Abstract:Bile acids are the main end products of cholesterol metabolism in the body, playing an important regulatory role in the digestion and metabolism of nutrients, especially lipids. The intestine is the main organ for digestion and absorption, as well as an important site for bile acids to achieve liver-intestinal circulation. Its function is closely related to the interaction between bile acids as signaling molecules and their receptors. Focusing on the metabolism and regulatory mechanism of bile acids, the author reviews the roles and underlying mechanisms of bile acids and their receptors in regulating intestinal barriers, immune function, and gut microbiota. The aim is to provide reference for related research on improving intestinal health by bile acids and their receptors.

Abstract:The CRISPR/Cas system can recognize foreign DNA, cut the target, and block or silence the expression of the foreign gene. Due to its precise targeting function, the CRISPR/Cas system has been developed as a highly efficient gene editing tool. With high specificity and operability, the CRISPR/Cas system has been combined with biosensors to improve the detection specificity and efficiency of the sensors. This review introduces the characteristics and applications of CRISPR/Cas system, analyzes the latest progress in the application of CRISPR/Cas system-based biosensors in the detection of foodborne pathogens, and summarizes and prospects the application of such biosensors.

Abstract:Objective The aim of this study is to explore the effects of dextran with a small molecular weight (SDex) on the viability of intestinal microorganisms.Method The probiotic effects of laboratory-designed SDex containing only α-1,6 glycosidic bonds was evaluated by in vitro fermentation with nine typical intestinal strains, including strains from the genera Bifidobacterium and Lactobacillus, as well as species such as Escherichia coli and Enterococcus faecalis. The microbial proliferation and organic acid production during fermentation were compared among SDex, commercial dextran 20 (Dex 20), and isomalto-oligosaccharides (IMOs).Result SDex and IMOs significantly promoted the proliferation of probiotics such as Bifidobacterium and Lactobacillus as well as the production of organic acids, while Dex 20 showed little effects. SDex did not promote the growth of the conditionally pathogenic bacterium Escherichia coli but showed a less positive effect on the neutral bacterium Enterococcus faecalis than IMOs. Neither SDex nor IMOs significantly affected the lactic acid production. However, SDex promoted acetic acid production while SDex enhanced formic acid production by probiotics.Conclusion SDex demonstrates selective probiotic effects of promoting the proliferation of Bifidobacterium and Lactobacillus while exerting little effect on Escherichia coli, which indicates SDex is essential for replenishing existing prebiotics.

Abstract:Objective This study aims to evaluate the antibiotic resistance and transmission risk of Enterococcus faecalis (E. faecalis) in the pork production chain.Method A total of 300 samples were collected from the surface of live pigs, carcasses, and the environment in 2 pig farms, 1 slaughterhouse, and 2 markets in Hangzhou, Zhejiang. The strain identification, antibiotic resistance phenotypes, resistance genotypes, and virulence genotypes were performed by 16S rDNA sequences, broth microdilution method (MIC), and PCR. In addition, multilocus sequencing typing (MLST) and conjugational transfer tests were conducted.Result The 125 isolates of E. faecalis showed high resistance to antibiotics such as tiamulin, tetracycline, and doxycycline, and the isolates from pig farms showed higher antibiotic resistance rates than those from the slaughterhouse and markets. Ten antibiotic resistance genes were detected in the 125 strains of E. faecalis, and the detection rates of optrA, tet(M), tet(L), and erm(B) were higher. The detection rate of antibiotic resistance genes followed the trend of pig farm > slaughterhouse > market. Among the virulence genes tested, efaA was the most prevalent, followed by gelE, asa1, ace, cylA, hyl, and esp. Overall, the detection rate of virulence genes was the highest in pig farms, moderate in the slaughterhouse, and the lowest in markets. MLST results indicated that horizontal transmission was the main mechanism for the prevalence of optrA in pig-derived E. faecalis in Hangzhou, Zhejiang. The results of conjugation transfer tests showed that optrA in 12 pig-derived strains could be transferred to the recipient strain JH2-2 by plasmid conjugation.Conclusion The antibiotic resistance and transmission risk of E. faecalis in the pork production chain are assessed, which provides a theoretical basis for ensuring the safety of meat products.

