First Report of Botrytis cinerea Causing Stem Rot on Hydrangea macrophylla in China.

Hydrangea (Hydrangea macrophylla), commonly referred to as big leaf hydrangea, is a species within the Hydrangeaceae family notable for its ornamental value. Characterized by its vividly colored sepals and lush, striking inflorescences, this species is globally esteemed as both a potted and landscape plant. Notably, in 2022, an alarming incidence of stem rot was observed in approximately 40% of H. macrophylla plants aged between six and twelve months within 16 greenhouses situated in Nanjing City (N 31°14', E 118°22'), Jiangsu Province, China. Initial symptoms of the disease manifested as wet gray-black spots at the base of the seedlings and stems, progressing to a necrotic gray-white discoloration in the stems and accompanied by the growth of gray mold on the affected parts. This infection ultimately led to the wilting of the leaves and the death of the seedlings. For pathogen identification, stem tissues at the interface of diseased and healthy sections were excised, surface-sterilized with 75 % ethanol for 30 s, followed by a 2 - 3 min treatment with 3% sodium hypochlorite, and subsequently rinsed three times with sterile water before air drying. Sections measuring 2 - 3 mm were then cultured on potato dextrose agar (PDA) medium, supplemented with 50 mg/mL rifampicin (RFP), and incubated at 25 ℃ for 3 - 5 d (Zhou et al. 2022). Upon 2 - 3 days of incubation, notable growth of fungal colonies was observed. Mycelial clusters from the periphery of these colonies were subsequently transferred to fresh PDA plates and incubated at 25 ℃ for an additional 5 - 7 d. A particular colony, designated JSNJ2022-2 and now preserved at the Jiangsu Academy of Agricultural Sciences, was selected for detailed examination. This colony exhibited a flocculent texture, with a coloration ranging from grey-white to light brown. It was characterized by the presence of irregularly formed, hard sclerotia within the hyphae. The conidiophores were observed to be slender and erect, featuring dendritic branches at their extremities. The conidia were clustered on the conidiophore like grapes. These conidia were generally colorless or grey, oval in shape, smooth and transparent, and measured between 6.4 - 12.2 × 7.3 - 18.2 µm (n = 50). For genetic analysis, genomic DNA (gDNA) was extracted using the DNA secure Plant Kit (Tiangen Biotech, Beijing, China). Polymerase chain reaction (PCR) amplification was performed using a set of universal primers of ITS1/ITS4 (White et al. 1990), primers corresponding to the specific sequences of glyceraldehyde-3-phosphate dehydrogenase (G3PDH), heat-shock protein 60 (HSP60), and DNA-dependent RNA polymerase subunit II (RPB2) (Yang et al. 2020). The resultant PCR products were sequenced, and the resulting sequences were submitted to the GenBank database, under the accession numbers OP131597, OP142320, OP142321, and OP142322, respectively. BLAST analysis of the sequences obtained from the isolate JSNJ2022-2 revealed a high degree of genetic similarity, ranging from 99 to 100%, with known sequences of Botrytis cinerea (accessions MK051124.1, MH796662.1, MH479931.1, and KU760986.1). To elucidate the phylogenetic position of the isolate, a phylogenetic tree was constructed using the maximum likelihood method, supported by 1,000 bootstrap replications, in the Mega7 software (Kumar et al. 2016). The results of this analysis confirmed that the strains under study clustered within the same branch as B. cinerea. To establish the pathogenicity of the isolate, Koch's postulates (Falkow 1988) were employed. Healthy potted H. macrophylla seedlings, approximately three months old, were wound inoculated at the base of the seedlings with a 6 mm diameter mycelium plug of JSNJ2002-2 cultivated on PDA for 3 days, which was subsequently covered with moistened degreasing cotton. Control plants were treated with moistened degreasing cloths minus the pathogen. Post-inoculation, these plants were placed in a growth chamber maintained at 25 ℃ with a relative humidity range of 60 - 80%. After a 3-d incubation period, the inoculated plants displayed symptoms identical to those initially observed in the greenhouse. The pathogen was successfully re-isolated from these inoculated plants and was morphologically re-confirmed as B. cinerea, thus satisfying the criteria of Koch's postulates. To our knowledge, this report represents the first documented incidence of B. cinerea causing stem rot in H. macrophylla in China.

