A serine carboxypeptidase-like acyltransferase catalyzes consecutive four-step reactions of hydrolyzable tannin biosynthesis in Camellia oleifera.

Hydrolyzable tannins (HTs), a class of polyphenolic compounds found in dicotyledonous plants, are widely used in food and pharmaceutical industries because of their beneficial effects on human health. Although the biosynthesis of simple HTs has been verified at the enzymatic level, relevant genes have not yet been identified. Here, based on the parent ion-fragment ion pairs in the feature fragment data obtained using UPLC-Q-TOF-/MS/MS, galloyl phenolic compounds in the leaves of Camellia sinensis and C. oleifera were analyzed qualitatively and quantitatively. Correlation analysis between the transcript abundance of serine carboxypeptidase-like acyltransferases (SCPL-ATs) and the peak area of galloyl products in Camellia species showed that SCPL3 expression was highly correlated with HT biosynthesis. Enzymatic verification of the recombinant protein showed that CoSCPL3 from C. oleifera catalyzed the four consecutive steps involved in the conversion of digalloylglucose to pentagalloylglucose. We also identified the residues affecting the enzymatic activity of CoSCPL3 and determined that SCPL-AT catalyzes the synthesis of galloyl glycosides. The findings of this study provide a target gene for germplasm innovation of important cash crops that are rich in HTs, such as C. oleifera, strawberry, and walnut.

Genome-wide identification and molecular evolution of Dof gene family in Camellia oleifera.

DNA binding with one finger(Dof) gene family is a class of transcription factors which play an important role on plant growth and development. Genome-wide identification results indicated that there were 45 Dof genes(ColDof) in C.oleifera genome. All 45 ColDof proteins were non-transmembrane and non-secretory proteins. Phosphorylation site analysis showed that biological function of ColDof proteins were mainly realized by phosphorylation at serine (Ser) site. The secondary structure of 44 ColDof proteins was dominated by random coil, and only one ColDof protein was dominated by α-helix. ColDof genes' promoter region contained a variety of cis-acting elements, including light responsive regulators, gibberellin responsive regulators, abscisic acid responsive regulators, auxin responsive regulators and drought induction responsive regulators. The SSR sites analysis showed that the proportion of single nucleotide repeats and the frequency of A/T in ColDof genes were the largest. Non-coding RNA analysis showed that 45 ColDof genes contained 232 miRNAs. Transcription factor binding sites of ColDof genes showed that ColDof genes had 5793 ERF binding sites, 4381 Dof binding sites, 2206 MYB binding sites, 3702 BCR-BPC binding sites. ColDof9, ColDof39 and ColDof44 were expected to have the most TFBSs. The collinearity analysis showed that there were 40 colinear locis between ColDof proteins and AtDof proteins. Phylogenetic analysis showed that ColDof gene family was most closely related to that of Camellia sinensis var. sinensis cv.Biyun and Camellia lanceoleosa. Protein-protein interaction analysis showed that ColDof34, ColDof20, ColDof28, ColDof35, ColDof42 and ColDof26 had the most protein interactions. The transcriptome analysis of C. oleifera seeds showed that 21 ColDof genes were involved in the growth and development process of C. oleifera seeds, and were expressed in 221 C. oleifera varieties. The results of qRT-PCR experiments treated with different concentrations NaCl and PEG6000 solutions indicated that ColDof1, ColDof2, ColDof14 and ColDof36 not only had significant molecular mechanisms for salt stress tolerance, but also significant molecular functions for drought stress tolerance in C. oleifera. The results of this study provide a reference for further understanding of the function of ColDof genes in C.oleifera.

Integrative metabolome and transcriptome analyses reveal the coloration mechanism in Camellia oleifera petals with different color.

