Inhibitory activities of five fungicides on Alternaria suffruticosae and their field control efficacy against tree peony black spot.

Tree peony black spot (TPBS), mainly caused by Alternaria suffruticosae, is a common leaf disease on the ornamental peony, which posed a great threat on the flower buds in the current year and the flowering quality in the next year. However, there was only one fungicide registered for the control of the disease, difenoconazole. In order to avoid the severe problem of pathogen resistance caused by long-term use of difenoconazole, it is necessary to screen more chemical fungicides for the prevention and control of TPBS. In the paper, the biological activities of flutolanil, phenamacril, pyraclostrobin, and boscalid on mycelial growth, conidial germination, germ tube elongation and sporulation quantity of A. suffruticosae were determined, and field control efficacy were conducted to evaluate the preventive and therapeutic activities. Difenoconazole, was used as a control simultaneously. The results showed that pyraclostrobin had the strongest inhibitory effects on the conidial germination, mycelium growth, germ tube elongation and sporulation quantity, with the average EC50 of 0.0517, 0.5343, 0.0008 and 0.8068 µg/mL respectively. The inhibitory activity of flutolanil on the four developmental stages of A. suffruticosae was weaker than the other three fungicides. Compared with flutolanil, boscalid, the other succinate dehydrogenase inhibitors, had more srtong inhibitory effects on the mycelial growth and sporulation quantity, with the average EC50 of 3.8603 and 1.4760 µg/mL respectively. Phenamacril had a moderate inhibitory level, which had more inhibitory activity on conidial germination and germ tube elongation, with the average EC50 of 31.5349 and 5.2597 µg/mL. All of the four fungicides had no significant effects on the shape of spores and germ tubes. The control fungicide difenoconazole had the strongest inhibitory activity on mycelial growth, and the average EC50 was only 0.3297 µg/ml. However, its inhibitory activity on the other three growth stages was not high. In the field trials, pyraclostrobin had high control efficacy on TPBS even at low concentrations, reaching a minimum of 62.6293%, which was higher than that of difenoconazole. The other three fungicides had higher control efficacy at high concentrations, but decreased significantly at low concentrations. Considering the dosage and control efficacy, pyraclostrobin was the first choice for the control of TPBS. Pyraclostrobin is the preferred alternative fungicide of difenoconazole for the prevention and control of TPBS in production.

Molecular mechanism of somatic embryogenesis in paeonia ostii 'Fengdan' based on transcriptome analysis combined histomorphological observation and metabolite determination.

BACKGROUND: Tree peony (Paeonia sect. Moutan DC.) is a famous flower native to China with high ornamental, medicinal, and oil value. However, the low regeneration rate of callus is one of the main constraints for the establishment of a genetic transformation system in tree peony. By histomorphological observation, transcriptomic analysis and metabolite determination, we investigated the molecular mechanism of somatic embryogenesis after the establishment of a culture system and the induction of somatic embryo(SE) formation. RESULTS: We found that SE formation was successfully induced when cotyledons were used as explants. A total of 3185 differentially expressed genes were screened by comparative transcriptomic analysis of embryogenic callus (EC), SE, and non-embryogenic callus (NEC). Compared to NEC, the auxin synthesis-related genes GH3.6 and PCO2 were up-regulated, whereas cytokinin dehydrogenase (CKX6) and CYP450 family genes were down-regulated in somatic embryogenesis. In SE, the auxin content was significantly higher than the cytokinin content. The methyltransferase-related gene S-adenosylmethionine synthase (SAMS) and the flavonoid biosynthesis-related gene (ANS and F3'5'H) were down-regulated in somatic embryogenesis. The determination of flavonoids showed that rhoifolin and hyperoside had the highest content in SE. The results of transcriptome analysis were consistent with the relative expression of 8 candidate genes by quantitative polymerase chain reaction analysis. CONCLUSION: The results revealed that auxin and cytokinin may play a key role in 'Fengdan' somatic embryogenesis. The genes related to somatic embryogenesis were revealed, which has partly elucidated the molecular mechanism of somatic embryogenesis in 'Fengdan'.

