification results showed that the contents of these sixteen investigated compounds varied greatly among the leaves from different locations. The developed new method would be a valuable reference for further study and development of this bioactive plant.

Insight into farming native microbiome by bioinoculant in soil-plant system.

Applying beneficial microorganisms (BM) as bioinoculants presents a promising soil-amendment strategy while impacting the native microbiome, which jointly alters soil-plant performance. Leveraging the untapped potential of native microbiomes alongside bioinoculants may enable farmers to sustainably regulate soil-plant systems via natural bioresources. This review synthesizes literature on native microbiome responses to BMs and their interactive effects on soil and plant performance. We highlight that native microbiomes harbor both microbial "helpers" that can improve soil fertility and plant productivity, as well as "inhibitors" that hinder these benefits. To harness the full potential of resident microbiome, it is crucial to elucidate their intricate synergistic and antagonistic interplays with introduced BMs and clarify the conditions that facilitate durable BM-microbiome synergies. Hence, we indicate current challenges in predicting these complex microbial interactions and propose corresponding strategies for microbiome breeding via BM bioinoculant. Overall, fully realizing the potential of BMs requires clarifying their interactions with native soil microbiomes and judiciously engineering microbiome to harness helpful microbes already present within agroecosystems.

Optimizing tradeoff strategies of soil microbial community between metabolic efficiency and resource acquisition along a natural regeneration chronosequence.

The tradeoff between community-level soil microbial metabolic efficiency and resource acquisition strategies during natural regeneration remains unclear. Herein, we examined variations in soil extracellular enzyme activity, microbial metabolic quotient (qCO2), and microbial carbon use efficiency (CUE) along a chronosequence of natural regeneration by sampling secondary forests at 1, 10, 20, 30, 40, and 100 years after rubber plantation (RP) clearance. The results showed that the natural logarithms of carbon (C)-, nitrogen (N)-, and phosphorus (P)-acquiring enzyme activities were 1:1.68:1.37 and 1:1.54:1.38 in the RP and secondary forests, respectively, thus demonstrating that microbial metabolism was co-limited by N and P. Moreover, the soil microbial C limitation initially increased (1-40 years) and later decreased (100 years). Overall, the qCO2 increased, decreased, and then increased again in the initial (< 10 years), middle (10-40 years), and late (100 years) successional stages, respectively. Except for specific P-acquiring enzyme activities, the changes in other indicators with natural regeneration were consistent in the dry and wet seasons. Both qCO2 and CUE were mainly predicted by microbial community composition and physiological traits. These results indicate that soil microbial communities could employ tradeoff strategies between metabolic efficiency and resource acquisition to cope with variations in resources. Our findings provide new information on tradeoff strategies between metabolic efficiency and resource acquisition during natural regeneration.

Genome assembly of two diploid and one auto-tetraploid Cyclocarya paliurus genomes.

Cyclocarya paliurus, an endemic species in the genus Juglandaceae with the character of heterodichogamy, is one of triterpene-rich medicinal plants in China. To uncover the genetic mechanisms behind the special characteristics, we sequenced the genomes of two diploid (protandry, PA-dip and protogyny, PG-dip) and one auto-tetraploid (PA-tetra) C. paliurus genomes. Based on 134.9 (~225x), 75.5 (~125x) and 271.8 Gb (~226x) subreads of PacBio platform sequencing data, we assembled 586.62 Mb (contig N50 = 1.9 Mb), 583.45 Mb (contig N50 = 1.4 Mb), and 2.38 Gb (contig N50 = 430.9 kb) for PA-dip, PG-dip and PA-tetra genome, respectively. Furthermore, 543.53, 553.87, and 2168.65 Mb in PA-dip, PG-dip, and PA-tetra, were respectively anchored to 16, 16, and 64 pseudo-chromosomes using over 65.4 Gb (~109x), 68 Gb (~113x), and 264 (~220x) Hi-C sequencing data. Annotation of PA-dip, PG-dip, and PA-tetra genome assembly identified 34,699, 35,221, and 34,633 protein-coding genes (90,752 gene models) or allele-defined genes, respectively. In addition, 45 accessions from nine locations were re-sequenced, and more than 10 × coverage reads were generated.

