Lactobacillus fermentum WC2020 increased the longevity of Caenorhabditis elegans via JNK-mediated antioxidant pathway.

Lactobacillus fermentum can exert antiaging effects, but their roles are strain-specific, and little is known about the molecular mechanisms in some strains. This study investigated the antiaging effects of L. fermentum WC2020 (WC2020) isolated from Chinese fermented pickles and the underlying mechanism of the action in Caenorhabditis elegans. WC2020 enhanced the mean lifespan of L1-stage and L4-stage worms by 22.67% and 12.42%, respectively, compared with Escherichia coli OP50 (OP50), a standard food source for C. elegans. WC2020-induced longevity was accompanied by an increase in body length and mitochondrial transmembrane potential and a reduction in lipid accumulation and the production of reactive oxygen species and malondialdehyde. Moreover, WC2020 increased the production of glutathione, superoxide dismutases, and catalases and altered the transcripts of many phenotype-related genes. Furthermore, WC2020-fed jnk-1 rather than akt-2 or pmk-1 loss-of-function mutants showed similar lifespans to OP50-fed worms. Correspondingly, WC2020 significantly upregulated the expression of jnk-1 rather than genes involved in insulin-like, p38 MAPK, bate-catenin, or TGF-beta pathway. Moreover, the increase in body length, mitochondrial transmembrane potential, and antioxidant capability and the decrease in lipid accumulation induced by WC2020 were not observed in jnk-1 mutants. Additionally, WC2020 increased the expression of daf-16 and the proportion of daf-16::GFP in the nucleus, and increased lifespan disappeared in WC2020-fed daf-16 loss-of-function mutants. In conclusion, WC2020 activated the JNK/DAF-16 pathway to improve mitochondria function, reduce oxidative stress, and then extend the longevity of nematodes, suggesting WC2020 could be a potential probiotic targeting JNK-mediated antioxidant pathway for antiaging in food supplements and bioprocessing. PRACTICAL APPLICATION: Aging has a profound impact on the global economy and human health and could be delayed by specific diets and nutrient resources. This study demonstrated that Lactobacillus fermentum WC2020 could be a potential probiotic strain used in food to promote longevity and health via the JNK-mediated antioxidant pathway.

Microbial Composition of Water Kefir Grains and Their Application for the Detoxification of Aflatoxin B1.

Water kefir grains (WKGs), the starter used to develop a traditional beverage named water kefir, consist of a symbiotic mixture of probiotics with diverse bioactivities, but little is known about their abilities to remove mycotoxins that have serious adverse effects on humans and animals. This study investigated the ability of WKGs to remove aflatoxin B1 (AFB1), one of the most toxic mycotoxins, under different settings, and determined the mechanism of absorption mediated by WKGs and the effect of WKGs on the toxicity induced by AFB1 and the reduction in AFB1 in cow milk and tea soups. The results showed the WKGs used herein were dominated by Lactobacillus, Acetobacter, Phenylobacterium, Sediminibacterium, Saccharomyces, Issatchenkia, and Kodamaea. HPLC analysis demonstrated that the WKGs effectively removed AFB1 at concentrations ranging from 1 to 5 µg/mL, pH values ranging from 3 to 9, and temperatures ranging from 4 to 45 °C. Additionally, the removal of AFB1 mainly depended on absorption, which was consistent with the Freundlich and pseudo-second-order kinetic models. Moreover, only 49.63% of AFB1 was released from the AFB1-WKG complex after four washes when the release of AFB1 was non-detectable. Furthermore, WKG treatment caused a dramatic reduction in the mutagenicity induced by AFB1 according to an Ames test and reduced more than 54% of AFB1 in cow milk and three tea soups. These results suggested that WKGs can act as a potential bio-absorbent with a high binding ability to detoxify AFB1 in food and feed via a chemical action step and multi-binding sites of AFB1 absorption in a wide range of scenarios.

Prevention of DSS-induced colitis in mice with water kefir microbiota via anti-inflammatory and microbiota-balancing activity.