Phyllodulcin from the hexane fraction of Hydrangea macrophylla inhibits glucose-induced lipid accumulation and reactive oxygen species generation in Caenorhabditis elegans.

We confirmed that the hexane layer of Hydrangea macrophylla leaf extract (HLH) is rich in phyllodulcin (PD), an alternative sweetener, through high performance liquid chromatography (HPLC) analysis. To investigate in vivo activity of HLH and its PD, acute toxicity and growth rate of Caenorhabditis elegans were tested and there are no clinical abnormalities at 125-500 µg/mL of HLH. HLH decreased the total lipid and triglyceride contents dose-dependently in glucose-induced obese worms. Also, HLH increased survival rates under oxidative and thermal stress and decreased body reactive oxygen species (ROS) contents significantly. Such antioxidant properties of HLH were attributed to the enhanced activity of the antioxidant enzyme catalase. To determine whether the effect of HLH was due to PD, worms were treated with PD (concentration contained in HLH), and inhibitory effects on total lipids and ROS were observed. Our results suggest that HLH and its PD as a natural alternative sweetener can be used as materials to improve metabolic diseases.

Regulation of blue infertile flower pigmentation by WD40 transcription factor HmWDR68 in Hydrangea macrophylla 'forever summer'.

BACKGROUND: WD40 transcription factors are crucial in plant growth and developmental, significantly impacting plant growth regulation. This study investigates the WD40 transcription factor HmWDR68's role in developing the distinctive blue infertile flower colors in Hydrangea macrophylla 'Forever Summer'. METHODS AND RESULTS: The HmWDR68 gene was isolated by PCR, revealing an open reading frame of 1026 base pairs, which encodes 341 amino acids. Characterized by four WD40 motifs, HmWDR68 is a member of the WD40 family. Phylogenetic analysis indicates that HmWDR68 shares high homology with PsWD40 in Camellia sinensis and CsWD40 in Paeonia suffruticosa, both of which are integral in anthocyanin synthesis regulation. Quantitative real-time PCR (qRT-PCR) analysis demonstrated that HmWDR68 expression in the blue infertile flowers of 'Forever Summer' hydrangea was significantly higher compared to other tissues and organs. Additionally, in various hydrangea varieties with differently colored infertile flowers, HmWDR68 expression was markedly elevated in comparison to other hydrangea varieties, correlating with the development of blue infertile flowers. Pearson correlation analysis revealed a significant association between HmWDR68 expression and the concentration of delphinidin 3-O-glucoside, as well as key genes involved in anthocyanin biosynthesis (HmF3H, HmC3'5'H, HmDFR, and HmANS) in the blue infertile flowers of 'Forever Summer' hydrangea (P < 0.01). CONCLUSION: These findings suggest HmWDR68 may specifically regulate blue infertile flower formation in hydrangea by enhancing delphinidin-3-O-glucoside synthesis, modulating expression of HmF3H, HmC3'5'H, HmDFR and HmANS. This study provides insights into HmWDR68's role in hydrangea's blue flowers development, offering a foundation for further research in this field.

First Reports of Cladosporium tenuissimum Causing Leaf spots on Hydrangea macrophylla in Anhui province in China.