Squalene epoxidase (SE) gene related to triterpenoid biosynthesis assists to select elite genotypes in medicinal plant: Cyclocarya paliurus (Batal.) Iljinskaja.

Triterpenoids, known for their anti-inflammatory, anticancer, and hypoglycemic properties, are the major bioactive components in Cyclocarya paliurus (Batal.) Iljinskaja. Selecting elite individuals with high triterpenoids content is the basis of C. paliurus industry for medicinal use. In this study, seasonal variation patterns of total triterpenoids and five triterpene monomers accumulation for three groups with different total triterpenoid contents (TTC; H: 59.74-64.03 mg g-1; M: 47.66-57.08 mg g-1, and L: 35.26-42.22 mg g-1) were surveyed. Seasonal expression dynamics of 6 key genes relevant to triterpenoids biosynthesis, including HMGR, DXR, SQS, SE, LUS, and ß-AS, were described by quantitative real-time PCR (qRT-PCR) for three groups. The expression levels of HMGR, SE, LUS, and ß-AS genes in group H were higher than in groups M and L. In addition, Pearson correlation analysis showed that they were significantly positively correlated with triterpene accumulation, and the expression level of SE gene not only was significantly correlated with downstream genes, but also exhibited a linear relationship with TTC, especially in September. These results suggest that SE gene could serve as an effective make for screening elite individuals with high TTC from the germplasm of C. paliurus for medicinal use. Further testing on randomly selected individuals in next September proved the feasibility and reliability of SE gene in assisted selection. Also, we successfully cloned the full-length cDNA of SE. Thus, our work provides an efficient way to attain superior genotypes to develop medicinal industry of C. paliurus in practice.

Two new seco-abietane type diterpenoids from the leaves of Isodon serra.

Two new seco-abietane type diterpenoids, named as isodonserra acid A and B (1-2), along with six known compounds, angustanoic acid A (3), epipalustric acid (4), raserrane (5), 7-methoxy coumarin (6), umbelliferone (7), and (-)-loliolide (8), were obtained from the leaves of Isodon serra. The new structures of compounds 1 and 2 were elucidated by analysing their 1D NMR, 2D NMR and HR-ESI-MS spectra. Compounds 1-8 showed moderate hepatoprotective activity against APAP-induced HepG2 cell injury with a cell survival rate from 50.4% to 78.7% at a concentration of 10 µM (p < .001, bicyclol as the positive drug, 71.7%).

Construction of the dynamic model of SCI rehabilitation using bidirectional stimulation and its application in rehabilitating with BCI.

Patients with complete spinal cord injury have a complete loss of motor and sensory functions below the injury plane, leading to a complete loss of function of the nerve pathway in the injured area. Improving the microenvironment in the injured area of patients with spinal cord injury, promoting axon regeneration of the nerve cells is challenging research fields. The brain-computer interface rehabilitation system is different from the other rehabilitation techniques. It can exert bidirectional stimulation on the spinal cord injury area, and can make positively rehabilitation effects of the patient with complete spinal cord injury. A dynamic model was constructed for the patient with spinal cord injury under-stimulation therapy, and the mechanism of the brain-computer interface in rehabilitation training was explored. The effects of the three current rehabilitation treatment methods on the microenvironment in a microscopic nonlinear model were innovatively unified and a complex system mapping relationship from the microscopic axon growth to macroscopic motor functions was constructed. The basic structure of the model was determined by simulating and fitting the data of the open rat experiments. A clinical rehabilitation experiment of spinal cord injury based on brain-computer interface was built, recruiting a patient with complete spinal cord injury, and the rehabilitation training and follow-up were conducted. The changes in the motor function of the patient was simulated and predicted through the constructed model, and the trend in the motor function improvement was successfully predicted over time. This proposed model explores the mechanism of brain-computer interface in rehabilitating patients with complete spinal cord injury, and it is also an application of complex system theory in rehabilitation medicine. Supplementary Information: The online version contains supplementary material available at 10.1007/s11571-022-09804-3.