Water kefir, a natural and stable functional microbiota system consisting of a symbiotic mixture of probiotics, shows multiple bioactivities but little is known about the effect of water kefir microbiota on the prevention of inflammatory bowel disease (IBD), which is one of the most common intestinal problems and has become a worldwide public health concern. Here, the main objectives of the present study were to investigate the preventative effects of water kefir microbiota, a probiotic consortium mainly consisting of bacteria belonging to Acetobacter, Lactobacillus, and Komagataeibacter and fungi belonging to Saccharomyces and Talaromyces, in a dextran sodium sulfate (DSS)-induced colitis mouse model and unveil the underlying mechanism of the action. Water kefir microbiota effectively improved the disease severity of DSS-induced colitis, including decreased body weight and colon length, increased spleen index and DAI score, and colonic tissue damage. Moreover, water kefir microbiota restored the abnormal expression of tight junction proteins (such as occludin, ZO-1, and claudin-1) and pro-inflammatory and anti-inflammatory cytokines (such as IL-1ß, IL-6, TNF-α, COX-2, iNOS, and IL-10) and inactivated TLR4-MyD88-NF-κB pathway induced by DSS. Water kefir microbiota also improved the composition and metabolism of intestinal microbiota. These findings demonstrated that water kefir microbiota could exert protective roles in the DSS-induced colitis mouse model by reducing inflammation and regulating microbial dysbiosis, which will be helpful for the development of water kefir microbiota-based microbial products as an alternative preventative strategy for IBD.

Beneficial Effects of Bacillus amyloliquefaciens D1 Soy Milk Supplementation on Serum Biochemical Indexes and Intestinal Health of Bearded Chickens.

This study investigated the effects of dietary supplementation with Bacillus amyloliquefaciens D1 (B. amyloliquefaciens D1) on growth performance, serum anti-inflammatory cytokines, and intestinal microbiota composition and diversity in bearded chickens. To investigate the effects of Bacillus amyloliquefaciensa and fermented soy milk, 7-day-old broilers were orally fed different doses of Bacillus amyloliquefaciens D1 fermented soy milk for 35 days, with the unfermented soy milk group as the Placebo group. This study found that B. amyloliquefaciens D1 fermented soy milk improved the intestinal microbiota of broilers, significantly increasing the abundance of beneficial bacteria and decreasing the abundance of harmful bacteria in the gut. B. amyloliquefaciens D1 fermented soy milk also significantly reduced the serum lipopolysaccharide (LPS) content. The body weight and daily weight gain of broilers were increased. In conclusion, the results of this study are promising and indicate that supplementing the diets of bearded chickens with B. amyloliquefaciens D1 fermented soy milk has many beneficial effects in terms of maintaining intestinal microbiota balance and reducing inflammation in chickens.

A micro-carbon nanotube transistor for ultra-sensitive, label-free, and rapid detection of Staphylococcal enterotoxin C in food.

Staphylococcal enterotoxin C (SEC) is an enterotoxin produced by Staphylococcus aureus, which can cause intestinal diseases. Therefore, it is of great significance to develop a sensitive detection method for SEC to ensure food safety and prevent foodborne diseases in humans. A field-effect transistor (FET) based on high-purity carbon nanotubes (CNTs) was used as a transducer, and a nucleic acid aptamer with high affinity was used for recognition to capture the target. The results indicated that the biosensor achieved an ultra-low theoretical detection limit of 1.25 fg/mL in PBS, and its good specificity was verified by detecting target analogs. Three typical food homogenates were used as the solution to be measured to verify that the biosensor had a swift response time (within 5 min after sample addition). An additional study with a more significant basa fish sample response also showed excellent sensitivity (theoretical detection limit of 8.15 fg/mL) and a stable detection ratio. In summary, this CNT-FET biosensor enabled the label-free, ultra-sensitive, and fast detection of SEC in complex samples. The FET biosensors could be further used as a universal biosensor platform for the ultrasensitive detection of multiple biological toxic pollutants, thus considerably stopping the spread of harmful substances.

