Consumption of indigestible saccharides and administration of Bifidobacterium pseudolongum reduce mucosal serotonin in murine colonic mucosa.

SCFA increase serotonin (5-hydroxytryptamine, 5-HT) synthesis and content in the colon in vitro and ex vivo, but little is known in vivo. We tested whether dietary indigestible saccharides, utilised as a substrate to produce SCFA by gut microbiota, would increase colonic 5-HT content in mice. Male C57BL/6J mice were fed a purified diet and water supplemented with 4 % (w/v) 1-kestose (KES) for 2 weeks. Colonic 5-HT content and enterochromaffin (EC) cell numbers were lower in mice supplemented with KES than those without supplementation, while monoamine oxidase A activity and mRNA levels of tryptophan hydroxylase 1 (Tph1), chromogranin A (Chga), Slc6a4 and monoamine oxidase A (Maoa) genes in the colonic mucosa, serum 5-HT concentration and total 5-HT content in the colonic contents did not differ between groups. Caecal acetate concentration and Bifidobacterium pseudolongum population were higher in KES-supplemented mice. Similar trends were observed in mice supplemented with other indigestible saccharides, that is, fructo-oligosaccharides, inulin and raffinose. Intragastric administration of live B. pseudolongum (108 colony-forming units/d) for 2 weeks reduced colonic 5-HT content and EC cell numbers. These results suggest that changes in synthesis, reuptake, catabolism and overflow of 5-HT in the colonic mucosa are not involved in the reduction of colonic 5-HT content by dietary indigestible saccharides in mice. We propose that gut microbes including B. pseudolongum could contribute to the reduction of 5-HT content in the colonic mucosa via diminishing EC cells.

Bacterial and fungal microbiota of total mixed ration silage stored at various temperatures.

AIMS: To obtain insights into how bacterial and fungal microbiota and fermentation products composition are affected by storage temperature for TMR silage, which can be manufactured year-round. METHODS AND RESULTS: TMR silage was stored at 10°C, 25°C, ambient temperature (AT; 20-35°C) and 40°C. Lactic acid production was delayed when stored at 10°C, and acid production stagnated after 2 weeks when stored at 40°C. The patterns of acetic acid and ethanol production were inversely related, with ethanol production promoted at 10°C and 25°C and acetic acid production promoted at AT and 40°C. The bacterial diversity was reduced in TMR silage with high lactic acid and acetic acid content, and the fungal diversity was reduced in TMR silage with high ethanol content. CONCLUSIONS: The intensity of lactic acid production was accounted for by the high abundance of Lactobacillus, and its stagnated production at a substantially high storage temperature was related to an increased abundance of Bacillus. The enhanced production of acetic acid or ethanol can be explained by differences in the fungal microbiota. SIGNIFICANCE AND IMPACT OF THE STUDY: The integrated analysis of bacterial and fungal microbiota can provide in-depth insights into the impact of storage temperature on TMR silage fermentation.

Expression of serotonin receptor HTR4 in glucagon-like peptide-1-positive enteroendocrine cells of the murine intestine.

Serotonin (5-hydroxytryptamine [5-HT]) synthesized and released in enterochromaffin (EC) cells participates in various functions in the gastrointestinal tract by acting on a diverse range of 5-HT receptors (HTRs) expressed on smooth muscle, enteric neurons, and epithelial cells. We previously observed that genes encoding HTR2A, HTR2B, and HTR4 are expressed in murine intestinal organoids, suggesting the expression of these HTRs in intestinal epithelial cells. The present study investigated the localization of these HTRs in the murine intestine by immunofluorescence staining. HTR2A, HTR2B, and HTR4 localized in individual solitary cells in the epithelium, while HTR2C was observed in the lamina propria. In the epithelium, HTR2A, HTR2B, and HTR4 colocalized with 5-HT, and HTR4 colocalized with glucagon-like peptide 1 (GLP-1) and peptide YY (PYY). Murine intestinal organoids show a colocalization pattern that is similar to in vivo HTR2A and HTR4 with 5-HT, GLP-1, and PYY. Intraperitoneal and intragastric administration of tegaserod, an HTR4 agonist, failed to alter plasma GLP-1 levels in fasted mice. However, intragastric but not intraperitoneal administration of tegaserod reduced dietary lipid-induced increases of plasma GLP-1 levels. This action of tegaserod was inhibited by co-administration of RS39604, an HTR4 antagonist. These results suggest that murine ileal GLP-1/PYY-producing enteroendocrine (EE) cells express HTR4, while 5-HT-producing EC cells express HTR2A, HTR2B, and HTR4. In addition, the observations regarding in vivo GLP-1 secretion suggest that HTR4 signaling in ileal EE cells suppresses dietary lipid-induced GLP-1 secretion. We thus propose that EC and EE cells may interact with each other through paracrine signaling mechanisms.

