Application of soy protein isolate-xylose conjugates for improving the viability and stability of probiotics microencapsulated by spray drying.

BACKGROUND: Production and consumption of probiotics need to meet many adverse stresses, which can reduce their health-promoting effects on humans. Microencapsulation is an effective technique to improve the biological activity of probiotics and wall materials are also required during encapsulation. Application of Maillard reaction products (MRPs) in probiotic delivery is increasing. RESULTS: This work aims to study the effects of soy protein isolate (SPI)-xylose conjugates heated at different times on the viability and stability of probiotics. SPI-xylose MRPs formed after heat treatment based on changes in the browning intensity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Fourier transform infrared spectroscopy. After heat treatment, α-helix and ß-sheet contents of SPI-xylose mixture shifted from 11.3% and 31.3% to 6.4-11.0% and 31.0-36.9%, respectively, and the thermal stability slightly changed. During spray drying, except for MRP240@LAB, probiotic viability was higher in the MRP-based probiotic microcapsules (21.36-25.31%) than in Mix0@LAB (20.17%). MRP-based probiotic microcapsules had smaller particle sizes (431.1-1243.0 nm vs. 7165.0 nm) and greater intestinal digestion tolerance than Mix0@LAB. Moreover, the MRP-based probiotic microcapsules showed better storability than Mix0@LAB and adequate growth and metabolism capacity. CONCLUSION: SPI-xylose Maillard reaction products are a promising wall material for probiotics microencapsulation, which can improve bacterial survivability during spray drying and enhance bacterial gastrointestinal digestion resistance. This study sheds light on preparing probiotic microcapsules with superior properties by spray drying. © 2023 Society of Chemical Industry.

Metatranscriptomics reveals the gene functions and metabolic properties of the major microbial community during Chinese Sichuan Paocai fermentation.

Chinese Sichuan Paocai (CSP) is one of the world's best-known fermented vegetables with a large presence in the Chinese market. The dynamic microbial community is the main contributor to Paocai fermentation. However, little is known about the ecological distribution and functional importance of these community members. In this study, metatranscriptomics was used to comprehensively explore the active microbial community members and key transcripts with significant functions in the Paocai fermentation process. Enterobacter, Leuconostoc, and Lactobacillus dominated the three-fermentation stages (Pre-, Mid- and Lat-), respectively. Carbon metabolism was the most abundant pathway. GH (glycoside hydrolase) and GT (lycosyl transferase) were the two most highly expressed carbohydrate-active enzymes. The most highly differentially expressed genes were grouped in the biosynthesis of amino acids, followed by glycolysis. Meta-pathways in the Sichuan Paocai fermentation ecosystem were reconstructed, Lactobacillaceae and Enterobacteriaceae were the two most important metabolic contributors. In addition, the nrfA and nirB were two genes referred to distinct nitrite reductase enzymes and 9 specialized genes, such as eclo, ron and ent were expressed to produce autoinducer 2 (AI-2) kinase in response to population density. The present study revealed functional enzymes and meta-pathways of the active microbial communities, which provide a deeper understanding of their contribution to CSP products.

Interactions between Lactobacillus plantarum NCU116 and its environments based on extracellular proteins and polysaccharides prediction by comparative analysis.

Lactic acid bacteria (LAB) play a significant role in food industry and artisan fermented-food. Most of the applicable LABs were commonly obtained from natural fermented food or human gut. And Lactobacillus plantarum NCU116 was screened from a LAB-dominated traditional Chinese sauerkraut (TCS). In order to comprehend the interaction between NCU116 and its environments, comparative genomics were performed to identify genes involved in extracellular protein biosynthesis and secretion. Four secretory pathways were identified, including Sec and FPE pathways, holins and efflux ABC transporter system. Then 348 potential secretory proteins were identified, including 11 alpha-amylases responsible for degradation of macromolecules, and 8 mucus binding proteins which attribute to adherence to intestine epithelium. Besides, EPS clusters of NCU116 (EPS116) were identified and analyzed by comparing to other strains, which suggested a novel genotype of EPS clusters. These findings could be critical to extend the application of NCU116 in food and pharmaceuticals industries.

Stability of potential prophages in commercial strain Lactobacillus plantarum NCU116 under various stressors.

