MDPI AG Acta Medica Okayama 1422-0067 26 12 2025 Specific Heat-Killed Lactic Acid Bacteria Enhance Mucosal Aminopeptidase N Activity in the Small Intestine of Aged Mice 5742 EN Takeshi Tsuruta Faculty of Environmental, Life, Natural Science and Technology, Okayama University Mami Wakisaka Faculty of Environmental, Life, Natural Science and Technology, Okayama University Takumi Watanabe Bio-Lab Co., Ltd. Aoi Nishijima Faculty of Environmental, Life, Natural Science and Technology, Okayama University Akihito Ikeda Faculty of Environmental, Life, Natural Science and Technology, Okayama University Mao Teraoka Faculty of Environmental, Life, Natural Science and Technology, Okayama University Tianyang Wang Faculty of Environmental, Life, Natural Science and Technology, Okayama University Kuiyi Chen Faculty of Environmental, Life, Natural Science and Technology, Okayama University Naoki Nishino Faculty of Environmental, Life, Natural Science and Technology, Okayama University Aminopeptidase N (APN), an enzyme expressed in the small intestinal mucosa, is involved in dietary protein digestion. Previous studies have shown that oral administration of fermented milk containing lactic acid bacteria (LAB) enhances mucosal APN activity in young mice. This study aimed to investigate whether LAB strains stimulate mucosal APN activity in aged mice and to evaluate its relevance to age-related changes in body composition. The underlying molecular mechanisms were also explored in vitro. Experiment 1: Aged C57BL/6J mice were fed diets supplemented with heat-killed LAB strains\Enterococcus faecalis OU-23 (EF), Leuconostoc mesenteroides OU-03 (LM), or Lactiplantibacillus plantarum SNK12 (LP). Compared to the aged Control group, the ileal APN activity was significantly higher in the LP group. LP administration also elevated serum Gla-osteocalcin levels and decreased serum CTX-1 levels. Experiment 2: IEC-6 cells were co-cultured with LP that had been treated with RNase, DNase, or lysozyme. APN activity was significantly lower in cells co-cultured with DNase- or lysozyme-treated LP compared to those co-cultured with untreated LP. A specific LAB strain may enhance mucosal APN activity in the aged intestine, potentially contributing to improved bone metabolism. This effect may be mediated by bacterial DNA and peptidoglycan. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2072-6643 16 17 2024 Cyclic Oligosaccharide-Induced Modulation of Immunoglobulin A Reactivity to Gut Bacteria Contributes to Alterations in the Bacterial Community Structure 2824 EN Taisei Miyamoto Graduate School of Environmental and Life Science, Okayama University Takeshi Tsuruta Graduate School of Environmental and Life Science, Okayama University Mao Teraoka Graduate School of Environmental and Life Science, Okayama University Tianyang Wang Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Graduate School of Environmental and Life Science, Okayama University Immunoglobulin A (IgA) is a major gut antibody that coats commensal gut bacteria and contributes to shaping a stable gut bacterial composition. Although previous studies have shown that cyclic oligosaccharides, including cyclic nigerosyl-1,6-nigerose (CNN) and cyclodextrins (CDs, including alpha CD, beta CD, and gamma CD), alter the gut bacterial composition, it remains unclear whether cyclic oligosaccharides modify the IgA coating of gut bacteria, which relates to cyclic oligosaccharide-induced alteration of the gut bacterial composition. To address this issue, mice were maintained for 12 weeks on diets containing CNN, alpha CD, beta CD, or gamma CD; the animals' feces were evaluated for their bacterial composition and the IgA coating index (ICI), a measure of the degree of IgA coating of bacteria. We observed that the intake of each cyclic oligosaccharide altered the gut bacterial composition, with changes in the ICI found at both the phylum and genus levels. The ICI for Bacillota, Lachnospiraceae NK4A136 group, UC Lachnospiraceae, and Tuzzerella were significantly and positively correlated with the relative abundance (RA) in total bacteria for these bacteria; in contrast, significant correlations were not seen for other phyla and genera. Our observations suggest that cyclic oligosaccharide-induced modulation of the IgA coating of gut bacteria may partly relate to changes in the community structure of the gut bacteria. No potential conflict of interest relevant to this article was reported.