Dietary soy, meat, and fish proteins modulate the effects of prebiotic raffinose on composition and fermentation of gut microbiota in rats.

Soy, meat (mixture of pork and beef), and fish proteins were fed to rats with and without prebiotic raffinose (RAF), and the composition and fermentation of gut microbiota were examined. Bifidobacterium spp. populations were higher, and propionic acid concentration was lower in soy protein-fed than meat protein-fed rats. Likewise, Enterobacteriaceae populations were higher in fish protein-fed rats than other rats. RAF feeding increased Bifidobacterium spp. and decreased Faecalibacterium prausnitzii populations regardless of the dietary protein source. Interactions between dietary proteins and RAF were shown for Lactobacillus spp. and Clostridium perfringens group; the increase of Lactobacillus spp. populations by RAF was seen only for soy protein-fed rats, whereas the reduction of C. perfringens group by RAF was evident in fish and meat protein-fed rats. It is concluded that dietary proteins may differentially modulate the effects of prebiotic oligosaccharides on gut fermentation and microbiota, with differences observed between plant and animal proteins.

Exosomes isolated from sera of mice fed Lactobacillus strains affect inflammatory cytokine production in macrophages in vitro.

Orally administered Lactobacillus strains, including L. plantarum No.14 and L. rhamnosus GG, reportedly reduce inflammatory cytokine production in mice. The present study tested our idea that circulating exosomes mediate the action of Lactobacillus strains. The lipopolysaccharide-induced production of TNF-α and IL-6 in vitro was attenuated in peritoneal exudate cells (PECs) isolated from C57BL/6N mice that had been fed L. plantarum No.14. When PECs were cultured for 24 h with exosomes isolated from the serum of mice fed L. plantarum No.14 or L. rhamnosus GG, accumulation of both TNF-α and of the corresponding mRNA was lowered. Growth in the presence of these exosomes also decreased the production of TNF-α and IL-6 by the murine macrophage cell line RAW264.7. In contrast, supplementation with exosome-depleted serum of mice fed L. plantarum No.14 or L. rhamnosus GG failed to affect the production of TNF-α and IL-6 by RAW264.7 cells. When PECs and RAW264.7 cells were cultured for 24 h with PKH67-labeled exosomes isolated from murine serum, fluorescent signal was observed inside the cells, suggesting that these cells incorporate serum exosomes. We propose that the anti-inflammatory activity of orally administered L. plantarum No.14 and L. rhamnosus GG is mediated, at least in part, by circulating exosomes.

Variability, stability, and resilience of fecal microbiota in dairy cows fed whole crop corn silage.

The microbiota of whole crop corn silage and feces of silage-fed dairy cows were examined. A total of 18 dairy cow feces were collected from six farms in Japan and China, and high-throughput Illumina sequencing of the V4 hypervariable region of 16S rRNA genes was performed. Lactobacillaceae were dominant in all silages, followed by Acetobacteraceae, Bacillaceae, and Enterobacteriaceae. In feces, the predominant families were Ruminococcaceae, Bacteroidaceae, Clostridiaceae, Lachnospiraceae, Rikenellaceae, and Paraprevotellaceae. Therefore, Lactobacillaceae of corn silage appeared to be eliminated in the gastrointestinal tract. Although fecal microbiota composition was similar in most samples, relative abundances of several families, such as Ruminococcaceae, Christensenellaceae, Turicibacteraceae, and Succinivibrionaceae, varied between farms and countries. In addition to the geographical location, differences in feeding management between total mixed ration feeding and separate feeding appeared to be involved in the variations. Moreover, a cow-to-cow variation for concentrate-associated families was demonstrated at the same farm; two cows showed high abundance of Succinivibrionaceae and Prevotellaceae, whereas another had a high abundance of Porphyromonadaceae. There was a negative correlation between forage-associated Ruminococcaceae and concentrate-associated Succinivibrionaceae and Prevotellaceae in 18 feces samples. Succinivibrionaceae, Prevotellaceae, p-2534-18B5, and Spirochaetaceae were regarded as highly variable taxa in this study. These findings help to improve our understanding of variation and similarity of the fecal microbiota of dairy cows with regard to individuals, farms, and countries. Microbiota of naturally fermented corn silage had no influence on the fecal microbiota of dairy cows.

