Differential effects of pectin-based dietary fibre type and gut microbiota composition on in vitro fermentation outcomes.

Interactions between human gut microbiota and dietary fibres (DF) are influenced by the complexity and diversity of both individual microbiota and sources of DF. Based on 480 in vitro fermentations, a full factorial experiment was performed with six faecal inocula representing two enterotypes and three DF sources with nanometer, micrometer, and millimeter length-scales (apple pectin, apple cell walls and apple particles) at two concentrations. Increasing DF size reduced substrate disappearance and fermentation rates but not biomass growth. Concentrated DF enhanced butyrate production and lactate cross-feeding. Enterotype differentiated final microbial compositions but not biomass or fermentation metabolite profiles. Individual donor microbiota differences did not influence DF type or concentration effects but were manifested in the promotion of different functional microbes within each population with the capacity to degrade the DF substrates. Overall, consistent effects (independent of donor microbiota variation) of DF type and concentration on kinetics of substrate degradation, microbial biomass production, gas kinetics and metabolite profiles were found, which can form the basis for informed design of DF for desired rates/sites and consequences of gut fermentation. These results add further evidence to the concept that, despite variations between individuals, the human gut microbiota represents a community with conserved emergent properties.

Protective Effects of Mulberry (Morus atropurpurea Roxb.) Leaf Protein Hydrolysates and Their In Vitro Gastrointestinal Digests on AAPH-Induced Oxidative Stress in Human Erythrocytes.

Mulberry leaf protein hydrolysates (HMP), and their in vitro gastrointestinal digests (GHMP), have shown favorable chemical antioxidant activities. The aim of this study is to investigate the potential protective effects of HMP and GHMP against 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH)-induced oxidative stress in human erythrocytes. The inhibition rate of hemolysis, the reactive oxygen species (ROS) level, the concentration of malondialdehyde (MDA), the reduced glutathione (GSH) and oxidized glutathione (GSSH), and the enzymatic activities of total superoxide dismutase (SOD), catalase (CAT), and cellular glutathione peroxidase (GSH-Px) were evaluated as the biomarkers of oxidative status in human erythrocytes. The results showed that HMP and GHMP effectively inhibit the occurrence of erythrocyte hemolysis in the range of 0.025-1.0 mg/mL, and the inhibition rates of HMP and GHMP reached 92% and 90% at concentrations of 0.4 mg/mL and 1.0 mg/mL, respectively. HMP and GHMP reduced the AAPH-induced oxidative hemolysis damage via suppressing the generation of ROS by inhibiting the formation of MDA, maintaining the balance of GSH/GSSG, and preserving the activities of the antioxidant enzymes, including SOD, GSH-Px, and CAT. Our findings revealed that both HMP and GHMP could be used as natural antioxidants, and have the potential for further application in the development of functional foods.

Edible Packaging as a Functional Carrier of Prebiotics, Probiotics, and Postbiotics to Boost Food Safety, Quality, and Shelf Life.

The safety limitations of chemical preservatives led to an increasing trend among industries and customers toward preservative-free foods; hence, the necessity has arisen for developing innovative, safe antimicrobial elements to prolong the shelf life. Beneficial microorganisms that are described as probiotics and also their metabolites are increasingly being considered as bioprotective agents. These microorganisms could be beneficial for extending food shelf-life and boosting human health. During distribution and storage (25 °C or 4 °C), they could contribute to suppressing unwanted microbes and then improving food safety and quality. Also, by tolerating the harsh conditions of gastrointestinal tract (low pH (~3), presence of bile salts, digestive enzymes, competition with other microbes, etc.), probiotics could exert several biological effects at the host. Besides inclusion in foods and supplements, probiotics and their functional metabolites could be delivered via edible packaging (EP). Recent studies have demonstrated the strong potential of pre/pro/post-biotic EP in food biopreservation. These packaging systems may show different potency of food biopreservation. Among others, postbiotics, as metabolic by-products of probiotics, have gained tremendous attention among researchers due to their unique properties like presenting a variety of antimicrobial activities, convenience in use in different industrial stages and commercialization, extended shelf life, and stability in a wide range of pH and temperature. In addition to antimicrobial activities, various bio-EP could differently influence physical or sensorial attributes of food commodities, impacting their acceptance by consumers. Hence, this study is aimed at presenting a comprehensive review of the application of bio-EP, not only by providing a protective barrier against physical damage but also by creating a controlled atmosphere to improve the health and shelf life of food.