Wiley Acta Medica Okayama 1613-4125 68 15 2024 Unabsorbed Fecal Fat Content Correlates with a Reduction of Immunoglobulin a Coating of Gut Bacteria in High]Lard Diet]Fed Mice 2400078 EN Emiko Katsumata Graduate School of Environmental and Life Science, Okayama University Takeshi Tsuruta Graduate School of Environmental and Life Science, Okayama University Kei Sonoyama Research Faculty of Agriculture, Hokkaido University Takashi Yoshida TAIYO YUSHI Corporation Mio Sasaki TAIYO YUSHI Corporation Mao Teraoka Graduate School of Environmental and Life Science, Okayama University Tianyang Wang Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Graduate School of Environmental and Life Science, Okayama University 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.<br> 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.<br> Conclusion: Unabsorbed fat content in the gut may result in attenuated IgA coating of bacteria in high-lard diet-fed mice.<br> No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2306-7381 10 3 2023 Monitoring the Milk Composition, Milk Microbiota, and Blood Metabolites of Jersey Cows throughout a Lactation Period 226 EN Peter Kiiru Gathinji Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Zabiallah Yousofi Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Karin Akada Animal Products Research Group, Institute of Livestock and Grassland Science, National Agriculture and Research Organization Ajmal Wali Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University This study aimed to determine how milk composition, milk microbiota, and blood metabolites may change during the lactation period in Jersey cows. Milk and jugular blood samples were collected from eight healthy cows every other month from the beginning to the end of their lactation period. Samples of airborne dust were also collected to determine whether the cowshed microbiota could affect milk microbiota. Milk yield peaked in the first two months and gradually decreased as the lactation period progressed. Milk fat, protein, and solids-not-fat contents were low in the first month, and then increased during the middle and late lactation periods. In the first month, plasma non-esterified fatty acids (NEFA), haptoglobin (Hp), and aspartate transaminase (AST) levels were elevated, and high abundances of Burkholderiaceae and Oxalobacteraceae were observed in milk and airborne dust microbiota. The finding that contamination of the environmental microbiota in milk was coupled with elevated plasma NEFA, Hp, and AST levels indicated that impaired metabolic function during the early lactation period may increase the invasion of opportunistic bacteria. This study can affirm the importance of feeding and cowshed management and should provide a helpful addition to improving Jersey cow farming. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2311-5637 8 1 2021 Bacterial and Fungal Microbiota of Guinea Grass Silage Shows Various Levels of Acetic Acid Fermentation 10 EN Jianjian Hou Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Graduate School of Environmental and Life Science, Okayama University This study aimed to gain insights into the bacterial and fungal microbiota associated with the acetic acid fermentation of tropical grass silage. Direct-cut (DC, 170 g dry matter [DM]/kg) and wilted (WT, 323 g DM/kg) guinea grass were stored in a laboratory silo at moderate (25 degrees C) and high (40 degrees C) temperatures. Bacterial and fungal microbiota were assessed at 3 days, 1 month, and 2 months after ensiling. Lactic acid was the primary fermentation product during the initial ensiling period, and a high Lactococcus abundance (19.7-39.7%) was found in DC silage. After two months, the lactic acid content was reduced to a negligible level, and large amounts of acetic acid, butyric acid, and ethanol were found in the DC silage stored at 25 degrees C. The lactic acid reduction and acetic acid increase were suppressed in the DC silage stored at 40 degrees C. Increased abundances of Lactobacillus, Clostridium, and Wallemia, as well as decreased abundances of Saitozyma, Papiliotrema, and Sporobolomyces were observed in DC silages from day three to the end of the 2 month period. Wilting suppressed acid production, and lactic and acetic acids were found at similar levels in WT silages, regardless of the temperature and storage period. The abundance of Lactobacillus (1.72-8.64%) was lower in WT than in DC silages. The unclassified Enterobacteriaceae were the most prevalent bacteria in DC (38.1-64.9%) and WT (50.9-76.3%) silages, and their abundance was negatively related to the acetic acid content. Network analysis indicated that Lactobacillus was involved in enhanced acetic acid fermentation in guinea grass silage. No potential conflict of interest relevant to this article was reported.

