In vitro effects of velvet antler water extracts from Formosan Sambar deer and red deer on barrier integrity in Caco-2 cell.

Background: The mucus integrity and abnormal inflammatory response are the crucial biomarker of inflammatory bowel disease (IBD). Velvet antler (VA) has been used as traditional Chinese medicines for many years. Anti-inflammatory property was demonstrated via suppression of cyclooxygenase-2 and cytokines protein expression. And it has further proved to promote wound healing in streptozotocin-induced diabetic rats model. The aforementioned functionalities of VA extracts may be associated with the treatment of IBD. Thus, the aim of present study was to evaluate the effect of velvet antler water extracts form Formosan Sambar deer (Rusa unicolor swinhoei, SVAE) and red deer (Cervus elaphus, RVAE) on the barrier function and to investigate the possible mechanism using in vitro model. Methods: Human colonic epithelial cell models (Caco-2) were co-cultured with various concentrations of both SVAE and RVAE (250-500 µg mL-1) in dextran sulfate sodium (DSS)-induced colitis model. Trans-epithelial electrical resistance (TEER) value and the macromolecule permeability of Fluorescein isothiocyanate (FITC)-labeled dextran were measured to evaluate the integrity of monolayer of Caco-2. Western blotting was performed for analysis of protein expressions of occludin, Zonula occludens-1 (ZO-1), claudin-1, claudin-2 and myosin light chain kinase (MLCK). The cytotoxicity was conducted by MTT assay. Results: Results indicated that both SVAE and RVAE could enhance integrity of monolayer in dextran sulfate sodium (DSS)-induced colonic epithelial cell model (Caco-2) through reducing the decline of trans-epithelial electrical resistance (TEER) and macromolecule permeability at the concentration of 250 µg mL-1. RVAE significantly increased the expression of tight junction proteins (occludin and ZO-1) while SVAE significantly reduced the activity of MLCK (P < 0.05.). Elevated C-C chemokine ligand 20 (CCL20) production suggested that both SVAE and RVAE could enhance the repair of epithelial cell. Besides, MTT assay revealed that both extracts showed no cytotoxicity. Conclusion: Thus, SVAE and RVAE supplementation may attenuate barrier damage by enhancing the occludin and ZO-1 protein expression, decreasing MLCK expression, promoting the CCL20 production. In the future, animal study is needed for further confirmation.

A novel immobilized cell system involving Taiwanese kefir microorganisms and sugar cane pieces for fermented milk production.

The objective of this study was to develop a novel immobilized system using kefir lactic acid bacteria and sugar cane pieces for the production of fermented milk. Lactobacillus kefiranofaciens HL1, Lactobacillus kefiri HL2, Leuconostoc mesenteroides HL3, and Lactococcus lactis HL4 were isolated from Taiwanese kefir grains and immobilized on pieces of sugar cane using adsorption. Scanning electron micrographs of the cell-immobilized sugar cane pieces (CISCP) showed that the microorganisms were embedded within the porous structures of the sugar cane pieces. During 28 cycles of repeated batch fermentation, viable cells on both sugar cane pieces and fermented products were maintained at 10 log cfu/g (cfu/mL). Random amplified polymorphic DNA PCR analysis revealed that Leu. mesenteroides HL3 (29-43%) and Lc. lactis HL4 (31-49%) were predominant on the CISCP, and the fermented samples had 79% Lc. lactis HL4. When tracking fermentation parameters, the data on the microbial, chemical, and physical properties of the fermented milk suggested that the CISCP had stable fermentative ability over the course of successive fermentations. We found an enhancement of the acid-producing ability of CISCP as the number of fermentations increased, with a significant growth in titratable acidity from 0.65 to 0.81% by the end.

Improving effect of a probiotic mixture on memory and learning abilities in d-galactose-treated aging mice.