Overexpression of ATP Synthase Subunit Beta (Atp2) Confers Enhanced Blast Disease Resistance in Transgenic Rice.

Previous research has shown that the pathogenicity and appressorium development of Magnaporthe oryzae can be inhibited by the ATP synthase subunit beta (Atp2) present in the photosynthetic bacterium Rhodopseudomonas palustris. In the present study, transgenic plants overexpressing the ATP2 gene were generated via genetic transformation in the Zhonghua11 (ZH11) genetic background. We compared the blast resistance and immune response of ATP2-overexpressing lines and wild-type plants. The expression of the Atp2 protein and the physiology, biochemistry, and growth traits of the mutant plants were also examined. The results showed that, compared with the wild-type plant ZH11, transgenic rice plants heterologously expressing ATP2 had no significant defects in agronomic traits, but the disease lesions caused by the rice blast fungus were significantly reduced. When infected by the rice blast fungus, the transgenic rice plants exhibited stronger antioxidant enzyme activity and a greater ratio of chlorophyll a to chlorophyll b. Furthermore, the immune response was triggered stronger in transgenic rice, especially the increase in reactive oxygen species (ROS), was more strongly triggered in plants. In summary, the expression of ATP2 as an antifungal protein in rice could improve the ability of rice to resist rice blast.

Proteomics Analysis Reveals the Molecular Mechanism of MoPer1 Regulating the Development and Pathogenicity of Magnaporthe oryzae.

Glycosylphosphatidylinositol (GPI) anchoring the protein GPI modification post-transcriptionally is commonly seen. In our previous study, MoPer1, a GPI anchoring essential factor, has a critical effect on Magnaporthe oryzae growth, pathogenicity, and conidiogenesis, but its molecular mechanism is not clear. Here, we extracted the glycoproteins from the ΔMoper1 mutant and wild-type Guy11 to analyze their differential levels by quantitative proteomic analysis of TMT markers. After background subtraction, a total of 431 proteins, with significant changes in expression, were successfully identified, and these differential proteins were involved in biological regulation, as well as cellular process and metabolic process, binding, catalytic activity, and other aspects. Moreover, we found that MoPer1 regulates the expression of 14 proteins involved in growth, development, and pathogenicity of M. oryzae. The above findings shed light on MoPer1's underlying mechanism in regulating growth, development, and pathogenicity of M. oryzae.

Characterization of Purified Mulberry Leaf Glycoprotein and Its Immunoregulatory Effect on Cyclophosphamide-Treated Mice.

Mulberry leaf protein is a potentially functional food component and health care agent with antioxidant and anti-inflammatory properties. However, its composition, immunoregulatory effects, and gut microbial regulatory effects are unclear. Herein, ultra-filtrated and gel-fractionated mulberry leaf protein (GUMP) was characterized. Its effects on cyclophosphamide-induced immunosuppressed mice were further investigated. The results indicated that GUMP is a glycoprotein mainly containing glucose, arabinose, and mannose with 9.23% total sugar content. Its secondary structure is mainly ß-sheet. LC-MS/MS analysis showed that GUMP closely matched with a 16.7 kDa mannose-binding lectin and a 52.7 kDa Rubisco's large subunit. GUMP intervention significantly improved serous TNF-α, IL-6, and IL-2 contents; increased serum immunoglobulins (IgA and IgG) levels; and reversed splenic damage prominently. Moreover, GUMP administration increased fecal shot-chain fatty acid concentration and up-regulated the relative abundance of Odoribacter, which was positively correlated with SCFAs and cytokine contents. Overall, GUMP alleviated immunosuppression through the integrated modulation of the gut microbiota and immune response. Therefore, GUMP could be a promising dietary supplement to help maintain gut health.

Anti-inflammatory effects and molecular mechanisms of bioactive small molecule garlic polysaccharide.