Whole-genome Duplication Reshaped Adaptive Evolution in A Relict Plant Species, Cyclocarya paliurus.

Cyclocarya paliurus is a relict plant species that survived the last glacial period and shows a population expansion recently. Its leaves have been traditionally used to treat obesity and diabetes with the well-known active ingredient cyclocaric acid B. Here, we presented three C. paliurus genomes from two diploids with different flower morphs and one haplotype-resolved tetraploid assembly. Comparative genomic analysis revealed two rounds of recent whole-genome duplication events and identified 691 genes with dosage effects that likely contribute to adaptive evolution through enhanced photosynthesis and increased accumulation of triterpenoids. Resequencing analysis of 45 C. paliurus individuals uncovered two bottlenecks, consistent with the known events of environmental changes, and many selectively swept genes involved in critical biological functions, including plant defense and secondary metabolite biosynthesis. We also proposed the biosynthesis pathway of cyclocaric acid B based on multi-omics data and identified key genes, in particular gibberellin-related genes, associated with the heterodichogamy in C. paliurus species. Our study sheds light on evolutionary history of C. paliurus and provides genomic resources to study the medicinal herbs.

Responses of Physiology, Photosynthesis, and Related Genes to Saline Stress in Cornus hongkongensis subsp. tonkinensis (W. P. Fang) Q. Y. Xiang.

Cornus hongkongensis subsp. tonkinensis (W. P. Fang) Q. Y. Xiang is a native evergreen species with high ornamental value for abundant variations in leaf, bract, fruit, and tree gesture. To broaden its cultivation in coastal saline soil, salt damage and survival rate, physiological responses, photosynthetic performance, and related genes were evaluated for annual seedlings exposed to 0.3% salt (ST) concentrations for 60 days. Syndromes of salt damage were aggravated, and the survival rate decreased with prolonged stress duration; all stressed seedlings displayed salt damage, and 58.3% survived. Under short-term saline stress (5 d), marked increases in malondialdehyde (MDA), relative electrical conductivity (REC), and decreases in superoxide dismutase (SOD), photosynthetic rate (Pn), stomatal conductance (gs), and internal carbon dioxide concentration (Ci) were recorded. The stable leaf water use efficiency (WUE) and chlorophyll content were positive physiological responses to ensure photosynthetic performance. Meanwhile, the expression levels of genes related to photosystem II (psbA) and photorespiration (SGAT and GGAT) were upregulated, indicating the role of photorespiration in protecting photosynthesis from photoinhibition. After 30 days of stress (≥30 d), there was a significant increase in MDA, REC, soluble sugar (SS), soluble protein (SP), and Ci, whereas descending patterns in Pn, gs, WUE, the maximal photochemical efficiency of photosystem II (Fv/Fm), and potential activities of PSII (Fv/F0) occurred in salt-stressed seedlings, compared with CK. Meanwhile, the expression levels of related genes significantly dropped, such as psbA, LFNR, GGAT, GLYK, and PGK, indicating photoinhibition and worse photosynthetic performance. Our results suggest that the moderate salt tolerance of C. hongkongensis subsp. tonkinensis mostly lies in a better photosynthetic system influenced by active photorespiration. Hence, these results provide a framework for better understanding the photosynthetic responses of C. hongkongensis subsp. tonkinensis to salt stress.

Hexachloro-1,3-butadiene as a Functional Additive for Constructing an Efficient Solid Electrolyte Interface Layer for Long-Life Stable Li Anodes.