Genetic stability of bacterium as a starter culture is vital for product quality in fermentation industry. The commercial strain Lactobacillus plantarum NCU116 widely used in fruit and vegetable fermentation was induced with various stressors to investigate the stability of potential prophages. PHAge Search Tool (PHAST) identified three potential prophages in bacterial genome. By spectrophotometric analysis, mitomycin C (MMC), lactic acid, and bile salt were found to inhibit the growth of L. plantarum NCU116 while ethanol and hydrogen peroxide had no notable impacts. Transcriptions of four phage-synthesizing genes (phaR, phacap, phaada, phatail) and four phage-resistant genes (cas116, helR, hsd1, hsd2) under stressors were investigated by quantitative reverse transcription PCR. MMC was found to most significantly upregulated transcriptions of phage-synthesizing genes, followed by lactic acid and bile salt. By transmission electron microscopy, no virus particles from the lysates of strain NCU116 treated by MMC were observed, corresponding to the result that no phage nucleic acids could be extracted from the supernatants of strain NCU116 treated by MMC. This study suggested that no prophages could be induced from L. plantarum NCU116 by strong inducer MMC, indicating its genetic stability, which supports the comprehensive application of strain NCU116 in industry without causing fermentation failure.

Evaluation and comparison of the microbial communities and volatile profiles in homemade suansun from Guangdong and Yunnan provinces in China.

BACKGROUND: Suansun is a traditional salt-free fermented bamboo shoot product that has been widely consumed as a cooking ingredient in south China for centuries. The aim of this study was to evaluate and compare the microbial and metabolic diversity in samples of two kinds of suansun, namely Guangdong suansun (GD) and Yunnan suansun (YN), using high-throughput sequencing (HTS) and headspace solid-phase microextraction-gas chromatograph-mass spectrometry (HS-SPME/GC-MS), respectively, and then to assess the influence of environmental factors on the microbial communities. RESULTS: The results showed that Lactobacillus and Serratia were the most abundant bacterial genera in both the GD and YN groups. For the fungi, Pichia, Candida, and Debaryomyces were the major genera in the GD group, whereas Pichia and Zygosaccharomyces were the dominant genera in the YN group. The canonical correlation analysis (CCA) results demonstrated that three environmental factors - temperature, longitude, and altitude - play a more important role in affecting the microbial community composition of suansun than physical and chemical factors. The fugal community composition was more influenced by environmental factors than the bacterial community. The volatile profile of the GD group differed from that of the YN group, and the difference was mainly reflected in the relative alcohol, aldehyde, ester, and aromatic compound content. CONCLUSIONS: This study provided insights into the microbial and metabolic profiles of suansun products. The findings might be useful for the improvement and standardization of suansun production. © 2020 Society of Chemical Industry.

GhCPK33 Negatively Regulates Defense against Verticillium dahliae by Phosphorylating GhOPR3.

Verticillium wilt, caused by the soil-borne fungus Verticillium dahliae, is a destructive vascular disease in plants. Approximately 200 dicotyledonous plant species in temperate and subtropical regions are susceptible to this notorious pathogen. Previous studies showed that jasmonic acid (JA) plays a crucial role in plant-V. dahliae interactions. V. dahliae infection generally induces significant JA accumulation in local and distal tissues of the plant. Although JA biosynthesis and the associated enzymes have been studied intensively, the precise mechanism regulating JA biosynthesis upon V. dahliae infection remains unknown. Here, we identified the calcium-dependent protein kinase GhCPK33 from upland cotton (Gossypium hirsutum) as a negative regulator of resistance to V. dahliae that directly manipulates JA biosynthesis. Knockdown of GhCPK33 by Agrobacterium tumefaciens-mediated virus-induced gene silencing constitutively activated JA biosynthesis and JA-mediated defense responses and enhanced resistance to V dahliae Further analysis revealed that GhCPK33 interacts with 12-oxophytodienoate reductase3 (GhOPR3) in peroxisomes. GhCPK33 phosphorylates GhOPR3 at threonine-246, leading to decreased stability of GhOPR3, which consequently limits JA biosynthesis. We propose that GhCPK33 is a potential molecular target for improving resistance to Verticillium wilt disease in cotton.

Bacterial community dynamics and physicochemical characteristics in natural fermentation of jiang-shui, a traditional food made in northwest China.