AIMS Press Acta Medica Okayama 2471-1888 7 1 2021 Cecum microbiota in rats fed soy, milk, meat, fish, and egg proteins with prebiotic oligosaccharides 1 12 EN Souliphone Sivixay Department of Animal Science, Graduate School of Life and Environmental Science, Okayama University Gaowa Bai Department of Animal Science, Graduate School of Life and Environmental Science, Okayama University Takeshi Tsuruta Department of Animal Science, Graduate School of Life and Environmental Science, Okayama University Naoki Nishino Department of Animal Science, Graduate School of Life and Environmental Science, Okayama University 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. No potential conflict of interest relevant to this article was reported.

Asian-Australasian Journal of Animal Sciences Acta Medica Okayama 1011-2367 33 11 2019 An investigation of seasonal variations in the microbiota of milk, feces, bedding, and airborne dust 1858 1865 EN Thuong Thi Nguyen Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Haoming Wu Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Objective</br> The microbiota of dairy cow milk varies with the season, and this accounts in part for the seasonal variation in mastitis-causing bacteria and milk spoilage. The microbiota of the cowshed may be the most important factor because the teats of a dairy cow contact bedding material when the cow is resting. The objectives of the present study were to determine whether the microbiota of the milk and the cowshed vary between seasons, and to elucidate the relationship between the microbiota.</br> Methods</br> We used 16S rRNA gene amplicon sequencing to investigate the microbiota of milk, feces, bedding, and airborne dust collected at a dairy farm during summer and winter.</br> Results</br> The seasonal differences in the milk yield and milk composition were marginal. The fecal microbiota was stable across the two seasons. Many bacterial taxa of the bedding and airborne dust microbiota exhibited distinctive seasonal variation. In the milk microbiota, the abundances of Staphylococcaceae, Bacillaceae, Streptococcaceae, Microbacteriaceae, and Micrococcaceae were affected by the seasons; however, only Micrococcaceae had the same seasonal variation pattern as the bedding and airborne dust microbiota. Nevertheless, canonical analysis of principle coordinates revealed a distinctive group comprising the milk, bedding, and airborne dust microbiota.</br> Conclusion</br> Although the milk microbiota is related to the bedding and airborne dust microbiota, the relationship may not account for the seasonal variation in the milk microbiota. Some major bacterial families stably found in the bedding and airborne dust microbiota, e.g., Staphylococcaceae, Moraxellaceae, Ruminococcaceae, and Bacteroidaceae, may have greater influences than those that varied between seasons. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2076-2607 8 9 2020 Bacterial and Fungal Microbiota Associated with the Ensiling of Wet Soybean Curd Residue under Prompt and Delayed Sealing Conditions 1334 EN Ajmal Wali Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Graduate School of Environmental and Life Science, Okayama University Wet soybean curd residue (SCR) obtained from two tofu factories (F1 and F2) was anaerobically stored with or without added beet pulp (BP). Sealing was performed on the day of tofu production (prompt sealing (PS)) or 2 days after SCR was piled and unprocessed (delayed sealing (DS)). Predominant lactic acid fermentation was observed regardless of the sealing time and BP addition.Acinetobacterspp. were the most abundant (>67%) bacteria in pre-ensiled SCR, regardless of the factory and sealing time. In PS silage, the abundances of typical lactic acid-producing bacteria, such asLactobacillus,Pediococcus, andStreptococcusspp. reached >50%. In DS silage,Acinetobacterspp. were the most abundant in F1 products, whereasBacillusspp. were the most abundant in long-stored F2 products. The fungal microbiota were highly diverse. AlthoughCandida,Aspergillus,Cladosporium,Hannaella, andWallemiaspp. were found to be the most abundant fungal microbiota, no specific genera were associated with factory, sealing time, or fermentation products. These results indicated that owing to preceding processing, including heating, distinctive microbiota may have participated in the ensiling of wet by-products. Lactic acid fermentation was observed even in DS silage, and an association ofBacillusspp. was suggested. No potential conflict of interest relevant to this article was reported.