Abstract:Objective Functional constipation is a chronic gastrointestinal disorder caused by multiple factors, with a relatively high and increasing prevalence in China. Previous studies have found that Bacillus coagulans (B. coagulans)CCFM1041 can alleviate symptoms of functional constipation in mice.Method A double-blind, randomized controlled clinical trial was conducted to evaluate the clinical efficacy of CCFM1041 in relieving functional constipation. A total of 73 participants were observed for changes in constipation-related indicators, serum levels of gastrointestinal regulatory neurotransmitters, and gut microbiota before and after the intervention.Result After taking CCFM1041, 84.2% of participants experienced increased bowel movement frequency, and 94.7% of participants showed improved stool consistency. Compared with the control group, CCFM1041 significantly raised the mass concentrations of motilin (MTL) and gastrin (GAS) in the serum and effectively regulated the gut microbiota. The correlation analysis indicates that CCFM1041 may exert the effect by modulating gut microbiota.Conclusion CCFM1041 can effectively alleviate symptoms of functional constipation in participants. The research results provide scientific evidence for the application of B. coagulans CCFM1041 in constipation relief and offer support for screening probiotics effective in alleviating constipation.

Abstract:Objective The effects of the transcription regulation system on menaquinone-7 (MK-7) production in Bacillus subtilis (B. subtilis) 168 were studied by excavating transcription regulators.Method With the signal molecule PhrG pentapeptide, RapG, and the transcription regulator DegU, a PhrG-RapG-DegU regulatory system was successfully constructed in B. subtilis 168 to fine-tune the expression of key genes, so that the expression of key genes in the MK-7 synthesis pathway can respond to the changes in cell density.Result The production of MK-7 in strain BS06 reached 102.47 mg/L through this system.Conclusion Fine tuning of quorum sensing can effectively improve the MK-7 production in B. subtilis 168.

Abstract:Objective Acid precipitation, a common method in plant protein processing, can efficiently achieve protein enrichment and removal of non-protein components (including off-flavor substances) on a large scale. This study aims to explore the causes for the abnormal increases in aldehydes of soymilk after acid precipitation.Method Changes in volatile compounds of soymilk before and after acid precipitation were analyzed by gas chromatography-mass spectrometry (GS-MS). Soymilk components were fractionated by centrifugation, and the contributions of different fractions to the increases in aldehydes were investigated. The changes in the molar concentrations of lipid hydroperoxides before and after acid precipitation were determined, and a molar ratio between these changes and the alterations in aldehyde content was established. Subsequently, the effects of critical factors (acids, acid precipitation temperature, and Fe²⁺ concentration) influencing lipid hydroperoxide decomposition on the mass fraction of hexanal in the product after acid precipitation were systematically examined.Result The changes in the mass fractions of alcohols, acids, ketones, and furans in soymilk after acid precipitation were as anticipated. The mass fractions of aldehydes (including hexanal, nonanal, and trans-2-heptenal) increased, while the mass molar concentration of lipid hydroperoxides demonstrated a remarkable decrease. The centrifugation results showed that the oil phase in the upper layer was the main source for the increases in the mass fractions of aldehydes, with the largest contribution. The subsequent analysis revealed that the ratio of lipid hydroperoxide molar concentration decrease to aldehyde molar concentration increase was 5∶1 following acid treatment, indicating partial decomposition of the former into aldehydes, which is consistent with previous findings. The analysis of influencing factors showed that compared with hydrochloric acid, citric acid or malic acid for acid precipitation reduced the decomposition of lipid hydroperoxides and increased the formation of hexanal. However, higher acid precipitation temperature and higher Fe²⁺ concentration promoted the decomposition of lipid hydroperoxides and the increase of hexanal accumulation.Conclusion During acid precipitation, the abnormal increases in aldehydes exhibited a distinct distribution pattern compared with other off-flavor substances. Replacing hydrochloric acid with citric acid or malic acid, lowering the precipitation temperature, and controlling the Fe²⁺ concentration within the system can effectively inhibit the decomposition of lipid hydroperoxides, thereby reducing the formation of aldehydes.