Forever Summer Hydrangea (Hydrangea macrophylla) is a common flowering plant in the Yangtze River Valley area of China, and it is widely cultivated globally (Chen et al. 2015). In July 2023, H. macrophylla leaves exhibiting visible diseased lesions were reported in a nursery in Wuhu, Anhui Province, China. The incidence reached 40% in a 0.2 ha area. The primary disease symptom was multiple irregular necrotic spots (0.5 to 1 mm in diameter) appearing on the leaves. These spots on the leaves were faded yellow around the perimeter and grayish brown in the center.). 15 leaf samples were sterilized with 75% alcohol and rinsed three times in sterile distilled water, then transferred to antibiotic-added potato dextrose agar (PDA) for incubation at 27°C. The colonies were fluffy, flocculent, or hairy, dark green, gray-green to gray-brown in color, and spreading or protruding punctate with a colorless halo on PDA. The conidiophores were brown to dark brown, smooth or rough surface, mostly unbranched, clearly differentiated, erect or curved. The conidia displayed a light brown to brown hue, lemon shape, fusiform, elongated ellipsoid or others with obvious spore markings and spore umbilicus. Genomic DNA was extracted from fungal colonies on infected leaves of three collections separately (Braun et al. 2003) and the internal transcribed spacer regions (ITS), actin (ACT) genes and partial translation elongation factor-l-alpha (EF) were amplified and sequenced using the primers ITS1/4 (Yin et al. 2012), ACT-512F/ACT-783R and EF 1-728F/986R (Carbone and Kohn 1999), respectively. DNA sequences of isolates were identical and deposited in GenBank (accession no. OR362754 for ITS, OR611929 for ACT and PP209106 for EF). The consensus sequences from ITS, EF and ACT showed 100%, 98.98% and 100% identical to Cladosporium strains (accession no. OQ186140.1, MT154169.1 and OL322092.1), respectively. To confirm the pathogenicity of the isolates, hydrangeas were planted in 15-cm pots containing commercial potting mix (one plant/pot). Three healthy plants were inoculated at the five to eight leaf stage by spraying 50 µL of the isolate conidial suspension (4 × 106 spores/mL) on healthy leaves. Three plants treated with sterile distilled water were used as controls. After inoculation, all plants were placed in a humidity chamber (>95% relative humidity, 26°C) for 48 h and then transferred to a greenhouse at 22/27°C. All inoculated leaves exhibited symptoms similar to those observed in the nursery 10 days after inoculation, while no symptoms were observed for control leaves. The fungus was re-isolated and confirmed to be C. tenuissimum. Based on the above morphological characterization and molecular identification, the causal agent for this leaf spot disease was identified as C. tenuissimum. Although C. tenuissimum has been reported to cause disease on H. paniculata in northern China (Li et al.2021), this is the first time that C. tenuissimum has been found on H. macrophylla in southern China. This new disease of H. macrophylla caused by C. tenuissimum is a threat to urban greening and is worth further investigation.

Production of Dihydroisocoumarins by Callus Induction from Hydrangea macrophylla var. thunbergii Leaves.

Dihydroisocoumarins, hydrangenol 8-O-ß-D-glucopyranoside (1), phyllodulcin 8-O-ß-D-glucopyranoside (2), hydrangenol (3), and phyllodulcin (4), are well-known as the major secondary metabolites in the leaves of Hydrangea macrophylla var. thunbergii. Dihydroisocoumarins are pharmaceutical compounds with diverse bioactivity. Although dihydroisocoumarins are commonly isolated from Hydrangea plants or via organic chemical synthesis, their production via callus induction is considered a promising alternative. In the present study, callus induction and proliferation of H. macrophylla var. thunbergii, and constituents 1-4 were quantified in calluses cultured in 17 different media. We found that the combination of the phytohormones 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BA) was useful for callus proliferation in H. macrophylla var. thunbergii. The balance and concentrations of indole-3-acetic acid (IAA) and BA greatly affected the contents of 1-4. Particularly, 1 (2.03-3.46% yield from the dry callus) was successfully produced from the callus induced by IAA (0.5 mg/L) and BA (1.0 mg/L) at yields comparable to isolated yields from plants. To the best of our knowledge, this is the first study to show that the calluses of H. macrophylla var. thunbergii contained 1. These findings may be useful for producing bioactive dihydroisocoumarins.

Quantitative analysis of taxiphyllin, a cyanogenic glycoside, in the leaves of Hydrangea macrophylla var. thunbergii.

The dried and fermented leaves of Hydrangea macrophylla var. thunbergii are currently used as crude drugs (Sweet Hydrangea Leaf) with a sweet taste for patients with diabetes. In recent years, cases of food poisoning with symptoms of vomiting etc. have been reported after drinking a decoction of this crude drug. Cyanogenic glycosides have been suggested as potential causative agents. However, cyanogenic glycosides from H. macrophylla var. thunbergii was ambiguous. In the present study, we found that the leaves contained the cyanogenic glycoside taxiphillin (1). Next, the content of 1 in leaves of different sizes, colors, parts, and growth periods was quantified. In addition, we prepared the leaves of plants grown in five types of soils with different pH values (pH 5.0-7.5). The content of 1 in the leaves of the plants grown in these soils was quantified. The content of 1 varied greatly, with more than a three-fold difference, depending on when the leaves were collected from the plants. Furthermore, we compared the content of 1 in the crude drug obtained under different processing conditions for H. macrophylla var. thunbergii. The results showed that 1 was mostly hydrolyzed during plant processing. It has been suggested that cyanogenic glycosides are not the causative constituents of food poisoning.