The aim of this study was to evaluate the antiaging effect of a probiotic mixture using an in vivo mouse model in which aging was induced with d-galactose. Results of the Morris water maze test indicated that long-term administration of the probiotic mixture improved memory and learning abilities and ameliorated the apoptosis pattern in the hippocampus of aging mice treated with d-galactose. An antioxidation experiment indicated that administration of the probiotic mixture could restore activities of the antioxidant enzymes superoxide dismutase and catalase and inhibit the production of malondialdehyde. The antioxidant-related proteins nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) were upregulated in liver after treatment of d-galactose-treated aging mice with probiotics. Finally, the probiotic treatment did affect the production of short-chain fatty acids in d-galactose-treated aging mice. Our results highlighted a possible antioxidative effect triggered by short-chain fatty acids that contributed to improving the memory and learning abilities following treatment with the probiotic mixture and suggested that probiotics could serve as a therapy to modulate physiological function.

Effects of heat, cold, acid and bile salt adaptations on the stress tolerance and protein expression of kefir-isolated probiotic Lactobacillus kefiranofaciens M1.

Lactobacillus kefiranofaciens M1 is a probiotic strain isolated from Taiwanese kefir grains. The present study evaluated the effects of heat, cold, acid and bile salt adaptations on the stress tolerance of L. kefiranofaciens M1. The regulation of protein expression of L. kefiranofaciens M1 under these adaptation conditions was also investigated. The results showed that adaptation of L. kefiranofaciens M1 to heat, cold, acid and bile salts induced homologous tolerance and cross-protection against heterologous challenge. The extent of induced tolerance varied depending on the type and condition of stress. Proteomic analysis revealed that 27 proteins exhibited differences in expression between non-adapted and stress-adapted L. kefiranofaciens M1 cells. Among these proteins, three proteins involved in carbohydrate metabolism (triosephosphate isomerase, enolase and NAD-dependent glycerol-3-phosphate dehydrogenase), two proteins involved in pH homeostasis (ATP synthase subunits AtpA and AtpB), two stress response proteins (chaperones DnaK and GroEL) and one translation-related protein (30S ribosomal protein S2) were up-regulated by three of the four adaptation treatments examined. The increased synthesis of these stress proteins might play a critical protective role in the cellular defense against heat, cold, acid and bile salt stresses.

Effects of Lactobacillus paracasei 01 fermented milk beverage on protection of intestinal epithelial cell in vitro.

BACKGROUND: Intestinal protection is an important function of probiotics. However, there is no evidence that the potential probiotic Lactobacillus paracasei 01 is able to improve intestinal function. In the present study, the protective effect and underlying mechanisms of L. paracasei 01 on intestinal epithelial cell in vitro were investigated. RESULTS: A fermented milk beverage containing L. paracasei 01 at 3 × 10(8) CFU mL(-1) was made from skim milk powder, glucose, sucrose and isomaltooligosaccharide. Results indicated that L. paracasei 01 fermented milk beverage could reduce macromolecule permeability caused by dextran sodium sulfate-induced damage. Besides, 50 mL L(-1) fermented milk beverage with live L. paracasei 01 in the cell medium could increase intestinal epithelial cell Caco-2 growth, decrease lipopolysaccharide/tumor necrosis factor-α (TNF-α)/interferon-γ (IFN-γ)-induced Caco-2 cell death and chemokine CCL-20 production. The protecting mechanisms involve promoting intestinal epithelial cell growth and intestinal epithelial integrity to strengthen the intestinal barrier against chemical and inflammatory stimuli-induced damage. Inhibition of inflammatory cytokines and chemokine such as IFN-γ, TNF-α and CCL-20 also contributes to the beneficial effects of this product on epithelial function. CONCLUSION: Lactobacillus paracasei 01 fermented milk beverage may be a new functional food with intestinal protecting effects in vitro.

Effects of a novel encapsulating technique on the temperature tolerance and anti-colitis activity of the probiotic bacterium Lactobacillus kefiranofaciens M1.

