Characteristics of the phyllosphere microbial community and its relationship with major aroma precursors during the tobacco maturation process.

Numerous bacteria, fungi and other microorganisms in the tobacco phyllosphere interstellar area participate in the physiological metabolism of plants by interacting with the host. However, there is currently little research on the characteristics of tobacco phyllosphere microbial communities, and the correlation between tobacco phyllosphere microbial communities and phyllosphere factor indicators is still unknown. Therefore, high-throughput sequencing technology based on the 16S rRNA/ITS1 gene was used to explore the diversity and composition characteristics of tobacco phyllosphere bacterial and fungal communities from different maturation processes, and to identify marker genera that distinguish phyllosphere microbial communities. In this study, the correlations between tobacco phyllosphere bacterial and fungal communities and the precursors of major aroma compounds were explored. The results showed that as the tobacco plants matured, the density of glandular trichomes on the tobacco leaves gradually decreased. The surface physicochemical properties of tobacco leaves also undergo significant changes. In addition, the overall bacterial alpha diversity in the tobacco phyllosphere area increased with maturation, while the overall fungal alpha diversity decreased. The beta diversity of bacteria and fungi in the tobacco phyllosphere area also showed significant differences. Specifically, with later top pruning time, the relative abundances of Acidisoma, Ralstonia, Bradyrhizobium, Alternaria and Talaromyces gradually increased, while the relative abundances of Pseudomonas, Filobassidium, and Tausonia gradually decreased. In the bacterial community, Acidisoma, Ralstonia, Bradyrhizobium, and Alternaria were significantly positively correlated with tobacco aroma precursors, with significant negative correlations with tobacco phyllosphere trichome morphology, while Pseudomonas showed the opposite pattern; In the fungal community, Filobasidium and Tausonia were significantly negatively correlated with tobacco aroma precursors, and significantly positively correlated with tobacco phyllosphere trichome morphology, while Alternaria showed the opposite pattern. In conclusion, the microbiota (bacteria and fungi) and aroma precursors of the tobacco phyllosphere change significantly as tobacco matures. The presence of Acidisoma, Ralstonia, Bradyrhizobium and Alternaria in the phyllosphere microbiota of tobacco may be related to the aroma precursors of tobacco.

In vitro study on antioxidant and lipid-lowering activities of tobacco polysaccharides.

Tobacco polysaccharides were extracted by hot water extraction, and purified and separated using DEAE-52 cellulose chromatography columns, and three purified polysaccharide fractions, YCT-1, YCT-2, and YCT-3, were finally obtained. The physicochemical properties of the three fractions were analyzed by ultraviolet spectroscopy, high-performance liquid chromatography and high-performance gel chromatography. The in vitro antioxidant activity of tobacco polysaccharides was compared among different fractions by using DPPH radical, hydroxyl radical scavenging assay and potassium ferricyanide method. The in vitro hypoglycemic activity was compared using α-amylase and α-glucosidase activity inhibition assay. And the in vitro hypolipidemic activity were investigated by using pancreatic lipase activity inhibition assay and HepG-2 intracellular lipid accumulation assay. All the results showed that the constituent monosaccharides of the three tobacco polysaccharide fractions were similar, but the molar percentages of each monosaccharide were different. The average molecular weights of the three components were 27,727 Da, 27,587 Da, and 66,517 Da, respectively, and the scavenging activities on DPPH radicals and hydroxyl radicals were at a high level with good quantitative-effect relationships. The reducing power were much lower than that of the positive control VC, and the three polysaccharide fractions had a weak inhibitory ability on α-amylase activity, but showed excellent inhibitory ability on α-glucosidase and pancreatic lipase activity. In addition, the results of cellular experiments showed that all three fractions were able to inhibit lipid over-accumulation in HepG-2 cells by increasing the mRNA expression levels of PPAR-α, CPT-1A, and CYP7A1 genes, and the tobacco polysaccharide YCT-3 showed the best effect. The mechanism by which YCT-3 ameliorated the over-accumulation of intracellular lipids in HepG-2 cells was found to be related to its influence on the expression of miR-155-3p and miR-17-3p in the exosomes of HepG-2 cells.

Synergistic Reduction of Arsenic Uptake and Alleviation of Leaf Arsenic Toxicity in Maize (Zea mays L.) by Arbuscular Mycorrhizal Fungi (AMF) and Exogenous Iron through Antioxidant Activity.