Characterization of Agronomic and Seed Oil Features for Different Cultivars of Tree Peony.

Tree peony is a unique oil plant resource in China, and tree peony seed oil is one of the healthy edible oils with a very promising future. However, the main oil tree peony cultivars promoted in China are Paeonia ostii 'Fengdan' and Paeonia rockii. In order to explore new oil tree peony cultivars, 68 tree peony cultivars were investigated and cultivars with oil potential were selected by cluster analysis and grey relational analysis (GRA) in this study. The results demonstrated that the 68 cultivars varied significantly in terms of agronomic characteristics (p < 0.05), with the coefficient of variation in seed yield per plant reaching a high of 75.36%. The oil content of 46 cultivars was higher than 'Fengdan' (20.87 ± 0.26%) and 'Zibanbai' (21.24 ± 1.01%), while the alpha-linolenic acids and total unsaturated fatty acid contents of 26 cultivars were higher than 'Fengdan' (39.79 ± 1.13% and 88.99 ± 0.47%) and 'Zibanbai' (40.51 ± 0.09% and 93.59 ± 0.09%). Finally, three cultivars with better integrated traits were selected by cluster analysis and grey relational analysis (GRA), comprising of 'Changshoule', 'Xianchizhenghui', and 'Yupantuojin'. The contents of alpha-linolenic acids and total unsaturated fatty acids in 'Changshoule' (47.98 ± 0.17% and 93.60 ± 0.08%), 'Xianchizhenghui' (49.44 ± 0.63% and 93.80 ± 0.06%), and 'Yupantuojin' (40.46 ± 0.26% and 93.58 ± 0.06%) were higher than that of 'Fengdan' (39.79 ± 1.13% and 88.99 ± 0.47%). In general, these cultivars can be used as hybrid parental materials for breeding new excellent oil tree peony cultivars.

Ascription of nosZ gene, pH and copper for mitigating N2O emissions in acidic soils.

Soil nitrous oxide (N2O) emissions are alarming for global warming and climate change. N2O reduction is carried out only by nosZ gene encoded N2O-reductase, which is highly sensitive to acidic pH and copper (Cu) contents. Therefore, a microcosm study was conducted to examine the attribution of soil pH management, Cu supply and nosZ gene abundance for N2O emission mitigation. Cu was applied at the dose of 0, 10, 25 and 50 mg kg-1 to three acidic soils (Soil 1, 2 and 3) without and with dolomite (0 and 5 g kg-1). Cu application and soil pH increment substantially enlarged the abundance of nosZ gene, and consequently mitigated soil N2O emissions; highest reduction with 25 Cu mg kg-1. Decline in NH4+ and subsequently accumulation of NO3-, and large contents of MBC and DOC in dolomite treated soils led to a substantial N2O reduction. The cumulative N2O emissions were lowest in the treatment of 25 Cu mg kg-1 with dolomite application for each soil. Results suggest that soil pH increment, an adequate Cu supply, and nosZ gene abundance can potentially lower soil N2O emissions in acidic soils.

Screening of Reference miRNA of Different Early- and Late-Flowering Tree Peony Varieties.

miRNA plays an important role in plant growth and development and in response to various stresses. Quantitative real-time PCR (qRT-PCR) technology is often used to detect the expression level of miRNAs and genes by comparing with reference genes. In order to screen out the optimal reference miRNAs in different tree peony varieties, the petals of 42 different early- and late-flowering tree peony varieties were used as experimental materials, and geNorm, NormFinder, Bestkeeper, and RefFinder software were used to evaluate the stability of 16 candidate reference miRNAs. The results showed that the average Ct values of all candidate reference miRNAs were between 15.34 ± 0.29 and 32.64 ± 0.38. The optimal number of reference miRNAs was four, which were PsPC-5p-19095, PsPC-3p-51259, PsmiR159a, and PsPC-3p-6660 in geNorm. The stability of PsPC-3p-6660 was the highest in the analysis results of NormFinder software. Among the analysis results of Bestkeeper software, PsMIR319-p5 has the highest stability. Among the results of comprehensive evaluation and analysis of several software using RefFinder, the candidate reference miRNA with the highest stability was PsPC-3p-6660. When PsPC-3p-6660 was used as the reference miRNA, the expression of PomiR171 and PomiR414 in response to different flowering times of tree peony was relatively stable in 42 tree peony varieties, indicating that PsPC-3p-6660 was stable and reliable. The results of this study provide a reference miRNA for studying the expression changes of miRNA in different tree peony varieties and further exploring the regulatory mechanism of miRNA in different peony varieties.