Lithium (Li) metal is considered as one of the attractive anodes for next-generation high-energy-density batteries due to its ultrahigh theoretical specific capacity and low potential. However, many great challenges including uncontrolled dendrite growth and undesired side reactions during repeated cycling still seriously hinder its practical application in Li metal secondary batteries. Herein, we report the hexachloro-1,3-butadiene (HCBD) molecule as a functional additive to stabilize the Li anode by forming a stable solid electrolyte interface (SEI) layer with high Li ion conductivity via in situ surface and electrochemical reactions. Density functional theory calculations demonstrate that HCBD can preferentially react with the Li anode, which generates an ionic conducting species (LiCl) into an SEI layer. The LiCl-rich SEI layer effectively regulates Li+ deposition/stripping kinetics and then induces uniform nucleation of Li+ and reduces the side reactions between the Li anode and electrolyte. With an optimal amount of HCBD in an ether-based electrolyte, an excellent cycling lifespan (7000 h) was achieved with a low hysteresis voltage of ∼10 mV at 1.0 mA cm-2 in a Li||Li symmetrical cell. Furthermore, the LiFePO4-based cell with the additive-functionalized Li anode displays obviously improved cycling stability (with a high specific capacity of 141.1 mAh g-1 after 350 cycles at 1 C).

BCI system with lower-limb robot improves rehabilitation in spinal cord injury patients through short-term training: a pilot study.

In the recent years, the increasing applications of brain-computer interface (BCI) in rehabilitation programs have enhanced the chances of functional recovery for patients with neurological disorders. We presented and validated a BCI system with a lower-limb robot for short-term training of patients with spinal cord injury (SCI). The cores of this system included: (1) electroencephalogram (EEG) features related to motor intention reported through experiments and used to drive the robot; (2) a decision tree to determine the training mode provided for patients with different degrees of injuries. Seven SCI patients (one American Spinal Injury Association Impairment Scale (AIS) A, three AIS B, and three AIS C) participated in the short-term training with this system. All patients could learn to use the system rapidly and maintained a high intensity during the training program. The strength of the lower limb key muscles of the patients was improved. Four AIS A/B patients were elevated to AIS C. The cumulative results indicate that clinical application of the BCI system with lower-limb robot is feasible and safe, and has potentially positive effects on SCI patients. Supplementary Information: The online version contains supplementary material available at 10.1007/s11571-022-09801-6.

Comparative analysis of the complete chloroplast genome between tetraploidy and diploidy of Cyclocarya paliurus (Batal.) Iljinskaja.

Cyclocarya paliurus (Batal.) Iljinskaja, a monotypic species in Cyclocarya of Juglandaceae, is regarded as one of important medical plants in China. In order to reveal the alterations in chloroplast (cp) genome with nuclear genome duplication, we presented the complete cp genomes of C. paliurus, and firstly analyzed on the basis of ploidy type (tetraploid and diploid C. paliurus). The total length of the cp genome of tetraploid and diploid C. paliurus is 160,938 and 161,105 bp, respectively. Both type genome consist of a large single-copy (LSC) region (90,221 and 90,391 bp), a small single-copy (SSC) region (18,593 and 18,590 bp), and an pair of invert repeats (IRs) regions (26,062 and 26,062 bp). Tetraploid and diploid plastid genome contain 132 and 137 genes, 87 and 88 protein-coding genes, 37 and 39 tRNA genes, and both eight rRNA genes, respectively. Closely phylogenetic relationship by analyzing 23 cp genomes suggests that tetraploid C. paliurus probably originated from diploid C. paliurus.

Effects of Anti-Vascular Endothelial Growth Factor Drugs Before and After Pars Plana Vitrectomy in Patients with Polypoidal Choroidal Vasculopathy and Vitreous Hemorrhage.