Lactobacillus kefiranofaciens M1 (M1) has been shown to possess many different beneficial health effects including anti-colitis activity. The purpose of this study was to develop a novel and easily scaled-up encapsulating technique that would improve the temperature tolerance of the bacterium and reduce the sensitivity of the organism to gastrointestinal fluid. A mixture of sodium alginate, gellan gum and skim milk powder was used as a coating material to entrap M1. The M1 gel was then directly freeze dried in order to dehydrate the covering and form microcapsules. The viable cell numbers of M1 present only dropped ten folds after the freeze-drying encapsulation process. The viable cell counts remained constant at 5 × 10(7) CFU/g after heating from 25 °C to 75 °C and holding at 75 °C for 1 min. The viable cell counts were reduced to 10(6) CFU/g and 10(5) CFU/g after 8-week storage at 4 °C and subsequent heat treatment with simulated gastrointestinal fluid test (SGFT) and bile salts, respectively. The effect of encapsulated M1 on the organism's anti-colitis activity was evaluated using the dextran sodium sulfate (DSS) induced colitis mouse model. An in vivo study indicated that administration of heat treated encapsulated M1 was able to ameliorate DSS-induced colitis producing a significant reduction in the bleeding score and an attenuation of inflammatory score. These findings clearly demonstrate that encapsulation of M1 using this novel technique is able to provide good protection from temperature changes and SGFT treatment and also does not affect the organism's anti-colitis activity.

Optimizing Production of Two Potential Probiotic Lactobacilli Strains Isolated from Piglet Feces as Feed Additives for Weaned Piglets.

Two probiotic strains, Lactobacillus johnsonii x-1d-2 and Lactobacillus mucosae x-4w-1, originally isolated from piglet feces, have been demonstrated to possess antimicrobial activities, antibiotic resistances and interleukin-6 induction ability in RAW 267.4 macrophages in our previous study. These characteristics make L. johnsonii x-1d-2 and L. mucosae x-4w-1 good candidates for application in feed probiotics. In this study, soybeal meal, molasses and sodium acetate were selected to optimize the growth medium for cultivation of L. johnsonii x-1d-2 and L. mucosae x-4w-1. These two strains were then freeze-dried and mixed into the basal diet to feed the weaned piglets. The effects of L. johnsonii x-1d-2 and L. mucosae x-4w-1 on the growth performance and fecal microflora of weaned piglets were investigated. The results showed that the bacterial numbers of L. johnsonii x-1d-2 and L. mucosae x-4w-1 reached a maximum of 8.90 and 9.30 log CFU/mL, respectively, when growing in optimal medium consisting of 5.5% (wt/vol) soybean meal, 1.0% (wt/vol) molasses and 1.0% (wt/vol) sodium acetate. The medium cost was 96% lower than the commercial de Man, Rogosa and Sharpe medium. In a further feeding study, the weaned piglets fed basal diet supplemented with freeze-dried probiotic cultures exhibited higher (p<0.05) body weight gain, feed intake, and gain/feed ratio than weaned piglets fed basal diet. Probiotic feeding also increased the numbers of lactobacilli and decreased the numbers of E. coli in the feces of weaned piglets. This study demonstrates that L. johnsonii x-1d-2 and L. mucosae x-4w-1 have high potential to be used as feed additives in the pig industry.

Effect of acid adaptation on the environmental stress tolerance of three strains of Vibrio parahaemolyticus.

Three strains of Vibrio parahaemolyticus (690, BCRC 13023, and BCRC 13025), involved in foodborne outbreaks in Taiwan, were subjected to acid adaptation at pH 5.5 for 90 min. The effects of acid adaptation on the tolerance of V. parahaemolyticus to various environmental stresses, including heat (47°C), cold (4°C and -20°C), ethanol (8%), high salt (20% NaCl), and hydrogen peroxide (20 ppm) were examined. Results showed that acid adaptation increased the thermal tolerance of the three test strains of V. parahaemolyticus, while it did not affect their cold tolerance. Acid adaptation also increased the ethanol tolerance in V. parahaemolyticus 690 and BCRC 13025, but not in BCRC 13023. Differences in the tolerance to high salts were noted among the three strains after prior acid adaptation. However, these acid-adapted V. parahaemolyticus strains were more susceptible to hydrogen peroxide than their nonadapted controls. These findings demonstrated that acid adaption responses of V. parahaemolyticus varied among strains and types of stress challenge.