Identification of Floral Volatile Components and Expression Analysis of Controlling Gene in Paeonia ostii 'Fengdan' under Different Cultivation Conditions.

In order to explore the release rule of floral volatile substances and the diurnal variation of different flower development stages of Paeonia ostii 'Fengdan' in potted and ground-planted conditions, dynamic headspace adsorption combined with gas chromatography-mass spectrometry(GC-MS) was used to analyze the dynamic changes in floral volatile components and contents. Quantitative real-time PCR (qRT-PCR) was used to analyze changes in flower fragrance-regulating genes PsPAL, PsTPSs, and PsbHLH at different flower development stages and a daily change process at the full-blooming stage. The results show that there were differences in aroma components and contents of Paeonia ostii 'Fengdan' at different flower development stages and different time quantum of every day. There were 25 and 28 aroma components identified in 7 flower development stages of tree peonies planted in pots and in the field, respectively, and 23 and 22 aroma components identified at different time quantum of the day, of which the largest and highest content was alkanes. The main characteristic aroma substances were (E)-ß-ocimene, 1,3,5-trimethoxybenzene, 2,4-di-tert-butylphenol, methyl jasmonate, nerol, and cinnamyl alcohol; released amounts of the abovementioned substances varied depending on the development stage and the time of the day. The expression of flower fragrance-controlling genes (PsPAL, PsTPSs, and PsbHLH) in tree peonies varied greatly in different conditions. The results of this study provide a valuable resource to investigate floral fragrance formation in tree peonies.

Exogenous Melatonin Enhances the Yield and Secondary Metabolite Contents of Prunella vulgaris by Modulating Antioxidant System, Root Architecture and Photosynthetic Capacity.

Melatonin (MT) plays a number of key roles in regulating plant growth and secondary metabolite accumulation. Prunella vulgaris is an important traditional Chinese herbal medicinal plant which is used for the treatment of lymph, goiter, and mastitis. However, the effect of MT on the yield and medicinal component content of P. vulgaris remains still unclear. In this research, we have examined the influence of different concentrations of MT (0, 50, 100, 200, 400 µM) on the physiological characteristics, secondary metabolite contents, and yield of P. vulgaris biomass. The results showed that 50-200 µM MT treatment had a positive effect on P. vulgaris. MT treatment at 100 µM greatly increased the activities of superoxide dismutase and peroxidase, the contents of soluble sugar and proline, and obviously decreased the relative electrical conductivity, the contents of malondialdehyde and hydrogen peroxide of leaves. Furthermore, it markedly promoted the growth and development of the root system, increased the content of photosynthetic pigments, improved the performance of photosystems I and II and the coordination of both photosystems, and enhanced the photosynthetic capacity of P. vulgaris. In addition, it significantly increased the dry mass of whole plant and spica and promoted the accumulation of total flavonoids, total phenolics, caffeic acid, ferulic acid, rosmarinic acid, and hyperoside in the spica of P. vulgaris. These findings demonstrated that the application of MT could effectively activate the antioxidant defense system of P. vulgaris, protect the photosynthetic apparatus from photooxidation damage, and improve the photosynthetic capacity and the root absorption capacity, thereby promoting the yield and accumulation of secondary metabolites in P. vulgaris.

