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Stroke is one of the leading causes of death and disability worldwide. However, information on stroke-related tongue coating microbiome (TCM) is limited, and whether TCM modulation could benefit for stroke prevention and rehabilitation is unknown. Here, TCM from stroke patients (SP) was characterized using molecular techniques. The occurrence of stroke resulted in TCM dysbiosis with significantly reduced species richness and diversity. The abundance of Prevotella, Leptotrichia, Actinomyces, Alloprevotella, Haemophilus, and TM7_[G-1] were greatly reduced, but common infection Streptococcusand Pseudomonas were remarkably increased. Furthermore, an antioxidative probiotic Lactiplantibacillus plantarumAR113 was used for TCM intervention in stroke rats with cerebral ischemia/reperfusion (I/R). AR113 partly restored I/R induced change of TCM and gut microbiota with significantly improved neurological deficit, relieved histopathologic change, increased activities of antioxidant enzymes, and decreased contents of oxidative stress biomarkers. Moreover, the gene expression of antioxidant-related proteins and apoptosis-related factors heme oxygenase-1 (HO-1), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), nuclear factor erythroid 2-related factor 2 (Nrf2), NAD(P)H:quinone oxidoreductase-1 (NQO-1), and Bcl-2 was significantly increased, but cytochrome C, cleaved caspase-3, and Bax were markedly decreased in the brain by AR113 treatment. The results suggested that AR113 could ameliorate cerebral I/R injury through antioxidation and anti-apoptosis pathways, and AR113 intervention of TCM may have the application potential for stroke prevention and control.

Probiotics could effectively eliminate excess reactive oxygen species (ROS) generated during aging or lipid metabolism disorders, but their mechanism is unclear. The major purpose of this study was to investigate the mechanism of Lactiplantibacillus plantarunAR113 alleviating oxidative stress injury in the D-galactose induced aging mice. The result showed that pretreatment with L. plantarunAR113 significantly relieving H2O2induced cytotoxicity in HepG2 cells by maintain cell membrane integrity and increasing antioxidant enzyme activities. In D-galactose induced aging mice, L. plantarunAR113 could significantly attenuate liver damage and inflammatory infiltration by promoting endogenous glutathione (GSH) synthesis and activating the Nrf2/Keap1 signaling pathway in mice, and increasing the expression of regulated phase II detoxification enzymes and antioxidant enzymes. Further analysis shown that gavage of L. plantarunAR113 could significantly reduce the expression of G protein-coupled receptor 78 (GPR78) and C/EBP homologous protein (CHOP) proteins, and promote the restoration of endoplasmic reticulum (ER) homeostasis, thereby activating cell anti-apoptotic pathways. These results were also confirmed in H2O2-treated HepG2 experiments. It indicated that L. plantarunAR113 could inhibit D-galactose-induced liver injury through dual inhibition of ER stress and oxidative stress. L. plantarunAR113 have good application potential in anti-aging and alleviating metabolic disorders.

Probiotics have great potential in regulating intestinal pain. In this study, the effects of Lactobacillus plantarumAR495 on the visceral sensitivity and gut microbiota of irritable bowel syndrome (IBS) rats were studied. The results showed that tryptase released after mast cell activation and degranulation plays a key role in visceral pain, and L. plantarumAR495 reduced the stimulation of colonic mast cells and the expression of protease-activated receptor 2 (PAR2) and TRPV1 in dorsal root ganglia. Research further showed that supplementation with L. plantarumAR495 increased the level of short-chain fatty acids (SCFAs) and enhanced the barrier function of the colon. In addition, the microbiota analysis of the colon indicated that L. plantarumAR495 promoted the proliferation of Bifidobacteriumand inhibited the proliferation of Lachnospiraceae, which alleviated the imbalance of the intestinal microbiota caused by IBS to a certain extent. In total, L. plantarumAR495 might reduce visceral sensitivity through the Mast cell-PAR2-TRPV1 signaling pathway by maintaining the homeostasis of the intestinal barrier.

