Pseudostellaria Heterophylla Research Articles

Determination of seven oligosaccharides and sucrose in Pseudostellaria heterophylla by pressurized liquid extraction and ultra-high performance liquid chromatography with charged aerosol detector and tandem mass spectrometry

A simple, rapid, and sensitive ultra-high performance liquid chromatography with charged aerosol detector (UPLC-CAD) method was developed for firstly simultaneous determination of seven oligosaccharides, including two pairs of linear oligosaccharides isomers (DP3-1, DP3-2 and DP4-1, DP4-2) and 3 high branched oligosaccharides (DP 5, 6 and 7), as well as sucrose in Pseudostellaria heterophylla. The separation was performed on a Waters BEH Amide column (2.1 × 150 mm i.d., 1.7 μm) with gradient elution within 20 min. All calibration curves for the investigated analytes showed good linearity (R2 > 0.9992). Their limits of detection (LOD) and quantification (LOQ) were in the ranges of 0.24–1.35 μg/mL and 0.78–4.96 μg/mL, respectively. Repeatability of all investigated components detected in samples was less than 2.9%. All the recoveries of each analyte ranged from 99.2% to 104.3% were acceptable. An ultra-high performance liquid chromatography–tandem quadrupole time-of-flight mass spectrometer (UPLC-QTOF MS) method was also applied to authenticate the oligosaccharides in different samples by comparing to the isolated standards. Oligosaccharides, especially linear oligosaccharides isomers and high branched oligosaccharides with different degree of polymerization (DP), were firstly identified and determined in P. heterophylla. Their contents in 33 samples of P. heterophylla from different locations was successfully compared, which is helpful to evaluate its quality.

The complete chloroplast genome of Prince Ginseng, Pseudostellaria heterophylla (Miq.) Pax (Caryophyllaceae)

We presented complete chloroplast genome of Prince Ginseng, Pseudostellaria heterophylla which is 149,795 bp long and has four subregions: 81,460 bp of large single copy (LSC) and 16,983 bp of small single copy (SSC) regions are separated by 25,676 bp of inverted repeat (IR) regions including 126 genes (81 CDS, 8 rRNAs, and 37 tRNAs). The overall GC content of the chloroplast genome is 36.5% and those in the LSC, SSC, and IR regions are 34.3%, 29.4%, and 42.3%, respectively. Phylogenetic trees of 25 Caryophyllaceae species present phylogenetic position of P. heterophylla among available Pseudostellaria species.

ITRAQ-Based Quantitative Proteomic Analysis of Pseudostellaria heterophylla from Geo-Authentic Habitat and Cultivated Bases

Background: Pseudostellaria heterophylla is an important tonic traditional Chinese medicine. However, the molecular changes in the herb from geo-authentic habitat and cultivated bases remain to be explored. Objective: The purpose of this research was to study differences in P. heterophylla from geo-authentic habitat and cultivated bases. Methods: High-throughput technologies of transcriptomic and proteomic were used to identify proteins. Isobaric Tags for Relative and Absolute Quantification (iTRAQ) MS/MS has been utilized to evaluate changes in P. heterophylla from geo-authentic habitat and cultivated bases. Results: In this study, a total of 3775 proteins were detected, and 140 differentially expressed proteins were found in P. heterophylla from geo-authentic habitat and cultivated bases. 44 significantly differential expressed proteins were identified based on functional analysis classified into nine categories. Five differentially expressed proteins were confirmed at the gene expression level by Quantitative realtime PCR. Catabolic metabolism, carbohydrate metabolism, and response to stress of oxidoreductases and transferases in P. heterophylla from geo-authentic habitat were stronger than in those from cultivated bases, but protein folding and response to stress of heat shock proteins, isomerases, rubisco large subunit-binding proteins, chaperone proteins, and luminal-binding proteins in herbs from cultivated bases were more active. ADG1 and TKTA could be the critical proteins to regulate sucrose; MFP2 and CYS may be the crucial proteins that control the metabolism of fatty acids and amino acids. Conclusion: These results will provide the basic information for exploring the differences in secondary metabolites in P. heterophylla from geo-authentic habitat and cultivated bases and the protein mechanism of its quality formation.