Comparative microbiota assessment of wilted Italian ryegrass, whole crop corn, and wilted alfalfa silage using denaturing gradient gel electrophoresis and next-generation sequencing.

The microbiota of pre-ensiled crop and silage were examined using denaturing gradient gel electrophoresis (DGGE) and next-generation sequencing (NGS). Wilted Italian ryegrass (IR), whole crop corn (WC), and wilted alfalfa (AL) silages stored for 2 months were examined. All silages contained lactic acid as a predominant fermentation product. Across the three crop species, DGGE detected 36 and 28 bands, and NGS identified 253 and 259 genera in the pre-ensiled crops and silages, respectively. The NGS demonstrated that, although lactic acid bacteria (LAB) became prevalent in all silages after 2 months of storage, the major groups were different between crops: Leuconostoc spp. and Pediococcus spp. for IR silage, Lactobacillus spp. for WC silage, and Enterococcus spp. for AL silage. The predominant silage LAB genera were also detected by DGGE, but the presence of diverse non-LAB species in pre-ensiled crops was far better detected by NGS. Likewise, good survival of Agrobacterium spp., Methylobacterium spp., and Sphingomonas spp. in IR and AL silages was demonstrated by NGS. The diversity of the microbiota described by principal coordinate analysis was similar between DGGE and NGS. Our finding that analysis of pre-ensiled crop microbiota did not help predict silage microbiota was true for both DGGE and NGS.

Organoids as an ex vivo model for studying the serotonin system in the murine small intestine and colon epithelium.

Intestinal organoids were recently established as an ex vivo model of the intestinal epithelium. The present study investigated the serotonin (5-hydroxytryptamine, 5-HT) system using organoids. Organoids from murine small intestinal and colonic crypts were successfully cultured. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that small intestinal and colonic organoids express mRNAs encoding tryptophan hydroxylase-1 (TPH1) (the rate-limiting enzyme of 5-HT synthesis), serotonin reuptake transporter (SERT), 5-HT receptor (HTR)2A, HTR2B, and HTR4. SERT mRNA levels were significantly higher in the small intestine than in the colon in both the mucosal tissues and organoids, as estimated by quantitative real-time RT-PCR. Although the 5-HT concentration and levels of chromogranin A (CgA) (an enteroendocrine cell marker), TPH1, and HTR4 mRNAs were significantly higher in the colonic mucosa than the small intestinal mucosa, they were the same in small intestinal and colonic organoids. There were no significant differences in HTR2A and HTR2B mRNA levels between the small intestine and colon in either the mucosal tissues or organoids. Immunofluorescence staining showed that the number of CgA-positive cells in the colonic organoids appeared to increase upon culturing with acetate. Acetate supplementation significantly increased CgA, TPH1, and HTR4 mRNA levels in the colonic organoids. We propose that organoids are useful for investigating the 5-HT system in the intestinal epithelium, even though colonic organoids may require gut microbiota-derived factors such as short-chain fatty acids.

Intestinal epithelial cells promote secretion of leptin and adiponectin in adipocytes.