BACKGROUND: Camellia olelfera petals are colorful, and have high ornamental value. However, the color formation mechanism of C. olelfera petals with different color is still unclear. In our study, WGCNA method was applied to integrate metabolites and transcriptomes to investigate the coloration mechanism of four C. olelfera cultivars with different petal colors. RESULTS: Here, a total of 372 flavonoids were identified (including 27 anthocyanins), and 13 anthocyanins were significantly differentially accumulated in C. olelfera petals. Among them, cyanidin-3-O-(6''-O-p-Coumaroyl) glucoside was the main color constituent in pink petals, cyanidin-3-O-glucoside, cyanidin-3-O-galactoside, cyanidin-3-O-rutinoside, and cyanidin-3-O-(6''-O-malonyl) glucoside were the main contributors to candy pink petals, and peonidin-3-O-glucoside was the important color substance responsible for the red petals of C. oleifera. Furthermore, six structural genes (Co4CL1, CoF3H1, CoF3'H, CoANS, CoUGT75C1-4, and CoUGT75C1-5), three MYBs (CoMYB1, CoMYB4, and CoMYB44-3), three bHLHs (CobHLH30, CobHLH 77, and CobHLH 79-1), and two WRKYs (CoWRKY7 and CoWRKY22) could be identified candidate genes related to anthocyanins biosynthesis and accumulation, and lead to the pink and red phenotypes. The regulatory network of differentially accumulated anthocyanins and the anthocyanins related genes in C. olelfera petals were established. CONCLUSIONS: These findings elucidate the molecular basis of the coloration mechanisms of pink and red color in C. olelfera petals, and provided valuable target genes for future improvement of petals color in C. olelfera.

High-yield hybrid breeding of Camellia oleifolia based on ISSR molecular markers.

BACKGROUND: C. Oleifera is among the world's largest four woody plants known for their edible oil production, yet the contribution rate of improved varieties is less than 20%. The species traditional breeding is lengthy cycle (20-30 years), occupation of land resources, high labor cost, and low accuracy and efficiency, which can be enhanced by molecular marker-assisted selection. However, the lack of high-quality molecular markers hinders the species genetic analysis and molecular breeding. RESULTS: Through quantitative traits characterization, genetic diversity assessment, and association studies, we generated a selection population with wide genetic diversity, and identified five excellent high-yield parental combinations associated with four reliable high-yield ISSR markers. Early selection criteria were determined based on kernel fresh weight and cultivated 1-year seedling height, aided by the identification of these 4 ISSR markers. Specific assignment of selected individuals as paternal and maternal parents was made to capitalize on their unique attributes. CONCLUSIONS: Our results indicated that molecular markers-assisted breeding can effectively shorten, enhance selection accuracy and efficiency and facilitate the development of a new breeding system for C. oleifera.

The complex hexaploid oil-Camellia genome traces back its phylogenomic history and multi-omics analysis of Camellia oil biosynthesis.

Oil-Camellia (Camellia oleifera), belonging to the Theaceae family Camellia, is an important woody edible oil tree species. The Camellia oil in its mature seed kernels, mainly consists of more than 90% unsaturated fatty acids, tea polyphenols, flavonoids, squalene and other active substances, which is one of the best quality edible vegetable oils in the world. However, genetic research and molecular breeding on oil-Camellia are challenging due to its complex genetic background. Here, we successfully report a chromosome-scale genome assembly for a hexaploid oil-Camellia cultivar Changlin40. This assembly contains 8.80 Gb genomic sequences with scaffold N50 of 180.0 Mb and 45 pseudochromosomes comprising 15 homologous groups with three members each, which contain 135 868 genes with an average length of 3936 bp. Referring to the diploid genome, intragenomic and intergenomic comparisons of synteny indicate homologous chromosomal similarity and changes. Moreover, comparative and evolutionary analyses reveal three rounds of whole-genome duplication (WGD) events, as well as the possible diversification of hexaploid Changlin40 with diploid occurred approximately 9.06 million years ago (MYA). Furthermore, through the combination of genomics, transcriptomics and metabolomics approaches, a complex regulatory network was constructed and allows to identify potential key structural genes (SAD, FAD2 and FAD3) and transcription factors (AP2 and C2H2) that regulate the metabolism of Camellia oil, especially for unsaturated fatty acids biosynthesis. Overall, the genomic resource generated from this study has great potential to accelerate the research for the molecular biology and genetic improvement of hexaploid oil-Camellia, as well as to understand polyploid genome evolution.