Although garlic polysaccharides have been found to possess anti-inflammatory activities, anti-inflammatory study on small molecule water-soluble garlic polysaccharide (WSGP) is few. In this study, a novel WSGP with a molecular weight of 1853 Da was isolated by DEAE-52 and Sephadex G-100 column and the chemical composition was identified by monosaccharide composition and methylation analysis. Furthermore, the antioxidant effects of WSGP and the potential molecular mechanisms on LPS-induced inflammatory responses in RAW264.7 macrophage cells were investigated. The results showed that WSGP has strong antioxidant activity, such as DPPH, hydroxyl, superoxide anion, ABTS radical scavenging capacity, Fe2+ chelating ability and reducing power. Meanwhile, WSGP could considerably suppress the manufacturing of NO and the mRNA and protein expression degrees of IL-6, TNF-α, and IL-1ß in LPS inspired RAW264.7 macrophages WSGP could significantly suppress the production of NO and the mRNA and protein expression levels of IL-1ß, IL-6, and TNF-α in LPS stimulated RAW264.7 macrophage cells (p < 0.05). In addition, the phosphorylated IκB-α, p65, and STAT3 proteins were significantly increased in LPS-induced macrophages, while this trend was significantly reversed by WSGP treatment in a concentration-dependent manner (p < 0.05). Consequently, WSGP supplementation might reduce LPS-induced inflammatory responses by suppressing proinflammatory cytokines and NF-κB and STAT3 pathway activation. The finding of this research would give scientific guidelines for the judicious use of small molecular garlic polysaccharide in anti-inflammatory treatments.

Effects of enzymatic hydrolysis on physicochemical property and antioxidant activity of mulberry (Morus atropurpurea Roxb.) leaf protein.

To improve the antioxidant efficiency of mulberry leaf protein (MLP), alcalase, protamex, papain, flavourzyme, neutrase, and trypsin were used to hydrolyze MLP. The yield of soluble peptides, secondary structures, molecular weight distributions, and antioxidant activities of MLP hydrolysates (MLPHs) were investigated. Results showed that the native MLP was rich in the fraction above 6.5 kDa and was mainly composed of ß-sheets, while MLPHs were abundant in the fractions of 0.3-0.6 kDa and 0.6-6.5 kDa and were mainly composed of disordered coils and ß-folds. Limited hydrolysis of MLP could lead to better antioxidant activity than extensive hydrolysis. After enzymatic hydrolysis, the content of total sugar and total phenol in MLP increased significantly. MLP hydrolysates prepared with neutrase, alcalase, and protamex were preferable to other enzymes. Meanwhile, an enzyme to substrate level of 1% and a hydrolysis time of 2 hr were the optimum conditions to obtain higher antioxidant hydrolysates using neutrase.

GroEL protein from the potential biocontrol agent Rhodopseudomonas palustris enhances resistance to rice blast disease.

BACKGROUND: GroEL, which is a chaperone, plays a key role in maintaining protein homeostasis and, among other functions, serves to prevent protein misfolding and aggregation. In addition, the GroEL protein also has a significant effect on enhancing plant resistance and inhibiting plant diseases. However, the function of the GroEL protein in the inhibition of rice blast remains unknown. RESULTS: Field experiment results show that photosynthetic bacteria PSB-06 have a good control effect on Magnaporthe oryzae. PSB-06 also can promote rice growth and enhance stress resistance. A GroEL protein which was separated and purified from photosynthetic bacteria had a significant antagonistic effect on appressorial formation and pathogenicity of Magnaporthe oryzae, meanwhile transcriptional analysis demonstrated that the GroEL protein could improve the expression of defense gene of rice. CONCLUSION: Our results show that the photosynthetic bacteria Rhodopseudomonas palustris significantly controls rice blast disease. Its action involves an extracellular GroEL protein, which inhibits appressoria formation, antagonizes the pathogenicity of Magnaporthe oryzae and promotes a host defense response. The research results provide evidence of the potential of this photosynthetic bacterium as a biocontrol agent at least for rice blast control. © 2021 Society of Chemical Industry.

One-step duplex RT-droplet digital PCR assay for the detection of norovirus GI and GII in lettuce and strawberry.