The efficient removal of congo red and ciprofloxacin by peony seeds shell activated carbon with ultra-high specific surface area.

Preparation of high-performance activated carbon from agroforestry waste biomass can effectively improve the shortcomings of traditional biomass carbon performance. Using the waste biomass peony seeds shell (PSS) as the precursors in this study, high performance activated carbon was prepared by the KOH two-step activation method and used to remove congo red (CR) and ciprofloxacin (CIP) in water pollution. The obtained PSS-based activated carbons (PSACs) were characterized by SEM, EDS, N2 adsorption-desorption isotherm, FTIR, and XRD methods. The results showed that the activated carbon at 700 °C (PSAC-700) had an ultra-high specific surface area (2980.96 m2/g), excellent micropore volume (1.12 cm3/g), and abundant surface functional groups. The results of adsorption performance revealed that PSAC-700 exhibited excellent adsorption capacity for CR (qmax = 2003.2 mg/g) and CIP (qmax = 782.3 mg/g), which was superior to the carbon-based adsorbents reported reviously in the literature. Langmuir model could well describe the adsorption process of PSACs for CR and CIP, indicating that the pollutant molecules were uniformly adsorbed on the surface monolayer. The regeneration experiment suggested that after three cycles, the adsorption capacities of PSAC-700 for CR and CIP reached 1814 mg/g and 697 mg/g, respectively, with good repeatability. The preparation of PSAC-700 in this study has high adsorption capacity and strong application, which is an ideal material for wastewater purification adsorbent and has broad application prospect.

Genome-wide association study of 23 flowering phenology traits and 4 floral agronomic traits in tree peony (Paeonia section Moutan DC.) reveals five genes known to regulate flowering time.

Tree peony is a unique traditional flower in China, with large, fragrant, and colorful flowers. However, a relatively short and concentrated flowering period limits the applications and production of tree peony. A genome-wide association study (GWAS) was conducted to accelerate molecular breeding for the improvement of flowering phenology traits and ornamental phenotypes in tree peony. A diverse panel of 451 tree peony accessions was phenotyped for 23 flowering phenology traits and 4 floral agronomic traits over 3 years. Genotyping by sequencing (GBS) was used to obtain a large number of genome-wide single-nucleotide polymorphisms (SNPs) (107 050) for the panel genotypes, and 1047 candidate genes were identified by association mapping. Eighty-two related genes were observed during at least 2 years for flowering, and seven SNPs repeatedly identified for multiple flowering phenology traits over multiple years were highly significantly associated with five genes known to regulate flowering time. We validated the temporal expression profiles of these candidate genes and highlighted their possible roles in the regulation of flower bud differentiation and flowering time in tree peony. This study shows that GWAS based on GBS can be used to identify the genetic determinants of complex traits in tree peony. The results expand our understanding of flowering time control in perennial woody plants. Identification of markers closely related to these flowering phenology traits can be used in tree peony breeding programs for important agronomic traits.

Expression Analysis of CoHMGS in Cornus officinalis and Subcellular Localization of the Enzyme It Encodes.

Cornus officinalis is a perennial deciduous tree or shrub. Its mature fruits are extracted and used in Traditional Chinese Medicine, called Shanzhuyu. The characteristic active components of C. officinalis include loganin and morroniside, which belong to iridoid glycosides. 3-Hydroxy-3-methylglutaryl-CoA synthase (HMGS) is a key enzyme in the cytoplasmic mevalonate pathway providing the precursor molecules isopentenyl pyrophosphate and dimethylallyl pyrophosphate for isoprenoid biosynthesis such as sterols, triterpenes, and their derivatives such as iridoid glycosides. Different concentrations of methyl jasmonate (MeJA) and ethephon (ETH) solutions were sprayed on C. officinalis seedlings, and the effect of hormones on CoHMGS gene expression was detected by real-time fluorescence quantitative PCR. The quantitative real-time PCR results showed that 750 mg/L ETH treatment had the most significant induction effect on CoHMGS gene expression. The HPLC analysis of extracts revealed that the treatment could also significantly increase the content of morroniside and loganin in the leaves of C. officinalis. By use of a CoHMGS-green fluorescent protein (GFP) fusion construct for heterologous expression in tobacco, laser scanning confocal microscopy revealed a cytoplasmic localization. This preliminary study of the CoHMGS gene could prepare the ground for more precisely elucidating the synthesis of secondary metabolite in C. officinalis.