Exploring the Preventive and Therapeutic Mechanisms of Probiotics in Chronic Kidney Disease through the Gut-Kidney Axis.

Gut dysbiosis contributes to deterioration of chronic kidney disease (CKD). Probiotics are a potential approach to modulate gut microbiota and gut-derived metabolites to alleviate CKD progression. We aim to provide a comprehensive view of CKD-related gut dysbiosis and a critical perspective on probiotic function in CKD. First, this review addresses gut microbial alterations during CKD progression and the adverse effects associated with the changes in gut-derived metabolites. Second, we conduct a thorough examination of the latest clinical trials involving probiotic intervention to unravel critical pathways via the gut-kidney axis. Finally, we propose our viewpoints on limitations, further considerations, and future research prospects of probiotic adjuvant therapy in alleviating CKD progression. Enhancing our understanding of host-microbe interactions is crucial for gaining precise insights into the mechanisms through which probiotics exert their effects and identifying factors that influence the effectiveness of probiotics in developing strategies to optimize their use and enhance clinical outcomes.

Investigating the Efficacy of Kidney-Protective Lactobacillus Mixture-Containing Pet Treats in Feline Chronic Kidney Disease and Its Possible Mechanism.

Microbiota-based strategies are a novel auxiliary therapeutic and preventative way of moderating chronic kidney disease (CKD). Lactobacillus mixture (Lm) was previously demonstrated to exert a renal-protective function in the CKD mice model. The efficacy of probiotics in pet foods is a relatively new area of study, and thus verifying the potential health benefits is necessary. This study evaluated the efficacy of Lm treats in feline CKD and elucidated the mechanisms underlying host-microbe interactions. CKD cats (2 and 3 stages) were administrated probiotic pet treats daily (10 g) for 8 weeks. The results demonstrated that during the eight weeks of Lm administration, creatinine was reduced or maintained in all cats with CKD. Similarly, gut-derived uremic toxin (GDUT), indoxyl sulfate (IS), were potential clinical significance in IS after Lm treatment (confidence intervals = 90%). The life quality of the cats also improved. Feline gut microbiome data, metabolic functional pathway, and renal function indicator analyses revealed the possible mechanisms involved in modulating CKD feline microbial composition. Further regulation of the microbial functions in amino acid metabolism after Lm administration contributed to downregulating deleterious GDUTs. The current study provides potential adjuvant therapeutic insights into probiotic pet foods or treats for pets with CKD.

Multiomics Strategy Reveals the Mechanism of Action and Ameliorating Effect of Deer Velvet Antler Water Extracts on DSS-Induced Colitis.

Velvet antler is a precious traditional Chinese medicine used for thousands of years. This study investigated the anti-colitis effects of water extracts of Formosan sambar deer (SVAE) and red deer (RVAE) to identify the possible mechanisms and the bioactive compounds using a dextran sulfate sodium (DSS)-induced colitis mouse model. The mechanism of action and the ameliorating effects of SVAE and RVAE on DSS-induced colitis were evaluated using a mouse model. Ultra-high performance liquid chromatography-mass/mass and gas chromatography-mass/mass were applied to identify the bioactive components of the SVAE and RVAE water extracts. The results revealed that both high-dose SVAE and RVAE could ameliorate the symptoms of colitis due to reduced systemic inflammatory responses, enhanced intestinal barrier integrity by restoration of tight junction proteins, and improved gut dysbiosis. The potentially bioactive components of SVAE and RVAE were identified as small molecules (<3 kDa). Further identification by untargeted metabolomics analysis suggested that l-carnitine, hypoxanthine, adrenic acid, creatinine, gamma-aminobutyric-lysine, oleic acid, glycine, poly-γ-glutamic acid, and eicosapentaenoic acid in VAWEs might be involved in ameliorating the symptoms of colitis. This study provided evidence for the potential usage of SVAE and RVAE as anti-colitis agents.

