Bacillus subtilis is a generally recognized as safe probiotic, which is used as a starter for natto fermentation. Natto is a functional food with antithrombus function due to nattokinase. Compared with natto, fermented milk is a more popular fermented food, which is commonly fermented by Lactobacillus bulgaricus and Streptococcus. However, there is no report on B. subtilis-fermented milk. In this study, to produce a functional fermented milk with antithrombus function, a B. subtilis strain (B. subtilis JNFE0126) that produced both nattokinase and milk-clotting enzyme was isolated from traditionally fermented natto and used as the starter for the functional fermented milk. In liquid fermentation culture, the peak values of thrombolytic activity and milk-clotting activity were 3,511 U/mL at 96 h and 874.5 Soxhlet unit/mL at 60 h, respectively. The optimal pH and temperature were pH 7.0 at 40°C for nattokinase and pH 6.5 and 55°C for milk-clotting enzyme, respectively. The thrombolytic activity in the fermented milk reached 215.1 U/mL after 8 h of fermentation. Sensory evaluation showed that the acceptance of the milk fermented by B. subtilis JNFE0126 was similar to the traditional milk fermented by L. bulgaricus and S. thermophilus. More importantly, oral intake of the fermented milk by the thrombosis-model mice prevented the development of thrombosis. Our results suggest that B. subtilis JNFE0126-fermented milk has potential as a novel, functional food in the prevention of thrombosis-related cardiovascular diseases.In our previous studies, we revealed the effect of lactose inclusion in calf starters on the growth performance and gut development of calves. We conducted the present study as a follow-up study to identify the shift in rumen microbiota and its relation to rumen fermentation when calves are fed a lactose-containing starter. Thirty Holstein bull calves were divided into 2 calf starter treatment groups texturized calf starter (i.e., control; n = 15) or calf starter in which starch was replaced with lactose at 10% (i.e., LAC10; n = 15) on a dry matter basis. All calves were fed their respective treatment calf starter ad libitum from d 7, and kleingrass hay from d 35. Rumen digesta were collected on d 80 (i.e., 3 wk after weaning) and used to analyze rumen microbiota and fermentation products. There was no apparent effect of lactose feeding on the α-diversity and overall composition of rumen microbiota. Amplicon sequencing and real-time PCR quantification of the 16S rRNA gene confirmed that the abundance of butyr in the rumen of lactose-fed calves partially explains the increase in the proportion of rumen acetate that was observed in our previous study.Extracellular vesicles (EV) in milk, particularly exosomes, have attracted considerable attention as bioactive food compounds and for their use in drug delivery. The utility of small EV in milk (sMEV) as an animal feed additive and in drug delivery would be enhanced by cost-effective large-scale protocols for the enrichment of sMEV from byproducts in dairy plants. Here, we tested the hypothesis that sMEV may be enriched from byproducts of cheesemaking by tangential flow filtration (EV-FF) and that the sMEV have properties similar to sMEV prepared by ultracentrifugation (sMEV-UC). Three fractions of EV were purified from the whey fraction of cottage cheese making by using EV-FF that passed through a membrane with a 50-kDa cutoff (50 penetrate; 50P), and subfractions of 50P that were retained (100 retentate; 100R) or passed through (100 penetrate; 100P) a membrane with a 100-kDa cutoff; sMEV-UC controls were prepared by serial ultracentrifugation. The abundance of sMEV ( sMEV-UC. More than 100 mature microRNA were detected in sMEV-UC by using sequencing analysis, compared with 36 to 60 microRNA in EV-FF. Only 100R and sMEV-UC yielded mRNA in quantities and qualities sufficient for sequencing analysis; an average of 276,000 and 838,000 reads were mapped to approximately 14,600 and 18,500 genes in 100R and sMEV-UC, respectively.  https://www.selleckchem.com/products/FK-506-(Tacrolimus).html  In principal component analysis, microRNA, mRNA, and protein in EV-FF preparations clustered separately from control sMEV-UC. We conclude that under the conditions used here, flow filtration yields a heterogeneous population of milk EV.The objective of the present study was to elucidate the effect of feeding either colostrum or milk-based formula on the mRNA abundance of genes related to pathogen recognition [toll-like receptors (TLR1-10)], antimicrobial defense [β-defensin 1 (DEFB1) and peptidoglycan recognition protein 1 (PGLYRP1)], and tight junctions (claudin 1 = CLDN1, claudin 4 = CLDN4, and occludin = OCLN) in different sections of the small intestine of neonatal calves at d 4 of life. Holstein dairy calves were fed either colostrum (COL; n = 7) or milk-based formula (FOR; n = 7) with comparable nutrient composition but lower contents of several bioactives in the formula than in the respective colostrum group until d 4 of life. Following euthanasia on d 4 (2 h after feeding), tissue samples from the duodenum, jejunum (proximal, middle, and distal), and ileum were collected. The mRNA abundance of the target genes was quantified by quantitative PCR. The mRNA abundance of TLR1, TLR6, TLR9, and TLR10 were greater in COL than in FOR calvesf CLDN1 in the middle and proximal jejunum compared with the other gut regions. Overall, the greater mRNA abundance of 5 different TLR, and CLDN1 in most intestinal sections of the COL calves may suggest that feeding colostrum improves immune responsiveness and epithelial barrier function in neonatal calves.Casein (CN) micelles will coagulate in the stomach after ingestion, which is similar to the cheesemaking process. Although genetic variants of bovine proteins, especially κ-CN, have been confirmed to influence the coagulation properties of the CN micelle, its influence on milk digestibility has not been revealed yet. This study aimed to investigate how genetic variants, glycosylation degree of κ-CN, and CN micelle size influence digestion rates during in vitro gastrointestinal digestion. Three milk pools, representing κ-CN phenotypes of either AA, BB, or AB composition were prepared from milk of individual Danish Holstein cows representing these different genotypes. In vitro digestion of the 3 milk pools, AA, BB, or AB, was investigated by sodium dodecyl sulfate-PAGE, liquid chromatography-mass spectrometry, and degree of hydrolysis. The results showed that κ-CN AA milk had faster digestion rate in the gastric phase compared with BB and AB milks, whereas only small differences were apparent in the intestinal digestion phase.