Membrane characteristics are determined by the fatty acids composition, which affects survival rates after freeze-drying. However, it is unknown which composition provides the greatest effect. In this study, we found that the addition of Tween 80 and Tween 20 significantly increased survival rates of Lactiplantibacillus plantarum, which reached a maximum of 93.1 %. Conversely, Tween 60 caused a significant decrease. We further found that the difference between the effects of adding Tween 80 and Tween 60 was the change in oleic acid contents. To verify the role of oleic acid, we used CRISPR-Cas9 to knock-out the key synthesis gene cla-er. The survival rates of L. plantarumAR113Δcla-erdeclined to 5.48 %. The addition of oleic acid restored the rates to those of wild type strains. Moreover, the membrane integrity and fluidity of knockout strains markedly decreased. This is the first confirmation that Tween 80 or Tween 20 increases the survival rate by increasing the content of oleic acid in the cell membrane. These findings also indicated that oleic acid in cell membranes has a substantial protective effect on L. plantarumduring freeze-drying.

Western diet is thought to increase susceptibility to inflammatory bowel disease (IBD), and probiotics are a potential therapeutic agent for IBD. This study revealed the effects of Lactobacillus plantarumAR113 and L. plantarumAR113Δbsh1 on a dextran sulfate sodium (DSS)-induced colitis mouse model under the Western diet (WD). After four weeks of WD and low-sugar and low-fat diet (LD) intervention, induction with 3% DSS, and intragastric administration of probiotics, we found that L. plantarumAR113 could regulate blood glucose and lipid levels and have a certain protective effect on hepatocytes. Our results suggested that the L. plantarumAR113 alleviated DSS-induced colitis under the Western diet by improving dyslipidemia, repairing intestinal barrier dysfunction, and inhibiting the TLR4/Myd88/TRAF-6/NF-κB inflammatory pathway. However, these changes were not demonstrated in the L. plantarumAR113Δbsh1, and therefore, we reasoned that the presence of bsh1 may play a crucial role in the L. plantarumAR113 exerting its anti-inflammatory function. The relationship between bile salt hydrolase (BSH) and colitis was worthy of further exploration.

Long-term exposure to adverse life events that provoke acute or chronic psychological stress (hereinafter “stress”) can negatively affect physical health and even increase susceptibility to psychological illnesses, such as anxiety and depression. As a part of the hypothalamic–pituitary–adrenal axis, corticotropin-releasing factor (CRF) released from the hypothalamus is primarily responsible for the stress response. Typically, CRF disrupts the gastrointestinal system and leads to gut microbiota dysbiosis, thereby increasing risk of functional gastrointestinal diseases, such as irritable bowel syndrome. Furthermore, CRF increases oxidative damage to the colon and triggers immune responses involving mast cells, neutrophils, and monocytes. CRF even affects the differentiation of intestinal stem cells (ISCs), causing enterochromaffin cells to secrete excessive amounts of 5-hydroxytryptamine (5-HT). Therefore, stress is often accompanied by damage to the intestinal epithelial barrier function, followed by increased intestinal permeability and bacterial translocation. There are multi-network interactions between the gut microbiota and stress, and gut microbiota may relieve the effects of stress on the body. Dietary intake of probiotics can provide energy for ISCs through glycolysis, thereby alleviating the disruption to homeostasis caused by stress, and it significantly bolsters the intestinal barrier, alleviates intestinal inflammation, and maintains endocrine homeostasis. Gut microbiota also directly affect the synthesis of hormones and neurotransmitters, such as CRF, 5-HT, dopamine, and norepinephrine. Moreover, the Mediterranean diet enhances the stress resistance to some extent by regulating the intestinal flora. This article reviews recent research on how stress damages the gut and microbiota, how the gut microbiota can improve gut health by modulating injury due to stress, and how the diet relieves stress injury by interfering with intestinal microflora. This review gives insight into the potential role of the gut and its microbiota in relieving the effects of stress via the gut–brain axis.

The short-circuit current calculation is of vital importance for the power system. With a high proportion of new energy sources, the setting of relay protection and the selection of electrical equipment have been severely degraded. An accurate short-circuit current calculation model of new energy source is necessary for the short-circuit calculation of power grid with new energy sources. However, due to the DFIG’s control system is a ”black box”, and an accurate fault characteristics and short-circuit current calculation models of DFIG is difficult to obtained. Currently, for the fault current of the DFIG, lots of research works is carried out under symmetrical fault or the crowbar protection is activated condition. However, under crowbar protection is not activated and unsymmetrical fault condition, there are no unified short-circuit calculation models of DFIG. In this paper, the transient process of DFIG under those condition is analyzed, and mathematical expressions are derived. Moreover, the comprehensive and unified short-circuit calculation models of DFIG have established under different low voltage ride through control conditions. The influence of GSC and slip rate of DFIGs can be considered. The test results show that the LVRT control strategies, the GSC and slip have an important influence on the DFIG’s short-circuit calculation model. The quantitative research results are of important significance for relay protection setting calculation, grid operation and control, and the selection of electrical equipment of power grid with DFIGs.

