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Influenza is an acute respiratory disorder caused by the influenza virus and is associated with prolonged hospitalization and high mortality rates in older individuals and chronically ill patients. Vaccination is the most effective preventive strategy for ameliorating seasonal influenza. However, the vaccine is not fully effective in cases of antigenic mismatch with the viral strains circulating in the community. The emergence of resistance to antiviral drugs aggravates the situation. Therefore, developing new vaccines and antiviral drugs is essential. Castanea crenata honey (CH) is an extensively cultivated food worldwide and has been used as a nutritional supplement or herbal medicine. However, the potential anti-influenza properties of CH remain unexplored. In this study, the in vitro and in vivo antiviral effects of CH were assessed. CH significantly prevented influenza virus infection in mouse Raw264.7 macrophages. CH pretreatment inhibited the expression of the viral proteins M2, PA, and PB1 and enhanced the secretion of proinflammatory cytokines and type-I interferon (IFN)-related proteins in vitro . CH increased the expression of RIG-1, mitochondrial antiviral signaling (MAVS) protein, and IFN-inducible transmembrane protein, which interferes with virus replication. CH reduced body weight loss by 20.9%, increased survival by 60%, and decreased viral replication and inflammatory response in the lungs of influenza A virus-infected mice. Therefore, CH stimulates an antiviral response in murine macrophages and mice by preventing viral infection through the RIG-1-mediated MAVS pathway. Further investigation is warranted to understand the molecular mechanisms involved in the protective effects of CH on influenza virus infection.

Ectomycorrhizal (ECM) fungal community of the European chestnut has been poorly investigated, and mostly by sporocarp sampling. We proposed the study of the ECM fungal community of 2-year-old chestnut hybrids Castanea × coudercii (Castanea sativa × Castanea crenata) using molecular approaches. By using the chestnut hybrid clones 111 and 125, we assessed the impact of grafting on ECM colonization rate, species diversity, and fungal community composition. The clone type did not have an impact on the studied variables; however, grafting significantly influenced ECM colonization rate in clone 111. Species diversity and richness did not vary between the experimental groups. Grafted and ungrafted plants of clone 111 had a different ECM fungal species composition. Sequence data from ITS regions of rDNA revealed the presence of 9 orders, 15 families, 19 genera, and 27 species of ECM fungi, most of them generalist, early-stage species. Thirteen new taxa were described in association with chestnuts. The basidiomycetes Agaricales (13 taxa) and Boletales (11 taxa) represented 36% and 31%, of the total sampled ECM fungal taxa, respectively. Scleroderma citrinum, S. areolatum, and S. polyrhizum (Boletales) were found in 86% of the trees and represented 39% of total ECM root tips. The ascomycete Cenococcum geophilum (Mytilinidiales) was found in 80% of the trees but accounted only for 6% of the colonized root tips. These results could help to unveil the impact of grafting on fungal symbionts, improving management of chestnut agro-ecosystems and production of edible fungal species.

This study isolated, determined, and quantified plant growth inhibitors in Japanese chestnut (Castanea crenata Sieb. et Zucc), a deciduous species native to Japan and Korea. In laboratory assays, C. crenata leaves showed strong inhibition on germination and seedling growth of Echinochloa crus-galli (barnyardgrass), Lactuca sativa (lettuce), and Raphanus sativus (radish). Laboratory and greenhouse trials showed that leaves of C. crenata appeared as a promising material to manage weeds, especially the dicot weeds. By GC-MS and HPLC analyses, gallic, protocatechuic, p-hydroxybenzoic, caffeic, ferulic, ellagic, and cinnamic acids were identified and quantified, of which ellagic acid was present in the highest quantity (2.36 mg/g dried leaves). By column chromatography and spectral data (1H- and 13C-NMR, IR, and LC-MS) analysis, a compound identified as 2α,3β,7β,23-tetrahydroxyurs-12-ene-28-oic acid (1) was purified from the methanolic leaf extract of C. crenata (0.93 mg/g dried leaves). This constituent showed potent inhibition on growth of E. crus-galli, a problematic weed in agricultural practice. The inhibition of the compound 1 (IC50 = 2.62 and 0.41 mM) was >5 fold greater than that of p-hydroxybenzoic acid (IC50 = 15.33 and 2.11 mM) on shoot and root growth of E. crus-galli, respectively. Results suggest that the isolated the compound 1 has potential to develop natural herbicides to manage E. crus-galli. This study is the first to isolate and identify 2α,3β,7β,23-tetrahydroxyurs-12-ene-28-oic acid in a plant and report its plant growth inhibitory potential.