The study was designed to develop a sensitive one-step duplex reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) to detect norovirus genogroup I and II (NoV GI and GII) in lettuce and strawberry. The specificity, sensitivity, repeatability and robustness of the assay was compared with RT-qPCR. The lowest concentration detected by RT-ddPCR for NoV GI and NoV GII were 4.68 and 8.47 copies/µL respectively, much lower than that of RT-qPCR with a number of 46.8 and 84.7 copies/µL, respectively. Lettuce and strawberry samples were artificially contaminated with NoV GI and GII suspensions, with inoculum size of 3.00 × 106 to 1.70 × 108 copies and 4.80 × 105 to 2.50 × 107 copies, respectively. Strawberry spiked with low inoculum size revealed positive results by RT-ddPCR, while recorded negative by RT-qPCR. Meanwhile, RT-ddPCR also showed a higher average recovery rate for NoV in lettuce and strawberry than RT-qPCR.The limit of detection (LoDs) of RT-ddPCR for NoVs in lettuce was 2.32 × 104 copies/25g (NoV GI) and 2.36 × 104 ciopies/25g (NoV GII), and that in strawberry was 2.56 × 104 copies/25g (NoV GI) and 2.64 × 104 ciopies/25g (NoV GII), which were 10 folds lower than that of RT-qPCR. The developed duplex RT-ddPCR assay exhibited stability and increased capacity to resist inhibitors in food samples with low concentration of NoV, making it a reliable method to avoid false negative result as opposed to RT-qPCR. In conclusion, one-step RT-ddPCR method developed in this study is pertinent in detecting foodborne virus such as NoV.

Structural characterization and functional activity of an exopolysaccharide secreted by Rhodopseudomonas palustris GJ-22.

One water exopolysaccharide, designated G-EPS, was secreted by Rhodopseudomonas palustris GJ-22 culture media. The structure of G-EPS was characterized with HPGPC, GC-MS, methylation, 1D and 2D NMR, along with UV and FT-IR spectrum. The G-EPS molecular weight was 10.026 kilodalton, and is composed of D-mannose (92.8%) and d-glucose (7.2%). The purified G-EPS promoted plant growth and induced systemic resistance against TMV in Nicotiana benthamiana. These results suggested that G-EPS is an important active component of the bio-control capacity of GJ-22.

The effects of bioactive compounds from blueberry and blackcurrant powders on the inhibitory activities of oat bran pastes against α-amylase and α-glucosidase linked to type 2 diabetes.

The α-amylase and α-glucosidase inhibitory activities by extracts of oat bran, blueberry and blackcurrant powders, as well as oat bran pastes supplemented 25% of blueberry and blackcurrant powder, were studied by measuring their half inhibitory (IC50) concentrations. Addition of blueberry or blackcurrant powder into oat bran paste increased α-amylase and α-glucosidase inhibitory activities with a decrease in IC50 values. The main anthocyanidin content was measured by pH differential method and the potential inhibitory mechanisms of these extracts were also investigated by detailed inhibition kinetics and docking simulations. The results showed that: (1) cyanidin and delphinidin were the main anthocyanidin profiles in extracts; (2) only blackcurrant powder was a competitive inhibitor, while other extracts were all mixed-type inhibitors against α-amylase; (3) both blueberry- and blackcurrant-enriched pastes were competitive inhibitors, while other extracts were all mixed-type inhibitors towards α-glucosidase; (4) the α-amylase and α-glucosidase inhibitory activities by extracts were potentially driven by hydrogen bonding, cyanidin-3-glucoside and delphinidin-3-glucoside had stronger binding affinity compared to malvidin-3-glucoside and cyanidin-3-rutinside. This study suggested supplementary of blueberry and blackcurrant with oat bran might be a potential source of bioactive products for antidiabetic activity.

Anti-Inflammatory and Intestinal Microbiota Modulation Properties of Jinxiang Garlic (Allium sativum L.) Polysaccharides toward Dextran Sodium Sulfate-Induced Colitis.