Carbon source is an essential factor influencing exopolysaccharide (EPS) production. However, there is still lack of systematic investigation how carbon sources affect EPS characterization in S. thermophilus. In this study, EPSs from S. thermophilusS-3 using lactose, glucose and galactose as the sole carbon source were characterized, and named LacS3EPS, GlcS3EPS and GalS3EPS, respectively. The yield of EPSs was significantly affected by the carbon source. The monosaccharide and linkage type of EPSs were independent of the carbon source. Compared with LacS3EPS, the glucose content of GlcS3EPS increased while galactose content decreased, but the opposite trend occurred for GalS3EPS. The molecular weight of EPSs changed significantly, with the highest for GlcS3EPS (1630 kDa) and the lowest for GalS3EPS (414 kDa). Combined with two wormlike cylinder models, the conformation of EPSs was characterized by random coils with descending rigidity in the order GlcS3EPS > LacS3EPS > GalS3EPS. Additionally, gene transcription and correlation analysis revealed that the variation in EPSs was mainly related to the genes encoding nucleotide sugars and glycosyltransferases. The results suggested that carbon sources could alter molecular characteristics of EPS by influencing gene expression related to EPS biosynthesis.

Lacticaseibacillus caseiis a common bacterium, due to its small genome and relatively simple metabolism, it will process the potential to become a cell factory. Although CRISPR has been successfully applied to the gene editing of L. casei, the technology is limited by the size of the knock-in gene and cannot achieve efficient integration of large fragments, which has impeded the application of L. caseiin synthetic biology. Here, we develop a genome integration tool applicable to L. casei. Three possible phage integrases were screened and introduced into L. caseiAR1088. A highly active integrase in L. caseiwas screened. Based on this, a single plasmid-based efficient gene integration tool for L. caseiwas established. With the assistance of CRISPR-Cas9 editing system, we realized multi-site integration of genome. Then, homologous modeling of integrase φA2 was conducted to explore its core amino acid site and minimum functional region. Two core sites of H334 and T366 were identified, and it was confirmed that the full-length protein of integrase was essential for integration. As a proof of concept, the bile salt hydrolase gene from Lactiplantibacillus plantarumwas integrated into AR1088 genome at multiple sites, which improved the acid and bile salt tolerance of the engineered strain. The system developed in this study overcomes the deficiencies of currently available genetic tools in the multi-site chromosomal integration of large DNA fragment in L. casei, and may be extended to other lactobacillus.

Bifidobacteriumcan be used as probiotic that confers various health benefits to its host. The functional analysis of specific genes can help elucidate the probiotic mechanism of Bifidobacterium. However, the low transformation efficiency has hindered the in-depth study of its function by genetic manipulation. In this study, we optimized transformation methods for Bifidobacterium animalisAR668 to improve the transformation efficiency up to 2.15 ×105 CFU/μg DNA. Additionally, the expression of competence gene ssb2could enhance the transformability which was the first attempt to apply it to B. animalis. Moreover, the transformation efficiency was increased after the plasmids were modified by gene0033and gene1509which limited the interference of the restriction-modification system in the uptake of foreign DNA by cells. Based on the above optimization, the transformation efficiency of 7.2 ×105 CFU/μg DNA for AR668 was obtained, which was more than 100 times higher than that of the original condition. Together, we successfully improved the transformation efficiency of B. animalisAR668, contributing to its genetic manipulation and modification.