A Pilot Study of Nephrogenic Probiotics to Further Improve an Already Stabilized Graft Function After Kidney Transplantation.

BACKGROUND: The majority of kidney recipients are a subset of chronic kidney disease. Our previous study demonstrated that the combination of Lactobacillus plantarum and Lactobacillus paracasei (Lm) had the highest clearance ability of uremic toxins and improved kidney function in a mouse model. This study aimed to evaluate Lm in improving graft function, effects on immunosuppressants, and safety in transplant recipients. METHODS: We retrospectively reviewed 24 patients. Twelve of them take Lm regularly; we compared the creatinine measurements and estimated glomerular filtration rate 3 months before and after Lm using a 2-tailed Wilcoxon matched-pairs signed-rank test while also evaluating the drug level of immunosuppressants and infection events. Other 12 patients who do not have Lm for evaluation of laboratory calibration and compared the proportion of improving creatinine using Fisher's exact test. RESULTS: The creatinine decreased by 0.06 mg/dL (P = .02), and the estimated glomerular filtration rate increased by 3.1 mL/min/1.73 m2 (P = .03) after Lm supplementation. This pilot study revealed the association of higher incidence (odds ratio 13.3, 95% CI 1.64-77.2, P = .01) of decreasing creatinine in transplant recipients using Lm. Furthermore, results showed a trend of higher trough levels of tacrolimus and sirolimus, which might provide a potential strategy for reducing the dosages of immunosuppressants. CONCLUSION: Our findings revealed an association between a higher incidence of decreasing creatinine in kidney transplant recipients using Lm, which may also provide a potential strategy for reducing the acquired dosages of immunosuppressants.

In vitro ruminal fermentation and cow-to-mouse fecal transplantations verify the inter-relationship of microbiome and metabolome biomarkers: potential to promote health in dairy cows.

Introduction: There are differences in the gut microbiome and metabolome when the host undergoes different physical or pathological conditions. However, the inter-relationship of microbiome and metabolome biomarkers to potentially promote the health of dairy cows needs to be studied. Further, the development of next-generation probiotics for dairy cattle health promotion has not been demonstrated. Objective: In the present study, we identified the microbiome and metabolome biomarkers associated with healthy cows. Methods: We analyzed the relationships of the ruminal microorganism profile and metabolites between healthy and mastitis lactating dairy cows. The roles of bacterial biomarker were further verified by in vitro fermentation and cow-to-mouse fecal microbiota transplantation (FMT). Results: Two species, Ruminococcus flavefaciens and Bifidobacterium longum subsp. longum, and six rumen metabolites were positively correlated with healthy cows by Spearman's correlation analysis. Through in vitro ruminal fermentation, inoculating R. flavefaciens and B. longum subsp. longum showed the upregulation of the levels of putrescine, xanthurenic acid, and pyridoxal in the mastitis ruminal fluid, which confirmed the inter-relationships between these microbiota and metabolites associated with healthy cows. Further, we verified the role of R. flavefaciens and B. longum subsp. longum in promoting health by FMT. The administration of R. flavefaciens and B. longum subsp. longum reduced the death rate and recovered the bodyweight loss of germ-free mice caused by FMT mastitis feces. Discussion: We provided evidence that the bacterial biomarkers alter downstream metabolites. This could indirectly indicate that the two bacterial biomarkers have the potential to be used as next-generation probiotics for dairy cattle, although it needs more evidence to support our hypothesis. Two species, R. flavefaciens and B. longum subsp. longum, with three metabolites, putrescine, xanthurenic acid, and pyridoxal, identified in the ruminal fluid, may point to a new health-promoting and disease-preventing approach for dairy cattle.