Isolation and Purification of Bacillus amyloliquefaciens D1 Protease and Its Application in the Fermentation of Soybean Milk to Produce Large Amounts of Free Amino Acids.

In this study, a strain of Bacillus amyloliquefaciens D1, with a notably high production of neutral protease, was isolated from Morchella crassipes. The protease was purified to 10.4-fold with a specific activity of 4542.9 U/mg and 2.7% recovery. The enzyme was purified by 70% (NH4)2SO4 and DEAE-Cellulose-52 column. The estimated molecular mass of the purified protease obtained by SDS-PAGE was approximately 40 kDa. The enzyme was optimally active at pH 6.0 and 50 °C. Furthermore, the maximum hydrolysis rate (Vmax) and apparent Michaelis-Menten constant (Km) values of the purified protease were 8.2 mg/mL and 65.7 µg/(min mL). The enzymatic properties and rapid and efficient purification of Bacillus amyloliquefaciens D1 provide the basis for its potential commercialization and industrial development. Moreover, more essential amino acids, such as isoleucine, leucine, and phenylalanine, would be released when the strain fermented soybean milk, and then a better amino acid profile would be formed in soybean milk. Results suggest that this strain exhibits great potential in fermented soybean milk, and the enzyme could lay a foundation for its industrial application and further research.

Transcriptomic Analysis Revealed Antimicrobial Mechanisms of Lactobacillus rhamnosus SCB0119 against Escherichia coli and Staphylococcus aureus.

Lactic acid bacteria were reported as a promising alternative to antibiotics against pathogens. Among them, Lactobacillus rhamnosus could be used as probiotics and inhibit several pathogens, but its antibacterial mechanisms are still less known. Here, L. rhamnosus SCB0119 isolated from fermented pickles could inhibit bacterial growth or even cause cell death in Escherichia coli ATCC25922 and Staphylococcus aureus ATCC6538, which was mainly attributed to the cell-free culture supernatant (CFS). Moreover, CFS induced the accumulation of reactive oxygen species and destroyed the structure of the cell wall and membrane, including the deformation in cell shape and cell wall, the impairment of the integrity of the cell wall and inner membrane, and the increases in outer membrane permeability, the membrane potential, and pH gradient in E. coli and S. aureus. Furthermore, the transcriptomic analysis demonstrated that CFS altered the transcripts of several genes involved in fatty acid degradation, ion transport, and the biosynthesis of amino acids in E. coli, and fatty acid degradation, protein synthesis, DNA replication, and ATP hydrolysis in S. aureus, which are important for bacterial survival and growth. In conclusion, L. rhamnosus SCB0119 and its CFS could be used as a biocontrol agent against E. coli and S. aureus.

Soy isoflavone-specific biotransformation product S-equol in the colon: physiological functions, transformation mechanisms, and metabolic regulatory pathways.

Epidemiological data suggest that regular intake of soy isoflavones may reduce the incidence of estrogen-dependent and aging-associated disorders. Equol is a metabolite of soy isoflavone (SI) produced by specific gut microbiota and has many beneficial effects on human health due to its higher biological activity compared to SI. However, only 1/3 to 1/2 of humans are able to produce equol in the body, which means that not many people can fully benefit from SI. This review summarizes the recent advances in equol research, focusing on the chemical properties, physiological functions, conversion mechanisms in vitro and vivo, and metabolic regulatory pathways affecting S-equol production. Advanced experimental designs and possible techniques in future research plan are also fully discussed. Furthermore, this review provides a fundamental basis for researchers in the field to understand individual differences in S-equol production, the efficiency of metabolic conversion of S-equol, and fermentation production of S-equol in vitro.

Metagenomic and metabolomic analyses show correlations between intestinal microbiome diversity and microbiome metabolites in ob/ob and ApoE-/- mice.