Dihydroisocoumarins of Hydrangea macrophylla var. thunbergia inhibit binding of the SARS-CoV-2 spike protein to ACE2.

Binding of the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to the cognate angiotensin-converting enzyme 2 (ACE2) receptor is the initial step in the viral infection process. In this study, we screened an in-house extract library to identify food materials with inhibitory activity against this binding using enzyme-linked immunosorbent assays and attempted to ascertain their active constituents. Hydrangea macrophylla var. thunbergia leaves were identified as candidate materials. Its active compounds were purified using conventional chromatographic methods and identified as naringenin, dihydroisocoumarins, hydrangenol, and phyllodulcin, which have affinities for the ACE2 receptor and inhibit ACE2 receptor-spike S1 binding. Given that boiled water extracts of H. macrophylla leaves are commonly consumed as sweet tea in Japan, we speculated that this tea could be used as a potential natural resource to reduce the risk of SARS-CoV-2 infection.

Study on the Flower Induction Mechanism of Hydrangea macrophylla.

The flower induction of Hydrangea macrophylla "Endless Summer" is regulated by a complex gene network that involves multiple signaling pathways to ensure continuous flowering throughout the growing season, but the molecular determinants of flower induction are not yet clear. In this study, genes potentially involved in signaling pathway mediating the regulatory mechanism of flower induction were identified through the transcriptomic profiles, and a hypothetical model for this regulatory mechanism was obtained by an analysis of the available transcriptomic data, suggesting that sugar-, hormone-, and flowering-related genes participated in the flower induction process of H. macrophylla "Endless Summer". The expression profiles of the genes involved in the biosynthesis and metabolism of sugar showed that the beta-amylase gene BAM1 displayed a high expression level at the BS2 stage and implied the hydrolysis of starch. It may be a signaling molecule that promotes the transition from vegetative growth to reproductive growth in H. macrophylla "Endless Summer". Complex hormone regulatory networks involved in abscisic acid (ABA), auxin (IAA), zeatin nucleoside (ZR), and gibberellin (GA) also induced flower formation in H. macrophylla. ABA participated in flower induction by regulating flowering genes. The high content of IAA and the high expression level of the auxin influx carrier gene LAX5 at the BS2 stage suggested that the flow of auxin between sources and sinks in H. macrophylla is involved in the regulation of floral induction as a signal. In addition, flowering-related genes were mainly involved in the photoperiodic pathway, the aging pathway, and the gibberellin pathway. As a result, multiple pathways, including the photoperiodic pathway, the aging pathway, and the gibberellin pathway, which were mainly mediated by crosstalk between sugar and hormone signals, regulated the molecular network involved in flower induction in H. macrophylla "Endless Summer".

Comparison of Growth in Hydrangea macrophylla var. thunbergii Grown in Different Soil pH and Quantitative Analysis of Its Sweetness-Related Constituents.

Dried and fermented (processed) leaves of Hydrangea macrophylla Seringe var. thunbergii Makino (Hydrangeae Dulcis Folium) are currently used as a crude drug with a sweet taste for diabetic patients and as an oral refrigerant. The sweet taste of this crude drug is primarily attributed to phyllodulcin. However, there are currently no standards for the cultivation of H. macrophylla var. thunbergii and the isolation and production of the primary constituents of this crude drug. In the present study, we prepared five types of soils with different pH values (pH 7.5-5.0) and investigated the effects of these soils on the growth of this plant. The contents of phyllodulcin and its glycoside, phyllodulcin 8-O-ß-D-glucopyranoside, in the leaves of plants grown in these soils were quantified. Furthermore, the correlation between the sweetness of Hydrangeae Dulcis Folium and phyllodulcin was investigated. The results showed that soils with pH ranging from 7.0 to 5.5 was not only suitable for plant growth but also increased the content of phyllodulcin and phyllodulcin 8-O-ß-D-glucopyranoside in the leaves. Altogether, these findings could be useful for the development of high-quality Hydrangeae Dulcis Folium.

Immunostimulatory activity of Lactococcus lactis LM1185 isolated from Hydrangea macrophylla.