Although leptin and adiponectin are the predominant adipokines, how their circulating levels are regulated is incompletely understood. The present study tested whether intestinal epithelial cells influence the expression and secretion of these adipokines by adipocytes. Leptin gene expression and secretion by cultured human primary adipocytes and Simpson-Golabi-Behmel Syndrome adipocytes increased upon coculture with human enterocytic Caco-2 cells or incubation in conditioned medium of Caco-2 cells. Although adiponectin secretion increased, its mRNA levels decreased. Tissue homogenate of the ileum (but not the jejunum, colon, or liver) of nonobese C57BL/6J mice also stimulated leptin and adiponectin secretion by cultured murine 3T3-L1 adipocytes. However, ileal homogenate of obese KK-Ay mice had no effect on leptin and adiponectin secretion. We propose that as yet unidentified humoral factors released from intestinal epithelial cells are involved in regulating circulating leptin and adiponectin levels. Decreased production of such factors may contribute to hyperphagia in KK-Ay mice.

Dual labeling with 5-bromo-2'-deoxyuridine and 5-ethynyl-2'-deoxyuridine for estimation of cell migration rate in the small intestinal epithelium.

Small intestinal epithelium is a self-renewing system in which the entire sequence of cell proliferation, differentiation, and removal is coupled to cell migration along the crypt-villus axis. We examined whether dual labeling with different thymidine analogues, 5-bromo-2'-deoxyuridine (BrdU) and 5-ethynyl-2'-deoxyuridine (EdU), can be used to estimate cell migration rates on the villi of small intestines in rats. Rats received a single intraperitoneal injection of BrdU and EdU within a time interval, and signals in tissue sections were examined by immunohistochemistry and the "click" reaction, respectively. We successfully observed BrdU- and EdU-positive cells on the epithelium with no cross-reaction. In addition, we observed an almost complete overlapping of BrdU- and EdU-positive cells in rats administered simultaneously with BrdU and EdU. By calculating the cell migration rate by dividing the distance between the median cell positions of the distribution of BrdU- and EdU-positive cells by the time between the injection of BrdU and EdU, we estimated approximately 9 and 5 µm/h for the cell migration rates on the villi in the jejunum and ileum, respectively. We propose that dual labeling with BrdU and EdU within a time interval, followed by detecting with immunohistochemistry and the click reaction, respectively, is useful to estimate accurately the cell migration rate in the intestinal epithelium in a single animal.

Adiponectin is partially associated with exosomes in mouse serum.

Exosomes are membrane vesicles 30-120 nm in diameter that are released by many cell types and carry a cargo of proteins, lipids, mRNA, and microRNA. Cultured adipocytes reportedly release exosomes that may play a role in cell-to-cell communication during the development of metabolic diseases. However, the characteristics and function of exosomes released from adipocytes in vivo remain to be elucidated. Clearly, adipocyte-derived exosomes could exist in the circulation and may be associated with adipocyte-specific proteins such as adipocytokines. We isolated exosomes from serum of mice by differential centrifugation and analyzed adiponectin, leptin, and resistin in the exosome fraction. Western blotting detected adiponectin but no leptin and only trace amounts of resistin in the exosome fraction. The adiponectin signal in the exosome fraction was decreased by proteinase K treatment and completely quenched by a combination of proteinase K and Triton X-100. Quantitative ELISA showed that the exosome fraction contains considerable amounts of adiponectin, but not leptin or resistin. The concentration of adiponectin in the serum and the ratio of adiponectin to total protein in the exosome fraction were lower in obese mice than in lean mice. These results suggest that a portion of adiponectin exists as a transmembrane protein in the exosomes in mouse serum. We propose adiponectin as a marker of exosomes released from adipocytes in vivo.

Unabsorbed Fecal Fat Content Correlates with a Reduction of Immunoglobulin a Coating of Gut Bacteria in High-Lard Diet-Fed Mice.

SCOPE: Immunoglobulin A (IgA) selectively coats gut bacteria and contributes to regulatory functions in gastrointestinal inflammation and glucose metabolism. Excess intake of lard leads to decrease in the IgA coating of gut bacteria, although the underlying mechanisms remain unknown. This study validates how unabsorbed fat derived from a high-lard diet in the gut affects the IgA coating of bacteria, as assessed in mouse models using three types of dietary fat (lard, medium-, and long-chain triglycerides [MLCTs], and medium-chain triglycerides [MCTs]) exhibiting different digestibilities. METHODS AND RESULTS: C57BL/6J mice are maintained on diets containing lard, MLCTs, or MCTs at 7% or 30% w/w for 10 weeks (n = 6 per group). The fecal fatty acid concentration is measured to quantify unabsorbed fat content. The ratio of IgA-coated bacteria to total bacteria (IgA coating ratio) in the feces is measured by flow cytometry. Compared to lard-fed mice, MLCT- and MCT-fed mice exhibit lower fecal concentrations of palmitic acid, stearic acid, and oleic acid and higher IgA coating ratios at both 7% and 30% dietary fat, and these parameters exhibit significant negative correlations. CONCLUSION: Unabsorbed fat content in the gut may result in attenuated IgA coating of bacteria in high-lard diet-fed mice.