Streptomyces camelliae sp. nov., isolated from rhizosphere soil of Camellia oleifera Abel.

A novel actinobacterium strain, designated HUAS 2-6 T, was isolated from the rhizosphere soil of Camellia oleifera Abel collected from Taoyuan County, Northwestern Hunan Province, South China. This strain was subjected to a polyphasic taxonomic study. Strain HUAS 2-6 T is characterized by morphology typical of members of the genus Streptomyces, with deep purplish vinaceous aerial mycelia and deep dull lavender substrate mycelia. Strain HUAS 2-6 T, based on the full-length 16S rRNA gene sequence analysis, exhibited the highest similarities to S. puniciscabiei S77T (99.31%), S. filipinensis NBRC 12860 T (99.10%), S. yaanensis CGMCC 4.7035 T (99.09%), S. fodineus TW1S1T (99.08%), S. broussonetiae CICC 24819 T (98.76%), S. achromogenes JCM 4121 T (98.69%), S. barringtoniae JA03T (98.69%), and less than 98.70% with other validly species. In phylogenomic tree, strain HUAS 2-6 T was clustered together with S. broussonetiae CICC 24819 T, suggesting that they were closely related to each other. However, average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) between them were much less than the species cutoff values (ANI 96.7% and dDDH 70%). Moreover, in phenotypic and chemotaxonomic characteristics, strain HUAS 2-6 T is distinct from S. broussonetiae CICC 24819 T. On the basis of the polyphasic data, strain HUAS 2-6 T is proposed to represent a novel species, Streptomyces camelliae sp. nov. (= MCCC 1K04729T = JCM 35918 T).

Identification and analysis of MAPK cascade gene families of Camellia oleifera and their roles in response to cold stress.

BACKGROUND: Low-temperature severely limits the growth and development of Camellia oleifera (C. oleifera). The mitogen-activated protein kinase (MAPK) cascade plays a key role in the response to cold stress. METHODS AND RESULTS: Our study aims to identify MAPK cascade genes in C. oleifera and reveal their roles in response to cold stress. In our study, we systematically identified and analyzed the MAPK cascade gene families of C. oleifera, including their physical and chemical properties, conserved motifs, and multiple sequence alignments. In addition, we characterized the interacting networks of MAPKK kinase (MAPKKK)-MAPK kinase (MAPKK)-MAPK in C. oleifera. The molecular mechanism of cold stress resistance of MAPK cascade genes in wild C. oleifera was analyzed by differential gene expression and real-time quantitative reverse transcription-PCR (qRT-PCR). CONCLUSION: In this study, 21 MAPKs, 4 MAPKKs and 55 MAPKKKs genes were identified in the leaf transcriptome of C. oleifera. According to the phylogenetic results, MAPKs were divided into 4 groups (A, B, C and D), MAPKKs were divided into 3 groups (A, B and D), and MAPKKKs were divided into 2 groups (MEKK and Raf). Motif analysis showed that the motifs in each subfamily were conserved, and most of the motifs in the same subfamily were basically the same. The protein interaction network based on Arabidopsis thaliana (A. thaliana) homologs revealed that MAPK, MAPKK, and MAPKKK genes were widely involved in C. oleifera growth and development and in responses to biotic and abiotic stresses. Gene expression analysis revealed that the CoMAPKKK5/CoMAPKKK43/CoMAPKKK49-CoMAPKK4-CoMAPK8 module may play a key role in the cold stress resistance of wild C. oleifera at a high-elevation site in Lu Mountain (LSG). This study can facilitate the mining and utilization of genetic resources of C. oleifera with low-temperature tolerance.

Phenolic and flavonoid compounds from the fruit shell of Camellia oleifera.