Abstract:Objective This study aims to identify honey authenticity by terahertz (THz) spectroscopy.Method Different honey samples were analyzed by stable isotope ratio mass spectrometry (IRMS), and the results served as a reference for terahertz spectroscopy. The sample requirements and analytical conditions of terahertz spectroscopy were verified by comparing the results of these two methods.Result Liquid honey samples exhibited no discernible absorption peaks. However, significant differences in refractive index were observed among samples. The presence of water in liquid honey, particularly at high levels, affected the refractive index, potentially leading to misjudgment. After removal of water from liquid honey via freeze-drying, terahertz attenuated total reflection was employed for identification. The freeze-dried honey samples still showed no discernible absorption peaks but exhibited distinct refractive index variations, being grouped into two categories: one with minor refractive index fluctuations and average values ranging from 2.04 to 2.29, and the other with considerable fluctuations and average values ranging from 1.03 to 2.22.Conclusion Consistent results from both analytical methods demonstrate that terahertz spectroscopy can be employed for authenticity identification of honey samples.

Abstract:Objective This study aims to explore the effect of palmitic acid (PA)-rich diet on prostate cancer (PCa) in mice.Method The TCGA database and immunohistochemistry (IHC) were used to analyze the differential expression of fatty acid synthase (FASN) between the PCa tissue and adjacent normal tissue. In addition, twelve nude mice were subcutaneously implanted with LNCaP cells and assigned into two groups. One group was fed a PA-rich diet, while the other group was fed a normal control (NC) diet. Twelve mice overexpressing the C-myc gene (C-myc^T) were assigned into two groups and fed the same diets as the nude mice. The growth of PCa was analyzed by tumor mass measurement, HE staining, and Ki67 staining. Furthermore, the expression of epithelial-mesenchymal transition (EMT)-related proteins, such as E-cadherin, N-cadherin, and transforming growth factor (TGF)-β1, was determined by IHC and Western blot. Finally, relevant gene data were retrieved from the TCGA database and analyzed to explore the relationship among CD36, TGF-β1, and PCa.Result The expression of FASN was markedly higher in the PCa tissue than in the adjacent normal tissue, and there was a positive correlation between FASN expression and the progression of PCa. In nude mice and C-myc^T mice, a PA-rich diet significantly promoted the progression of PCa, induced the EMT process, and activated the TGF-β1 signal. The expression of TGF-β1 was significantly associated with the decreased survival rate of individuals diagnosed with PCa. The expression of CD36 was positively correlated with TGF-β1. Highly expressed TGF-β1, TGF-βR1, and CD36 were risk factors for PCa, and TGF-β1 was an independent risk factor for PCa.Conclusion The PA-rich diet promoted the tumor growth, activated the TGF-β1 signal pathway, and induced EMT in the mouse model of PCa.

Abstract:Objective The aim of this study is to explore the effects of edible fungal polysaccharides on the proliferation and metabolism of Lactobacillus rhamnosus GG (LGG).Method Flarnmulina velutipes polysaccharides (FVP), Tremella fueiformls polysaccharides (TFP), Auricularia auricula polysaccharides (AAP), lentinan (LNT), Hericium erinaceus polysaccharides (HEP), and Poria cocos polysaccharides (PCP) were selected as carbon sources for the culture of LGG. The viable bacterial count and the concentration of short-chain fatty acids (SCFAs) were determined. Two polysaccharides were selected from the six polysaccharides and added in different mass ratios to the LGG-fermented milk, and then the pH and textural properties of the fermented milk samples were determined.Result The TFP and PCP groups had the higher polysaccharide utilization rate and the highest production of SCFAs. When the two polysaccharides were added into the fermented milk, different mass ratios of polysaccharides had little effect on the acid production of the fermented milk. The hardness, viscosity, cohesiveness, and adhesion of the fermented milk decreased when the mass ratio of TFP to PCP was higher (P<0.05), and the viscosity of the fermented milk reached the maximum when the mass ratio of TFP to PCP was 1∶1.Conclusion TFP and PCP can promote the proliferation and metabolism of LGG. The taste of LGG-fermented milk can be improved by adding TFP and PCP in a mass ratio of 1∶1.