In this study, different plant parts (barks, flowers, inner skins, kernels and leaves) of Castanea crenata (Japanese chestnut) were analyzed for total phenolic, flavonoid, and tannin contents. Antioxidant properties were evaluated by using 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), reducing power, and β-carotene bleaching methods. The highest total phenolic and tannin contents were found in the inner skins (1034 ± 7.21 mg gallic acid equivalent/g extract and 253.89 ± 5.59 mg catechin equivalent/g extract, respectively). The maximum total flavonoid content was observed in the flowers (147.41 ± 1.61 mg rutin equivalent/g extract). The inner skins showed the strongest antioxidant activities in all evaluated assays. Thirteen phenolic acids and eight flavonoids were detected and quantified for the first time. Major phenolic acids were gallic, ellagic, sinapic, and p-coumaric acids, while the principal flavonoids were myricetin and isoquercitrin. The inner skin extract was further fractionated by column chromatography to yield four fractions, of which fraction F3 exhibited the most remarkable DPPH scavenging capacity. These results suggest that C. crenata provides promising antioxidant capacities, and is a potential natural preservative agent in food and pharmaceutical industries.

The inner shell of the chestnut (Castanea crenata) has long been used in Asia as a medicinal herb for improving digestion and blood circulation, and treating diarrhea. However, most chestnut shells are now treated as waste materials in industrial peeling processes. In this study, we examined the metabolite variation among major cultivars of C. crenata shells using mass spectrometry. Among five representative cultivars, Okkwang, Porotan, and Ishizuuchi had higher levels of bioactive compounds, such as ellagic acid derivatives, ellagitannins, flavonoids, and gallic acid derivatives. Their antioxidant capacity was positively correlated with their chemical composition. The byproducts (whole shells) from the industrial peeling process were re-evaluated in comparison with the inner shell, a rich source of phenolic compounds. The phenolic acids and flavonoid glucoside derivatives were significantly higher in the whole shells, whereas the levels of flavonoids were higher in the inner shells. In addition, the whole shell extracts significantly reduced cellular reactive oxygen species production compared to the inner shell extracts. This study demonstrated the different biochemical benefits of different C. crenata cultivars through metabolic profiling and suggests that the whole shell could be used as a functional ingredient, as it has the highest levels of bioactive products and antioxidant effects.

Chestnuts are an important food crop commonly used as a food ingredient due to their nutritional properties and potential health benefits. In Korea, chestnuts have been crossbred to develop cultivars with insect resistance and high productivity, producing multiple chestnut varieties. This study classified 17 Castanea crenata cultivars produced in Korea according to origin and harvest time and determined the metabolites in chestnut kernels using 1H nuclear magnetic resonance spectroscopy. The 17 C. crenata cultivars were divided into four groups based on their geographic origin: Korean native, Korean hybrid, Japanese native, and Japanese hybrid. The cultivars were also divided into three groups depending on their harvest period: early-ripening cultivar, mid-ripening cultivar, and late-ripening cultivar. The partial least squares-discriminant analysis score plot revealed differences among the groups. Identified metabolites, including amino acids, organic acids, and sugars, contributed to discriminating the origin and harvest time of the C. crenata chestnut kernels. Significant differences were observed, mainly in amino acids, which suggests that the composition of amino acids is one factor influenced by both the origin and harvest time of C. crenata. These results are useful to both growers and breeders because they identify the nutritional and metabolic characteristics of each C. crenata cultivar.

Castanea crenata (Fagaceae) is a species of chestnut tree that is endemic to the Republic of Korea and Japan. While its kernels are consumed, chestnut by-products such as shells and burs, which account for 10–15% of the total weight, are discarded as waste. Phytochemical and biological studies have been carried out to eliminate this waste and develop high-value products from its by-products. In this study, five new compounds (1–2, 6–8) along with seven known compounds were isolated from the shell of C. crenata. This is the first study to report diterpenes from the shell of C. crenata. Comprehensive spectroscopic data including 1D, 2D NMR, and CD spectroscopy were used to determine the compound structures. All isolated compounds were examined for their ability to stimulate dermal papilla cell proliferation using a CCK-8 assay. In particular, 6β,7β,16α,17-Tetrahydroxy-ent-kauranoic acid, isopentyl-α-L-arabinofuranosyl-(1→6)-β-D-glucopyranoside, and ellagic acid exhibited the most potent proliferation activity of all.