Arbuscular mycorrhizal fungi (AMF) play key roles in enhancing plant tolerance to heavy metals, and iron (Fe) compounds can reduce the bioavailability of arsenic (As) in soil, thereby alleviating As toxicity. However, there have been limited studies of the synergistic antioxidant mechanisms of AMF (Funneliformis mosseae) and Fe compounds in the alleviation of As toxicity on leaves of maize (Zea mays L.) with low and moderate As contamination. In this study, a pot experiment was conducted with different concentrations of As (0, 25, 50 mgꞏkg-1) and Fe (0, 50 mgꞏkg-1) and AMF treatments. Results showed that under low and moderate As concentrations (As25 and As50), the co-inoculation of AMF and Fe compound significantly increased the biomass of maize stems and roots, phosphorus (P) concentration, and P-to-As uptake ratio. Moreover, the co-inoculation of AMF and Fe compound addition significantly reduced the As concentration in stem and root, malondialdehyde (MDA) content in leaf, and soluble protein and non-protein thiol (NPT) contents in leaf of maize under As25 and As50 treatments. In addition, co-inoculation with AMF and Fe compound addition significantly increased the activities of catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) in the leaves of maize under As25 treatment. Correlation analysis showed that stem biomass and leaf MDA content were very significantly negatively correlated with stem As content, respectively. In conclusion, the results indicated that the co-inoculation of AMF and Fe compound addition can inhibit As uptake and promote P uptake by maize under low and moderate As contamination, thereby mitigating the lipid peroxidation on maize leaves and reducing As toxicity by enhancing the activities of antioxidant enzymes under low As contamination. These findings provide a theoretical basis for the application of AMF and Fe compounds in the restoration of cropland soil contaminated with low and moderate As.

Differences in soil physicochemical properties and rhizosphere microbial communities of flue-cured tobacco at different transplantation stages and locations.

Rhizosphere microbiota play an important role in regulating soil physical and chemical properties and improving crop production performance. This study analyzed the relationship between the diversity of rhizosphere microbiota and the yield and quality of flue-cured tobacco at different transplant times (D30 group, D60 group and D90 group) and in different regions [Linxiang Boshang (BS) and Linxiang ZhangDuo (ZD)] by high-throughput sequencing technology. The results showed that there were significant differences in the physicochemical properties and rhizosphere microbiota of flue-cured tobacco rhizosphere soil at different transplanting times, and that the relative abundance of Bacillus in the rhizosphere microbiota of the D60 group was significantly increased. RDA and Pearson correlation analysis showed that Bacillus, Streptomyces and Sphingomonas were significantly correlated with soil physical and chemical properties. PIGRUSt2 function prediction results showed that compared with the D30 group, the D60 group had significantly increased metabolic pathways such as the superpathway of pyrimidine deoxyribonucleoside salvage, allantoin degradation to glyoxylate III and pyrimidine deoxyribonucleotides de novo biosynthesis III metabolic pathways. The D90 group had significantly increased metabolic pathways such as ubiquitol-8 biosynthesis (prokaryotic), ubiquitol-7 biosynthesis (prokaryotic) and ubiquitol-10 biosynthesis (prokaryotic) compared with the D60 group. In addition, the yield and quality of flue-cured tobacco in the BS region were significantly higher than those in the ZD region, and the relative abundance of Firmicutes and Bacillus in the rhizosphere microbiota of flue-cured tobacco in the BS region at the D60 transplant stage was significantly higher than that in the ZD region. In addition, the results of the hierarchical sample metabolic pathway abundance map showed that the PWY-6572 metabolic pathway was mainly realized by Paenibacillus, and that the relative abundance of flue-cured tobacco rhizosphere microbiota (Paenibacillus) participating in PWY-6572 in the D60 transplant period in the BS region was significantly higher than that in the ZD region. In conclusion, different transplanting periods of flue-cured tobacco have important effects on soil physical and chemical properties and rhizosphere microbial communities. There were significant differences in the rhizosphere microbiota and function of flue-cured tobacco in different regions, which may affect the performance and quality of this type of tobacco.

Induction mechanism of cigarette smoke components (CSCs) on dyslipidemia and hepatic steatosis in rats.