Modulatory Effects of A1 Milk, A2 Milk, Soy, and Egg Proteins on Gut Microbiota and Fermentation.

Milk can be divided into A1 and A2 types according to ß-casein variants, and there is a debate about whether A1 milk consumption exacerbates gut environments. This study examined the cecum microbiota and fermentation in mice fed A1 casein, A2 casein, mixed casein (commercial casein), soy protein isolate, and egg white. The cecum acetic acid concentration was higher, and the relative abundances of Muribaculaceae and Desulfovibrionaceae were greater in mice fed A1 versus A2 casein. The other parameters of cecum fermentation and microbiota composition were similar among the mice fed A1, A2, and mixed caseins. The differences were more distinctive among the three caseins, soy, and egg feedings. Chao 1 and Shannon indices of the cecum microbiota were lowered in egg white-fed mice, and the microbiota of mice fed milk, soy, and egg proteins were separately grouped by principal coordinate analysis. Mice fed the three caseins were characterized by a high abundance of Lactobacillaceae and Clostridiaceae, those fed soy were characterized by Corynebacteriaceae, Muribaculaceae, and Ruminococcaceae, and those fed egg white were characterized by Eggerthellaceae, Rikenellaceae, and Erysipelatoclostridiaceae. Thus, although several differences can arise between A1 and A2 caseins in terms of their modulatory effects on gut environments, the differences between milk, soy, and egg proteins can be more distinctive and are worth further consideration.

Cyclic Nigerosylnigerose Attenuates High-Fat Diet-Induced Fat Deposition, Colonic Inflammation, and Abnormal Glucose Metabolism and Modifies Gut Immunoglobulin a Reactivity to Commensal Bacteria.

SCOPE: High-fat diet (HFD) intake induces gut dysbiosis, inflammation in the peripheral tissues, and a reduction in immunoglobulin A (IgA) coating of gut bacteria, which is related to HFD-induced insulin resistance (IR). This study evaluates the effect of cyclic nigerosylnigerose (CNN), a dietary fiber that prevents gut inflammation and promotes IgA coating of gut bacteria, on the above-mentioned HFD-induced disorders. METHODS AND RESULTS: Balb/c mice are fed an HFD and administered CNN for 20 weeks. CNN administration reduces mesenteric adipose tissue weight, colonic tumor necrosis factor α (TNFα) mRNA expression, and serum endotoxin levels and ameliorates HFD-induced abnormal glucose metabolism. Additionally, CNN administration promotes gut bacteria-specific IgA secretion and alters IgA reactivity to gut bacteria. The alterations of IgA reactivity to specific bacteria such as Erysipelatoclostridium, Escherichia, Faecalibaculum, Lachnospiraceae genera, and Stenotrophomonas are correlated with mesenteric adipose tissue weight, colonic TNFα mRNA expression, serum endotoxin levels, and a homeostasis model assessment for IR. CONCLUSION: CNN-induced alterations in IgA reactivity to gut bacteria may be related to the suppression of HFD-induced fat deposition, colonic inflammation, endotoxemia, and IR. These observations indicate that dietary fiber that modulates IgA reactivity to gut bacteria may be useful in preventing HFD-induced disorders.

Local free fatty acids trigger the expression of lipopolysaccharide-binding protein in murine white adipose tissue.