BACKGROUND: Chinese jiang-shui is fermented food without salt. In consideration of the few studies of the bacterial community and dynamics of jiang-shui fermentation, the aim of this study was to investigate the microbial diversity and dynamics of jiang-shui fermentation using high-throughput sequencing (HTS), denaturing gradient gel electrophoresis (DGGE) and the culture-dependent method. RESULTS: The culture-dependent method showed that pathogenic bacteria (Staphylococcus aureus, Salmonella and Escherichia coli) and yeast mainly existed in the early stage and lactic acid bacteria dominated until the end stage. HTS and DGGE revealed that Serratia marcescens, Serratia sp., Lactobacillus curvatus, Lactococcus lactis, uncultured bacterium and Bacillus thuringiensis started the fermentation, followed by the middle stage with Lactococcus lactis, Weissella sp. and Bacillus arthracis as the predominant species. The end stage was characterized with Lactobacillus pentosus, Weissella cibaria and Weissella sp. as the major bacteria. CONCLUSIONS: The results showed that genera Serratia, Lactococcus, Weissella and Lactobacillus dominated the whole process of jiang-shui fermentation. This study provided a good analysis of the bacterial changes of jiang-shui fermentation, and future studies should consider the relationships among the sensory characteristics, microbial communities and metabolites. © 2019 Society of Chemical Industry.

The synthesis of benzimidazoles via a recycled palladium catalysed hydrogen transfer under mild conditions.

An efficient synthesis of benzimidazoles was developed by virtue of a recycled palladium catalyzed hydrogen transfer. The reaction can be carried out smoothly under mild conditions to give rise to a variety of benzimidazoles with good to excellent yields. The palladium catalyst could be recovered easily and reused six times with great catalytic activity.

Muscle Fiber Characteristics and Transcriptome Analysis in Slow- and Fast-Growing Megalobrama amblycephala.

Growth is an important trait in aquaculture that is influenced by various factors, among which genetic regulation plays a crucial role. Megalobrama amblycephala, one of the most important freshwater species in China, exhibits wide variations in body mass among individuals of the same age within the same pool. But the molecular mechanisms underlying wide variation in body mass remain unclear. Here, we performed muscle histological and transcriptome analysis of muscle tissues from Fast-Growing (FG) and Slow-Growing (SG) M. amblycephala at the age of 4 months old (4 mo) and 10 months old (10 mo) to elucidate its muscle development and growth mechanism. The muscle histological analysis showed smaller diameter and higher total number of muscle fibers in FG compared to SG at 4 mo, while larger diameter and total number of muscle fibers were detected in FG at 10 mo. The transcriptome analysis of muscle tissue detected 1171 differentially expressed genes (DEGs) between FG and SG at 4 mo, and 718 DEGs between FG and SG at 10 mo. Furthermore, 44 DEGs were consistently up-regulated in FG at both 4 mo and 10 mo. Up-regulated DEGs in FG at 4 mo were mainly enriched in the pathways related to cell proliferation, while down-regulated DEGs were significantly enriched in cell fusion and muscle contraction. Up-regulated DEGs in FG at 10 mo were mainly enriched in the pathways related to cell proliferation and protein synthesis. Therefore, these results provide novel insights into the molecular mechanism of M. amblycephala muscle growth at different stages, and will be of great guiding significance to promote the fast growth of M. amblycephala.

Characterization, mechanism and in vivo validation of Helicobacter pylori antagonism by probiotics screened from infants' feces and oral cavity.

Helicobacter pylori (H. pylori) infection is a major cause of chronic gastritis, intestinal metaplasia, and gastric carcinoma. Antibiotics, the conventional regimen for eliminating H. pylori, cause severe bacterial resistance, gut dysbiosis and hepatic insufficiency. Here, fifty lactic acid bacteria (LAB) were initially screened out of 266 strains obtained from infants' feces and oral cavity. The antagonistic properties of these 50 strains against H. pylori were investigated. Based on eight metrics combined with principal component analysis, three LAB with probiotic function and excellent anti-H. pylori capacity were affirmed. Combining dynamics test, metabolite assays, adhesion assays, co-cultivation experiments, and SEM and TEM observations, LAB were found to antagonize H. pylori by causing coccoid conversion and intercellular adhesion. Furthermore, it was found that LAB antagonized H. pylori by four pathways, i.e., production of anti-H. pylori substances, inhibition of H. pylori colonization, enhancement of the gastric mucosal barrier, and anti-inflammatory effect. In addition, animal model experiments verified that the final screened superior strain L. salivarius NCUH062003 had anti-H. pylori activity in vivo. LAB also reduced IL-8 secretion, ultimately alleviating the inflammatory response of gastric mucosa. Whole genome sequencing (WGS) data showed that the NCUH062003 genome contained the secondary metabolite biosynthesis gene cluster T3PKS. Furthermore, NCUH062003 had a strong energy metabolism and substance transport capacity, and produced a small molecule heat stable peptide (SHSP, 4.1-6.5 kDa). Meanwhile, LAB proved to be safe through antibiotic susceptibility testing and CARD database comparisons.

