Mulberry (Morus alba L.) is a flowering tree traditionally used in Chinese herbal medicine. Mulberry leaf flavonoids (MLFs) have been reported to exert important anti-inflammatory and antioxidant properties. The purpose of this study was to select the MLF with the best anti-inflammatory and antioxidative activities from MLFs eluted by different ethanol concentrations (30%, 50%, and 75%) and explore its pharmacological properties. Three types of MLFs inhibited the production of nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and inflammatory cytokines in lipopolysaccharide (LPS)-induced RAW 264.7 cells. All MLFs boosted the antioxidative capacity by decreasing the reactive oxygen species (ROS) production and the scavenging of 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals and improving the metal ion chelating activity and reducing power. The results revealed that the MLFs eluted by 30% ethanol exhibited the best anti-inflammatory and antioxidative activities. A nontargeted metabolomic analysis was used to analyze 24 types of differential flavonoids between the MLFs. Quercetin, kaempferol, and their derivatives in 30%MLF were more abundant than the other two MLFs. Furthermore, we evaluated the pharmacological activities of 30%MLF in dextran sodium sulfate (DSS)-induced ulcerative colitis (UC) mice. The 30%MLF could alleviate the clinical symptoms, reduce the secretion of inflammatory cytokines, and inhibit the activation of the inflammatory pathway in DSS-induced colitis mice. This study will provide valuable information for the development of MLFs eluted by 30% ethanol as a functional food.

Mulberry, an important woody tree, has strong tolerance to environmental stresses, including salinity, drought, and heavy metal stress. However, the current research on mulberry resistance focuses mainly on the selection of resistant resources and the determination of physiological indicators. In order to clarify the molecular mechanism of salt tolerance in mulberry, the physiological changes and proteomic profiles were comprehensively analyzed in salt-tolerant (Jisang3) and salt-sensitive (Guisangyou12) mulberry varieties. After salt treatment, the malondialdehyde (MDA) content and proline content were significantly increased compared to control, and the MDA and proline content in G12 was significantly lower than in Jisang3 under salt stress. The calcium content was significantly reduced in the salt-sensitive mulberry varieties Guisangyou12 (G12), while sodium content was significantly increased in both mulberry varieties. Although the Jisang3 is salt-tolerant, salt stress caused more reductions of photosynthetic rate in Jisang3 than Guisangyou12. Using tandem mass tags (TMT)-based proteomics, the changes of mulberry proteome levels were analyzed in salt-tolerant and salt-sensitive mulberry varieties under salt stress. Combined with GO and KEGG databases, the differentially expressed proteins were significantly enriched in the GO terms of amino acid transport and metabolism and posttranslational modification, protein turnover up-classified in Guisangyou12 while down-classified in Jisang3. Through the comparison of proteomic level, we identified the phenylpropanoid biosynthesis may play an important role in salt tolerance of mulberry. We clarified the molecular mechanism of mulberry salt tolerance, which is of great significance for the selection of excellent candidate genes for saline-alkali soil management and mulberry stress resistance genetic engineering.

Phyllosphere and rhizosphere are unique and wide‐ranging habitats that harbor various microbial communities, which influence plant growth and health, and the productivity of the ecosystems. In this study, we characterized the shared microbiome of the phyllosphere and rhizosphere among three plants (Ipomoea pes‐caprae, Wedelia chinensis, and Cocos nucifera), to obtain an insight into the relationships between bacteria (including diazotrophic bacteria) and fungi, present on these host plants. Quantitative PCR showed that the abundances of the microbiome in the soil samples were significantly higher than those in the phyllosphere samples, though there was an extremely low abundance of fungi in bulk soil. High‐throughput sequencing showed that the alpha‐diversity of bacteria and fungi was higher in the rhizosphere than the phyllosphere samples associated with the same plant, while there was no obvious shift in the alpha‐diversity of diazotrophic communities between all the tested phyllosphere and soil samples. Results of the microbial composition showed that sample‐specific bacteria and fungi were found among the phyllosphere and rhizosphere of the different host plants. About 10%–27% of bacteria, including diazotrophs, and fungi overlapped between the phyllosphere and the rhizosphere of these host plants. No significant difference in microbial community structure was found among the tested rhizosphere samples, and soil properties had a higher influence on the soil microbial community structures than the host plant species.