The lactic acid bacteria, Lactococcus lactis subsp. lactis LM1185 was isolated from Hydrangea macrophylla. Strain LM1185 showed 50.5% of acid tolerance at pH 2.5 for 2 h and 30.4% of 0.3% (w/v) bile salt tolerance for 24 h. The antioxidant activity of this strain was measured at 99.4% of 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging activity. When RAW 264.7 macrophage cells were treated with strain LM1185, there was no observed cytotoxicity. This strain showed high nitric oxide production and mRNA expression levels of cytokines such as tumor necrosis factor-α and inducible nitric oxide synthase (iNOS). The nuclear factor-kB signaling pathway was activated by this strain resulting in the production of iNOS and cyclooxygenase-2 determined by western blotting. The present results indicated that L. lactis subsp. lactis LM1185 could be used as potential probiotics and may play a crucial role in the immunostimulatory effect on macrophages. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01199-5.

Exploring the Molecular Mechanism of Sepal Formation in the Decorative Flowers of Hydrangea macrophylla 'Endless Summer' Based on the ABCDE Model.

With its large inflorescences and colorful flowers, Hydrangea macrophylla has been one of the most popular ornamental plants in recent years. However, the formation mechanism of its major ornamental part, the decorative floret sepals, is still not clear. In this study, we compared the transcriptome data of H. macrophylla 'Endless Summer' from the nutritional stage (BS1) to the blooming stage (BS5) and annotated them into the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) databases. The 347 identified differentially expressed genes (DEGs) associated with flower development were subjected to a trend analysis and a protein-protein interaction analysis. The combined analysis of the two yielded 60 DEGs, including four MADS-box transcription factors (HmSVP-1, HmSOC1, HmAP1-2, and HmAGL24-3) and genes with strong connectivity (HmLFY and HmUFO). In addition, 17 transcription factors related to the ABCDE model were screened, and key candidate genes related to the development of decorative floret sepals in H. macrophylla were identified by phylogenetic and expression pattern analysis, including HmAP1-1, HmAP1-2, HmAP1-3, HmAP2-3, HmAP2-4, and HmAP2-5. On this basis, a gene regulatory network model of decorative sepal development was also postulated. Our results provide a theoretical basis for the study of the formation mechanism of decorative floret sepals and suggest a new direction for the molecular breeding of H. macrophylla.

Comparative physiology and transcriptome analysis reveals that chloroplast development influences silver-white leaf color formation in Hydrangea macrophylla var. maculata.

BACKGROUND: Hydrangea macrophylla var. Maculata 'Yinbianxiuqiu' (YB) is an excellent plant species with beautiful flowers and leaves with silvery white edges. However, there are few reports on its leaf color characteristics and color formation mechanism. RESULTS: The present study compared the phenotypic, physiological and transcriptomic differences between YB and a full-green leaf mutant (YM) obtained from YB. The results showed that YB and YM had similar genetic backgrounds, but photosynthesis was reduced in YB. The contents of pigments were significantly decreased at the edges of YB leaves compared to YM leaves. The ultrastructure of chloroplasts in the YB leaves was irregular. Transcriptome profiling identified 7,023 differentially expressed genes between YB and YM. The expression levels of genes involved in photosynthesis, chloroplast development and division were different between YB and YM. Quantitative real-time PCR showed that the expression trends were generally consistent with the transcriptome data. CONCLUSIONS: Taken together, the formation of the silvery white leaf color of H. macrophylla var. maculata was primarily due to the abnormal development of chloroplasts. This study facilitates the molecular function analysis of key genes involved in chloroplast development and provides new insights into the molecular mechanisms involved in leaf coloration in H. macrophylla.

Separation of Dihydro-Isocoumarins and Dihydro-Stilbenoids from Hydrangea macrophylla ssp. serrata by Use of Counter-Current Chromatography.

Previously, different Hydrangea macrophylla ssp. serrata cultivars were investigated by untargeted LC-MS analysis. From this, a list of tentatively identified and unknown compounds that differ significantly between these cultivars was obtained. Due to the lack of reference compounds, especially for dihydro-isocoumarins, we aimed to isolate and structurally characterise these compounds from the cultivar 'Yae-no-amacha' using NMR and LC-MS methods. For purification and isolation, counter-current chromatography was used in combination with reversed-phase preparative HPLC as an orthogonal and enhanced purification workflow. Thirteen dihydro-isocoumarins in combination with other metabolites could be isolated and structurally identified. Particularly interesting was the clarification of dihydrostilbenoid glycosides, which were described for the first time in H. macrophylla ssp. serrata. These results will help us in further studies on the biological interpretation of our data.