Immunomodulatory effects of Pseudostellaria heterophylla peptide on spleen lymphocytes via a Ca2+/CaN/NFATc1/IFN-γ pathway.

Pseudostellaria heterophylla has been becoming a popular research topic because of its functionally active components. The immunomodulatory activity of P. heterophylla peptide (PPH) derived from protein hydrolysate and the molecular mechanism underlying its immunomodulatory effect were investigated in this study. Immunomodulatory PPH achieved the highest stimulation index of 1.53 at a concentration of 100 μg mL-1 for 48 h in spleen lymphocytes and promoted the secretions of tumor necrosis factor-α, interferon-γ, and interleukin-10. Moreover, PPH could elevate the intracellular Ca2+ concentration, calcineurin activity and nuclear factor of activated T cell (NFAT) c1 mRNA expression. Meanwhile these effects could be diminished by the treatment of verapamil and cyclosporin A, suggesting that PPH may activate spleen lymphocytes via the Ca2+/CaN/NFATc1/IFN-γ signaling pathway. These findings demonstrate that the P. heterophylla peptide has the potential to be utilized as a nutraceutical supplement to strengthen the immune system in the human body.

Plasticity in the reproductive strategy of a clonal cleistogamous species, Pseudostellaria heterophylla

The presence of cleistogamous and chasmogamous flowers on the same plant individual is considered to represent a “mixed” reproductive strategy. If a cleistogamous species also exhibits clonal propagation, then competition for limited resources is assumed to exist among the three reproductive modes. To date, however, the relationships and interactions among cleistogamous, chasmogamous, and clonal modes of reproduction have received little attention. In this study, we performed manipulative experiments to investigate the interactions among these different types of reproduction in the perennial herbaceous plant species Pseudostellaria heterophylla. The results showed that 66.4%–87.6% of individuals produce chasmogamous flowers and that the fruiting rates of these flowers in each surveyed population were between 23.5% and 77.4%. Furthermore, we found that 8.3% of the individuals of this species show inbreeding depression. We also detected significant negative correlations between the production of chasmogamous and cleistogamous flowers and between cleistogamous flower production and root tuber mass. However, chasmogamous flower production in an individual plant was found to have little influence on its subsequent clonal propagation. We propose that the plasticity of reproductive strategies observed in P. heterophylla is due to changes in the resource pool and resource allocation.

Physico-Chemical and Antifungal Properties of a Trypsin Inhibitor from the Roots of Pseudostellaria heterophylla.

Plant peptidase inhibitors play essential roles in the defense systems of plants. A trypsin inhibitor (PHTI) with a molecular mass of 20.5 kDa was isolated from the fresh roots of the medicinal herb, Pseudostellaria heterophylla. The purification process involved ammonium sulfate precipitation, gel filtration chromatography on Sephadex G50, and ion-exchange chromatography on DEAE 650M. The PHTI contained 3.7% α-helix, 42.1% β-sheets, 21.2% β-turns, and 33% disordered structures, which showed similarity with several Kunitz-type trypsin inhibitors. Inhibition kinetic studies indicated that PHTI was a competitive inhibitor, with a Ki value of 3.01 × 10−9 M, indicating a high affinity to trypsin. The PHTI exhibited considerable stability over a broad range of pH (2–10) and temperatures (20–70 °C); however, metal ions, including Fe3+, Ba2+, Mn2+, and Al3+, could inactivate PHTI to different degrees. Results of fluorescence spectroscopy and circular dichroism showed that Fe3+ could bind to TI with an association constant of 2.75 × 105 M−1 to form a 1:1 complex, inducing conformation changes and inactivation of PHTI. In addition, PHTI could inhibit the growth of the phytopathogens, Colletotrichum gloeosporioides and Fusarium oxysporum, through disruption of the cell membrane integrity. The present study extended research on Pseudostellaria heterophylla proteins and makes PHTI an exploitable candidate as an antifungal protein for further investigation.