Mulberry sclerotiniose caused by Ciboria shiraiana is a devastating disease of mulberry (Morus alba L.) fruit in Northwest China. At present, no disease-resistant varieties are used in production, as the molecular mechanisms of this disease are not well understood. In this study, to explore new prevention methods and provide direction for molecular breeding, transcriptomic sequencing and un-targeted metabolomics were performed on healthy (CK), early-stage diseased (HB1), and middle-stage diseased (HB2) mulberry fruits. Functional annotation, gene ontology, a Kyoto encyclopedia of genes and genomes (KEGG) analysis, and a Mapman analysis of the differentially expressed genes revealed differential regulation of genes related to plant hormone signal transduction, transcription factors, and phenylpropanoid biosynthesis. A correspondence between the transcript pattern and metabolite profile was observed in the phenylpropanoid biosynthesis pathway. It should be noted that the log2 ratio of eugenol (isoeugenol) in HB1 and HB2 are 85 times and 23 times higher than CK, respectively. Our study shows that phenylpropanoid biosynthesis may play an essential role in response to sclerotiniose pathogen infection and eugenol(isoeugenol) enrichment in mulberry fruit, which may provide a novel method for mulberry sclerotiniose control.

Mulberry leaves (ML) are a promising alternative fodder source due to their high protein content and the abundance of active components. A test of three inoculants in various combinations revealed that high-quality ML silage was produced at an inoculum ratio of 1:1:0 (50% Saccharomyces cerevisiae, 50% Lactobacillus plantarum, and 0% Bacillus subtilis). Using dry matter (DM) loss, pH, ammonia-N and amino acid contents, total antioxidant activity, and total flavonoids content to evaluate silage quality, this inoculant mixture was shown to produce high-quality silage within a range of inoculum size (5–15%), moisture contents (50–67%), ensiling temperatures (27–30°C), and ensiling duration (14–30 days). A third trial comparing silages produced after 30 days at 28°C and 50% moisture content revealed that silage E, prepared using an L. plantarum inoculant alone, displayed the lowest DM loss and pH, and low bacterial diversity, and it was dominated by Lactobacillus (88.6%), with low abundance of Enterobacter (6.17%). In contrast, silage B5, prepared with equal ratios of L. plantarum and S. cerevisiae, was dominated by Enterococcus (67.16%) and Lactobacillus (26.94%), with less marked yeast persistence, and reducing the DM content from 50 to 40% altered these relative abundances to 5.47 and 60.61, respectively. Control silages produced without an inoculant had the highest pH and ammonia-N content (indicative of poor quality), had the lowest antioxidant activity, had higher bacterial diversity, and were dominated by Carnobacterium (74.28%) and Enterococcus (17.3%). In summary, ensiling of ML conditions with proper inoculants yielded high-quality silage with a favorable microbial community composition.

The improvement of a plant's tolerance to drought is a major endeavor in agriculture. Polyploid plants often exhibit enhanced stress tolerance relative to their diploid progenitor, but the matching stress tolerance is still little understood. Own-rooted stem cuttings of mulberry (Morus alba L.) cultivar Shinichinose (2n = 2x = 28) and Shaansang-305 (2n = 3x = 42) were used in this study, of which the latter (triploid) has more production and application purposes. The responses of triploid Shaansang-305 and diploid progenitor ShinIchinose under drought stress were compared through an investigation of their physiological traits, RNA-seq, and secondary metabolome analysis. The results showed that the triploid exhibited an augmented abscisic acid (ABA) content and a better stress tolerance phenotype under severe drought stress. Further, in the triploid plant some genes (TSPO, NCED3, and LOC21398866) and ATG gene related to ABA signaling showed significantly upregulated expression. Interestingly, the triploid accumulated higher levels of RWC and SOD activity, as well as more wax on the leaf surface, but with less reductive flavonoid than in diploid. Our results suggest triploid plants may better adapt to with drought events. Furthermore, the flavonoid metabolism involved in drought resistance identified here may be of great value to medicinal usage of mulberry. The findings presented here could have substantial implications for future studies of crop breeding.