Biochemistry and transcriptome analyses reveal key genes and pathways involved in high-aluminum stress response and tolerance in hydrangea sepals.

Hydrangea [Hydrangea macrophylla (Thunb.) Ser.] is a high aluminum-tolerant ornamental plant species, which has a specific characteristic of color change, ie. some cultivars' floral color will change from red to blue or blue-violet planted in acidic soil containing aluminum. This study aims to understand the complex molecular mechanisms of floral color change under Al stress, through comparative biochemistry and transcriptome analyses between an Al3+-sensitive cultivar 'Bailer' and insensitive cultivar 'Ruby' under Al-stress. The results of biochemistry analysis showed that 'Bailer' displayed higher contents of Al3+ and delphinium-3-O-glucoside than that of 'Ruby' after Al2(SO4)3 treating. Meanwhile, the transcriptome analysis of different tissues identified 12,321 differentially expressed genes (DEGs) in 'Bailer' and 6,703 in 'Ruby'. Transcriptome analysis showed that changes in genes' expression pattern in several genes and pathways [such as including metal transporters, reactive oxygen species (ROS) scavenging enzyme, plant hormone signal transduction and favonoid biosynthesis pathway] were the key contributors to the Al3+-sensitive cultivar 'Bailer'. Besides, gene co-expression network analysis (WGCNA) demonstrated that five hub genes, including ABC transporters (TRINITY_DN1053_c0_g1, TRINITY_DN3377_c0_g2), cationic amino acid transporter (TRINITY_DN9684_c0_g2), oligopeptide transporter (TRINITY_DN1147_c0_g2) and flavonol synthase (TRINITY_DN15902_c0_g1), played vital roles in the networks regulating Al tolerance in hydrangea. Furthermore, HmABCI17's (TRINITY_DN1053_c0_g1) expression enhanced Al tolerance in yeast. The conclusions of this study are helpful to elucidate the differences and molecular mechanisms of different hydrangea cultivars on Al tolerance, and provide new insights into molecular assisted-screening for breeding blue flowers in hydrangea and other ornamental plants.

A New Ilarvirus Found in French Hydrangea.

In this study, a new virus was identified in French hydrangea plants, exhibiting chlorotic vein banding and necrotic ring spots on older leaves. The virus was mechanically transmitted to herbaceous hosts, in which it induced local and systemic or only local symptoms. The genome of the new virus was characterized and consisted of three RNA sequences that were 3422 (RNA 1), 2905 (RNA 2) and 2299 (RNA 3) nucleotides long, with five predicted open reading frames; RNA2 was bicistronic and contained conserved domains and motifs typical of ilarviruses. The phylogenetic analysis of the predicted proteins-p1, p2a, p3a and p3b-revealed its close relationship to recognized members of subgroup 2 within the genus Ilarvirus. Homologous antiserum was effective in the detection of the virus in plant extracts and no cross reactions with two other distinct members of subgroup 2 were observed. Overall, the biological features, phylogenetic relationships and serological data suggest that this virus is a new member of the genus, for which we propose the name hydrangea vein banding virus (HdVBV).

Hydrangea macrophylla and Thunberginol C Attenuate Stress-Induced Anxiety in Mice.

Stress is an important neurological input for successful life. However, chronic stress and stress hormones could be a cause of various neurological disorders including anxiety disorders. Therefore, there have been many efforts to find effective materials for curing stress-induced neurological disorders. In this study, we examined the effect of Hydrangea macrophylla (HM) on corticosterone-induced neurotoxicity, stress-induced anxiety in mice and suggested a possible active ingredient of HM. HM protected cortical neurons against neurotoxicity of corticosterone (CORT), a stress hormone. HM also blocked CORT-induced hippocampal synaptic deficit via regulating Akt signaling. Oral administration of HM improved chronic restraint stress-induced anxiety in Elevated Plus maze test along with reduction of plasma corticosterone and TNF-α levels. Moreover, HM reduced stress-induced neuroinflammation and oxidative stress. Thunberginol C, an active ingredient of HM, also prevented CORT-induced neuronal cell death and restraint stress-induced anxiety. Moreover, thunberginol C reduced plasma TNF-α level and neuroinflammation and oxidative stress. Collectively, HM could be a good candidate for preventing stress-induced neurological disorders and thunberginol C may be an active ingredient of HM for this purpose.

Current and future distribution of the deciduous shrub Hydrangea macrophylla in China estimated by MaxEnt.