Qualitation and quantification of water soluble non-starch polysaccharides from Pseudostellaria heterophylla in China using saccharide mapping and multiple chromatographic methods

Qualitative and quantitative analysis of non-starch polysaccharide (NSP) from Pseudostellaria heterophylla in China (Guizhou, Anhui and Fujian) were performed using high performance size exclusion chromatography coupled with multi-angle laser light scattering and refractive index detector (HPSEC-MALLS-RID), saccharide mapping based on polysaccharide analysis by using carbohydrate gel electrophoresis (PACE) and high performance thin layer chromatography (HPTLC). Results showed that PACE and HPTLC fingerprints of enzymatic hydrolysates of NSP were similar. Moreover, the results indicated that 1,4-β-d-Galp, 1,5-α-Araf, 1,4-α-d-GalAp and 1,4-β-d-Glcp linkages existed in NSP. Based on the results of HPSEC-MALLS/RI, the Mw, Rg, and contents of NSP in P. heterophylla were ranging from 4.37 × 104 to 1.70 × 105 Da, 46.4 to 65.9 nm, and 0.43% to 1.41%, respectively. There were no significant difference (p>0.05) observed among the three main producing areas, which suggested that polysaccharides could be used as marker for quality control of P. heterophylla.

Cloning and analyzing a squalene synthase gene in Pseudostellaria heterophylla

Triterpene saponins are primary bioactive compounds in the tuberous roots of Pseudostellaria heterophylla, which is one of well-known traditional Chinese medicines. Squalene synthase (SQS) is the key speed-limiting enzyme of saponin biosynthesis pathway. Here, we cloned and analyzed a SQS gene in P. heterophylla. The full length cDNA of SQS was cloned and designated PhSQS (GenBank accession no. KY436585). The open reading frame (ORF) of PhSQS contains 1245 bp and encodes a putative protein of 414 amino acids. In-silico analysis of amino acid sequence of PhSQS showed a conserved isoprenoid family domain and catalytic sites and the three-dimensional (3D) structure of PhSQS was not deviated from that of the SQSs from other species. The phylogenetic relationship of PhSQS was more closely related to that from Beta vulgaris. The semi-quantitative RT-PCR revealed the expression level of PhSQS was significantly higher in root tissue than that of other tissues. Our results suggest that PhSQS is a candidate of functional gene of squalene synthase and may be involved in triterpene saponin biosynthesis.

Rapid authentication of Pseudostellaria heterophylla (Taizishen) from different regions by Raman spectroscopy coupled with chemometric methods

The quality of Pseudostellaria heterophylla depends on the growing area of the plants greatly. Compared with near infrared spectroscopy employed to discriminate the geographic regions before, the Raman signal is more obvious. So far, discrimination of P. heterophylla from different regions by Raman spectroscopy has not yet been realized. Hence, Raman spectroscopy coupled with chemometric methods to rapidly and effectively discriminate P. heterophylla from different regions was studied. Original spectra of P. heterophylla in wavenumber range of 4000–100 cm−1 were acquired. Then, a steady and exact model, partial least squares discriminant analysis (PLS-DA), was constructed. And competitive adaptive reweighted sampling (CARS) was further used to extract effective wavelength spectral characteristic variables. Results show that CARS-PLS-DA model is an appropriate model to discriminate the P. heterophylla, with determination coefficient of calibration (R2C), root mean square error of cross validation (RMSECV), determination coefficient of prediction (R2P), and root mean squared error of prediction (RMSEP) are 0.9468, 0.1979, 0.9665, and 0.1120, respectively. These results demonstrated that the built method is useful and effective to discriminate P. heterophylla from different regions, which provide a new idea in the application of rapid field test.

Heterophyllin B Ameliorates Lipopolysaccharide-Induced Inflammation and Oxidative Stress in RAW 264.7 Macrophages by Suppressing the PI3K/Akt Pathways.