Purpose: To compare the clinical effects of postoperative versus perioperative injection of anti-vascular endothelial growth factor (VEGF) drugs before and after pars plana vitrectomy (PPV) in patients with vitreous hemorrhage secondary to polypoidal choroidal vasculopathy (PCV). Methods: This was a retrospective study of patients who underwent PPV due to vitreous hemorrhage between October 2013 and June 2019 at Ningbo Eye Hospital. The patients who underwent PPV surgery due to PCV-secondary vitreous hemorrhage were included. The primary outcome was the changes in best-corrected visual acuity. The secondary outcome was the central macular thickness. Results: Compared with the postoperative group (n = 20), the perioperative group (n = 18) showed a smaller number of postoperative anti-VEGF injections (5.1 ± 0.8 vs. 8.0 ± 1.5, P < 0.05) and lower frequencies of early hyphema (5.6% vs. 30.0%, P < 0.05), and recurrent vitreous hemorrhage (11.1% vs. 30.0%, P < 0.05). The logarithm of minimal angle resolution (LogMAR) was smaller in the perioperative group compared with the postoperative group at 1 week, 1 month, and 3 months after PPV (P < 0.05), but there were no differences thereafter. Compared with the postoperative group, the perioperative group had thinner fovea at 1 week, 1 month, and 3 months (P < 0.05), but the differences disappeared after 3 months. Conclusion: In patients with PCV and vitreous hemorrhage, compared with postoperative anti-VEGF, perioperative anti-VEGF could reduce the difficulty of surgery and reduce the occurrence of postoperative complications, but there were no differences in long-term vision and macular thickness after surgery.

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Understanding the dynamics of phosphate-solubilizing and N2-fixing bacteria on soil nutrient and related enzyme activity under different organic fertilizer proportions (OFP) could provide references for screening appropriate inoculant type, OFP, and fertilization period. Here, we set four OFP levels (mass ratio: 0%, 4%, 8%, 12%) and inoculated two phosphate-solubilizing bacteria (Bacillus megaterium, Pseudomonas fluorescens) and two N2-fixing bacteria (Azotobacter chroococcum, Azospirillum brasilence) in the subtropical yellow-brown barren soil. After a 60-day soil incubation under controlled conditions (28 ℃, darkness), we examined the impacts of single/mixed applications of beneficial bacteria on soil available nutrients and related enzyme activities at different OFP levels and different sampling times (3rd, 8th, 16th, 30th, 45th, 60th day). The results showed that soil available nutrient contents increased with the elevated OFP levels, and exhibited as 12%>8%>4%>0%. With the extension of culture time, soil nutrient contents in all treatments first increased and then decreased. Compared with the single application of organic fertilizer, combined application of organic fertilizer and bacterial inoculants resulted in higher and longer improvement of soil nutrient contents and enzyme activities. The effects of inoculants on soil nutrient properties varied across four OFP levels. When the OFP was low (0-4%), inoculation significantly increased soil available nutrient contents, with no the differences between inoculants at the initial stage. However, with the extension of the culture time and the elevation of OFP, phosphate-solubilizing bacteria (especially for B. megaterium) significantly increased available phosphorus content while N2-fixing bacteria (especially for A. brasilence) significantly increased available nitrogen content. The mixed inoculant with four strains showed phosphate-solubilizing effect on soil and performed better than the single application of phosphate-solubilizing bacteria, but without prominent effect on nitrogen fixation. Soil nutrient contents were positively correlated with enzyme activity, which was affected by both cultural time and carbon-nitrogen ratio. Bacterial inoculations could significantly increase nutrient contents in the short term, but the specific functions of beneficial bacteria on soil were highly dependent on organic carbon input and carbon-nitrogen ratio. Coupled application of inoculants and organic fertilizer at an appropriate OFP level (8%-12%) could increase and extend the soil-remediating effects, while the inoculation should be conducted with an interval of 45-60 days to ensure the survival rate and the consecutive effect on soil.