Obesity and atherosclerosis are the most prevalent metabolic diseases. ApoE-/- and ob/ob mice are widely used as models to study the pathogenesis of these diseases. However, how gut microbes, gut bacteriophages, and metabolites change in these two disease models is unclear. Here, we used wild-type C57BL/6J (Wt) mice as normal controls to analyze the intestinal archaea, bacteria, bacteriophages, and microbial metabolites of ob/ob and ApoE-/- mice through metagenomics and metabolomics. Analysis of the intestinal archaea showed that the abundances of Methanobrevibacter and Halolamina were significantly increased and decreased, respectively, in the ob/ob group compared with those in the Wt and ApoE-/- groups (p < 0.05). Compared with those of the Wt group, the relative abundances of the bacterial genera Enterorhabdus, Alistipes, Bacteroides, Prevotella, Rikenella, Barnesiella, Porphyromonas, Riemerella, and Bifidobacterium were significantly decreased (p < 0.05) in the ob/ob mice, and the relative abundance of Akkermansia was significantly decreased in the ApoE-/- group. The relative abundances of A. muciniphila and L. murinus were significantly decreased and increased, respectively, in the ob/ob and ApoE-/- groups compared with those of the Wt group (p < 0.05). Lactobacillus_ prophage_ Lj965 and Lactobacillus _ prophage _ Lj771 were significantly more abundant in the ob/ob mice than in the Wt mice. Analysis of the aminoacyl-tRNA biosynthesis metabolic pathway revealed that the enriched compounds of phenylalanine, glutamine, glycine, serine, methionine, valine, alanine, lysine, isoleucine, leucine, threonine, tryptophan, and tyrosine were downregulated in the ApoE-/- mice compared with those of the ob/ob mice. Aminoacyl-tRNA synthetases are considered manifestations of metabolic diseases and are closely associated with obesity, atherosclerosis, and type 2 diabetes. These data offer new insight regarding possible causes of these diseases and provide a foundation for studying the regulation of various food nutrients in metabolic disease models.

Kefir microbiota and metabolites stimulate intestinal mucosal immunity and its early development.

Kefir consists of a large number of probiotics, which can regulate or shape the balance of intestinal microbiota, and enhance the host's immune response. Kefir microbiota can shape the mucosal immunity of the body through SCFAs, EPS, polypeptides, lactic acid, and other metabolites and microbial antigens themselves, and this shaping may have time windows and specific pathways. Kefir can regulate antibody SIgA and IL-10 levels to maintain intestinal homeostasis, and its secreted SIgA can shape the stable microbiota system by wrapping and binding different classes of microorganisms. The incidence of intestinal inflammation is closely linked to the development and maturation of intestinal mucosal immunity. Based on summarizing the existing research results on Kefir, its metabolites, and immune system development, this paper proposes to use Kefir, traditional fermented food with natural immune-enhancing components and stable functional microbiota, as an intervention method. Early intervention in the immune system may seize the critical window period of mucosal immunity and stimulate the development and maturation of intestinal mucosal immunity in time.

Comparative analysis of aroma components and quality of Geotrichum candidum after space mutation breeding.

Aim: The aroma-producing strain of Geotrichum candidum GDMCC60675 was taken as the research object, the composition of aroma-producing substances of G. candidum was studied, and the target strains of G. candidum suitable for food additives were screened out by mutagenesis. Methods: Mutants were obtained by space breeding. The colony morphology and cell morphology of the mutant strain were identified, the phylogenetic tree of the two strains was constructed, and the whole-genome sequences of the wild strain and the mutant strain were compared. The aroma components and key odor compounds of the two strains were analyzed and compared by HS-SPME-GC-MS and E-nose detection, and the data were processed by using the relative odor activity value (ROAV) analysis method. Results: A mutant strain of G. candidum was found with different characteristics of aroma production compared with wild-type G. candidum. It was found that its colony morphology and cell morphology were similar. However, it was found that the aroma-producing substances produced by the two strains were different, and the key difference compound was phenyl ethyl alcohol, which also proved that the two strains were different, and the main aroma note was different.

Detoxification of Aflatoxin B1 by a Potential Probiotic Bacillus amyloliquefaciens WF2020.