Heterophyllin B (HB), an active cyclic peptide, is a compound existing in the ethyl acetate extract of Pseudostellaria heterophylla (Miq.) Pax and exhibited the activity of inhibiting the production of NO and cytokines, such as IL-1β and IL-6, in LPS-stimulated RAW 264.7 macrophages. In addition, HB suppressed the production of ROS and the apoptosis induced by LPS in RAW 264.7 macrophages. The underlying mechanism was investigated in the LPS-induced RAW 264.7 cells. The results showed that HB decreased the level of IL-1β and IL-6 expression by qRT-PCR analysis. HB up-regulated the relative ratio of p-AKT/AKT and p-PI3K/PI3K as indicated by western blot analysis. In summary, HB inhibited the LPS-induced inflammation and apoptosis through the PI3K/Akt signaling pathways and represented a potential therapeutic target for treatment of inflammatory diseases.

Two Nitrogen-containing Compounds from Pseudostellaria heterophylla

Two new nitrogen-containing compounds, pseudosteheterins A (1) and B (2), were isolated from the glycosides-enriched cardioprotective fraction of the 70% EtOH extract of the roots of Pseudostellaria heterophylla. Their structures were elucidated by spectroscopic methods, and the absolute configuration of 1 was established by quantum chemical TDDFT calculation of the ECD spectrum of its model compound. Pseudosteheterin B showed significant protective effects in H9c2 myocardial cells against CoCl2-induced injury.

Novel Polysaccharide H-1-2 from Pseudostellaria Heterophylla Alleviates Type 2 Diabetes Mellitus

Background/Aims: To investigate the potential therapeutic effect of novel polysaccharide H-1-2 from pseudostellaria heterophylla against type 2 Diabetes Mellitus (T2DM) and elucidate the underling molecular mechanisms. Methods: Relative expression of HIF1α and Sirt1 in T2DM patients was determined via real-time PCR. The direct binding of HIF1α on Sirt1 promoter was validated by ChIP assay. The inhibitory regulation of Sirt1 by HIF1α was analyzed using luciferase reporter assay. The endogenous protein of HIF1α and Sirt1 in response to H-1-2 treatment was quantified by western blotting. The blood glucose, secreted insulin and serous lipid profiles were measured with ELISA kits. Results: We consolidated that HIF1α and Sirt1 was dysregulated in T2DM patients and subjected to H-1-2 modulation. H-1-2 significantly inhibited hypoxia and up-regulated Sirt1 expression in EndoC-βH1 cells. Accordingly, H-1-2 enhanced glucose-stimulation insulin secretion and improved blood glucose and lipid profiles in T2DM cells, and elevated the glucose and insulin tolerance simultaneously. Furthermore, we demonstrated that H-1-2 alleviated T2DM via inhibition of hypoxia and up-regulation of Sirt1 in isolated pancreatic β-cells from T2DM rats. Conclusion: Our data unambiguously demonstrated H-1-2 administration alleviated T2DM by enhancing Sirt1 expression through inhibition of hypoxia.

Transcriptome analysis of Pseudostellaria heterophylla in response to the infection of pathogenic Fusarium oxysporum

BackgroundPseudostellaria heterophylla (P. heterophylla), a herbaceous perennial, belongs to Caryophyllaceae family and is one of the Chinese herbal medicine with high pharmacodynamic value. It can be used to treat the spleen deficiency, anorexia, weakness after illness and spontaneous perspiration symptoms. Our previous study found that consecutive monoculture of Pseudostellaria heterophylla could lead to the deterioration of the rhizosphere microenvironment. The specialized forms of pathogenic fungus Fusarium oxysporum f.Sp. heterophylla (F. oxysporum) in rhizosphere soils of P. heterophylla plays an important role in the consecutive monoculture of P. heterophylla.ResultsIn this study, F. oxysporum was used to infect the tissue culture plantlets of P. heterophylla to study the responding process at three different infection stages by using RNA-sequencing. We obtained 127,725 transcripts and 47,655 distinct unigenes by de novo assembly and obtained annotated information in details for 25,882 unigenes. The Kyoto Encyclopedia of Genes and Genomes pathway analysis and the real-time quantitative PCR results suggest that the calcium signal system and WRKY transcription factor in the plant-pathogen interaction pathway may play an important role in the response process, and all of the WRKY transcription factor genes were divided into three different types. Moreover, we also found that the stimulation of F. oxysporum may result in the accumulation of some phenolics in the plantlets and the programmed cell death of the plantlets.ConclusionsThis study has partly revealed the possible molecular mechanism of the population explosion of F. oxysporum in rhizosphere soils and signal response process, which can be helpful in unraveling the role of F. oxysporum in consecutive monoculture problems of P. heterophylla.