Reference genes selection of Paeonia ostii 'Fengdan' under osmotic stresses and hormone treatments by RT-qPCR.

BACKGROUND: Tree peony possess significant ornamental, medicinal and oil values. Osmotic stresses including dehydratiuon and salinity limit the expansion of cultivation area of tree peony. Information on reference genes selection under osmotic stress and hormone stimulation of tree peony still limited. This study aimed to determine the stable reference genes suitable for tree peony under osmotic stresses and hormone treatments, and provide a theoretical basis for the molecular biology research. METHODS AND RESULTS: Twelve candidate reference genes were evaluated in Paeonia ostii 'Fengdan' under osmotic stress and hormone treatments by RT-qPCR. Delta Ct method, geNorm, and NormFinder were used for the comprehensive expression stability ranking comparison. The results revealed that tubulin-α was the preferred internal reference genes for drought and ABA treatment, tubulin-ß was identified as the most suitable reference gene under drought and OPDA induction, 18s-rRNA was regarded as the most stable gene for salinity and JA treatment, eIF-5 A was listed as the most stable gene for JA and MeJA treatments. The experiments also displayed that EF1-α were comparatively unstable under ABA and BR hormone treatments. CONCLUSION: These preferred reference genes could be useful in qPCR studies involving osmotic or hormonal stresses in Paeonia ostii 'Fengdan'. It is anticipated that the results will benefit tree peony functional genomics studies and molecular breeding research in the future.

Inhibitory Effect of Monoterpenoid Glycosides Extracts from Peony Seed Meal on Streptococcus suis LuxS/AI-2 Quorum Sensing System and Biofilm.

Streptococcus suis LuxS/AI-2 quorum sensing system regulates biofilm formation, resulting in increased pathogenicity and drug resistance, and diminished efficacy of antibiotic treatment. The remaining peony seed cake after oil extraction is rich in monoterpenoid glycosides, which can inhibit the formation of bacterial biofilm. In this study, we investigated the effect of seven major monocomponents (suffruticosol A, suffruticosol B, suffruticosol C, paeonifloin, albiflorin, trans-ε-viniferin, gnetin H) of peony seed meal on minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of S. suis. The results showed that the MICs of the seven single components were all greater than 200 µg/mL, with no significant bacteriostatic and bactericidal advantages. Crystal violet staining and scanning electron microscope observation showed that the seven single components had a certain inhibitory effect on the biofilm formation ability of S. suis at sub-MIC concentration. Among them, the ability of paeoniflorin to inhibit biofilm was significantly higher than that of the other six single components. AI-2 signaling molecules were detected by bioreporter strain Vibrio harvey BB170. The detection results of AI-2 signal molecules found that at 1/2 MIC concentration, paeoniflorin significantly inhibited the production of S. suis AI-2 signal, and the inhibitory effect was better than that of the other six single components. In addition, molecular docking analysis revealed that paeoniflorin had a significant binding activity with LuxS protein compared with the other six single components. The present study provides evidence that paeoniflorin plays a key role in the regulation of the inhibition of S. suis LuxS/AI-2 system and biofilm formation in peony seed meal.

Characteristics of PoVIN3, a Key Gene of Vernalization Pathway, Affects Flowering Time.