Climate change has a significant impact on the growth and distribution of vegetation worldwide. Hydrangea macrophylla is widely distributed and considered a model species for studying the distribution and responses of shrub plants under climate change. These results can inform decision-making regarding shrub plant protection, management, and introduction of germplasm resources, and are of great importance for formulating ecological countermeasures to climate change in the future. We used the maximum entropy model to predict the change, scope expansion/reduction, centroid movement, and dominant climate factors that restrict the growth and distribution of H. macrophylla in China under current and future climate change scenarios. It was found that both precipitation and temperature affect the distribution of suitable habitat for H. macrophylla. Akaike information criterion (AICc) was used to select the feature combination (FC) and the regularization multiplier (RM). After the establishment of the optimal model (FC = QP, RM = 0.5), the complexity and over-fitting degree of the model were low (delta AICc = 0, omission rate = 0.026, difference between training and testing area under the curve values = 0.0009), indicating that it had high accuracy in predicting the potential geographical distribution of H. macrophylla (area under the curve = 0.979). Overall, from the current period to future, the potential suitable habitat of this species in China expanded to the north. The greenhouse effect caused by an increase in CO2 emissions would not only increase the area of high-suitability habitat in Central China, but also expand the area of total suitable habitat in the north. Under the maximum greenhouse gas emission scenario (RCP8.5), the migration distance of the centroid was the longest (e.g., By 2070s, the centroids of total and highly suitable areas have shifted 186.15 km and 89.84 km, respectively).

Exploring the Molecular Mechanism of Blue Flower Color Formation in Hydrangea macrophylla cv. "Forever Summer".

Hydrangea macrophylla has a large inflorescence and rich colors, which has made it one of the most popular ornamental flowers worldwide. Thus far, the molecular mechanism of flower color formation in H. macrophylla flowers is unknown. By comparing the pigment content and transcriptome data of the bud period (FSF1), discoloration period (FSF2) and full-bloom stage (FSF3) of infertile blue flowers of H. macrophylla cv. "Forever Summer," we found that genes associated with anthocyanin production were most associated with the formation of blue infertile flowers throughout development. The anthocyanin biosynthesis pathway is the main metabolic pathway associated with flower color formation, and the carotenoid biosynthesis pathway appeared to have almost no contribution to flower color. There was no competition between the flavonoid and flavonol and anthocyanin biosynthesis pathways for their substrate. At FSF1, the key genes CHS and CHI in the flavonoid biosynthesis pathway were up-regulated, underlying the accumulation of a substrate for anthocyanin synthesis. By FSF3, the downstream genes F3H, C3'5'H, CYP75B1, DFR, and ANS in the anthocyanin biosynthesis pathway were almost all up-regulated, likely promoting the synthesis and accumulation of anthocyanins and inducing the color change of infertile flowers. By analyzing protein-protein interaction networks and co-expression of transcription factors as well as differentially expressed structural genes related to anthocyanin synthesis, we identified negatively regulated transcription factors such as WER-like, MYB114, and WDR68. Their site of action may be the key gene DFR in the anthocyanin biosynthesis pathway. The potential regulatory mechanism of flower color formation may be that WER-like, MYB114, and WDR68 inhibit or promote the synthesis of anthocyanins by negatively regulating the expression of DFR. These results provide an important basis for studying the infertile flower color formation mechanism in H. macrophylla and the development of new cultivars with other colors.

Insight into chemical mechanisms of sepal color development and variation in hydrangea.

Hydrangea (Hydrangea macrophylla) is a unique flower because it is composed of sepals rather than true petals that have the ability to change color. In the early 20th century, it was known that soil acidity and Al3+ content could intensify the blue hue of the sepals. In the mid-20th century, the anthocyanin component 3-O-glucosyldelphinidin (1) and the copigment components 5-O-caffeoylquinic, 5-O-p-coumaroylquinic, and 3-O-caffeoylquinic acids (2-4) were reported. Interestingly, all hydrangea colors from red to purple to blue are produced by the same organic components. We were interested in this phenomenon and the chemical mechanisms underlying hydrangea color variation. In this review, we summarize our recent studies on the chemical mechanisms underlying hydrangea sepal color development, including the structure of the blue complex, transporters involved in accumulation of aluminum ion (Al3+), and distribution of the blue complex and aluminum ions in living sepal tissue.