Thermostable alkaline protease (TAP) harbored by Pseudomonas fluorescensdecomposes protein in milk and dairy products, leading to milk and dairy product spoilage during storage. Thus, a specific, sensitive, rapid, and simple method is required to detect TAP-harboring P. fluorescens. Two sets of primers targeting the aprXand gyrBgenes of P. fluorescenswere designed. The detection system and conditions were optimized, and a real-time loop-mediated isothermal amplification (real-time LAMP) method was developed for the simultaneous detection of TAP-harboring P. fluorescensin two separate reaction tubes. The phylogenetic tree targeting aprXshowed that P. fluorescensand Pseudomonas luridaclustered on the same branch. The phylogenetic tree targeting gyrBshowed that P. fluorescensclustered on the same branch with 95% confidence value, whereas P. luridaclustered on different branches. DNA of 16 strains of P. fluorescensand 34 strains of non–P. fluorescenswas detected by real-time LAMP. TAP-harboring P. fluorescenscan only be identified when the real-time LAMP detection results of both aprXand gyrBare positive. The dissociation temperatures of aprXand gyrBin the real-time LAMP–amplified products were approximately 90.0 and 88.0°C, respectively. The detection limits of the real-time LAMP targeting aprXand gyrBwere 4.9 CFU per reaction in pure culture and 2.2 CFU per reaction in skimmed milk. The coefficient of variation of the repeatability test was less than 2%, indicating that the established real-time LAMP of P. fluorescenstargeting gyrBand aprXhas good stability and repeatability. Real-time LAMP was used to test 200 raw milk samples for the presence of TAP-harboring P. fluorescensin 3 h, and the coincidence rate of the results with those obtained using the traditional method, which takes at least 5 to 7 days, was 100%. Real-time LAMP will be a practical and effective method for accurate and rapid identification of TAP-harboring P. fluorescensin raw milk.

Hyperlipidemia significantly increases the risk of cardiovascular disease, and the number of patients is gradually increasing and tends to be younger. The prevalence factors and pathogenesis of hyperlipidemia are the hotspots of current research. Research has shown that abnormal blood lipids are closely related to gut microbiota and dietary habits. We investigated the association between gut microbiota typing and lipid indices in hyperlipidemic patients by 16s rRNA and resolved the differences with gut microbiota in healthy populations, and compared the experimental results using Meta-analysis. The results displayed that the gut microbiota of hyperlipidemic patients could be classified into two enterotypes (Bac and Bla) and there was a significant difference such as Faecalibacterium, Bacteroides, and Agathobacterbetween the two groups. There was a high similarity in the composition of gut microbiota between Bac enterotype patients and healthy individuals, and the TG levels of the patients in the Bac group were also less different from those of the healthy population. However, the relative abundance of Faecalibacteriumand Bacteroideswas lower appreciably in patients with Bla enterotypes compared to patients with Bac enterotypes and healthy populations, suggesting that TG levels may influence changes in gut microbiota. Furthermore, the results of Mate-analysis did not reveal a significant correlation between Bacteroidesand TG, whereas this study has detected a negative correlation between the abundance of Bacteroidesand TG content. And the changes in Faecalibacteriumabundance in the gut microbiota of hyperlipidemic patients and healthy people in this cohort population were consistent with the results of Meta-analysis. Therefore, they may be potential strains for the treatment of hyperlipidemia caused by elevated TG.

This study investigated the intervention effects of Lactobacillus caseiLC2W in murine (SPF C57BL/c) challenge infection models induced by Escherichia coliO157: H7. Mice were fed streptomycin with water for 3 days prior to intragastric gavage by E. coliO157:H7 (Control) or L. caseiLC2W together with E. coliO157:H7 (Intervention) to explore the role of L. caseiLC2W by biochemical indicators, histological evaluation, expression of colonic pro-inflammatory and intestinal barrier factors related to enteritis. Results showed that the administration of L. caseiLC2W was able to alleviate the symptoms of colitis induced by E. coliO157:H7, exhibiting lower weight loss as well as more intact colon tissue. Furthermore, L. caseiLC2W could down-regulate the expression of pro-inflammatory cytokines (TNF-α, IL-6, and IL-1β) and protect the intestinal barrier function by improving the level of MUC2, ZO-1 and E-cadherin 1 compared to the control group. These results demonstrate that L. caseiLC2W can reduce the severity of E. coliO157:H7 infection, and suggest L.caseiLC2W may maintain the immune balance and intestinal barrier to reduce colitis. In addition, we found the effect of intervention is similar to that of prevention, which is better than that of treatment.

A novel BL312 milk-clotting enzyme (MCE) exhibited high-level expression and remarkable milk-clotting activity (MCA) (865 ± 20 SU/mL) that was 3.3-fold higher than the control by optimizing induction conditions in recombinant Escherichia. coliharboring pET24a-proMCE. Through substrate-binding region analyses and modification, MCE-G165A was identified from nine mutants and showed a proteolytic activity of 49.4 ± 2.4 U/mL and an MCA/PA ratio of 18.2, which were respectively 1.9-fold lower and 2.0-fold higher than those of the control. The purified MCE-G165A (28 kDa) exhibited weak αs-casein, β-casein, and strong κ-casein (κ-CN) hydrolysis levels as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reversed-phase high-performance liquid chromatography. The milk-clotting mechanism for MCE-G165A was the primary hydrolysis of Met106-Ala107 and Asn123-Thr124 bonds in κ-CN, as determined by mass spectrometry. MCE-G165A showed different hydrolysis sites in casein, leading to various functional peptides. Feasible methods for obtaining MCEs suitable as calf rennet substitutes are presented.