OBJECTIVE: The purpose of this study was to explore the effect of cigarette smoke component (CSC) exposure on serum lipid levels in rats and the underlying molecular mechanism. METHODS: Male SPF-grade SD rats were randomly divided into a control group and a CSC exposure group, with the CSC group being exposed to CSC for 6 weeks. RT-PCR and Western blotting methods were used to detect lipid metabolism gene expression in rats, and 16S RNA gene sequencing was used to detect the gut microbiota in the rat cecum. Rat serum exosomes were prepared and identified, and the interaction of exosomal miR-291a-3p and miR-126a-5p with AMPK and CYP7A1 was detected by a dual luciferase reporter gene assay (DLRG). RESULTS: Serum indicators, including cholesterol levels and trimethylamine oxide (TMAO) content, were significantly affected in the CSC exposure group compared with the control group (P < 0.05), and the expression levels of adenylate-activated protein kinase (AMPK), acetyl-coenzyme A carboxylase (ACC) and HMG-CoA reductase (HMG-CoAR) genes were significantly increased (P < 0.05) in the liver, while the expression level of cholesterol 7α-hydroxylase (CYP7A1) was markedly decreased (P < 0.01). 16S rRNA gene sequencing of the gut microbiota in the rat cecum showed that the abundance of Firmicutes in the CSC group increased significantly at the phylum level, while the abundances of Bacteroidota and Spirochaetota were reduced significantly (P < 0.01). The relative abundance of Romboutsia, Turicibacter, and Clostridium sensu stricto increased significantly (P < 0.01), and the relative abundance of Prevotella, Muribaculaceae_norank, Lachnospiraceae NK4A136 group, Roseburia, Treponema, and Ruminococcus significantly decreased (P < 0.01) at the genus level. In addition, the exosome miR-291a-3p and miR-126a-5p levels were markedly regulated by CSC exposure (P < 0.01). The interactions of miR-291a-3p and miR-126a-5p with AMPK and CYP7A1 mRNA were also validated by the DLRG method. CONCLUSIONS: In summary, the rat dyslipidemia induced by CSC exposure may be related to the interference of gut microbiota structure and interaction of miRNAs from serum exosomes with target mRNAs, which further regulated AMPK-ACC/CYP7A1 signaling in rats.

Homeostatic regulation of flavonoid and lignin biosynthesis in phenylpropanoid pathway of transgenic tobacco.

Flavonoids and lignin consist of a large number of secondarymetabolites which are derived from the phenylpropanoid pathway, and they act as a significant role in plant growth, development, and stress response. However, few reports have documented that how different subbranches of phenylpropanoid metablolic pathway mutually interact. In Arabidopsis, AtCPC (AtCAPRICE) is known to play a negative role in anthocyanin accumulation. Nonetheless, whether AtCPC could control the biosynthesis of lignin is largely unknown. Additionally, whether the RrFLS and RrANR, flavonol synthase and anthocyanidin reductase, from Rosa rugosa regulate different branches of phenylpropanoid pathway is unclear. Here, we performed a series of transgenic experiments with short life cycle tobacco and RNA-Seq analysis. Finally, a series of assays related to biological, physiological, and phenotypic characteristics were undertaken. Our results indicated that ectopic expression of AtCPC in tobacco not only decreased the flavonoid compound accumulation, but also up-regulated several lignin biosynthetic genes, and significantly increased the accumulation of lignin. Our results also revealed that although they respectively improved the flavonol and proanthocyanidin contents, the overexpression of RrFLS and RrANR plays positive roles in lignin biosynthesis in transgenic tobacco plants. Our findings provide a novel insight into the mechanism underlying homeostatic regulation of flavonoid and lignin biosynthesis in phenylpropanoid pathway of plants.

RNA-seq analysis of the response of plant-pathogenic oomycete Phytophthora parasitica to the fungicide dimethomorph / Análisis RNA-seq de la respuesta del oomiceto fitopatógeno Phytophthora parasitica al fungicida dimetomorf

Phytophthora parasitica is an important oomycete that causes disease in a variety of plants, dimethomorph fungicides being specific for oomycetes. The aim of this study was to use RNA-seq to rapidly discover the mechanism by which dimethomorph acts in the treatment of P. parasitica. We found that the expression of 832 genes changed significantly after the dimethomorph treatment, including 365 up-regulated genes and 467 down-regulated genes. According to the Gene Ontology (GO) enrichment analysis, pathway enrichment and verification test results, the following conclusions are obtained: (i) the treatment of P. parasitica with dimethomorph causes changes in the expression levels of genes associated with the cell wall and cell wall synthesis; (ii) dimethomorph treatment results in reduced permeability of the cell membrane and changes in the expression of certain transport-related proteins; (iii) dimethomorph treatment increased reactive oxygen species and reduced the expression of genes related to the control of oxidative stress.