Two-stage fermentation of corn and soybean meal mixture by Bacillus subtilis and Lactobacillus acidophilus to improve feeding value: optimization, physicochemical property, and microbial community.

Microbial treatment can reduce the antinutritional factors and allergenic proteins in corn-soybean meal mixture (CSMM), but the role of the microbial community in hypoallergenicity and digestibility during the fermentation process remains unclear. Therefore, the fermentation strains of Bacillus and LAB were determined, and the compatibility and fermentation process of two-stage solid fermentation composite bacteria were optimized, and the dynamic changes in physicochemical property and microbial community during two-stage fermentation were investigated. Results showed that Bacillus subtilis NCUBSL003 and Lactobacillus acidophilus NCUA065016 were the best fermentation combinations. The optimal fermentation conditions were inoculum 7.14%, solid-liquid ratio of 1:0.88 and fermentation time of 74.30 h. The contents of TI, ß-conglycinin and glycinin decreased significantly after fermentation. Besides, TCA-SP, small peptides and FAA increased. Bacillus and Lactobacillus were the main genera. Pathogenic bacteria genera were inhibited effectively. This study suggests the feasibility of two-stage fermentation in improving the nutrient values and safety of the CSMM. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-023-01426-7.

Isolation and characterization of potential Lactobacillus acidophilus strains isolated from pig feces.

The aim of this study was to isolate and characterize Lactic Acid Bacteria (LAB) from 16 feces samples of pig farm, and to evaluate the probiotic potential of these isolates as potential oral probiotic candidates. The selection process was based on the isolation, identification, and a series of experiments for the selection of appropriate candidates with beneficial properties. The results demonstrated that most of LAB showed relatively strong resistance to pH 2.5 and high bile salts (1%), and had good survival in simulated gastric and intestinal juice. 9 isolates displayed antimicrobial activities against Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Escherichia coli, Salmonella typhimurium, Pseudomonas aeruginosa and Enterobacter sakazakii. Almost all isolates were sensitive to ampicillin, chloramphenicol, vancomycin and amoxicillin, and most of isolates exhibited resistance against tetracycline and vancomycin. The adhesion rates of LAB varied greatly. The results of the study suggested that the Lactobacillus acidophilus NCUA065001 have the important functional property of probiotic candidates to enhance gut integrity and could considered to be the potential antibiotic alternatives in the pig feed industry.

The microbial succession and their correlation with the dynamics of flavor compounds involved in the natural fermentation of suansun, a traditional Chinese fermented bamboo shoots.

Suansun is a typical fermented bamboo shoot product widely consumed in southern China. Complex microorganisms are involved during spontaneous fermentation of suansun. However, little is known about the succession and roles of these microbiota throughout the fermentation process, especially for the functional microbiota involved in aroma formation. Thus, in this study, the dynamic changes in physicochemical parameters, flavor compounds, and microbiota community during the natural production of suansun were investigated. The principal component analysis (PCA) results showed that physicochemical parameters and flavor compounds varied substantially during the 30-day fermentation. Moreover, the microbial flora exhibited a dynamic change from the acid-sensitive genera of Lactococcus, Leuconostoc, Weissella, and Enterobacter to an acid-resistant genus of Lactobacillus due to lactic acid and acetic acid accumulation. Furthermore, bidirectional orthogonal partial least squares (O2PLS) indicated that four bacterial genera, namely, Lactobacillus, Clostridium_sensu_stricto_1, Enterobacter, and Leuconostoc, showed strong connections with various characteristic flavors, thus playing crucial roles in the unique flavor formation of suansun. These results may shed light on the development of effective starter cultures for the industrial manufacture of high-quality suansun products.