BMFH Press Acta Medica Okayama 2186-3342 39 3 2020 Cyclic nigerosylnigerose ameliorates DSS-induced colitis with restoration of goblet cell number and increase in IgA reactivity against gut microbiota in mice 188 196 EN Takeshi Tsuruta Laboratory of Animal Nutrition, Graduate School of Environmental and Life Science, Okayama University Emiko Katsumata Laboratory of Animal Nutrition, Graduate School of Environmental and Life Science, Okayama University Akiko Mizote Hayashibara Co., Ltd. Hou Jian Jian Laboratory of Animal Nutrition, Graduate School of Environmental and Life Science, Okayama University Teresia Aluoch Muhomah Laboratory of Animal Nutrition, Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Laboratory of Animal Nutrition, Graduate School of Environmental and Life Science, Okayama University 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. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2076-2615 9 12 2019 The Relationship between Uterine, Fecal, Bedding, and Airborne Dust Microbiota from Dairy Cows and Their Environment: A Pilot Study 1007 EN Thuong T. Nguyen Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Ayumi Miyake Okayama Prefecture Livestock Research Institute Tu T. M. Tran Faculty of Agriculture and Food Technology, Tien Giang University Takeshi Tsuruta Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Naoki Nishino Department of Animal Science, Graduate School of Environmental and Life Science, Okayama University Simple Summary After calving, dairy cows face the risk of negative energy balance, inflammation, and immunosuppression, which may result in bacterial infection and disruption of the normal microbiota, thus encouraging the development of metritis and endometritis. This study characterized uterine, fecal, bedding, and airborne dust microbiota from postpartum dairy cows and their environment during summer and winter. The results clarify the importance of microbiota in cowshed environments, i.e., bedding and airborne dust, in understanding the postpartum uterine microbiota of dairy cows. <br/><br/> Abstract The aim of this study was to characterize uterine, fecal, bedding, and airborne dust microbiota from postpartum dairy cows and their environment. The cows were managed by the free-stall housing system, and samples for microbiota and serum metabolite assessment were collected during summer and winter when the cows were at one and two months postpartum. Uterine microbiota varied between seasons; the five most prevalent taxa were Enterobacteriaceae, Moraxellaceae, Ruminococcaceae, Staphylococcaceae, and Lactobacillaceae during summer, and Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Moraxellaceae, and Clostridiaceae during winter. Although Actinomycetaceae and Mycoplasmataceae were detected at high abundance in several uterine samples, the relationship between the uterine microbiota and serum metabolite concentrations was unclear. The fecal microbiota was stable regardless of the season, whereas bedding and airborne dust microbiota varied between summer and winter. With regards to uterine, bedding, and airborne dust microbiota, Enterobacteriaceae, Moraxellaceae, Staphylococcaceae, and Lactobacillaceae were more abundant during summer, and Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, and Clostridiaceae were more abundant during winter. Canonical analysis of principal coordinates confirmed the relationship between uterine and cowshed microbiota. These results indicated that the uterine microbiota may vary when the microbiota in cowshed environments changes. No potential conflict of interest relevant to this article was reported.

Acta Medica Okayama 00220302 91 6 2008 Ensiling of soybean curd residue and wet brewers grains with or without other feeds as a total mixed ration 2380 2387 EN F. Wang Naoki Nishino <p>Wet brewers grains and soybean curd residue were stored in laboratory-scale silos without (BG and SC silages, respectively) or with other ingredients as total mixed rations (BGT and SCT silages, respectively). Silages were opened after 14 and 56 d, and microbial counts, fermentation products, and aerobic stability were determined. Denaturing gradient gel electrophoresis was carried out to examine bacterial communities, and several bacteria that appeared to be involved in fermentation were identified. Lactic acid content was greater in SCT than in BGT silage, but lower in SC than in BG silage. Ethanol content was greater in BG than in SC regardless of silage type. Aerobic deterioration occurred promptly in ensiling materials (nonensiled by-products and total mixed ration mixtures) and in silages stored alone; however, SCT and BGT silages resisted deterioration and no heating was found for more than 5.5 d regardless of storage period. Silages were stable even with high yeast populations at silo opening, whereas prolonged ensiling decreased yeast counts in the 2 total mixed ration silages. The denaturing gradient gel electrophoresis profiles appeared similar between SCT and BGT silages but not between SC and BG silages. Weissella spp. and Lactobacillus brevis were common in aerobically stable SCT and BGT silages, and Lactobacillus buchneri was detected only in BGT silage. Both L. brevis and L. buchneri were found in silage but not in ensiling materials. Several other lactic acid bacteria were also identified in SCT and BGT silages, but did not appear to be related to fermentation and aerobic stability.</p> No potential conflict of interest relevant to this article was reported.