A new phenolic compound oleiphenol (1), and a new dihydrochalcone oleifechalcone (2) along with seven known compounds (3-9) were isolated from the fruit shell of Camellia oleifera Abel. The planar structures of compounds 1 and 2 were determined on the basis of extensive spectroscopic analyses (IR, UV, NMR, and HR-ESI-MS) and comparison with literature data. The absolute configurations of the new structures were determined by ECD calculations and chemical methods. In addition, compounds 1-9 underwent a series of pharmacological activity tests, including cytotoxic, anti-inflammatory, anti-RSV and antioxidant activities.

High-Density Genetic Map Construction and Quantitative Trait Locus Analysis of Fruit- and Oil-Related Traits in Camellia oleifera Based on Double Digest Restriction Site-Associated DNA Sequencing.

Camellia oleifera, an important tree species and source of edible oil in China, has received significant attention owing to the oil's high unsaturated fatty acid content, which has benefits for human health. However, the mechanisms underlying C. oleifera yield and oil quality are largely unknown. In this study, 180 F1 progenies were obtained from two parents with obvious differences in fruit- and oil-related traits. We constructed a high-density genetic map using a double digest restriction site-associated DNA sequencing (ddRAD-Seq) strategy in C. oleifera. This map spanned 3327 cM and anchored 2780 markers in 15 linkage groups (LGs), with an average marker interval of 1.20 cM. A total of 221 quantitative trait loci (QTLs) associated with fruit- and oil-related traits were identified across three years' worth of phenotypic data. Nine QTLs were detected simultaneously in at least two different years, located on LG02, LG04, LG05, LG06, and LG11, and explained 8.5-16.6% of the phenotypic variation in the corresponding traits, respectively. Seventeen major QTLs were obtained that explained 13.0-16.6% of the phenotypic variance. Eleven and five flanking SNPs of major QTLs for fruit- and oil-related traits were detected which could be used for marker-assisted selection in C. oleifera breeding programs. Furthermore, 202 potential candidate genes in QTL regions were identified based on the collinearity of the genetic map and the C. oleifera "CON" genome. A potential regulatory network controlling fruit development and oil biosynthesis was constructed to dissect the complex mechanism of oil accumulation. The dissection of these QTLs will facilitate the gene cloning underlying lipid synthesis and increase our understanding in order to enhance C. oleifera oil yield and quality.

An energy-efficient solution to sludge drying and combustion process through Camellia oleifera shells amended foaming.

Foaming pretreatment has been proven effective in promoting sludge drying, however, the variation in sludge properties significantly influences the foaming efficiency. Inspired by foam stabilizer of solid particles, Camellia oleifera shells (COS) was screened out from various biomasses as an additive incorporated with the CaO for promoting the sludge foaming. For the introduction of COS, this study analyzed the drying behaviors of foamed sludge, quantified the surface cracks information, characterized the combustion performance, and evaluated the energy consumption. The results indicated that 46.72-50.10% of time could be saved in foaming the sludge to 0.70 g/mL by addition of 3.0 wt% COS. Compared with the original sludge (OS), the 0.70 g/mL foamed sludge saved 47.43% of time for sludge drying at 80 °C, and this value further increased to 53.14% with 3.0 wt% COS addition. Combining the multifractal spectra and drying kinetics analysis, the foaming promoted the formation of complex surface cracks in the warm-up period, while COS further improved the complexity of cracks in the constant rate period, and the shrinkage of isolated sludge blocks in the falling rate period, thus enhanced the moisture diffusion and heat transfer. Furthermore, the appropriate porous structure and additional volatile matters promoted the combustion performance. The 0.90 g/mL foamed sludge with COS presented the lowest activation energy of 180.362 kJ/mol in combustion. Overall, compared with OS, the 0.70 g/mL foamed sludge with COS saved 40.65% energy consumption during the foaming, drying and combustion processes, providing an energy-efficient solution for the sludge treatment and disposal.

Efficient Preparation of Biodiesel Using Sulfonated Camellia oleifera Shell Biochar as a Catalyst.