Structure of a pectic polysaccharide from Pseudostellaria heterophylla and stimulating insulin secretion of INS-1 cell and distributing in rats by oral

A water-soluble, pectic polysaccharide named as 0.5MSC-F isolated from Pseudostellaria Heterophylla with a molecular weight of 4.8×104Da that was composed of rhamnose, galactose, arabinose, and galacturonic acid which the major monosaccharide contents range up to 63.20%. Where the main chain was consists of 1,4-linked galacturonic acid and a small amount of 1,2-linked rhamnose was embedded into backbone to connect alternative galacturonic acid in the form of Rhamnogalacturonan I (RG-I) structure. 1, 5-linked arabinose through C-4 of 1, 2-linked rhamnose, another 1, 3 or 1,6-linked galactose through C-4 of 1, 2-linked rhamnose, was interconnected to branch chain. 0.5MSC-F could obviously stimulated insulin secretion of islet cells cultured in high glucose are of potential practical value in the hypoglycemic action. Radioisotopes 99mTc-labeled-0.5MSC-F was analyzed by SPECT/CT image after oral in rats. At 2h post ingestion, above 40% of the radioactivity was observed in the intestine, but no found in the heart, liver, and kidney. Conjecturing absorption of 99mTc-labeled 0.5MSC-F might via intestinal mucosa absorption into the systemic circulation.

Stimulatory effect of lanthanum nitrate on the root tuber yield of Pseudostellaria heterophylla via improved photosynthetic characteristics

The stimulatory effects and associated mechanism of lanthanum nitrate on the root tuber yield of the traditional Chinese medicinal plant Pseudostellaria heterophylla, by means of improved photosynthetic characteristics, were investigated. Field experiments were conducted and the foliage was sprayed with varying concentrations of lanthanum nitrate. The results indicated that growth indexes, chlorophyll and carotenoid contents initially increased, and then decreased in a dose-dependent manner with increasing concentrations of lanthanum nitrate. The fresh and dry weights of root tuber yield per unit area significantly increased by 58.34% and 56.87% with a lanthanum nitrate concentration of 100 mg/L, respectively, compared with the control. Appropriate concentrations of lanthanum nitrate were found to alleviate photosynthetic depression at midday, as well as improve the mean values of photosynthetic rate (Pn), stomatal conductance (Gs) and transpiration rate (Tr) associated with the diurnal variation in photosynthetic parameters, while reducing intercellular CO2 concentration (Ci). During the experiment, the chlorophyll fluorescence parameters Fv/Fm, Fv‘/Fm’, ΦPSII and qP increased initially and then decreased along with the increasing concentration of lanthanum nitrate, while NPQ followed the opposite trend. Correlation analysis revealed that Pn had a significantly positive relationship with seedling height, ground diameter, chlorophyll and carotenoid contents, Gs and ΦPSII, while a negative relationship was observed with Ci and NPQ. Grey relational analysis (GRA) indicated that photosynthetic pigments, ΦPSII and qP were the primary factors impacting photosynthesis. These results suggested that the application of an appropriate concentration of lanthanum nitrate (100 mg/L) was effective in improving growth, root tuber yield and photosynthetic characteristics of Psendoste llaria (P) heterophylla. The regulatory effect of lanthanum nitrate on photosynthesis was related to the promotion of light energy absorption and conversion, improvement in photochemical efficiency and the alleviation of photoinhibition. The improvement of the non-stomatal limitation factor was the primary basis for the mitigation of midday photosynthetic depression.