Effect of the Velvet Antler of Formosan Sambar Deer (Cervus unicolor swinhoei) on the Prevention of an Allergic Airway Response in Mice.

Two mouse models were used to assay the antiallergic effects of the velvet antler (VA) of Formosan sambar deer (Cervus unicolor swinhoei) in this study. The results using the ovalbumin- (OVA-) sensitized mouse model showed that the levels of total IgE and OVA-specific IgE were reduced after VA powder was administrated for 4 weeks. In addition, the ex vivo results indicated that the secretion of T helper cell 1 (Th1), regulatory T (Treg), and Th17 cytokines by splenocytes was significantly increased (P < 0.05) when VA powder was administered to the mice. Furthermore, OVA-allergic asthma mice that have been orally administrated with VA powder showed a strong inhibition of Th2 cytokine and proinflammatory cytokine production in bronchoalveolar fluid compared to control mice. An increase in the regulatory T-cell population of splenocytes in the allergic asthma mice after oral administration of VA was also observed. All the features of the asthmatic phenotype, including airway inflammation and the development of airway hyperresponsiveness, were reduced by treatment with VA. These findings support the hypothesis that oral feeding of VA may be an effective way of alleviating asthmatic symptoms in humans.

Investigation of microorganisms involved in biosynthesis of the kefir grain.

The purpose of this study was to understand the significance of each microorganism in grain formation by evaluating their microbial aggregation and cell surface properties during co-aggregation of LAB and yeasts together with an investigation of biofilm formation. Non-grain forming strains from viili were also evaluated as a comparison. Results indicated that the kefir grain strains, Lactobacillus kefiranofaciens and Saccharomyces turicensis possess strong auto-aggregation ability and that Lactobacillus kefiri shows significant biofilm formation properties. Significant co-aggregation was noted when S. turicensis and kefir LAB strains (Lb. kefiranofaciens and Lb. kefiri) were co-cultured. Most of the tested LAB strains are hydrophilic and had a negative charge on their cell surface. Only the kefir LAB strains, Lb. kefiranofaciens HL1 and Lb. kefiri HL2, possessed very high hydrophobicity and had a positive cell surface charge at pH 4.2. In contrast, the LAB and yeasts in viili did not show any significant self-aggregation or biofilm formation. Based on the above results, we propose that grain formation begins with the self-aggregation of Lb. kefiranofaciens and S. turicensis to form small granules. At this point, the biofilm producer, Lb. kefiri, then begins to attach to the surface of granules and co-aggregates with other organisms and components in the milk to form the grains. On sub-culturing, more organisms attach to the grains resulting in grain growth. When investigated by scanning electron microscopy, it was found that short-chain lactobacilli such as Lb. kefiri occupy the surface, while long-chain lactobacilli such as Lb. kefiranofaciens have aggregated towards the center of the kefir grains. These findings agree with the above hypothesis on the formation of grains. Taken together, this study demonstrates the importance of cell surface properties together with fermentation conditions to the formation of grains in kefir.

Adaptive acid tolerance response of Vibrio parahaemolyticus as affected by acid adaptation conditions, growth phase, and bacterial strains.