This study prepared sulfonated Camellia oleifera shell biochar using Camellia oleifera shell agricultural waste as a carbon source, and evaluated its performance as a catalyst for preparing biodiesel. The biochar obtained from carbonizing Camellia oleifera shells at 500 °C for 2 h serves as the carbon skeleton, and then the biochar is sulfonated with chlorosulfonic acid. The sulfonic acid groups are mainly grafted onto the surface of Camellia oleifera shell biochar through covalent bonding to obtain sulfonic acid type biochar catalysts. The catalysts were characterized by Scanning Electron Microscope (SEM), X-ray diffraction (XRD), Nitrogen adsorption-desorption Brunel-Emmett-Taylor Theory (BET), and Fourier-transform infrared spectroscopy (FT-IR). The acid density of the sulfonated Camellia oleifera fruit shell biochar catalyst is 2.86 mmol/g, and the specific surface area is 2.67 m2/g, indicating high catalytic activity. The optimal reaction conditions are 4 wt% catalyst with a 6:1 alcohol to oil ratio. After esterification at 70 °C for 2 h, the yield of biodiesel was 91.4%. Under the optimal reaction conditions, after four repeated uses of the catalyst, the yield of biodiesel still reached 90%. Therefore, sulfonated Camellia oleifera shell biochar is a low-cost, green, non-homogeneous catalyst with great potential for biodiesel production by esterification reaction in future development.

The Antioxidant, Anti-Inflammatory and Moisturizing Effects of Camellia oleifera Oil and Its Potential Applications.

Camellia oleifera oil (CO oil) extracted from C. oleifera seeds has a 2300-year consumption history in China. However, there is relatively little research regarding its non-edible uses. This study determined the physicochemical properties of CO oil extracted via direct pressing, identified its main components using GC-MS, and evaluated its antioxidant, moisturizing, and anti-inflammatory activities. The results revealed that CO oil's acid, peroxide, iodine, and saponification values were 1.06 ± 0.031 mg/g, 0.24 ± 0.01 g/100 g, 65.14 ± 8.22 g/100 g, and 180.41 ± 5.60 mg/g, respectively. CO oil's tocopherol, polyphenol, and squalene contents were 82.21 ± 9.07 mg/kg, 181.37 ± 3.76 mg/kg, and 53.39 ± 6.58 mg/kg, respectively; its unsaturated fatty acid (UFA) content was 87.44%, and its saturated fatty acid (SFA) content was 12.56%. CO oil also demonstrated excellent moisture retention properties, anti-inflammatory effects, and certain free radical scavenging. A highly stable CO oil emulsion with competent microbiological detection was developed using formulation optimization. Using CO oil in the emulsion significantly improved the formulation's antioxidant and moisturizing properties compared with those of the emulsion formulation that did not include CO oil. The prepared emulsion was not cytotoxic to cells and could reduce cells' NO content; therefore, it may have potential nutritional value in medicine and cosmetics.

Integrated analysis of miRNAs, transcriptome and phytohormones in the flowering time regulatory network of tea oil camellia.

Camellia oleifera is a crucial cash crop in the southern region of China. Timely flowering is a crucial characteristic for maximizing crop productivity. Nevertheless, the cold temperature and wet weather throughout the fall and winter seasons in South China impact the timing of flowering and the yield produced by C. oleifera. This study examined the miRNAs, transcriptomes, and phytohormones that are part of the flowering time regulatory networks in distinct varieties of C. oleifera (Sep, Oct, and Nov). This study provides evidence that phytohormones significantly impact the timing of flowering in C. oleifera leaves. There is a positive correlation between the accumulation variations of zeatin (cZ), brassinolide (BL), salicylic acid (SA), 1-amino cyclopropane carboxylic acid (ACC), and jasmonic acid (JA) and flowering time. This means that blooming occurs earlier when the quantity of these substances in leaves increases. Abscisic acid (ABA), trans-zeatin-riboside (tZR), dihydrozeatin (dh-Z), and IP (N6-Isopentenyladenine) exhibit contrasting effects. Furthermore, both miR156 and miR172 play a crucial function in regulating flowering time in C. oleifera leaves by modulating the expression of SOC1, primarily through the miR156-SPL and miR172-AP2 pathways. These findings establish a strong basis for future research endeavors focused on examining the molecular network associated with the flowering period of C. oleifera and controlling flowering time management through external treatments. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-024-01473-2.