Abstract:Objective This study aimed to investigate the effects of Lactobacillus plantarum (L. plantarum) fermentation on the metabolism of flavonoids and organic acids in mango juice.Method Six strains of L. plantarum were selected for fermentation of the juice samples from two mango varieties ‘Tainong' and ‘Guiqi'. The mass concentrations of organic acids and flavonoids in the juice samples were determined by ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Orthogonal partial least squares discriminant analysis was employed to compare the metabolites in the mango juice samples fermented with different strains.Result L. plantarum fermentation significantly enhanced the total antioxidant capacity (T-AOC) of mango juice. Furthermore, the composition of organic acids changed significantly, with a significant increase in the mass concentration of lactic acid and decreases of over 99% in the mass concentration of malic acid and over 10% in the mass concentration of citric acid. In addition, a total of 142 flavonoids were detected in both unfermented mango juice and fermented mango juice, with 42 significantly differential metabolites identified after fermentation. The KEGG pathway enrichment analysis of differential metabolites showed that L. plantarum fermentation promoted the enrichment of products related to flavonoid biosynthesis and anthocyanin biosynthesis pathways. Compared with unfermented juice, the fermented juice samples of both mango varieties showed significantly increased mass concentrations of phlorizin, 3,4,2',4',6'-pentahydroxychalcone, apigenin, cyanidin-3,5-O-diglucoside, and pelargonidin-3-O-glucoside and significantly decreased mass concentrations of eriodictyol, luteolin, pinocembrin, pinobanksin, phloretin, quercetin, and 3,4,2',4',6'-pentahydroxychalcone-4'-O-glucoside.Conclusion L. plantarum fermentation significantly affects the levels of organic acids, anthocyanins, and flavonoids in mango juice, enhancing the mass concentrations of bioactive components in fermented mango juice.

Abstract:Objective This study aims to investigate the application of a novel thermophilic lytic polysaccharide monooxygenase (LPMO) in the efficient cellulase-mediated hydrolysis of lignocellulosic substrates.Method The thermophilic enzyme mgLPMO10, derived from a compost metagenome, was heterologously expressed in Escherichia coli (E. coli). Its expression level was improved by modifying the signal peptide of the expression vector and induction conditions (temperature, IPTG concentration, and induction time). The enzyme was then characterized in terms of the optimal temperature, optimal pH, thermal stability, pH stability, and the effects of metal ions on its enzyme activity. Subsequently, the synergistic hydrolysis efficiency of mgLPMO10 with two thermophilic endoglucanases (TnCelB and DtCelB) at different molar ratios toward phosphoric acid-swollen cellulose (PASC) and other cellulose substrates was evaluated at 70 ℃.Result The coding sequence of the pelB signal peptide in the expression vector significantly improved the expression level of mgLPMO10. After optimization (induction with 0.50 mmol/L IPTG at 28 ℃ for 18 h), the expression level reached 12.9 mg/L, representing a 90% increase compared with the pre-optimized level. The optimal reaction conditions of mgLPMO10 were 70 ℃ and pH 6.0, and this enzyme showed the maximum enzyme activity of 25.5 U/g. After adding 5 mmol/L Mn²⁺, its enzyme activity increased by 75%. When synergistically hydrolyzing PASC with TnCelB or DtCelB, the enzyme exhibits synergy degrees of 123% and 138%, respectively. Moreover, mgLPMO10 exhibited significant promoting effects on other cellulose substrates.Conclusion The combination of mgLPMO10 and thermophilic endoglucanases can significantly enhance cellulose hydrolysis efficiency under high-temperature conditions. The study provides an important enzyme and a theoretical basis for the development of high-temperature lignocellulose biorefining processes.