The tree peony (Paeonia section Moutan DC.) is the candidate flower in China, with abundant germplasm resources and high ornamental value. However, the short and concentrated flowering period severely restricted the improvement of the economic value of tree peonies. Based on the full-length transcriptome database of tree peonies, the PoVIN3 (GenBank ID: OP341879), involved in the flowering regulation of tree peonies were identified and cloned for the first time. The PoVIN3 was also characterized by bioinformatics methods, quantitative real-time PCR (qRT-PCR), and the establishment of a transgenic system. The expression levels of PoVIN3 in seven different petals developmental stages were the highest at the initial flowering stage of the variant cultivar of Paeonia ostii 'Fengdan,' the initial decay stage of the normal flowering Paeonia ostii 'Fengdan,' and the half opening stage of the late flowering Paeonia suffruticosa 'Lianhe.' Tissue-specific expression analysis showed that the relative expression levels of PoVIN3 were the highest in sepals of both normal flowering Paeonia ostii 'Fengdan' and the late flowering Paeonia suffruticosa 'Lianhe,' and the highest expression was in stamens of early flowering mutant Paeonia ostii 'Fengdan.' In addition, the flowering time of pCAMBIA2300-PoVIN3 transgenic plants was significantly earlier than that of the wild-type, indicating that PoVIN3 could promote plant flowering. The results provide a theoretical basis for exploring the role of PoVIN3 in the regulation of flowering in tree peonies.

Phenolic profiles and antioxidant activity in different organs of Sinopodophyllum hexandrum.

Sinopodophyllum hexandrum is a perennial anti-cancer medicinal plant as unique phytochemical composition podophyllotoxin, and it has special effects on the treatments of pneumonic, cervical and testicular cancers. Besides the podophyllotoxin, phenolic substances play a key role in the clinical practice. However, few reports were available in terms of the phenolic compositions and antioxidant activity. In this work, main phenolic compounds were quantified by RP-HPLC in seven organs from S. hexandrum. Simultaneously, the sodium borohydride/chloranil-based (SBC) method and the Folin-Ciocalteau colorimetric method were used to determine total flavonoids and total phenols contents, respectively. The antioxidant activity of the different organs was further assessed by three methods (DPPH method, ABTS method and FRAP method). Phenolic compositions/total flavonoids contents/total phenols contents/antioxidant activity was observed to have significant differences among different organs (P<0.05), but have a consistent changing rule viz. rhizome>root>fruit>flower>leaf>stem>petiole. Furthermore, a correlation analysis was employed and indicated that a positive correlation existed between phenolic compositions contents and antioxidant activity. Obviously, rhizome had high phenolic compositions contents and strong antioxidant activity with the low DPPHIC50 value of 23.52 µg/mL, high ABTS value of 1137.82 µmol equiv. Trolox/g and high FRAP value of 685.76 µmol equiv. Trolox/g. Therefore, rhizome is recommended as a preponderant medicinal part, and root is proposed as an alternative raw material resource for natural antioxidant agents in functional food, medicine and chemicals. This study can provide a new insight into the utilization extension of S. hexandrum resources.

Characterization of PsmiR319 during flower development in early- and late-flowering tree peonies cultivars.

The flowering period is the most important ornamental trait of tree peony, while industrial development of tree peony has been limited by short flowering period. miR319 plays an important regulatory role in plant flowering. In the current study, the expression characteristics and evolution of PsmiR319 in tree peony flowering was explored using 'Feng Dan' and 'Lian He', which are early-flowering and late-flowering varieties of tree peony, respectively. The structure, evolution, and target(s) of PsmiR319 were analyzed by bioinformatics. Evolution analysis showed that pre-PsmiR319 was distributed in 41 plant species, among which the length of the precursor sequence exhibited marked differences (between 52 and 308 bp). Pre-PsmiR319 of tree peony was located close to the corresponding sequences of Linum usitatissimum and Picea abies in the phylogenetic tree, and in addition, could form a typical hairpin structure including a mature body with a length of 20 bp located on the 3p arm and part of the loop sequence. The mature sequence of miR319 was highly conserved among different species. Target genes of PsmiR319 include MYB-related transcription factor in tree peony. Expression of PsmiR319, assayed by qRT-PCR, differed between 'Feng Dan' and 'Lian He' during different flower development periods. PsmiR319 and its target gene showed a negative expression regulation relationship during the periods of CE (color exposure), BS (blooming stage), IF (initial flowering), and HO (half opening) in the early-flowering 'Feng Dan', and the same in FB (Full blooming) periods of late-flowering 'Lian He'. Findings from this study provide a reference for further investigation into the mechanism of miR319 in the development of different varieties of tree peony.