Garlic polysaccharides are great potential agents because of their anti-inflammation, antioxidation, and immunomodulation properties. However, few studies have reported their anti-inflammatory effects on improving the colon system and corresponding intestinal microbiota. Herein, a water-soluble garlic polysaccharide (WSGP) was extracted from Jinxiang garlic to evaluate its effects on ameliorating dextran sulfate sodium (DSS)-induced colitis in a mouse model. The results showed that (1) after administration of the WSGP (200 or 400 mg/kg/day), the feed intake, body weight, and colon length of colitic mice were increased, while the disease activity index and the histological score of colitic mice were decreased; (2) the WSGP reduced the colonic tissue damage and inhibited the expression of inflammatory factors (interleukin 6, interleukin 1 beta , and tumor necrosis factor alpha); and (3) the WSGP enhanced the production of short-chain fatty acids and improved the composition of intestinal microbiota. The key microorganisms, including Muribaculaceae, Lachnospiraceae, Lachnospiraceae_NK4A136_group, Mucispirillum, Helicobacter, Ruminococcus_1, and Ruminiclostridium_5, were identified to be associated with inflammatory bowel diseases. Taken together, this study proved that WSGP supplementation could alleviate DSS-induced colitis by improving mucosal barriers, blocking proinflammatory cytokines, and modulating gut microbiota.

Two novel polysaccharides from Solanum nigrum L. exert potential prebiotic effects in an in vitro fermentation model.

In this research, two novel polysaccharides (S1 and S2) from Solanum nigrum L were extracted and purified. Then homogeneity, molecular weights, major chemical contents and monosaccharide compositions of S1 and S2 were determined. Then, the effects of S1 and S2 on human faecal microbial community and short-chain fatty acid production were investigated using an in vitro fermentation model. Results showed that S1 and S2 have different impacts on human gut microbiota in vitro. S1 selectively promoted the abundance of 9 genera and the production of propionic acid, butyric acid, isobutyric acid, valeric acid and isovaleric acid; while S2 selectively promoted the abundance of 8 genera and the production of acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid and succinic acid. Also, S1 group had higher abundance of genera Butyricimonas and Megamonas and higher levels of lactic acid than S2; while S2 group had higher abundance of Megaphaera and higher levels of butyric acid, valeric acid, isobutyric acid, isovaleric acid and succinic acid comparably. We concluded that S1 and S2 may have potential prebiotic functions.

Substrate Use Prioritization by a Coculture of Five Species of Gut Bacteria Fed Mixtures of Arabinoxylan, Xyloglucan, ß-Glucan, and Pectin.

Dietary fiber provides growth substrates for bacterial species that belong to the colonic microbiota of humans. The microbiota degrades and ferments substrates, producing characteristic short-chain fatty acid profiles. Dietary fiber contains plant cell wall-associated polysaccharides (hemicelluloses and pectins) that are chemically diverse in composition and structure. Thus, depending on plant sources, dietary fiber daily presents the microbiota with mixtures of plant polysaccharides of various types and complexity. We studied the extent and preferential order in which mixtures of plant polysaccharides (arabinoxylan, xyloglucan, ß-glucan, and pectin) were utilized by a coculture of five bacterial species (Bacteroides ovatus, Bifidobacterium longum subspecies longum, Megasphaera elsdenii, Ruminococcus gnavus, and Veillonella parvula). These species are members of the human gut microbiota and have the biochemical capacity, collectively, to degrade and ferment the polysaccharides and produce short-chain fatty acids (SCFAs). B. ovatus utilized glycans in the order ß-glucan, pectin, xyloglucan, and arabinoxylan, whereas B. longum subsp. longum utilization was in the order arabinoxylan, arabinan, pectin, and ß-glucan. Propionate, as a proportion of total SCFAs, was augmented when polysaccharide mixtures contained galactan, resulting in greater succinate production by B. ovatus and conversion of succinate to propionate by V. parvula Overall, we derived a synthetic ecological community that carries out SCFA production by the common pathways used by bacterial species for this purpose. Systems like this might be used to predict changes to the emergent properties of the gut ecosystem when diet is altered, with the aim of beneficially affecting human physiology.IMPORTANCE This study addresses the question as to how bacterial species, characteristic of the human gut microbiota, collectively utilize mixtures of plant polysaccharides such as are found in dietary fiber. Five bacterial species with the capacity to degrade polymers and/or produce acidic fermentation products detectable in human feces were used in the experiments. The bacteria showed preferential use of certain polysaccharides over others for growth, and this influenced their fermentation output qualitatively. These kinds of studies are essential in developing concepts of how the gut microbial community shares habitat resources, directly and indirectly, when presented with mixtures of polysaccharides that are found in human diets. The concepts are required in planning dietary interventions that might correct imbalances in the functioning of the human microbiota so as to support measures to reduce metabolic conditions such as obesity.