Abstract:Objective Horse milk has a high nutritional value and a unique flavor, which are closely related to its complex microbial composition, and different geographical locations and climates will affect the microbial community structure of horse milk. This study aims to explore the effects of different geographical locations on the microbial community structure in horse milk.Method Metagenomic technology was employed to analyze the microbial diversity of 14 batches of horse milk samples from 3 different regions in northern Xinjiang, and the functional genes were excavated.Result A total of 26 species of microorganisms were identified, belonging to 9 phyla, 14 classes, 25 orders, 28 families, and 45 genera. The dominant species were Lactobacillus helveticus, Acetobacter pasteurianus, Lactobacillus crispatus, and Brettanomyces bruxellensis. There were 11 common genera in the samples from different regions, with the dominant genus being Lactobacillus and the dominant species being L. helveticus, while the relative abundance was different in the samples from different regions. Compared with fresh horse milk, fermented horse milk showed increased relative abundance of Lactobacillus, Acetobacter, Lentilactobacillus, and Brettanomyces, and decreased relative abundance of Streptococcus, Terrisporobacter, Staphylococcus, Enterococcus, Campylobacter, and Klebsiella. The annotation in the KEGG database revealed 23 547 genes, which were involved in carbohydrate metabolism, amino acid metabolism, translation, membrane transport, energy metabolism, nucleotide metabolism, and cofactor and vitamin metabolism. The annotation results with the eggNOG database showed that the bacteria in horse milk contained abundant genes related to replication, recombination and repair, carbohydrate transport and metabolism, amino acid transport and metabolism, etc. Glucoside hydrolase genes (12 155 454) and glycosyl transferase genes (1 130 442) were abundant in horse milk bacteria, followed by auxiliary activity genes (184 638) and carbohydrate esterase genes (465 134).Conclusion Different geographical locations can affect the microbial community structure in horse milk, and the microbial community structure varies among different types of horse milk.

Abstract:Objective This study aims to reveal the interactions between collagen and volatile compounds during the cooking process.Method The relative molecular weight, particle size, solubility, secondary structure, and surface hydrophobicity of collagen and the droplet size of the collagen emulsion were studied. Furthermore, the binding ability of collagen to volatile compounds of the aqueous and emulsion systems of collagen was measured.Result As the heating time was extended, the triple-helix structure of collagen was destroyed and the polypeptide chains were degraded, which were accompanied with the decrease in surface hydrophobicity, increase in solubility, and heat-induced aggregation. The binding ability of collagen to volatile compounds gradually decreased with the extension of heating time. In the aqueous system, the release of volatile compounds was promoted after heating for 4 h. In the emulsion system, collagen exhibited a higher binding ability to volatile compounds and did not exhibit the effect of promoting release of volatile compounds even after heating for 4 h.Conclusion The triple-helix structure of collagen is beneficial to the binding to volatile compounds and the heating time has a negative correlation with the binding ability of collagen to volatile compounds. Collagen exhibits the strongest binding ability to volatile compounds in the case of short-term heating and the coexistence with lipids.

Abstract:Objective Staphylococcus aureus protein A (SPA) is widely used for the purification of monoclonal antibodies due to its high specificity for immunoglobulin G (IgG). However, the high costs of commercial media have limited the further development of SPA. To address this issue, this paper aims to prepare a high-performance affinity chromatography medium based on SPA as the substitute of introduced affinity chromatography media.Method SPA expressed by the recombinant strain was purified via Ni²⁺ affinity chromatography and then coupled to the Rigose HF matrix to prepare an self-prepared affinity chromatography medium.Result The self-prepared medium had a static saturated adsorption capacity of 115.5 mg/mL (based on the volume of the medium) and a dynamic adsorption capacity of 58.6 mg/mL (based on the volume of the medium). When being used for the separation and purification of monoclonal antibody samples, the self-prepared affinity chromatography medium achieved the purity of over 95%, with a recovery rate of no less than 90%. The medium demonstrated comparable or superior alkali tolerance, reusability, and purification performance for monoclonal antibody samples to commercial media.Conclusion A recombinant strain expressing SPA is successfully constructed, and the optimal preparation conditions of the self-prepared affinity chromatography medium are determined by optimization of the coupling conditions. The self-prepared affinity chromatography medium demonstrates research significance and commercial application value to a certain extent.