Lactobacillus salivariusis an important probiotic lactic acid bacterium that plays a major role in innate and acquired immunity. It has gained much attention in metabolic engineering and synthetic biology owing to its probiotic properties. However, few well-characterized constitutive promoters are available to modulate L. salivariusgene expression. Here, a pool of L. salivariusendogenous constitutive promoters was identified using RNA sequencing analysis. The promoters were sorted based on their transcript levels. The application potential of the promoters was evaluated using a fluorescence reporting system. This study provides the first well-characterized endogenous constitutive promoter library from L. salivarius, which has a potential for use as a basic toolbox in various lactic acid bacteria applications.

The gut microbiota plays a vital role in human health. Many studies have shown that an imbalance of in the intestinal flora not only can cause intestinal diseases; but also is closely related to obesity, diabetes and other chronic metabolic diseases. Probiotics are defined as live microorganisms, that can colonize the human intestinal tract to improve the microbiological balance and migigate diabetes and other metabolic diseases. Therefore, approaches that aim to target the intestinal flora and thus alleviate diabetes have unique advantages, although the underlying mechanisms are complex and diverse. From this perspective, this review summarize here the relationship between the intestinal flora, probiotics and diabetes. Particularly, it focuses on the mechanism by which probiotics relieve diabetes by targeting the intestinal microbiota from the aspects of oxidative stress, immune responses, amino acid metabolism, intestinal permeability and short-chain fatty acids.

The force-displacement data during loading and unloading process is obtained by wire rope stretch experiment, which includes 7 different diameters ropes. Based on the data, deformation curves of elastic extension are established. It is shown that the loading and unloading curves compose of the elastic hysteresis loop because the wire rope during unloading shows obvious anelasticity. Analysing the elastic deformation curve characteristics, the values of loading and unloading curve parameters are provided by fitting the experimental data. The loading elastic modulus is constant independent of initial load, and the unloading elastic modulus is non-linear which is related with the maximum load. The elastic modulus is applied to the calculation of freight ropeway bearing cable. Compared the result with the actual project data, it shows that the method considering the wire rope anelasticity can improve the precision of bearing cable calculation.

There are many factors affecting the resistance loss of wire rope over pulley, such as tension of wire rope, diameter of pulley, envelop angle and so on. In this paper, based on the mechanical model of wire rope over pulley, the bending effect of wire rope is considered. And the non-linear equations of exit tension with entry tension, pulley radius, envelop angle and bending stiffness are constructed. At last, the influence of various factors on resistance coefficient is analysed.

Graphene nanosheets (GNS) anchored on Mn3O4 nanoparticles have been successfully synthesized through in situ thermal decomposition of Mn (NO3)2 without the use of any templates or surfactants. Mn3O4 particles were coordinately distributed on the GNS surface. This was achieved by forming 3D nanostructures to avoid detrimental graphene layer stacking, and was characterized using a scanning electron microscope. The Mn3O4/GNS nanocomposite delivers an initial capacity of 1450 mAh g-1 at a current density of 100 mA g-1. It also maintains a high reversible capacity of 930 mAh g-1 even after 60 charge-discharge cycles without showing any apparent decay.

A vector finite element method is presented for the dynamic analysis of cable structures based on the vector form intrinsic finite element (VFIFE) and mechanical properties of suspension cable. Firstly, the suspension cable is discretized into different elements by space points, the mass and external forces of suspension cable are transformed into space points. The structural form of cable is described by the space points at different time. The equations of motion for the space points are established according to the Newton's second law. Then, the element internal forces between the space points are derived from the flexible truss structure. Finally, the motion equations of space points are solved by the central difference method with reasonable time integration step. The tangential tension of the bearing rope in a test ropeway with the moving concentrated loads is calculated and compared with the experimental data. The results show that the tangential tension of suspension cable with moving loads is consistent with the experimental data. This method has high calculated precision and meets the requirements of engineering application.