Novel Antioxidant Peptides Purified from Mulberry (Morus atropurpurea Roxb.) Leaf Protein Hydrolysates with Hemolysis Inhibition Ability and Cellular Antioxidant Activity.

Neutrase-hydrolysates hydrolyzed from mulberry leaf proteins were separated by ion exchange chromatography, gel filtration chromatography, and semipreparative reverse-phase HPLC. Purified fractions were analyzed for their radical scavenging activity, hemolysis inhibition ability, and cellular antioxidant activity (CAA). Three new antioxidant peptides, P1 (SVL, 317 Da), P2 (EAVQ, 445 Da), and P3 (RDY, 452 Da), were obtained from the most active HPLC fraction (R1) and identified using UPLC-QTOF-MS. These three peptides were then synthesized, and their antioxidant activities were analyzed. P1 and P2 had no ability to inhibit hemolysis of erythrocytes but did show antioxidant activity on HepG2 cells. P3 showed the highest hemolysis inhibition ability (92%) and CAA value (2204 µM QE/100 g peptide). The Tyr residues at the C-terminal region play an important role in the antioxidant activity in P3. Thus, the natural peptide R1 and synthesized P3 could be used as antioxidants and might be promising components of functional foods.

A high-throughput and ultrasensitive identification methodology for unauthorized GMP component based on suspension array and logical calculator.

To solve the problem of the unauthorized GMP components within import and export goods, the LI-US (Logic Identification of unauthorized GMP content by Universal-primer Suspension-array) system, which takes advantage of suspension array and logic calculator, was developed in the present study. Seventeen signal input channels have been optimized and validated in our research to ensure the multiplex practicality of the LI-US system. Three LI-US logic gates, including a YES gate, an OR gate and an AND gate, were designed as different detection strategies for GMP identification. The feasibility and specificity of the LI-US system were validated in the present study. Combining the optimization and evaluation of the signal input procedure, the sensitivity of this LI-US system reached 0.05% of the GMP mass concentration. The practicability evaluation of LI-US demonstrated its application within different substrates and varieties. In conclusion, the LI-US system was developed with extremely high specificity, sensitivity and practicability among different substrates and varieties, which could meet the demands of unauthorized GMP contents for both import and export goods.

Destiny of Dendrobium officinale Polysaccharide after Oral Administration: Indigestible and Nonabsorbing, Ends in Modulating Gut Microbiota.

Polysaccharides from functional foods have been proved to have diverse bioactivities, but little is known about what exactly happens to these polysaccharides after oral administration and even less about the underlying mechanism of action. Taking the marker polysaccharide (DOP) of Dendrobium officinale as an example, this study aims to demonstrate the dynamic distribution and degradation of orally dosed DOP in mice and in vitro using near-infrared fluorescence imaging and a kind of chromatographic analysis. The results indicate that (1) neither DOP nor fluorescence-labeled DOP (FDOP) was absorbed, (2) both DOP and FDOP were undigested and were quickly degraded to short-chain fatty acids in the large intestine, (3) DOP modulated gut microbiota, which could be associated with DOP's suppression of 4T1 tumor growth in mice. All of these findings suggest that some (maybe not all) bioactive polysaccharides share a common destiny: indigestible and nonabsorbing, ends in modulating bioactivities-associated gut microbiota.

Unintended effects of transgenic rice revealed by transcriptome and metabolism.

Genetically modified (GM) organisms have been developed for decades. However, unintended effects are the main concerns of safety assessment that needs to be carefully investigated. Here, eight varieties of GM rice that were developed in China were selected to assess the unintended effects through transcriptome and metabolism. There are 2892-8758 differentially expressed genes (DEGs) and 7-50 metabolites at significant level between GM varieties and their isogenic counterparts, which were far fewer than that between traditional rice varieties. The function enrichment analysis showed altered transcription in stress-related pathway and starch and sucrose metabolism. DEGs shared among eight GM samples constitute less than 1% of the genes in the genome, and none of them is reported more than four times. The insertion effect on the nearby gene expression and the associated metabolism is only restricted to 50 genes. All the results provide a comprehensive analysis of unintended effects and indication of difference in Chinese transgenic rice based on their backgrounds, transformation, and insertion elements.