Abstract:Objective This study aims to investigate the immunoregulatory mechanisms of lactoferrin formula and its four nutritional components (α-lactalbumin, N-acetylneuraminic acid, yeast β-glucan, and lactoferrin) in RAW264.7 macrophages.Method M1 and M2 macrophage polarization were induced by lipopolysaccharides (LPS) and interleukin (IL)-4, respectively. Quantitative real-time PCR, flow cytometry, and Western blot were employed to analyze macrophage polarization and related signaling pathways.Result Lactoferrin formula enhanced the phagocytic activity of macrophages and dose-dependently inhibited M1 polarization while promoting M2 polarization within the range of 10~50 μg/mL. A combination of four nutritional components exhibited similar effects, with greater effects than the three-component combination. Additionally, the lactoferrin formula upregulated antigen-presenting molecules and antioxidant enzyme activity. It modulated macrophage polarization by inhibiting the mitogen-activated protein kinase (MAPK)/nuclear factor (NF)-κB pathway and activating the protein kinase B (Akt)/signal transducer and activator of transcription 6 (STAT6) pathway, thereby alleviating inflammation.Conclusion Lactoferrin formula exerts immunoregulatory effects by modulating macrophage polarization.

Abstract:Objective This study aims to assess the effects of Fusarium on the malting process and malt quality of barley, screen out the microorganism with inhibitory activity against Fusarium, and explore the potential of this microorganism for degrading deoxynivalenol (DON).Method The mycotoxin-producing strain was isolated from domestic Dan’er barley contaminated with pathogenic fungi, and the interaction between the pathogen and the host was studied. The germination rate, water sensitivity, and DON mass fraction were compared between the barley infected for 5 d and standard barley. The DON mass fraction, activities of protease, β-glucanase, and amylase, and physicochemical indicators (free amino acids, soluble nitrogen, and β-glucan mass concentration) of malt produced from infected barley were measured. The microorganism capable of inhibiting the viability of the mycotoxin-producing strain was screened from barley and malt. The effects of the freeze-dried cell-free supernatant powder (cfsP) of the microorganism on the growth and development of the mycotoxin-producing strain were studied, and the antifungal effects and mechanisms of cfsP at different mass fractions (0.020~0.100 μg/kg) were tested. During the steeping stage, the antifungal strain was inoculated and its regulatory effects on the DON mass concentration and the processing performance of malt were evaluated.Result The mycotoxin-producing strain was identified as Fusarium culmorum (F. culmorum). After being infected with this strain for 5 d, the barley showed a germination rate decrease of 45.8%, significantly increased water sensitivity, and the DON accumulation reaching 276 μg/kg. Compared with the control group, the malt produced from infected barley showed an increase of 6 times in DON mass fraction, a decreased activity of amylase, increased activities of total protease and β-glucanase, increased levels of free amino acids and soluble nitrogen, and a reduced mass concentration of β-glucan. Lactobacillus plantarum (L. plantarum) NM-6 capable of inhibiting F. culmorum was screened out. The L. plantarum NM-6 cfsP with a mass concentration ≥ 0.020 μg/kg changed the mycelial morphology of F. culmorum, and the cfsP with a mass concentration ≥ 0.050 μg/kg inhibited the growth of the germination tube of conidia. The cfsP with a mass concentration of 0.100 μg/kg completely blocked the growth of the strain, and its antifungal activity was stronger at low pH conditions. When 2×10? CFU/g of L. plantarum NM-6 was inoculated into barley during the steeping stage, the DON mass fraction in the malt was approximately 50% of that of the control malt, and the filtration and solubility properties of the malt were significantly improved.Conclusion The Fusarium species infecting Dan’er barley was identified as F. culmorum and the microorganism with inhibitory activity against F. culmorum was identified as L. plantarum NM-6. The infection of F. culmorum led to a decrease in the germination ability and accumulation of DON in barley. L. plantarum NM-6 can efficiently inhibit the spore germination and mycelial growth of F. culmorum by secreting acid-tolerant active substances. Its application in the malting process can simultaneously achieve toxin control and quality improvement, providing a biocontrol strategy for the safe application of barley exposed to fungal infections.