Cloning and expression analysis on full length CDS of zeaxanthin epoxidase gene in Pseudostellaria heterophylla

Zeaxanthin epoxidase plays an important role in indirect pathway of plant abscisic acid biosynthesis. According to the data of Pseudostellaria heterophylla transcriptome, zeaxanthin epoxidase gene was isolated and named as PhZEP. The results of bioinformatics analysis showed that the coding sequence of PhZEP was 1 263 bp long and encoded 420 amino acids. The putative protein molecular weight was 47.34 kDa and its theoretical isoelectric point was 6.64. The characteristic structure domains were predicted, including binding site of lipoprotein and flavoprotein monooxyenase. A signal peptide was discovered at the N-terminal of amino acids. The Real-time PCR revealed that PhZEP had a higher expression level in leaves than other tissues of P.heterophylla. Highly expressed PhZEP was also observed at 10 d and 40 d tuberous root after flowering. PhZEP presented a different expression after treatment with ABA, fluridone and ABA +fluridone compared to the control. The expression of PhZEP in tuberous root after ABA treatment was close to that in control group, while PhZEP showed significant up-regulation in the fluridone treatment group. In this study, the PhZEP gene from P. heterophylla was cloned and this result has important significance for its functional identification. This research provides a basis for the further analysis on functional mechanism of ABA during development of P. heterophylla.

Immunomodulatory effects of Pseudostellaria heterophylla (Miquel) Pax on regulation of Th1/Th2 levels in mice with atopic dermatitis.

Pseudostellaria heterophylla (PH) has various pharmacological effects that include immunologic enhancement and anti-oxidation. However, it remains unclear whether PH exerts beneficial effects in dermatological diseases. The present study examined the effects of PH on a 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD) mouse model and elucidated its underlying mechanism of action. PH extract (1 and 100 mg/ml) was applied topically to DNCB-treated dorsal skin of mice every day for 11 days. The immunomodulatory effects of PH were evaluated by measuring skin thickness, mast cell infiltration, serum levels of immunoglobulin E (IgE), and mRNA expression levels of T helper (h)1/Th2 and pro-inflammatory cytokines in dorsal skin. In addition, cluster of differentiation (CD)4+ T cells were detected in dorsal skin by immunohistochemistry. Topical application of PH significantly reduced the thickness of dermis, epidermis and serum IgE production compared with the DNCB group. PH treatment inhibited infiltration of inflammatory cells, including mast cells and CD4+ T cells, and suppressed the mRNA expression levels of cytokines (interferon-γ, interleukin-4, −6, −8 and −1β, and tumor necrosis factor-α) associated with the immune response. Furthermore, PH treatment significantly downregulated the protein expression levels of nuclear factor-κB, phosphorylated inhibitor of κBα and mitogen-activated protein kinases. The results suggested that PH may be a potential therapeutic strategy for the treatment of AD via the modulation of Th1 and Th2 levels.

Effects of different amendments on contents of phenolic acids and specific microbes in rhizosphere of Pseudostellaria heterophylla.

Pseudostellaria heterophylla is a perennial herbaceous plant in the family Caryophyllaceae. The tuberous roots of P. heterophylla are highly valued in traditional Chinese medicine and have a high market demand. However, extended monoculture of P. heterophylla results in a significant decline in the biomass and quality, and escalates disease and pest problems. Therefore, it is important to understand the underlying mechanism and biocontrol methods for consecutive monoculture problems. With "Zheshen 2" as an experimental material, the changes in the contents of main nutrients in soil, phenolic acids and specific microbes under monoculture and different amendments were analyzed by using high performance liquid chromatography (HPLC) and qPCR. The results showed that consecutive monoculture of P. heterophylla led to a decrease in yield by 43.5% while the microbial fertilizer treatment and the paddy-upland rotation could relieve the consecutive monoculture problems. Available nitrogen, available phosphorus, available potassium and total potassium were significantly higher in the consecutively monocultured soils than in the newly planted soils. But consecutive monoculture resulted in soil acidification. HPLC analysis showed that conse-cutive monoculture of this plant did not lead to a consistent accumulation of soil phenolic acids. At middle stage of root expansion and at harvest stage, most of phenolic acids were even higher in the newly planted soils than in the consecutively monocultured soils. Furthermore, qPCR analysis showed that the amounts of three specific pathogens identified previously (i.e. Fusarium oxysporum, Talaromyces helicus, Kosakonia sacchari) were significantly higher in the consecutively monocultured soils than in the newly planted soils. However, the microbial fertilizer treatment and the paddy-upland rotation resulted in a significant decline in the population of these specific pathogens and improved the soil environment. In conclusion, the consecutive monoculture problems of P. heterophylla may be due to the rapid proliferation of host-specific pathogens, rather than the deficiency of soil nutrients and the autotoxicity of allelochemicals in root exudates. The results in this study could provide the theoretical basis to explore the underlying mechanism of replanting disease of P. heterophylla and its biocontrol strategies.