Microbial degradation is considered as an attractive method to eliminate exposure to aflatoxin B1 (AFB1), the most toxic mycotoxin that causes great economic losses and brings a serious threat to human and animal health, in food and feed. In this study, Bacillus amyloliquefaciens WF2020, isolated from naturally fermented pickles, could effectively degrade AFB1 ranging from 1 to 8 µg/ml, and the optimum temperature and pH value were 37-45°C and 8.0, respectively. Moreover, B. amyloliquefaciens WF2020 was considered to be a potential probiotic due to the synthesis of active compounds, absence of virulence genes, susceptibility to various antibiotics, and enhanced lifespan of Caenorhabditis elegans. Extracellular enzymes or proteins played a major role in AFB1 degradation mediated by B. amyloliquefaciens WF2020 into metabolites with low or no mutagenicity and toxicity to C. elegans. AFB1 degradation by the cell-free supernatant was stable up to 70°C, with an optimal pH of 8.0, and the cell-free supernatant could still degrade AFB1 by 37.16% after boiling for 20 min. Furthermore, B. amyloliquefaciens WF2020 caused a slight defect in fungal growth and completely inhibited AFB1 production when co-incubated with Aspergillus flavus. Additionally, B. amyloliquefaciens WF2020 suppressed the expression of 10 aflatoxin pathway genes and 2 transcription factors (alfR and alfS), suggesting that B. amyloliquefaciens WF2020 might inhibit AFB1 synthesis in A. flavus. These results indicate that B. amyloliquefaciens WF2020 and/or its extracellular enzymes or proteins have a promising potential to be applied in protecting food and feed from AFB1 contamination.

Correlation between the regulation of intestinal bacteriophages by green tea polyphenols and the flora diversity in SPF mice.

Green tea polyphenols (GTP) play an important role in shaping the gut microbiome, comprising a range of densely colonizing microorganisms, including bacteriophages. Previous studies focused on the effect of GTP on the bacteria in the gut microbiota. However, little is known about the role of GTP in the bacteriophage composition of healthy intestines. In this study, SPF male C57BL/6J mice were divided into a polyphenol-free diet group and a tea polyphenol diet group where drinking water was supplemented with 0.1% GTP for 28 days. The ultra-deep metagenomic sequencing of virus-like particle preparations and bacterial 16S rRNA sequencing were performed on mouse stool samples. Changes in the gut bacteriome, bacteriophages, and bacterial-bacteriophage correlations were then compared between the groups. The results revealed an abundance of Firmicutes, a significant decrease in Bacteroidetes, and a significant increase in the ratio of F/B after GTP exposure. The GTP altered the abundance (relative abundance > 1.00%) of Bifidobacterium (regulation rate of 89.78% and the abundance up-regulated by 0.89%) and Akkermansia (regulation rate of 99.70% and the abundance down-regulated by 1.77%). The abundance of Faecalibaculum (regulation rate of 60.17%) increased by 24.38% following GTP treatment. The GTP also altered the abundance of Salmonella phage (regulation rate of 98.64% and the abundance up-regulated by 3.16%) and that of Gordonia_phage_Yakult (regulation rate of 99.99% and the abundance down-regulated by 5.44%). It significantly increased the intestine's lytic phages and reduced the temperate phages by 29.22%. The dominant microorganisms (relative abundance >1.00%) of Bifidobacterium and Dubosiella had a significantly negative relationship with the Faecalibacterium phage and a significantly positive relationship with the Lactobacillus prophage. Exposure to GTP positively promoted changes in the gut bacteriophage community and interaction network in the microbial community of the SPF mice. These findings highlight the importance of "profitable" bacteriophage-bacteria relationships and reveal a potential mechanism of GTP towards the regulation of intestinal flora via intestinal phage communities.

Lipopolysaccharides derived from gram-negative bacterial pool of human gut microbiota promote inflammation and obesity development.