Aiming the selection of rootstocks to produce seedlings, the objective of this study was to quantify the seminiferous propagation in chestnut seeds, with or without incisions. Nine cultivars were used (‘Ibuki’, ‘Isumo’, ‘Kinshu’, ‘Moriwase’, ‘Okuni’, ‘Senri’, ‘Taishowase’, ‘Tamatsukuri’ and ‘Tiodowase’) and two selections (‘KM-1’ and ‘KM-2’) of hybrid chestnut trees (Castanea crenata x Castanea sp.). An incision was held in half of chestnuts and the other half remained intact. Percentage of germination, emergence and root length were quantified. The germinated nuts were transplanted and were quantified the diameter and height of seedlings. Then, the chestnut selections ‘KM-2’ and ‘Jacutinga’ were grafted by the cleft grafting method, and were quantified the percentage of sprouting, besides the diameter and height of grafts. Incisions increase the germination and seedling emergence in most cultivars and selections. ‘Taishowase’ and ‘Tamatsukuri’ stood out over the others to germination, growth of the rootstocks. RESUMO: Visando a seleção de porta-enxertos para a produção de mudas, o objetivo do trabalho foi quantificar a propagação seminífera em castanhas com ou sem a realização de incisões. Foram utilizadas nove cultivares (‘Ibuki’, ‘Isumo’, ‘Kinshu’, ‘Moriwase’, ‘Okuni’, ‘Senri’, ‘Taishowase’, ‘Tamatsukuri’ e ‘Tiodowase’) e duas seleções (‘KM-1’ e ‘KM-2’) de castanheira híbridas (Castanea crenata x Castanea sp.). Em metade das castanhas, realizou-se uma incisão e a outra metade das castanhas permaneceu intacta. Foram quantificadas a porcentagem de germinação, emergência e o comprimento das raízes. As castanhas germinadas foram transplantadas onde se quantificou o diâmetro e a altura das mudas e ao final, a massa seca da parte aérea. Em seguida foram enxertadas pelo método de garfagem as seleções de castanheira ‘KM-2’ e ‘Jacutinga’, se quantificando a porcentagem de brotação, além do diâmetro e altura dos enxertos. Concluiu-se que as incisões aumentam a germinação e a emergência das plântulas, na maioria das cultivares e seleções. ‘Taishowase’ e ‘Tamatsukuri’ se destacam em relação as demais quanto a germinação e crescimento dos porta-enxertos.

The complete chloroplast genome sequence of Castanea crenata was sequenced and assembled using PacBio Sequel data. The cpDNA was 160,787 bp in length, containing a pair of inverted repeats (IRs) of 25,654 bp each separated by a large and small single copy (LSC and SSC) regions of 90,645 bp and 18,836 bp, respectively. The cpDNA contained 102 genes, including 65 protein-coding genes, 8 ribosomal RNA genes and 37 transfer RNA genes. Phylogenetic analysis indicated that C. crenata was closest to C. pumila var. pumila, which is known as a typical variety of American chinquapin or dwarf chestnut.

Background MYBs are among the largest transcription factor families in plants. Consequently, members of this family are involved in a plethora of processes including development and specialized metabolism. The MYB families of many plant species were investigated in the last two decades since the first investigation looked at Arabidopsis thaliana . This body of knowledge and characterized sequences provide the basis for the identification, classification, and functional annotation of candidate sequences in new genome and transcriptome assemblies. Results A pipeline for the automatic identification and functional annotation of MYBs in a given sequence data set was implemented in Python. MYB candidates are identified, screened for the presence of a MYB domain and other motifs, and finally placed in a phylogenetic context with well characterized sequences. In addition to technical benchmarking based on existing annotation, the transcriptome assembly of Croton tiglium and the annotated genome sequence of Castanea crenata were screened for MYBs. Results of both analyses are presented in this study to illustrate the potential of this application. The analysis of one species takes only a few minutes depending on the number of predicted sequences and the size of the MYB gene family. This pipeline, the required bait sequences, and reference sequences for a classification are freely available on github: https://github.com/bpucker/MYB_annotator . Conclusions This automatic annotation of the MYB gene family in novel assemblies makes genome-wide investigations consistent and paves the way for comparative studies in the future. Candidate genes for in-depth analyses are presented based on their orthology to previously characterized sequences which allows the functional annotation of the newly identified MYBs with high confidence. The identification of orthologs can also be harnessed to detect duplication and deletion events.