Insights into the Regulation of Rhizosphere Bacterial Communities by Application of Bio-organic Fertilizer in Pseudostellaria heterophylla Monoculture Regime.

The biomass and quality of Pseudostellariae heterophylla suffers a significant decline under monoculture. Since rhizosphere miobiome plays crucial roles in soil health, deep pyrosequencing combined with qPCR was applied to characterize the composition and structure of soil bacterial community under monoculture and different amendments. The results showed compared with the 1st-year planted (FP), 2nd-year monoculture of P. heterophylla (SP) led to a significant decline in yield and resulted in a significant increase in Fusarium oxysporum but a decline in Burkholderia spp. Bio-organic fertilizer (MT) formulated by combining antagonistic bacteria with organic matter could significantly promote the yield by regulating rhizosphere bacterial community. However, organic fertilizer (MO) without antagonistic bacteria could not suppress Fusarium wilt. Multivariate statistics analysis showed a distinct separation between the healthy samples (FP and MT) and the unhealthy samples (SP and MO), suggesting a strong relationship between soil microbial community and plant performance. Furthermore, we found the application of bio-organic fertilizer MT could significantly increase the bacterial community diversity and restructure microbial community with relatively fewer pathogenic F. oxysporum and more beneficial Burkholderia spp. In conclusion, the application of novel bio-organic fertilizer could effectively suppress Fusarium wilt by enriching the antagonistic bacteria and enhancing the bacterial diversity.

De Novo Sequencing and Assembly Analysis of the Pseudostellaria heterophylla Transcriptome.

Pseudostellaria heterophylla (Miq.) Pax is a mild tonic herb widely cultivated in the Southern part of China. The tuberous roots of P. heterophylla accumulate high levels of secondary metabolism products of medicinal value such as saponins, flavonoids, and isoquinoline alkaloids. Despite numerous studies on the pharmacological importance and purification of these compounds in P. heterophylla, their biosynthesis is not well understood. In the present study, we used Illumina HiSeq 4000 sequencing platform to sequence the RNA from flowers, leaves, stem, root cortex and xylem tissues of P. heterophylla. We obtained 616,413,316 clean reads that we assembled into 127, 334 unique sequences with an N50 length of 951 bp. Among these unigenes, 53,184 unigenes (41.76%) were annotated in a public database and 39, 795 unigenes were assigned to 356 KEGG pathways; 23,714 unigenes (8.82%) had high homology with the genes from Beta vulgaris. We discovered 32, 095 DEGs in different tissues and performed GO and KEGG enrichment analysis. The most enriched KEGG pathway of secondary metabolism showed up-regulated expression in tuberous roots as compared with the ground parts of P. heterophylla. Moreover, we identified 72 candidate genes involved in triterpenoids saponins biosynthesis in P. heterophylla. The expression profiles of 11 candidate unigenes were analyzed by quantitative real-time PCR (RT-qPCR). Our study established a global transcriptome database of P. heterophylla for gene identification and regulation. We also identified the candidate unigenes involved in triterpenoids saponins biosynthesis. Our results provide an invaluable resource for the secondary metabolites and physiological processes in different tissues of P. heterophylla.