Phytophthora parasitica es un importante oomiceto que origina enfermedades en una variedad de plantas; el fungicida dimetomorf es específico contra oomicetos. El objetivo de este estudio fue utilizar la tecnología de RNA-seq para descubrir rápidamente el mecanismo por el que el dimetomorf actúa en el tratamiento de P. parasitica. Descubrimos que la expresión de 832 genes se modificaba significativamente tras el tratamiento con dimetomorf, incluyendo 365 genes que son sobrerregulados y 467 genes que son subrregulados. El análisis de enriquecimiento de ontología de genes (GO), análisis de enriquecimiento de las vías y pruebas de verificación permitieron extraer las conclusiones siguientes: 1) el tratamiento de P. parasitica con dimetomorf origina cambios en los niveles de expresión de los genes relacionados con la pared celular y su síntesis; 2) el tratamiento con dimetomorf origina una reducción de la permeabilidad de la membrana celular, así como cambios en la expresión de ciertas proteínas relacionadas con el transporte, y 3) el tratamiento con dimetomorf incrementó las especies reactivas del oxígeno y redujo la expresión de los genes relacionados con el control del estrés oxidativo.

RNA-seq analysis of the response of plant-pathogenic oomycete Phytophthora parasitica to the fungicide dimethomorph.

Phytophthora parasitica is an important oomycete that causes disease in a variety of plants, dimethomorph fungicides being specific for oomycetes. The aim of this study was to use RNA-seq to rapidly discover the mechanism by which dimethomorph acts in the treatment of P. parasitica. We found that the expression of 832 genes changed significantly after the dimethomorph treatment, including 365 up-regulated genes and 467 down-regulated genes. According to the Gene Ontology (GO) enrichment analysis, pathway enrichment and verification test results, the following conclusions are obtained: (i) the treatment of P. parasitica with dimethomorph causes changes in the expression levels of genes associated with the cell wall and cell wall synthesis; (ii) dimethomorph treatment results in reduced permeability of the cell membrane and changes in the expression of certain transport-related proteins; (iii) dimethomorph treatment increased reactive oxygen species and reduced the expression of genes related to the control of oxidative stress.

Dissecting quantitative trait loci for boron efficiency across multiple environments in Brassica napus.

High yield is the most important goal in crop breeding, and boron (B) is an essential micronutrient for plants. However, B deficiency, leading to yield decreases, is an agricultural problem worldwide. Brassica napus is one of the most sensitive crops to B deficiency, and considerable genotypic variation exists among different cultivars in response to B deficiency. To dissect the genetic basis of tolerance to B deficiency in B. napus, we carried out QTL analysis for seed yield and yield-related traits under low and normal B conditions using the double haploid population (TNDH) by two-year and the BQDH population by three-year field trials. In total, 80 putative QTLs and 42 epistatic interactions for seed yield, plant height, branch number, pod number, seed number, seed weight and B efficiency coefficient (BEC) were identified under low and normal B conditions, singly explaining 4.15-23.16% and 0.53-14.38% of the phenotypic variation. An additive effect of putative QTLs was a more important controlling factor than the additive-additive effect of epistatic interactions. Four QTL-by-environment interactions and 7 interactions between epistatic interactions and the environment contributed to 1.27-4.95% and 1.17-3.68% of the phenotypic variation, respectively. The chromosome region on A2 of SYLB-A2 for seed yield under low B condition and BEC-A2 for BEC in the two populations was equivalent to the region of a reported major QTL, BE1. The B. napus homologous genes of Bra020592 and Bra020595 mapped to the A2 region and were speculated to be candidate genes for B efficiency. These findings reveal the complex genetic basis of B efficiency in B. napus. They provide a basis for the fine mapping and cloning of the B efficiency genes and for breeding B-efficient cultivars by marker-assisted selection (MAS).

[Effects of soil moisture and boron supply on the plant growth and leaf water content and boron forms of oilseed rape cultivars with different boron use efficiency at their seedling stage].

The study with pot experiment showed that under the stress of low soil moisture content, the oilseed rape cultivars with high and low boron (B) use efficiency had a higher fresh mass per plant at their seedling stage under high B supply, compared with those under low B supply. The increment was 43.1% and 31.7%, respectively, but no significant difference was observed between the cultivars. Under two B supply levels, the bound water content and semi-bound B distribution proportion in the leaves of the cultivar with high B use efficiency were 11.4% and 1.7%, and 6.9% and 23.8% higher than those of the cultivar with low B use efficiency, respectively. Under normal soil moisture condition, high B supply induced an increase of fresh mass per plant by 11.1% for the cultivar with high B use efficiency and by 27.4% for the cultivar with low B use efficiency, compared with low B supply. Under these two B supply levels, the cultivar with high B use efficiency had a higher content of free water in its leaves, and its leaf free B content was 2 times as much as that of the cultivar with low B use efficiency, which would benefit the B transportation in plant.