Development and Characterization of Microsatellite Markers Based on the Chloroplast Genome of Tree Peony.

Tree peony (Paeonia suffruticosa Andr.) is a famous ornamental and medicinal flowering species. However, few high-efficiency chloroplast microsatellite markers have been developed for it to be employed in taxonomic identifications and evaluation of germplasm resources to date. In the present study, a total of 139 cpSSR loci were identified across eleven tree peony plastomes. Dinucleotide repeat SSRs (97.12%) were most abundantly repeated for the AT motif (58.27%), followed by the TA motif (30.94%) and the TC motif (7.91%). Twenty-one primer pairs were developed, and amplification tests were conducted for nine tree peony individuals. Furthermore, 19 cpSSR markers were amplified on 60 tree peony accessions by a capillary electrophoresis test. Of 19 cpSSR markers, 12 showed polymorphism with different alleles ranging from 1.333 to 3.000. The Shannon's information index and polymorphism information content values ranged from 0.038 to 0.887 (mean 0.432) and 0.032 to 0.589 (mean 0.268), respectively. The diversity levels for twelve loci ranged from 0.016 (at loci cpSSR-8 and cpSSR-26) to 0.543 (at locus cpSSR-15), averaging 0.268 for all loci. A total of 14 haplotypes (23.33%) were detected in the three populations. The haplotypic richness ranged from 0.949 to 1.751, with a mean of 1.233 per population. The genetic relationship suggested by the neighbor-joining-based dendrogram divided the genotypes into two clusters. The Jiangnan population was allotted to Cluster II, and the other two populations were distributed into both branches. These newly developed cpSSRs can be utilized for future breeding programs, population genetics investigations, unraveling the genetic relationships between related species, and germplasm management.

Paeoniflorin combined with norfloxacin ameliorates drug-resistant Streptococcus suis infection.

BACKGROUND: The increased resistance of bacterial pathogens to fluoroquinolones (FQs), such as norfloxacin and ciprofloxacin, supports the need to develop new antibacterial drugs and combination therapies using conventional antibiotics. The LuxS/AI-2 quorum sensing (QS) system can regulate the complex group behaviour of Streptococcus suis and impact its susceptibility to FQs. OBJECTIVES: We investigated the combination of paeoniflorin and norfloxacin as a novel and effective strategy against FQ-resistant S. suis. METHODS: FIC, AI-2 activity assay, real-time RT-PCR and biofilm inhibition assays were performed to investigate the in vitro effect of paeoniflorin combined with norfloxacin. Mouse protection and mouse anti-infection assays were performed to investigate the in vivo effect of paeoniflorin combined with norfloxacin. RESULTS: FIC results showed that paeoniflorin and norfloxacin exert a synergistic bactericidal effect. Evidence was brought that paeoniflorin reduces the S. suis AI-2 activity and significantly down-regulates the transcription of the FQ efflux pump gene. In addition, paeoniflorin can inhibit biofilm formation, thereby promoting the ability of norfloxacin to kill S. suis. Finally, we showed in a mouse model that paeoniflorin in association with norfloxacin is effective to treat S. suis infections. CONCLUSIONS: This study highlighted the inhibitory potential of paeoniflorin on the LuxS/AI-2 QS system of S. suis, and provided evidence that it can inhibit the FQ efflux pump and prevent biofilm formation to cooperate with norfloxacin in the treatment of resistant S. suis-related infections.

Supplementary honey bee (Apis mellifera L.) pollination enhances fruit growth rate and fruit yield in Paeonia ostii (family: Paeoniaceae).