Vibrio parahaemolyticus strain 690 was isolated from gastroenteritis patients. Its thermal and ethanol stress responses have been reported in our previous studies. In this study, we further investigated the effects of various acid adaptation conditions including pH (5.0-6.0) and time (30-90 min) on the acid tolerance in different growth phases of V. parahaemolyticus 690. Additionally, the adaptive acid tolerance among different V. parahaemolyticus strains was compared. Results indicated that the acid tolerance of V. parahaemolyticus 690 was significantly increased after acid adaptation at pH 5.5 and 6.0 for 30-90 min. Among the various acid adaptation conditions examined, V. parahaemolyticus 690 acid-adapted at pH 5.5 for 90 min exhibited the highest acid tolerance. The acid adaptation also influenced the acid tolerance of V. parahaemolyticus 690 in different growth phases with late-exponential phase demonstrating the greatest acid tolerance response (ATR) than other phases. Additionally, the results also showed that the induction of adaptive ATR varied with different strains of V. parahaemolyticus. An increase in acid tolerance of V. parahaemolyticus was observed after prior acid adaptation in five strains (556, 690, BCRC 13023, BCRC 13025, and BCRC 12864), but not in strains 405 and BCRC 12863.

Development of Next-Generation Probiotics by Investigating the Interrelationships between Gastrointestinal Microbiota and Diarrhea in Preruminant Holstein Calves.

(1) Background: We aimed to isolate and identify potential next-generation probiotics (NGP) by investigating the interrelationships between gastrointestinal microbiota and diarrhea in preruminant Holstein calves. (2) Material and methods: Twenty preruminant Holstein calves were divided into healthy and diarrheic groups after the combination outcomes of veterinary diagnosis and fecal scores. The fecal microbiome, plasma cytokines, plasma immunoglobulin (Ig) G and haptoglobin were analyzed. The potential probiotic bacteria were identified by comparing the microbiota difference between healthy and diarrheic calves and correlation analysis with fecal scores and inflammatory markers. The identified bacteria were also isolated for further evaluation for antimicrobial activities and immunoregulatory effects. (3) Results: Microbiota analysis suggested that Ruminococcaceae_UCG_014, Bifidobacterium and Pseudoflavonifractor positively correlated with bovine IgG and negatively correlated with fecal score; inflammatory factors, bovine HP, and IL-8 were classified as beneficial bacteria contributing to the health of the calves. The alternation of gut microbial composition also induced changes in the functional gene enrichment of gut microbiota in calves. The gathering of microbiomic data strongly indicated the possible beneficial effects of Bifidobacterium longum subsp. longum, expected to develop as NGP. After isolation and evaluation of the potential functionality in vitro, two specific bifidobacterial strains demonstrated antimicrobial activities and immunoregulatory effects. (4) Conclusions: The results provide a new probiotic searching approach for preventing gastrointestinal disorders in preruminant calves. Further animal study is necessary to verify the results.

Characterization of exopolysaccharide-producing lactic acid bacteria from Taiwanese ropy fermented milk and their application in low-fat fermented milk.

OBJECTIVE: The aim of this study was to characterize the exopolysaccharides (EPS)-producing lactic acid bacteria from Taiwanese ropy fermented milk (TRFM) for developing a clean label low-fat fermented milk. METHODS: Potential isolates from TRFM were selected based on the Gram staining test and observation of turbid suspension in the culture broth. Random amplified polymorphic DNA-polymerase chain reaction, 16S rRNA gene sequencing, and API CHL 50 test were used for strain identification. After evaluation of EPS concentration, target strains were introduced to low-fat milk fermentation for 24 h. Fermentation characters were checked: pH value, acidity, viable count, syneresis, and viscosity. Sensory evaluation of fermented products was carried out by 30 volunteers, while the storage test was performed for 21 days at 4°C. RESULTS: Two EPS-producing strains (APL15 and APL16) were isolated from TRFM and identified as Lactococcus (Lc.) lactis subsp. cremoris. Their EPS concentrations in glucose and lactose media were higher than other published strains of Lc. lactis subsp. cremoris. Low-fat fermented milk separately prepared with APL15 and APL16 reached pH 4.3 and acidity 0.8% with a viable count of 9 log colony-forming units/mL. The physical properties of both products were superior to the control yogurt, showing significant improvements in syneresis and viscosity (p<0.05). Our low-fat products had appropriate sensory scores in appearance and texture according to sensory evaluation. Although decreasing viable cells of strains during the 21-day storage test, low-fat fermented milk made by APL15 exhibited stable physicochemical properties, including pH value, acidity, syneresis and sufficient viable cells throughout the storage period. CONCLUSION: This study demonstrated that Lc. lactis subsp. cremoris APL15 isolated from TRFM had good fermentation abilities to produce low-fat fermented milk. These data indicate that EPS-producing lactic acid bacteria have great potential to act as natural food stabilizers for low-fat fermented milk.