Role of soil nutrient elements transport on Camellia oleifera yield under different soil types.

BACKGROUND: Most of Camellia oleifera forests have low fruit yield and poor oil quality that are largely associated with soil fertility. Soil physical and chemical properties interact with each other affecting soil fertility and C. oleifera growing under different soil conditions produced different yield and oil composition. Three main soil types were studied, and redundancy, correlation, and double-screening stepwise regression analysis were used for exploring the relationships between C. oleifera nutrients uptake and soil physical and chemical properties, shedding light on the transport law of nutrient elements from root, leaves, and kernel, and affecting the regulation of fruit yield and oil composition. RESULTS: In the present study, available soil elements content of C. oleifera forest were mainly regulated by water content, pH value, and total N, P and Fe contents. Seven elements (N, P, K, Mg, Cu, Mn and C) were key for kernel's growth and development, with N, P, K, Cu and Mn contents determining 74.0% the yield traits. The transport characteristics of these nutrients from root, leaves to the kernel had synergistic and antagonistic effects. Increasing oil production and unsaturated fatty acid content can be accomplished in two ways: one through increasing N, P, Mg, and Zn contents of leaves by applying corresponding N, P, Mg, Zn foliar fertilizers, while the other through maintaining proper soil moisture content by applying Zn fertilizer in the surface layer and Mg and Ca fertilizer in deep gully. CONCLUSION: Soil type controlled nutrient absorption by soil pH, water content and total N, P and Fe content. There were synergistic and antagonistic effects on the inter-organ transport of nutrient elements, ultimately affecting N, P, K, Cu and Mn contents in kernel, which determined the yield and oil composition of C. oleifera.

Ectopic expression of Camellia oleifera Abel. gibberellin 20-oxidase gene increased plant height and promoted secondary cell walls deposition in Arabidopsis.

MAIN CONCLUSION: Ectopic expression of Camellia oleifera Abel. gibberellin 20-oxidase 1 caused a taller phenotype, promoted secondary cell wall deposition, leaf enlargement, and early flowering, and reduced chlorophyll and anthocyanin accumulation and seed enlargement phenotype in Arabidopsis. Plant height and secondary cell wall (SCW) deposition are important plant traits. Gibberellins (GAs) play important roles in regulating plant height and SCWs deposition. Gibberellin 20-oxidase (GA20ox) is an important enzyme involved in GA biosynthesis. In the present study, we identified a GA synthesis gene in Camellia oleifera. The total length of the CoGA20ox1 gene sequence was 1146 bp, encoding 381 amino acids. Transgenic plants with CoGA20ox1 had a taller phenotype; a seed enlargement phenotype; promoted SCWs deposition, leaf enlargement, and early flowering; and reduced chlorophyll and anthocyanin accumulation. Genetic analysis showed that the mutant ga20ox1-3 Arabidopsis partially rescued the phenotype of CoGA20ox1 overexpression plants. The results showed that CoGA20ox1 participates in the growth and development of C. oleifera. The morphological changes in CoGA20ox1 overexpressed plants provide a theoretical basis for further exploration of GA biosynthesis and analysis of the molecular mechanism in C. oleifera.

First Report of Leaf Spot Blight of Camellia oleifera Caused by Diaporthe mahothocarpus in China.