Insufficient pollination leads to low and unstable production of oil tree peony. Supplementary managed honeybees (Apis mellifera L.) in agricultural ecosystems is a common practice for addressing the problem. At this study site (N 34°38'30″ and E 112°39'43″, with an altitude of 125.5 m), we set up four pollination areas (low-density bee pollination group (LDBP), high-density bee pollination group (HDBP), blank control group (CK1) and field control group (CK2)) to examine the pollination effectiveness of different densities of honeybee supplementation on oil tree peony (Paeonia ostii). Our work demonstrated that bee-pollination increased fruit size and growth rate. On average, bee-pollinated (LDBP) plants produced 63.16% more number of seeds per plant, showed also 53.47% more weight of seeds per plant than those in CK2. Also, seeds of LDBP contained, on average, 26.15% more oil content than CK2. Kernel percent and seed oil fatty acid content, however, were unaffected (F = 1.759, p = 0.074). Compared with LDBP, weight of seeds per plant and oil content with HDBP decreased by 21.89% and 2.63%, respectively. Following the same trend, compared with LDBP, HDBP slowed fruit growth and reduced fruit size. Our results showed that insufficient pollination limits fruit set in oil tree peony, while supplementary reasonable bee density in the field for pollination is an important strategy to maximize fruit yield.

Ultrasonic-assisted extraction and adsorption separation: Large-scale preparation of trans-ε-Viniferin, suffruficosol B and trans-Gnetin H for the first time.

In this study, a Standard Operating Procedure (SOP) for the large-scale extraction, enrichment, and separation of suffruticosol B (SB), trans-ε-Viniferin (TV), trans-gnetin H (TG) from oil tree peony seeds shell (PSS) was successfully constructed. The ultrasonic-assisted extraction (UAE), macroporous adsorption resin (MAR), and column chromatography (CC) were employed to extract, enrich and separate SB, TV and TG from PSS, and the conditions were optimized. The results implied that SB (1.6937 g), TV (0.5884 g) and TG (3.8786 g) with the purity of 99.67 %, 99.32 % and 98.54 %, respectively, were obtained after the extraction, enrichment and separation. The total yields of the SB, TV and TG were 0.61 mg/g, 0.02 mg/g and 6.64 mg/g with the total extraction rates at 70.55 %, 69.77 % and 78.36 %, respectively. This is the first report on the large-scale extraction, enrichment and separation of oligostilbenes. The SOP in this paper could produce high purity SB, TV and TG, and provide a new idea for PSS as a new oligostilbene resource. The study expands the new development and research field of PSS and provides theoretical support for the green utilization of oil tree peony.

Characterization and bioinformatics analysis of ptc-miR396g-5p in response to drought stress of Paeonia ostii.

Drought is one of the main abiotic stress factors affecting yield of Paeonia ostii. In this study, we conducted bioinformatics and differential expression analyses of P. ostii 'Feng Dan' ptc-miR396g-5p in leaf samples under different drought stress. ptc-miR396g-5p belongs to the miR396 family. Among the 271 plant species registered in the miRBase database, at least one miR396 member was found in 48 Angiospermae species, 3 in Gymnospermae species, and 1 in Pteridophy. Mature sequence alignment showed that P. ostii 'Feng Dan' ptc-miR396g-5p had high sequence similarity with miR396 from other species. Secondary structure prediction showed that the precursor sequence of 'Feng Dan' ptc-miR396g-5p could form a stable stem-loop structure, and the mature sequence was located on the 5' arm of the secondary structure. Phylogenetic tree analysis showed that 'Feng Dan' was closely related to 20 species such as Glycine max, Medicago truncatula, Populus trichocarpa, Citrus sinensis, Vitis vinifera, and Theobroma cacao. The predicted target gene of the 'Feng Dan' ptc-miR396g-5p encodes a Signal Transducer and Activator of Transcription (STAT) transcription factor. The negative correlation of expression between the miRNA and its target gene was confirmed by qRT-PCR. Our data indicate that 'Feng Dan' ptc-miR396g-5p's expression decreases under drought, leading to an expression increase of the STAT transcription factor.