Subspecies Classification and Comparative Genomic Analysis of Lactobacillus kefiranofaciens HL1 and M1 for Potential Niche-Specific Genes and Pathways.

(1) Background: Strains HL1 and M1, isolated from kefir grains, have been tentatively identified, based on their partial 16S rRNA gene sequences, as Lactobacillus kefiranofaciens. The two strains demonstrated different health benefits. Therefore, not only the genetic factors exerting diverse functionalities in different L. kefiranofaciens strains, but also the potential niche-specific genes and pathways among the L. kefiranofaciens strains, should be identified. (2) Methods: Phenotypic and genotypic approaches were employed to identify strains HL1 and M1 at the subspecies level. For the further characterization of the probiotic properties of both strains, comparative genomic analyses were used. (3) Results: Both strains were identified as L. kefiranofaciens subsp. kefirgranum. According to the COG function category, dTDP-rhamnose and rhamnose-containing glycans were specifically detected in the L. kefiranofaciens subsp. Kefirgranum genomes. Three unique genes (epsI, epsJ, and epsK) encoding glycosyltransferase in the EPS gene cluster, and the ImpB/MucB/SamB family protein encoding gene were found in HL1 and M1. The specific ability to degrade arginine via the ADI pathway was found in HL1. The presence of the complete glycogen metabolism (glg) operon in the L. kefiranofaciens strains suggested the importance of glycogen synthesis to enable colonization in kefir grains and extend survival under environmental stresses. (4) Conclusions: The obtained novel information on the potential genes and pathways for polysaccharide synthesis and other functionalities in our HL1 and M1 strains could be applied for further functionality predictions for potential probiotic screening.

Probiotic Lactiplantibacillus plantarum Tana Isolated from an International Weightlifter Enhances Exercise Performance and Promotes Antifatigue Effects in Mice.

Exercise causes changes in the gut microbiota, and in turn, the composition of the gut microbiota affects exercise performance. In addition, the supplementation of probiotics is one of the most direct ways to change the gut microbiota. In recent years, the development and application of human-origin probiotics has gradually attracted attention. Therefore, we obtained intestinal Lactiplantibacillus plantarum "Tana" from a gold-medal-winning weightlifter, who has taken part in various international competitions such as the World Championships and the Olympic Games, to investigate the benefits of Tana supplementation for improving exercise performance and promoting antifatigue effects in mice. A total of 40 male Institute of Cancer Research (ICR) mice were divided into four groups (10 mice/group): (1) vehicle (0 CFU/mice/day), (2) Tana-1× (6.15 × 107 CFU/mice/day), (3) Tana-2× (1.23 × 108 CFU /mice/day), and (4) Tana-5× (3.09 × 108 CFU/mice/day). After four weeks of Tana supplementation, we found that the grip strength, endurance exercise performance, and glycogen storage in the liver and muscle were significantly improved compared to those in the vehicle group (p < 0.05). In addition, supplementation with Tana had significant effects on fatigue-related biochemical markers; lactate, ammonia, and blood urea nitrogen (BUN) levels and creatine kinase (CK) activity were significantly lowered (p < 0.05). We also found that the improved exercise performance and antifatigue benefits were significantly dose-dependent on increasing doses of Tana supplementation (p < 0.05), which increased the abundance and ratio of beneficial bacteria in the gut. Taken together, Tana supplementation for four weeks was effective in improving the gut microbiota, thereby enhancing exercise performance, and had antifatigue effects. Furthermore, supplementation did not cause any physiological or histopathological damage.