Camellia oleifera Abel. is one of the native and important natural edible oil species in China. The cultivation of C. oleifera has vigorously increased in Guizhou Province in recent years. From June to August 2022, a severe leaf spot blight was observed on C. oleifera in Longli Plantation, with an incidence of 53.5% (n=200), which caused severe defoliation, negatively affected plant growth, and led to significant economic losses. Pale yellow and sub-circular leaf spots of 2-5 mm in diameter first appeared in the margin. The center of the spots then turned grey, and the edges turned brown. The symptomatic leaves gradually developed symptoms of blight with some brown acervular conidiomata, died, and fell off, with many deep black spots on the leaves (Fig. 1A-B). The fungal isolates GZU-Y2 and GZU-Y3 were obtained from the infected leaves of five-year-old symptomatic C. oleifera trees using the tissue isolation method, and a voucher specimen was deposited in the Forest Protection Laboratory, Guizhou University. Cultures grown on potato dextrose agar medium (PDA) were incubated at 28℃, 16L/8D. A round cream-like colony was formed on PDA, with a white surface, while the back gradually turned brown (Fig. 1C-E). The aerial hypha grew vigorously with an initial milky white color before turning grayish white. At 10 days after incubation, the pycnidia were dark brown to black and spherical, with a diameter of 563.3 µm (500 to 700) (n=20). The alpha conidia were unicellular, hyaline, aseptate, oval or fusiform and measured 6.1 µm (4.1 to 8.0) × 2.6 µm (1.9 to 3.6) (n=50). However, no beta conidia were observed (Fig. 1I). For further identification, total DNA from the pure culture was extracted using a DNA extraction kit (Sangon, Shanghai, China), and the internal transcribed spacer (ITS), translation elongation factor 1-α (TEF-1α), and beta-tubulin (TUB2) were amplified by PCR using the primers ITS1/ITS4 (White et al., 1990), EF1-728F/EF1-986R (Carbone and Kohn, 1999) and Bt2a/Bt2b (Glass and Donaldson, 1995), respectively, and sequenced for a BLASTn analysis and phylogenetic tree construction. The sequences of ITS, TEF-1α and TUB2 were deposited in GenBank as accession numbers OQ168242 (99.25%), OQ689451 (99.71%) and OQ689453 (100.00%) for GZU-Y2 and OQ674554 (99.25%), OQ689452 (99.71%) and OQ689454 (100.00%) for GZU-Y3, respectively. A phylogenetic tree (Fig. 2) was constructed with the software MEGA X using the Neighbor-Joining algorithm (Felsenstein, 1985). Based on its morphological and molecular characteristics, the pathogen was identified as Diaporthe mahothocarpus, one of the synonyms of D. eres and the teleomorph of Phomopsis mahothocarpi (Gao et al., 2014 and 2015; Chaisiri et al., 2021). A pathogenicity test was conducted by spraying spore suspensions (2 × 107 spores/mL) of isolate GZU-Y2 on the leaves of 20 pots of annual C. oleifera seedlings in vivo. The same number of control seedings were sprayed with sterile water. The seedlings were placed at a constant room temperature of 28°C, with the inoculation points wrapped in Parafilm for 5 d to retain moisture. After 10 d, typical symptoms appeared on the inoculated leaves (Fig. 1F-H), and the re-isolated fungal culture was identical in morphology and ITS sequence to that originally obtained, fulfilling Koch's postulates. To our knowledge, this is the first report of D. mahothocarpus causing leaf spot blight of C. oleifera in China. In our future work, we tend to study the green prevention and control of this disease.

High-Quality Genome Assembly and Annotation Resource of Elsinoë annonae, Causing Fruit Scab on Camellia oleifera.

Elsinoë annonae is a fungal pathogen that causes fruit scab disease in the edible-oil (tea oil) plant (Camellia oleifera Abel). The absence of genome resources for this fungus hampers functional genetic studies of the pathogenesis mechanism of E. annonae. This study reports the genome assembly of E. annonae strain SM-YC-2 collected from tea oil tree fruit with scab disease in Fujian Province, China. Combining 16.44 Gb of PacBio Sequel II long reads and 5.13 Gb of Illumina NovaSeq reads, we generated a 25.93-Mb (99.19% of expected genome size) high-quality genome assembly with 52.66% GC content, 5.05% repeats, and over 98% Benchmarking Universal Single-Copy Orthologs completeness for E. annonae strain SM-YC-2. These high-quality genome assembly resources will facilitate functional genomic characterization studies, enhance insights into the pathogenicity mechanism of E. annonae, and support the development of molecular-based control strategies.

Total Flavonoids from Camellia oleifera Alleviated Mycoplasma pneumoniae-Induced Lung Injury via Inhibition of the TLR2-Mediated NF-κB and MAPK Pathways.