Although lipopolysaccharide (LPS)-binding protein (LBP) is an acute-phase protein mainly produced by hepatocytes, it has also been proposed to be a pro-inflammatory adipokine. Obesity and the consumption of a high-fat diet (HFD) are reportedly associated with elevated levels of LPS in plasma and free fatty acids (FFAs) in white adipose tissue (WAT). We examined whether circulating LPS or local FFAs are responsible for the HFD-induced increase of LBP in WAT. Male C57BL/6J mice were fed either a normal-fat diet (NFD) or an HFD. The mRNA levels in the liver and mesenteric WAT (mWAT), total FFA content in mWAT, and LBP and LPS concentrations in plasma were determined. The Lbp mRNA level in mWAT was higher in mice fed the HFD than in those fed the NFD for 3, 7, or 28 days or 14 weeks, whereas the hepatic Lbp mRNA level did not differ between the groups. The Lbp mRNA level in mWAT was also increased by the HFD in germ-free mice, which do not have gut microbiota, the source of LPS. The plasma LPS level did not show a significant correlation with the mWAT Lbp mRNA level. The total FFA content in mWAT was higher in mice fed the HFD than in those fed the NFD and positively correlated with the Lbp mRNA level. Supplementation with palmitic acid increased the Lbp mRNA level in 3T3-L1 adipocytes. We propose that local FFAs, but not circulating LPS, are the trigger for increased Lbp expression in mWAT of mice fed the HFD.

Cecum microbiota in rats fed soy, milk, meat, fish, and egg proteins with prebiotic oligosaccharides.

Diet is considered the most influential factor in modulating the gut microbiota but how dietary protein sources differ in their modulatory effects is not well understood. In this study, soy, meat (mixture of beef and pork), and fish proteins (experiment 1) and soy, milk (casein), and egg proteins (experiment 2) were fed to rats with cellulose (CEL) and raffinose (RAF); the microbiota composition and short-chain fatty acid concentration in the cecum were determined. Egg protein feeding decreased the concentration of acetic acid and the richness and diversity of the cecum microbiota. RAF feeding increased the concentrations of acetic and propionic acids and decreased the richness and diversity of the cecum microbiota. When fed with CEL, the abundance of Ruminococcaceae and Christensenellaceae, Akkermansiaceae and Tannerellaceae, and Erysipelotrichaceae enhanced with soy protein, meat and fish proteins, and egg protein, respectively. The effects of dietary proteins diminished with RAF feeding and the abundance of Bifidobacteriaceae, Erysipelotrichaceae, and Lachnospiraceae increased and that of Ruminococcaceae and Christensenellaceae decreased regardless of the protein source. These results indicate that, although the effect of prebiotics is more robust and distinctive, dietary protein sources may influence the composition and metabolic activities of the gut microbiota. The stimulatory effects of soy, meat, and egg proteins on Christensenellaceae, Akkermansiaceae, and Erysipelotrichaceae deserve further examination to better elucidate the dietary manipulation of the gut microbiota.

Aicda deficiency exacerbates high-fat diet-induced hyperinsulinemia but not gut dysbiosis in mice.

Immunoglobulin A (IgA) is a major antibody in the gut. We previously observed that a high-fat diet (HFD) reduces IgA reactivity to gut microbiota, but the physiological implications have yet to be elucidated. We hypothesized that a reduction of IgA reactivity to gut microbiota induced by a HFD may contribute to development of gut dysbiosis and inflammation that accompanies HFD feeding. To test our hypothesis, we used Aicda deficient mice, which have a deficiency in IgA production. Aicda deficient mice and wild-type mice were fed normal-fat diet or HFD for 12 weeks. We found that HFD feeding but not Aicda deficiency altered the fecal microbiota composition. Meanwhile, Aicda deficiency significantly increased gene expression of inflammatory cytokines in the ileum, but not in the colon despite no significant difference between diets. These results suggest that a reduction of IgA reactivity to gut microbiota induced by HFD partly contributes to development of inflammation in the ileum, but not to gut dysbiosis. We also found that the fasting blood insulin level was significantly increased by Aicda deficiency only under HFD feeding. Furthermore, the gene expression of monocyte chemoattractant protein1, a major chemokine responsible for the onset of hyperinsulinemia, in the liver was significantly increased by HFD feeding and tended to be increased by Aicda deficiency. These findings suggest that a reduction of IgA reactivity to gut microbiota induced by HFD contributes to hyperinsulinemia partly via increasing monocyte chemoattractant protein-1 expression in the liver.