Screening, Safety Evaluation, and Mechanism of Two Lactobacillus fermentum Strains in Reducing the Translocation of Staphylococcus aureus in the Caco-2 Monolayer Model.

Staphylococcus aureus is a common commensal of humans, and its translocation from gastrointestine to peripheral organs and tissues could cause severe diseases and complications. This study focuses on the screening and characterization of Lactobacillus strains with significant inhibitory effect on the translocation of S. aureus through Caco-2 monolayers. First, strains with strong affinity for mucin and Caco-2 cells were obtained, via microtiter plate assay and adhesion assay, respectively. Obtained bacteria were further tested for their inhibitory effects on the growth of S. aureus by well diffusion assay. Subsequently, two strains preincubated with Caco-2 monolayers were found to inhibit the translocation of S. aureus CMCC26003 by 80.95 and 43.96%, respectively, via the transcellular translocation assay. These two strains were then identified to be Lactobacillus fermentum NCU3087 and L. fermentum NCU3088. Second, the mechanism of inhibition was investigated by analyzing the relative concentration of tight junction proteins and proinflammatory cytokines of Caco-2 cells, by Western blot and enzyme-linked immunosorbent assay, respectively. Results showed that both NCU3087 and NCU3088 significantly attenuated the degradation of occludin, claudin-1, ZO-1, and JAM-1 and suppressed the secretion of interleukin 6 and tumor necrosis factor-α induced by S. aureus, to different extent. Moreover, two Lactobacillus strains could barely translocate the Caco-2 monolayers, had no hemolytic activity, displayed strong resistance to gastrointestinal fluids, and were sensitive or moderate sensitive to nine clinically relevant antibiotics. Collectively, this study identified two Lactobacillus strains with significant inhibitory effect on the translocation of S. aureus, and their safeness for application was evaluated, thereby providing potential solutions for prevention of S. aureus and prophylaxis of related diseases.

Comparison of microbial communities and physiochemical characteristics of two traditionally fermented vegetables.

Guangxi Suansun (Guangxi SS) and Guangxi Suancai (Guangxi SC) are two kinds of traditionally fermented vegetables consumed as cooking ingredients in Guangxi Province, China, for thousands of years. However, little is known about their microbial communities as well as the differences between them. Thus, this study was aimed to investigate and compare the physicochemical indexes as well as the bacterial and fungal profiles of Guangxi SS and Guangxi SC. Results showed that the titratable acidity, lactic acid and acetic acid content in SS were significantly higher than those in SC, while the salinity of SS was significantly lower than that in SC, and the nitrite contents in all samples were are far lower than the limit of nitrite contents in fermented vegetables. Firmicutes, Proteobacteria, Ascomycota and Basidiomycota were the main phyla observed in both SS and SC samples. Lactobacillus, Serratia, Stenotrophomonas and Lactococcus were the major bacterial genera in both SS and SC samples, the predominant fungal genera in SS group were Kazachstania, Debaryomyces and Pichia, while the major genera in SC group were Kazachstania, Debaryomyces and Nakaseomyces. At the species level, Lactobacillus acetotolerans, Pichia kudriavzevii, Debaryomyces hansenii, Pichia norvegensis, Kazachstania exigua and Kazachstania humilis were the major species observed in SS, while L. delbrueckii, L. fermentum, L. aviarius, and Pichia kudriavzevii and Debaryomyces hansenii were the predominant species in SC. Salinity was found to be more strongly correlated to the bacterial and fungal communities of both SS and SC than other physicochemical factors (pH, the titratable acidity, lactic acid and acetic acid content). This study provided detailed insight into the microbial communities of Guangxi SS and Guangxi SC, and the findings may help understand the microbial structures of Chinese traditional fermented vegetables.

Copper-Catalyzed Geminal Difunctionalization of Terminal Alkynes by Splitting Sulfonyl Hydrazones into Two Parts.

A copper(I)-catalyzed geminal difunctionalization of terminal alkynes was developed via a carbene migratory insertion and an addition of a sulfonyl anion to the triple bond at the same time under mild reaction conditions, providing a variety of vinyl sulfones with good yields and excellent stereoselectivities.