Lipopolysaccharide (LPS) is the major component of the outer membrane of Gram-negative bacteria. It is found from intestinal microbes in the circulatory system and considered a trigger factor for low-grade inflammation in obesity. High-fat diet intake and its related obesity can cause gut microbiota disorder, leading to increased gut permeability, paracellular absorption and transcellular transport of endogenous endotoxin in the cardiovascular system. High-fat diet intake can also increase plasma LPS levels, and causing chronic or "low-grade" inflammation. In this review article, we summarize the recent research advancements on the mechanism of low-grade inflammation and its related obesity. We also propose several approaches that can be used to reduce endogenous endotoxin absorption.Supplemental data for this article is available online at https://doi.org/10.1080/08830185.2021.1996573 .

Corrigendum: Effect of a Humanized Diet Profile on Colonization Efficiency and Gut Microbial Diversity in Human Flora-Associated Mice.

[This corrects the article DOI: 10.3389/fnut.2021.633738.].

Vital roles of Pks11, a highly reducing polyketide synthase, in fungal conidiation, antioxidant activity, conidial cell wall integrity, and UV tolerance of Beauveria bassiana.

Fungal polyketide synthases play important and differential roles in synthesizing secondary metabolites and regulating several cell events, including asexual development, environmental adaptation, and pathogenicity. This study shows the important functions of a highly reducing polyketide synthase, Pks11, in Beauveria bassiana, a filamentous fungal insect pathogen used worldwide for pest biocontrol. The deletion of pks11 led to severe defects in conidial yields on different media and a decrease of 36.27% in the mean thickness of conidial cell wall under normal conditions. Compared with the wild-type, Δpks11 showed higher tolerance to oxidation and increased sensitivity to high temperature during colony growth. Moreover, the lack of pks11 caused a decrease in conidial germination after exposure to UV radiation but did not affect the virulence of B. bassiana against Galleria mellonella larvae via typical cuticle infection. These findings concurred with the alteration in the transcript levels of some phenotype-related genes. These data suggested that pks11 played vital roles in the asexual development, cell wall integrity, and fungal responses to oxidation, high temperature, and UV irradiation of B. bassiana.

Effect of a Humanized Diet Profile on Colonization Efficiency and Gut Microbial Diversity in Human Flora-Associated Mice.

Human flora-associated (HFA) mouse models allow us to design interventions for human disease research to test specific hypotheses and explore the complex commensal microbiome while avoiding the ethical limitations of using humans as models to directly study intestinal flora diseases. However, few studies have investigated the effect of a humanized diet profile (coarse-feed diet; CFD) on colonization efficiency and gut microbial diversity in HFA mice. We tested the colonization efficiency and gut microbial diversity in germ-free Kunming (KM) mice fed a CFD or a purified feed diet (PFD) at 1, 2, and 4 weeks. Although the colonization efficiencies differed significantly (67.50-70.00% vs. 72.69-85.96%) in the HFA mice, the colonization efficiency of the PFD-fed HFA mice (85.96%) was significantly higher than that of the CFD-fed mice (69.61%) at 2 weeks. At 4 weeks, the colonization efficiency of the PFD-fed mice (72.69%) was comparable to that of the CFD-fed mice (70.00%). Additionally, the gut microbial diversity of the CFD-fed HFA mice was similar to that of a human fecal donor. Regarding the Kyoto Encyclopedia of Genes and Genomes colonic microbiota metabolic pathways, the CFD-fed HFA mice showed more similarities to the human donor than to the PFD-fed mice in amino sugar and nucleotide sugar metabolism, biosynthesis of amino acids, carbon metabolism, purine metabolism, and phosphotransferase systems. In conclusion, the humanized diet profiles of the CFD and PFD could help establish human microbiotas in mice. Constructing HFA mouse models fed a CFD for 4 weeks may be useful in researching human-derived intestinal diseases.

Degradation and Detoxification of Aflatoxin B1 by Tea-Derived Aspergillus niger RAF106.