Development of a method for the identification of S-IgA-coated bacterial composition in mouse and human feces.

Some commensal intestinal bacteria in humans and mice are coated with secretory immunoglobulin A (S-IgA). It has been suggested that the S-IgA coating of commensal bacteria does not occur at random and thus identification of S-IgA-coated bacterial genera/species should help in elucidating the interaction between S-IgA and commensal intestinal bacteria, but no method of identifying the genera/species of S-IgA-coated bacteria has been established. To identify S-IgA-coated bacterial composition, we developed a method combining immunohistochemical detection of S-IgA and subsequent 16S rRNA targeted fluorescence in situ hybridization (FISH) analysis. Human and mouse fecal S-IgA coated bacterial composition was evaluated by this newly developed method with 10 frequently-used FISH probes. Fecal S-IgA-coated bacterial composition was successfully analyzed by this method, and this suggests that Enterobacteriaceae is preferably coated with S-IgA, whereas Bacteroides/Prevotella and Lactobacillus/Enterococcus groups appear to be poorly coated with S-IgA.

Examination of milk microbiota, fecal microbiota, and blood metabolites of Jersey cows in cool and hot seasons.

Microbiota of individual cow milk, bulk tank milk, and feces of Jersey cows were examined. Samples were collected from two farms (F1 and F2) in cool (November, Nov) and hot (July, Jul) seasons. Milk yield and milk composition were similar between the two farms and between the two seasons. Prevalent taxa of the fecal microbiota, i.e. Ruminococcaceae, Bacteroidaceae, Lachnospiraceae, Rikenellaceae, and Clostridiaceae, were unaffected by the farm and season. Relative abundance of milk microbiota for Pseudomonadaceae, Enterobacteriaceae, and Streptococcaceae (F1 > F2) and Lactobacillaceae, Bifidobacteriaceae, and Cellulomonadaceae (F1 < F2) were different between the two farms, and those for Staphylococcaceae, Bacillaceae, Ruminococcaceae, and Veillonellaceae (Nov < Jul) and Methylobacteriaceae and Moraxellaceae (Nov > Jul) were different between the two seasons. The microbiota of bulk tank milk was numerically different from that of individual cow milk. Principal coordinate analysis indicated that the milk microbiota was unrelated to the fecal microbiota. The finding that relative abundance of Pseudomonadaceae and Moraxellaceae appeared greater than those reported for Holstein milk suggested that higher protein and fat content may result in a greater abundance of proteolytic and lipolytic taxa in Jersey cow milk.

Cyclic nigerosylnigerose ameliorates DSS-induced colitis with restoration of goblet cell number and increase in IgA reactivity against gut microbiota in mice.

Cyclic nigerosylnigerose (CNN) is a cyclic oligosaccharide. Oral administration of CNN promotes immunoglobulin A (IgA) secretion in the gut. IgA is a major antibody secreted into the gut and plays a crucial role in suppressing gut inflammation due to commensal gut microbiota. To investigate the effect of administration of CNN to promote IgA secretion on gut inflammation, experimental colitis was induced with dextran sulfate sodium (DSS) in Balb/c mice after 6 weeks of CNN pre-feeding. The severity of colitis was evaluated based on a disease activity index (DAI), the gene expression of inflammatory cytokines, and a histological examination. The CNN-treated mice with DSS-induced colitis (CNN-DSS group) showed significantly lower DAI scores and mRNA levels of interleukin-1 compared with the CNN-untreated mice with DSS-induced colitis (DSS group). Histological examination of the colon revealed that the pathological score was significantly lower in the CNN-DSS group compared with the DSS group due to the reduced infiltration of immune cells. The number of goblet cells was significantly higher in the CNN-DSS group compared with the DSS group. The IgA concentration and the ratio of microbiota coated with IgA were evaluated in the cecal content. Although there was no difference in the IgA concentration among groups, a higher proportion of cecal microbiota were coated with IgA in the CNN-DSS group compared with that in the DSS group. These results suggest that CNN might preserve goblet cells in the colon and promote IgA coating of gut microbiota, which synergistically ameliorate gut inflammation in mice with DSS-induced colitis.