Mycoplasma pneumoniae (M. pneumoniae) is an atypical bacterial pathogen responsible for community-acquired pneumonia primarily among school-aged children and young adults. Camellia oleifera (C. oleifera) has been used as a medicinal and edible plant in China for centuries, the constituents from which possessed various bioactivities. Notably, flavonoids existing in residues of C. oleifera defatted seeds exhibited significant anti-inflammatory activities. In the present study, we investigated the impact of total flavonoids from C. oleifera (TFCO) seed extract on M. pneumoniae pneumonia. TFCO was obtained using multiple column chromatography methods and identified as kaempferol glycosides via UPLC-HRESIMS. In a M. pneumoniae pneumonia mouse model, TFCO significantly reduced the lung damage, suppressed IL-1ß, IL-6, and TNF-α production, and curbed TLR2 activation triggered by M. pneumoniae. Similarly, in RAW264.7 macrophage cells stimulated by lipid-associated membrane proteins (LAMPs), TFCO suppressed the generation of proinflammatory cytokines and TLR2 expression. Moreover, TFCO diminished the phosphorylation of IκBα, JNK, ERK, p38, and p65 nuclear translocation in vitro. In conclusion, TFCO alleviated M. pneumoniae-induced lung damage via inhibition of TLR2-mediated NF-κB and MAPK pathways, suggesting its potential therapeutic application in M. pneumoniae-triggered lung inflammation.

Rapid Screening of Phenolic Compounds with Anti-Enteritis Activity from Camellia oleifera Oil Using a Smurf Drosophila Model and Molecular Docking Methods.

Screening and identifying the active compounds in foods are important for the development and utilization of functional foods. In this study, the anti-enteritis activity of ethanol extract from Camellia oleifera oil (PECS) was quickly evaluated using a Smurf Drosophila model and the metabolomics approach, combined with molecular docking techniques, were performed to rapidly screen and identify compounds with potential anti-enteritis activity in PECS. PECS showed good anti-enteritis activity and inhibited the activity of 5-lipoxygenase (LOX), cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS). In particular, wighteone and p-octopamine were newly identified in C. oleifera oil and were proven to have good anti-enteritis activity. The inhibitory activity of kaempferitrin (IC50 = 0.365 mmol L-1) was higher than that of wighteone (IC50 = 0.424 mmol L-1) and p-octopamine (IC50 = 0.402 mmol L-1). Of note, the IC50 value of salazosulfapyridine was 0.810 mmol L-1. Inhibition of LOX activity is likely one of the anti-enteritis mechanisms of PECS. These new findings lay the foundation for further investigations into the underlying mechanisms of anti-enteritis activity in C. oleifera oil.

Quantitative Analysis of Camellia oleifera Seed Saponins and Aqueous Two-Phase Extraction and Separation.

At present, the technology used for the extraction and purification of Camellia oleifera saponins generally has the problems of high cost and low purity, and the quantitative detection of Camellia oleifera saponins also has the problems of low sensitivity and easy interference from impurities. To solve these problems, this paper aimed to use liquid chromatography for the quantitative detection of Camellia oleifera saponins, and to adjust and optimize the related conditions. In our study, the average recovery of Camellia oleifera saponins obtained was 100.42%. The RSD of precision test was 0.41%. The RSD of the repeatability test was 0.22%. The detection limit of the liquid chromatography was 0.06 mg/L, and the quantification limit was 0.2 mg/L. In order to improve the yield and purity, the Camellia oleifera saponins were extracted from Camellia oleifera Abel. seed meal by methanol extraction. Then, the extracted Camellia oleifera saponins were extracted with an ammonium sulfate/propanol aqueous two-phase system. We optimized the purification process of formaldehyde extraction and aqueous two-phase extraction. Under the optimal purification process, the purity of Camellia oleifera saponins extracted by methanol was 36.15%, and the yield was 25.24%. The purity of Camellia oleifera saponins obtained by aqueous two-phase extraction was 83.72%. Thus, this study can provide a reference standard for rapid and efficient detection and analysis of Camellia oleifera saponins for industrial extraction and purification.