Microbial degradation is an effective and attractive method for eliminating aflatoxin B1 (AFB1), which is severely toxic to humans and animals. In this study, Aspergillus niger RAF106 could effectively degrade AFB1 when cultivated in Sabouraud dextrose broth (SDB) with contents of AFB1 ranging from 0.1 to 4 µg/mL. Treatment with yeast extract as a nitrogen source stimulated the degradation, but treatment with NaNO3 and NaNO2 as nitrogen sources and lactose and sucrose as carbon sources suppressed the degradation. Moreover, A. niger RAF106 still degraded AFB1 at initial pH values that ranged from 4 to 10 and at cultivation temperatures that ranged from 25 to 45 °C. In addition, intracellular enzymes or proteins with excellent thermotolerance were verified as being able to degrade AFB1 into metabolites with low or no mutagenicity. Furthermore, genomic sequence analysis indicated that the fungus was considered to be safe owing to the absence of virulence genes and the gene clusters for the synthesis of mycotoxins. These results indicate that A. niger RAF106 and its intracellular enzymes or proteins have a promising potential to be applied commercially in the processing and industry of food and feed to detoxify AFB1.

Colonization Potential to Reconstitute a Microbe Community in Pseudo Germ-Free Mice After Fecal Microbe Transplant From Equol Producer.

Human intestinal microbiota plays a crucial role in the conversion of isoflavones into equol. Usually, human microbiota-associated (HMA) animal models are used, since it is difficult to establish the mechanism and causal relationship between equol and microbiota in human studies. Currently, several groups have successfully established HMA animal models that produce equol through germ-free mice or rats; however, the HMA model of producing equol through pseudo germ-free mice has not been established. The objective of this study is to establish an HMA mice model for equol production through pseudo germ-free mice, mimicking the gut microbiota of an adult human equol producer. First, a higher female equol producer was screened as a donor from 15 volunteers. Then, mice were exposed to vancomycin, neomycin sulfate, metronidazole, and ampicillin for 3 weeks to obtain pseudo germ-free mice. Finally, pseudo germ-free mice were inoculated with fecal microbiota of the equol producer for 3 weeks to establish HMA mice of producing equol. The results showed that (i) the ability to produce equol was partially transferred from the donor to the HMA mice. (ii) Most of the original intestinal microbiota of mice were eliminated after broad-spectrum antibiotic administration. (iii) The taxonomy data from HMA mice revealed similar taxa to the donor sample, and the species richness returned to the level close to the donor. (iv) The family Coriobacteriaceae and genera Collinsella were successfully transferred from the donor to HMA mice. In conclusion, the HMA mice model for equol production, based on pseudo germ-free mice, can replace the model established by germ-free mice. The model also provides a basis for studying microbiota during the conversion from isoflavones into equol.

Effect of Kefir on Soybean Isoflavone Aglycone Content in Soymilk Kefir.

Kefir is a traditional fermented milk originating in the Caucasus area and parts of Eastern Europe. In this study, the kefir culture, which is modified upon the addition of lactic acid bacteria (LAB) cells, specifically for soymilk kefir fermentation with the highest capacity of isoflavone biotransformation, was successfully produced, and the metagenomics composition of soymilk or milk fermented using these kefir cultures was investigated. The metagenome analysis showed that the microbiota of kefir in M-K (milk inoculated with kefir), SM-K (equal volumes of soymilk and milk inoculated with kefir), and S-K (pure milk inoculated with kefir) were related to the addition of soymilk or not. Furthermore, the HPLC chromatogram revealed that Guixia 2 (Guangzhou, China) may be a good source of soymilk kefir fermentation due to its high isoflavone aglycone content (90.23 ± 1.26 µg/g in daidzein, 68.20 ± 0.74 µg/g in genistein). Importantly, the starter culture created by adding 1.5 g probiotics (Biostime®, Guangzhou, China) to Chinese kefir showed a significant increase in the levels of isoflavone aglycones (72.07 ± 0.53 µg/g in isoflavone aglycones). These results provided insight into understanding the suitable soybean cultivar and starter cultures, which exhibit promising results of isoflavone biotransformation and flavor promotion during soymilk kefir fermentation.